Diagnostic Specificity of Serum Carcinoembryonic Antigen for Canine Mammary Neoplasms: The Impact of Non-Neoplastic Diseases

Diagnostic Specificity of Serum Carcinoembryonic Antigen for Canine Mammary Neoplasms: The Impact of Non-Neoplastic Diseases | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Short Report Diagnostic Specificity of Serum Carcinoembryonic Antigen for Canine Mammary Neoplasms: The Impact of Non-Neoplastic Diseases Leonardo Willian Amite Lababrín, Lara Coslop Comércio, Ana Carolina Teixeira Brandão Fagundes, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7481550/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Carcinoembryonic antigen (CEA) is a glycoprotein produced by normal mucosal cells of the gastrointestinal tract and has been utilized as a tumor biomarker in humans. In veterinary medicine, canine patients with mammary neoplasms present elevated CEA levels. However, CEA is also known to increase in other pathological conditions. This study compared serum CEA levels in dogs with mammary carcinoma and those with non-neoplastic diseases. Blood serum samples were collected from 118 dogs (101 females and 17 males) and were divided into five experimental groups: control (G1), mammary neoplasia (G2), ehrlichiosis (G3), gastroenteritis (G4), and non-neoplastic diseases (G5). CEA levels were measured using a human ELISA kit, and the results were analyzed and compared across the experimental groups. Statistical analyses included Tukey's test, ROC curve analysis, and Pearson’s correlation, with a significance level of 5%. The results demonstrated significant differences in mean CEA levels between groups G1 and G2 (P < 0.0001), G1 and G4 (P = 0.0267), and G1 and G5 (P = 0.0070), with elevated values in mammary carcinomas and non-neoplastic diseases. Greater sensitivity and specificity were noted in G2 compared to the other groups. However, specificity significantly dropped when non-neoplastic diseases were compared to mammary carcinomas. Furthermore, a moderate positive correlation was found between CEA levels and platelet count, indicating a possible influence of inflammation on the variation of this biomarker. The results demonstrated that CEA has good diagnostic value for dogs with mammary carcinomas; however, this was not observed for non-neoplastic diseases. However, non-neoplastic diseases can drastically affect CEA specificity. Future studies evaluating additional diseases will be necessary for a better understanding and clinical application of this biomarker in veterinary practice. mammary tumor ehrlichiosis gastroenteritis CEA biomarker Figures Figure 1 Introduction The ongoing search for methods and strategies to improve patient diagnosis, monitoring, and prognosis is essential in clinical practice (Senhorello et al. 2024 ; Soares et al. 2023 ; Chen et al. 2020 ). In human medicine, serological biomarkers have been introduced to assist in patient management from diagnosis through the completion of treatment. These biomarkers are invaluable for the early detection of cancer recurrence, monitoring inflammatory diseases, and determining patient prognosis (Wang et al. 2014 ; Hall et al. 2019 ; Chen et al. 2020 ; Anjan et al. 2024 ). Carcinoembryonic antigen (CEA) is one of the most commonly used serological biomarkers in human medicine. It is a glycoprotein involved in inflammatory and neoplastic processes (Egawa et al. 1996 ; Kelleher et al. 2019 ) and is primarily produced by the gastrointestinal mucosa under normal physiological conditions (Zhou et al. 1993 ). In humans, excessive production of this glycoprotein occurs in cases of colorectal carcinoma and is also expressed in other malignant and benign tumors within normal mucosal epithelium (Egawa et al. 1996 ). Consequently, its most common clinical application is as a tumor biomarker, particularly for monitoring various organs, especially colorectal and breast tumors (Hall et al. 2019 ). Additionally, elevated CEA levels have been noted in patients with hepatic, pancreatic, and gastrointestinal diseases, often associated with inflammation (Loewenstein et al. 1978; Kelleher et al. 2019 ). Hence, CEA is not exclusively elevated in malignant conditions (Peña et al. 2021). In veterinary medicine, the interest in CEA as a tumor biomarker, particularly for canine mammary neoplasms, has been growing. Studies have shown that CEA can be detected in healthy female dogs, and its levels are elevated in those with mammary neoplasms (Senhorello et al. 2020 ; Fan et al. 2021 ). Furthermore, CEA can be detected in serum via ELISA and in mammary tissue using Western blot and PCR, utilizing both canine and human antibodies (Campos et al. 2012 ; Senhorello et al. 2020 ; Fan et al. 2021 ; Jain et al. 2021 ). However, despite its potential, CEA is still considered poorly studied in veterinary medicine and cannot be reliably used for primary diagnosis of mammary neoplasms (Pinheiro et al. 2022 ). Recent literature also highlights that CEA levels may be elevated in other non-mammary neoplastic conditions, such as ovarian cancer (Kim et al. 2024 ), reinforcing its lack of absolute specificity in canine oncology. The broader consensus indicates that no single ideal biomarker that meets all diagnostic and prognostic criteria has been identified in veterinary medicine, particularly due to the persistent lack of specificity (Çolakoğlu et al. 2025 ). Although some studies have evaluated CEA levels in healthy female dogs and those with mammary neoplasms, demonstrating its sensitivity and specificity for monitoring mammary tumors (Senhorello et al. 2020 ), no research has yet assessed CEA in male dogs or compared CEA levels with other non-neoplastic diseases in dogs. Given that infectious and neoplastic diseases comprise a significant portion of veterinary clinical cases, identifying potential coexisting conditions that may influence biomarker levels is crucial for accurately interpreting results (Vieira et al. 2018 ; Fan et al. 2021 ). In human medicine, the impact of various diseases on CEA levels is well understood, enabling a more precise evaluation of this biomarker in clinical practice (Peña et al. 2021). This highlights a critical gap in veterinary literature, as the behavior of CEA in the presence of common non-neoplastic conditions or across sexes in healthy animals remains largely unexplored, potentially impacting diagnostic accuracy and clinical interpretation. This study aimed to evaluate serum CEA levels in healthy dogs, dogs with mammary neoplasms, dogs with gastroenteritis of unknown origin, and dogs with ehrlichiosis. In addition to comparing mean values, we aimed to evaluate the diagnostic value of CEA across all disease groups, assess the impact of non-neoplastic diseases on its diagnostic accuracy, and investigate the correlation between CEA levels and hematological and biochemical variables in dogs with non-neoplastic diseases. We hypothesize that CEA levels will be elevated in both neoplastic and non-neoplastic diseases in dogs, mirroring observations in humans. If confirmed, non-neoplastic diseases may interfere with the specificity of CEA in dogs with mammary neoplasms. Materials and Methods Ethical Approval This study was submitted to and approved by the Animal Experimentation Ethics Committee (protocol no. 589-2021) at Vila Velha University, Espírito Santo, Brazil, as well as the Animal Experimentation Ethics Committee of the Faculty of Agricultural and Veterinary Sciences at UNESP-Jaboticabal (protocol no. 10801/15). Owners who participated in the study received detailed guidance and signed an informed consent form. Additionally, a declaration verifying the origin of the samples was obtained to confirm their provenance when the clinical laboratories of the institutions provided serum samples. Sample Collection Samples were obtained from animals treated at the Veterinary Hospital of UVV (Vila Velha, ES), Seres Veterinary Hospital (Vila Velha, ES), and VetSaúde Veterinary Hospital (Vitória, ES). These animals underwent clinical examinations at the request of the attending veterinarians. After confirming the diagnoses and reviewing the medical records, blood serum samples were selected and stored in the clinical pathology laboratory's sample bank at the UVV Veterinary Hospital. All blood samples were centrifuged following standard procedures to separate the serum. The serum samples were individually aliquoted and stored at -20°C for further analysis of the CEA biomarker. Additionally, some control group samples and samples from dogs with mammary neoplasms were obtained from a separate research project. The CEA measurements from these samples were incorporated into this study, as the same methodology was employed. A total of 118 dogs were selected, with the mammary neoplasm group consisting solely of female dogs. Animals were included regardless of breed or age. The collected biological material was categorized into five experimental groups: Group 1 (G1): 29 serum aliquots from healthy dogs. Group 2 (G2): 56 serum aliquots from female dogs diagnosed with mammary carcinomas. Group 3 (G3): 17 serum aliquots from dogs diagnosed with ehrlichiosis. Group 4 (G4): 16 serum aliquots from dogs with gastroenteritis of unknown origin. Group 5 (G5): 33 serum aliquots from dogs with non-neoplastic diseases, comprising samples from G3 and G4. This group was specifically created to evaluate the combined impact of distinct non-neoplastic inflammatory and infectious conditions on CEA levels, allowing for a broader assessment of biomarker specificity. All animals underwent a comprehensive clinical evaluation, which included a physical examination, a complete blood count, and a serum biochemistry analysis. Dogs with mammary carcinoma (group 2) additionally had imaging studies (thoracic radiography and abdominal ultrasonography) for clinical staging based on the TNM classification according to the World Health Organization (WHO) guidelines. Definitive diagnosis was confirmed through histopathological analysis of tissue samples obtained following total, unilateral, or bilateral mastectomy, adhering to the established consensus guidelines for canine mammary tumors (Cassali et al. 2020). Dogs in Group 3 (ehrlichiosis) were diagnosed based on compatible clinical signs and laboratory findings, confirmed by the SNAP 4Dx test (IDEXX® Laboratories) for antibody detection and/or RT-PCR for detection of Ehrlichia canis or Ehrlichia ewingii antigens. For the purpose of this study, ehrlichiosis was considered as a systemic inflammatory non-neoplastic condition impacting CEA levels, without focusing on species-specific clinical or biochemical differentiations, which were beyond the scope of this investigation. Patients with gastroenteritis of unknown origin (group 3) were diagnosed based on clinical symptoms, exclusion of other possible causes, and ultrasonographic findings. Healthy dogs (group 1) were selected from routine consultations at the "Professor Ricardo Alexandre Rippler" Veterinary Hospital, UVV, Espírito Santo, Brazil, and the "Governador Laudo Natel" Veterinary Hospital, UNESP, Jaboticabal, São Paulo, Brazil. These animals were evaluated as part of elective neutering procedures and underwent a thorough physical examination, complete blood count, and serum biochemistry to confirm their health status. Exclusion criteria included the presence of concurrent diseases unrelated to those listed in this study, incomplete clinical records, absence of diagnostic tests confirming disease status, and serum samples that were hemolyzed, lipemic, or icteric after centrifugation. Serological Marker A human ELISA kit (AccuBind ELISA Kits, MONOBIND INC, catalog #1825-300) was used to determine serum CEA levels. The use of human-based assays for canine CEA determination is supported by previous studies, which demonstrate sufficient cross-reactivity and clinical utility for this biomarker in dogs (Campos et al. 2012; Pinheiro et al. 2022). The readings were performed using an ELISYS UNO Human analyzer, following the manufacturer's recommended protocol and previously established methodology (Senhorello et al. 2020) Statistical Analysis Each dog was considered an experimental unit for analyzing CEA levels and laboratory variables. The data were subjected to analysis of variance, and since they did not follow a normal distribution, the CEA values were transformed by raising them to the power of 0.25. When significant differences were detected in the F-test, means were compared using Tukey’s test. Specifically, hematological variables, including leukocytes, neutrophils, bands, lymphocytes, monocytes, eosinophils, and platelet counts, along with biochemical parameters such as creatinine, alanine aminotransferase (ALT), urea, and alkaline phosphatase, were correlated with CEA levels in the non-neoplastic disease groups using Pearson’s correlation coefficient. The area under the curve (AUC) of the receiver operating characteristic (ROC) curve was utilized to assess the diagnostic value of the serum marker for all diseases. The cutoff point was determined by comparing the control group with dogs diagnosed with mammary carcinoma. Sensitivity and specificity were calculated for all groups. Crucially, to evaluate the impact of non-neoplastic diseases on CEA specificity, an additional sensitivity and specificity analysis was performed by comparing the mammary carcinoma group (G2) against the non-neoplastic disease groups (G3, G4, and the combined G5). This comparative approach aimed to determine the potential for false-positive results resulting from these confounding conditions. Analyses were conducted using GraphPad Prism v. 10.4 software with a significance level of 5%. Results A total of 118 animals participated in the study, consisting of 17 males (14.4%) and 101 females (85.6%), with ages ranging from 3 months to 15 years. No significant difference was found in CEA levels between males (0.89 ± 0.84 ng/mL) and females (1.00 ± 0.75 ng/mL) without mammary neoplasia (P = 0.6224). Mean CEA levels were notably highest in the mammary carcinoma group (G2; 1.94 ng/mL) and lowest in the control group (G1; 0.65 ng/mL). Significant elevations in CEA were observed in both the mammary carcinoma group (G2) compared to healthy controls (G1) (P < 0.0001), and importantly, in the non-neoplastic gastroenteritis group (G4) (p = 0.0237) and the combined non-neoplastic diseases group (G5) (P = 0.0120) when compared to healthy controls (G1). Furthermore, a significant difference was detected between the mammary carcinoma group (G2) and the combined non-neoplastic diseases group (G5) (P = 0.0466), indicating a distinct profile of CEA elevation in these conditions. No significant differences were noted among the other group comparisons (P > 0.05) (Table 1, Figure 1). Table 1 – Mean and standard deviation of CEA levels in experimental groups. Group (n) CEA (ng/mL) G1 (29) 0.65 (±0.32) a G2 (56) 1.94 (±0.89) b G3 (17) 1.29 (±1.06) abc G4 (16) 1.50 (±0.98) bc G5 (33) 1.39 (±1.02) c CEA: Carcinoembryonic Antigen. Differing letters signify statistically significant differences between groups (P 0.05). The diagnostic performance of serum CEA was evaluated using ROC curve analysis (Table 2). Employing an optimal cutoff value of 1.08 ng/mL, derived from the comparison between healthy controls (G1) and mammary carcinoma cases (G2), CEA demonstrated high sensitivity (82.14%) and specificity (93.10%) for detecting mammary carcinomas. However, when this established cutoff was applied to assess sensitivity across other groups, CEA's performance was markedly lower for G4 (50.00%), G5 (48.48%), and G3 (47.06%). Crucially, to elucidate the interference of non-neoplastic diseases on CEA's diagnostic utility for mammary carcinomas, we conducted a comparative analysis between the mammary carcinoma group (G2) and the non-neoplastic disease groups (G3, G4, and G5). While sensitivity for mammary carcinomas remained constant at 82.14% across these comparisons, specificity decreased dramatically to 52.94% for G3, 50.00% for G4, and 51.52% for G5. This stark reduction unequivocally demonstrates a significant compromise in CEA's specificity in the presence of common non-neoplastic conditions, highlighting its limitations as a standalone biomarker in a broader clinical context. Table 2 – Sensitivity and specificity values for diseased animals compared to the control group and non-neoplastic disease groups compared to mammary carcinoma cases. Group Sensitivity (%) IC (%) Specificity (%) IC (%) AUC P-value G1 vs. G2 82.14 70.16-90.00 93.10 78.04-98.77 0.94 <0.0001 G1 vs. G3 47.06 26.17-69.04 93.10 78.04-98.77 0.69 0.0298 G1 vs. G4 50.00 28.00-72.00 93.10 78.04-98.77 0.77 0.0028 G1 vs. G5 48.48 32.50-64.78 93.10 78,04-98,77 0.73 0.0018 G3 vs. G2 82.14 70.16-90.00 52.94 30.96-73.83 0.71 0.0065 G4 vs. G2 82.14 70.16-90.00 50.00 28.00-72.00 0.66 0.0415 G5 vs. G2 82.14 70.16-90.00 51.52 35.22-67.50 0.69 0.0023 AUC: Area under the curve; IC: Confidence interval Among the hematological parameters evaluated, a significant moderate positive correlation was observed between platelet counts and CEA levels in both the ehrlichiosis group (G3; r = 0.68, P = 0.01) and the combined non-neoplastic diseases group (G5; r = 0.37, P = 0.01). No other significant correlations were found for leukocyte differentials (leukocytes, neutrophils, bands, lymphocytes, monocytes, or eosinophils) (P > 0.05). Furthermore, evaluation of correlations between CEA levels and serum biochemical parameters (creatinine, ALT, urea, and alkaline phosphatase) in animals with non-neoplastic diseases (n = 28) yielded no significant associations (P > 0.05) (Table 3). Table 3 – Pearson correlation coefficients between CEA levels and hematological and biochemical parameters. G3 G4 G5 Variable r P-value r P-value r P-value Leukocytes -0.44 0.125 -0.22 0.41 -0.30 0.06 Bands -0.34 0.17 -0.20 0.46 -0.23 0.14 Neutrophils -0.40 0.25 -0.05 0.84 -0.18 0.27 Lymphocytes -0.43 0.14 -0.37 0.17 -0.31 0.05 Monocytes -0.20 0.51 -0.06 0.82 -0.14 0.39 Eosinophils -0.13 0.66 -0.19 0.17 -0.07 0.65 Platelets 0.68 0.01 -0.37 0.16 0.37 0.01 RBCs 0.48 0.09 0.28 0.29 0.25 0.11 Hemoglobin 0.39 0.18 0.30 0.26 0.25 0.11 Hematocrit 0.39 0.18 0.27 0.31 0.25 0.12 Creatinine - - - - -0.07 0.70 ALT - - - - -0.08 0.68 Urea - - - - -0.16 0.39 Alkaline Phosphatase - - - - -0.21 0.26 ALT: alanine aminotransferase Discussion Current studies on CEA in veterinary medicine primarily focus on its association with mammary neoplasms in female dogs (Senhorello et al. 2020; Fan et al. 2021; Jain et al. 2021). However, no studies have compared CEA levels between male and female dogs; existing research has only evaluated this biomarker in healthy females and those with mammary neoplasia (Ledecky et al. 2013; Senhorello et al. 2020; Fan et al. 2021). In the present study, serum CEA levels were assessed in both males and females, revealing no significant differences between the sexes. Similarly, human studies have reported no correlation between CEA levels and sex but rather with underlying diseases. These include neoplastic conditions such as gastric, colorectal, and breast carcinomas (Deng et al. 2015; Reiter et al. 2000; Wang et al. 2014), as well as non-neoplastic conditions such as inflammatory bowel disease, hepatitis, pancreatitis, and COVID-19 (Hao et al. 2019; Chen et al. 2021; Anjan et al. 2024). The highest mean CEA concentrations were found in female dogs with mammary neoplasms (G2) in comparison to healthy females (G1) and dogs with non-neoplastic diseases (G5). These findings confirm the potential of this biomarker for monitoring female dogs with mammary neoplasms, supporting the studies by Senhorello et al. (2020) and Fan et al. (2021), which noted elevated CEA levels in patients with mammary tumors. This observation is consistent with recent findings by Singh et al. (2025), who reported significantly higher serum CEA levels in canines with malignant mammary tumors compared to healthy controls. Furthermore, following mastectomy, the serum concentration of this biomarker significantly dropped and remained low in patients evaluated in follow-up assessments (Senhorello et al. 2020). However, it is crucial to recognize that despite its prognostic potential in mammary neoplasms, CEA alone exhibits limited specificity and may not be a standalone indicator for early diagnosis or definitive assessment, as also highlighted by Singh et al. (2025), who found that CEA alone was not highly specific and showed no significant difference between benign tumors and healthy controls In humans, elevated serum CEA concentrations are associated with both neoplastic and non-neoplastic diseases (Hao et al. 2019; Peña et al. 2021). Addressing a critical gap in veterinary literature, this study is the first to comprehensively investigate CEA levels in dogs affected by common non-neoplastic conditions. While no significant difference was observed between the control group (G1) and animals diagnosed solely with ehrlichiosis (G3), a significant elevation in mean CEA levels was noted in the gastroenteritis group (G4) and, notably, in the combined group of non-neoplastic diseases (G5) when compared to healthy controls (G1). This pivotal finding strongly suggests that CEA levels in dogs are not exclusively indicative of neoplastic processes and can be significantly influenced by inflammatory and infectious non-neoplastic conditions, mirroring observations in human medicine and contrasting with the absence of significant CEA elevation in benign mammary tumors reported by Singh et al. (2025). This difference highlights the diverse etiologies that can influence CEA concentrations in canines, underscoring the need for careful differential diagnosis. Our results align with findings in human medicine, where elevated serum CEA levels have been reported in patients diagnosed with ulcerative colitis, inflammatory bowel disease, and other inflammation-associated conditions (Rule et al. 1973; Loewenstein et al. 1978; Hao et al. 2019; Kelleher et al. 2019). According to Tchoupa et al. (2014), carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) are part of a subfamily of immunoglobulin-related proteins that possess a high affinity for cellular adhesion to specific pathogens, such as Escherichia coli (Kelleher et al. 2019). Since this bacterium is one of the principal agents involved in the increased secretion of pro-inflammatory cytokines in dogs (Honneffer et al. 2014), it may explain the rise in CEA levels noted in gastroenteritis cases. Furthermore, Escherichia coli is the primary microorganism isolated from human patients with inflammatory bowel disease, further reinforcing this similarity (Tchoupa et al. 2014). Cellular adhesion can stimulate an innate immune response, activate specific kinases, promote the stimulation of small G proteins, and induce new gene expression events (Tchoupa et al. 2014). These processes can lead to cellular damage and inflammation, ultimately resulting in epithelial injury, which may contribute to tumorigenesis, tissue fibrosis, and increased serum CEA levels (Hao et al. 2019). Similarly, ehrlichiosis, a common hemoparasitic infection in dogs, can induce the gene expression of various cytokines, including interleukin-6 (IL-6), a key mediator of acute-phase responses (Gröne et al. 1998; Brachelente et al. 2005). The known correlation between circulating CEA levels and IL-6 in human patients with certain cancers (Nakagoe and Sawai, 2003; Holmer et al. 2015) could provide a plausible explanation for the observed elevations in CEA in some dogs with ehrlichiosis. These specific findings, alongside those from gastroenteritis, highlight that common inflammatory and infectious conditions can indeed elevate CEA levels in canines, thus complicating the interpretation of this biomarker for cancer screening or monitoring. This highlights the importance of a comprehensive diagnostic approach, rather than relying solely on CEA as an indicator. Further studies are indeed necessary to fully elucidate these specific pathophysiological mechanisms. It is important to note that, in addition to evaluating whether non-neoplastic diseases can elevate CEA levels in dogs, this study plays a crucial role in the clinical application of CEA for animals diagnosed with mammary cancer, as previous research supports this use (Senhorello et al. 2020; Fan et al. 2021; Jain et al. 2021). However, we still do not fully comprehend which diseases may affect marker values during patient follow-up. The optimal CEA cutoff point (1.08 ng/mL), established through a comparison between healthy controls and patients with mammary carcinoma, served as a crucial reference for evaluating the biomarker’s diagnostic utility. Our findings, consistent with previous research (Senhorello et al. 2020), demonstrated good sensitivity (82.14%) and specificity (93.10%) for CEA in detecting mammary carcinomas. However, it is noteworthy that even in studies confirming CEA elevation in malignant mammary tumors, the individual specificity of CEA has been reported as relatively low (Singh et al. 2025). This further underscores the importance of our investigation into confounding factors. Moreover, when this established cutoff was applied to assess sensitivity across other non-neoplastic groups, CEA's performance was considerably lower, aligning with human studies on CEA in non-malignant conditions (Hao et al. 2019). This disparity in performance highlights the inherent challenge of biomarker interpretation in diverse clinical presentations, a concern further amplified by recent discussions on the general inspecificity of tumor biomarkers in veterinary medicine (Çolakoğlu et al. 2025). Crucially, our study demonstrated that common non-neoplastic conditions significantly compromise CEA's specificity for mammary carcinomas, to a greater extent than the inherent limitations observed in other studies (Singh et al. 2025). When CEA measurement was evaluated against non-neoplastic diseases (G3, G4, and G5), the specificity for mammary carcinomas decreased dramatically (to 52.94% for G3, 50.00% for G4, and 51.52% for G5). This stark reduction unequivocally indicates that non-neoplastic diseases, such as gastroenteritis and ehrlichiosis, significantly heighten the risk of false-positive CEA results for mammary carcinomas. Consequently, a patient with mammary tumors undergoing CEA-based follow-up, who subsequently develops one of these common non-neoplastic conditions, may exhibit altered CEA values directly attributable to the intercurrent disease, thereby severely undermining the reliability of the biomarker for neoplastic monitoring. This finding provides direct evidence supporting the necessity of considering confounding non-neoplastic conditions when interpreting CEA levels in canine patients, a previously unaddressed clinical challenge." These findings align with those reported in human medicine, as CEA is a non-specific biomarker that can be elevated in non-cancerous conditions, including gastrointestinal diseases, inflammatory processes, and other disorders like renal and hepatic diseases, which may increase CEA levels even without neoplasia. As a result, CEA is not a reliable general tool for cancer detection, and the interpretation of results should take into account the patient's comorbidities (Trape et al. 2011; Hao et al. 2019; Peña et al. 2021). Our study extends this crucial understanding to veterinary medicine, emphasizing that clinical context and thorough differential diagnostics are paramount for accurate CEA interpretation in dogs, particularly given the inherent limitations of single biomarkers as noted by Singh et al. (2025). Our correlation analyses aimed to identify potential influences of hematological and biochemical changes, commonly associated with non-neoplastic diseases, on CEA levels. While most variables did not show significant correlations, a notable moderate positive correlation was identified between platelet counts and CEA levels in both the ehrlichiosis group (G4) and the combined non-neoplastic diseases group (G5). This finding provides indirect evidence of the intricate relationship between inflammatory processes and CEA elevation, supporting the broader understanding that CEA can be influenced by systemic inflammation. In inflammatory conditions, thrombocytosis can occur due to IL-6 production, which stimulates thrombopoietin production (Ceresa et al. 2007; Athanasiou et al. 2017). As IL-6 has been linked to increased CEA levels in humans (Nakagoe and Sawai, 2003; Holmer et al. 2015), our results suggest a similar underlying mechanism in dogs, where inflammation, mediated by cytokines like IL-6, could contribute to the observed CEA elevations. These correlations, particularly with platelets, further support the notion that CEA levels are not solely dictated by neoplastic disease, highlighting a crucial area for future mechanistic investigations in veterinary species. The findings are also consistent with the call from Singh et al. (2025) for more comprehensive studies on CEA's diagnostic value, particularly given its lower individual specificity. Regarding serum biochemical parameters (creatinine, ALT, urea, and alkaline phosphatase), no significant correlations were observed with CEA levels. This assessment was conducted in light of reports in human medicine linking elevated CEA levels to hepatic and renal diseases (Rule et al. 1973; Hao et al. 2019). While ehrlichiosis and gastroenteritis can induce systemic effects, they are not primarily defined by direct hepatic or renal dysfunction leading to consistent enzyme elevations. Thus, the limited number of animals exhibiting significant alterations in these specific parameters within our study groups might explain the lack of observed correlations. This finding suggests that CEA elevation in these specific non-neoplastic conditions may be more strongly associated with inflammatory processes (as indicated by the platelet correlation) rather than direct organ damage, as reflected by these biochemical markers. Future investigations involving a larger cohort of animals with confirmed hepatic or renal pathologies would be beneficial for fully elucidating these relationships. Despite the significant contributions of this study, certain limitations should be acknowledged. Firstly, while novel data on CEA levels in ehrlichiosis and gastroenteritis were provided, the range of non-neoplastic conditions investigated was limited. Further research encompassing a broader spectrum of inflammatory, infectious, and systemic diseases would provide a more comprehensive understanding of CEA's behavior in canine health and disease. Secondly, although our mechanistic discussion suggests the role of inflammatory mediators, such as interleukin-6 (IL-6), in CEA elevation, these were not directly measured in the study; thus, our explanations remain inferential. Future studies could incorporate direct cytokine quantification to solidify these proposed pathways. Lastly, although the overall sample size was robust, the individual sample sizes for specific non-neoplastic subgroups (G3 and G4) were relatively modest, and the inherent clinical heterogeneity within these conditions (e.g., varying severity or specific etiologies of gastroenteritis) could introduce variability. These considerations highlight avenues for future research aimed at further refining the diagnostic and prognostic utility of CEA in veterinary medicine. Conclusion This study provides crucial insights into the behavior of serum Carcinoembryonic Antigen (CEA) in canine patients. CEA levels were confirmed to be significantly elevated in dogs with mammary carcinomas, highlighting their potential for disease monitoring. Importantly, no significant difference was observed in CEA levels between healthy male and female dogs, thereby expanding the scope for CEA's clinical utility across both sexes. However, a key finding demonstrates that common non-neoplastic conditions, such as gastroenteritis and ehrlichiosis, also lead to a marked increase in CEA concentrations. This elevation significantly compromises CEA's specificity for mammary neoplasms, thereby increasing the risk of false-positive results. Furthermore, a positive correlation was noted between CEA levels and platelet count, suggesting that inflammatory processes significantly influence the variability of this biomarker. Therefore, while CEA holds value in canine mammary tumor assessment, its accurate interpretation requires a comprehensive clinical evaluation of the patient, taking into account concurrent non-neoplastic conditions. Future research should expand this knowledge by investigating other confounding variables that may affect CEA values and prioritizing the development of multi-biomarker panels to enhance diagnostic accuracy and specificity in veterinary oncology. Declarations Funding This work was funded by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), process number 2016/00128-5. Data Availability Statement The data supporting the conclusions of this study are available from the author upon reasonable request. Conflict of Interest Statement The authors declare that there is no conflict of interest. Author Contributions L.W.A.L: Conceptualization, investigation, methodology, writing – original draft. L.C.C: Investigation, methodology. A.C.T.B.F: Investigation, methodology. G.C.V: Investigation, methodology. C.G: methodology, data analysis. F.A.R.S: Investigation, methodology, data analysis. R.P.S: Investigation, methodology, data analysis. M.T.C: Conceptualization, methodology, resources, funding acquisition, supervision, writing – original draft. I.L.S.S: Conceptualization, data curation, investigation, methodology, data analysis, supervision, writing – original draft. All authors read and approved the final manuscript. Ethics approval Approval was granted by the Animal Experimentation Ethics Committee at the Faculty of Agricultural and Veterinary Sciences of UNESP-Jaboticabal (protocol no. 10801/15) and the Committee on Animal Experimentation Ethics (protocol no. 589-2021) at Vila Velha University (UVV). References Anjan MAH, Ahmed QMU, Masum AA, Sami CA, Matin MA, Islam MS, Chowdhury FR, Arafat SM, Rahman M, Hasan MN (2024) Role of Carcinoembryonic Antigen in Severity Assessment and Mortality Prediction in COVID-19 Patients. Cureus 16(9):e69894. https://doi.org/10.7759/cureus.69894 Athanasiou LV, Polizopoulou ZS, Papavasileiou EG, Mpairamoglou EL, Kantere MC, Rousou XA (2017) Magnitude of reactive thrombocytosis and associated clinical conditions in dogs. Vet Rec 181(10):267. https://doi.org/10.1136/vr.104042 de la Ayala F, Ortiz-Muñoz B, Quintanar-Verdúguez T et al (2021) Consensus of the Spanish society of laboratory medicine and the Spanish society of medical oncology on the methodology and criteria for evaluation of circulating tumour markers in breast cancer. Clin Transl Oncol 23(7):1272–1280. https://doi.org/10.1007/s12094-020-02529-x Brachelente C, Müller N, Doherr MG, Sattler U, Welle M (2005) Cutaneous leishmaniasis in naturally infected dogs is associated with a T helper-2-biased immune response. Vet pathol 42(2):166–175. https://doi.org/10.1354/vp.42-2-166 Campos LC, Lavalle GE, Estrela-Lima A, Melgaço de Faria JC, Guimarães JE, Dutra ÁP, Ferreira E, de Sousa LP, Rabelo ÉM, Vieira da, Costa AF, Cassali GD (2012) CA15.3, CEA and LDH in dogs with malignant mammary tumors. J Vet Intern Med 26(6):1383–1388. https://doi.org/10.1111/j.1939-1676.2012.01014.x Carlos RS, Muniz Neta ES, Spagnol FH, Oliveira LL, de Brito RL, Albuquerque GR, Almosny NR (2007) Frequency of antibodies anti-Ehrlichia canis, Borrelia burgdorferi and Dirofilaria immitis antigens in dogs from microrregion Ilhéus-Itabuna, State of Bahia. Brazil Rev Bras Parasitol Vet 16(3):117–120. https://doi.org/10.1590/s1984-29612007000300001 Cassali GD, Jark PC, Gamba C et al (2020) Consensus Regarding the Diagnosis, Prognosis and Treatment of Canine and Feline Mammary Tumors – 2019. Braz J Vet Pathol 13(3):555–574. https://doi.org/10.24070/bjvp.1983-0246.v13i3p555-574 Ceresa IF, Noris P, Ambaglio C, Pecci A, Balduini CL (2007) Thrombopoietin is not uniquely responsible for thrombocytosis in inflammatory disorders. Platelets 18(8):579–582. https://doi.org/10.1080/09537100701593601 Chen Q, Kong H, Qi X, Ding W, Ji N, Wu C, Huang C, Wu W, Huang M, Xie W, Liu Y, Tang J (2020) Carcinoembryonic Antigen: A Potential Biomarker to Evaluate the Severity and Prognosis of COVID-19. Front Med 7:579543. https://doi.org/10.3389/fmed.2020.579543 Çolakoğlu HE, Fidan Ç, Alçığır ME, Göbekçi AE, Devrim E, Anteplioğlu T, Yazlık MO, Özmen Ö, Reçber T, Nemutlu E, Kaya U (2025) Evaluation of serum CA15-3, CEA, Ki-67, PDL-1 and VEGF as a potential diagnostic and prognostic biomarkers in canine mammary tumors. Theriogenology 247:117579. https://doi.org/10.1016/j.theriogenology.2025.117579 Deng K, Yang L, Hu B, Wu H, Zhu H, Tang C (2015) The prognostic significance of pretreatment serum CEA levels in gastric cancer: a meta-analysis including 14651 patients. PLoS ONE 10(4):e0124151. https://doi.org/10.1371/journal.pone.0124151 Egawa K, Honda Y, Kuroki M, Inaba Y, Ono T (1996) Carcinoembryonic antigen and related antigens expressed on keratinocytes in inflammatory dermatoses. Br J Dermatol 134(3):451–459 Fan Y, Ren X, Liu X, Shi D, Xu E, Wang S, Liu Y (2021) Combined detection of CA15-3, CEA, and SF in serum and tissue of canine mammary gland tumor patients. Sci Rep 11(1):6651. https://doi.org/10.1038/s41598-021-85029-4 Gröne A, Frisk AL, Baumgärtner W (1998) Cytokine mRNA expression in whole blood samples from dogs with natural canine distemper virus infection. Vet Immunol Immunopathol 65(1):11–27. https://doi.org/10.1016/s0165-2427(98)00170-6 Hall C, Clarke L, Pal A, Buchwald P, Eglinton T, Wakeman C, Frizelle F (2019) A Review of the Role of Carcinoembryonic Antigen in Clinical Practice. Ann Coloproctol 35(6):294–305. https://doi.org/10.3393/ac.2019.11.13 Hao C, Zhang G, Zhang L (2019) Serum CEA levels in 49 different types of cancer and noncancer diseases. Prog Mol Biol Transl Sci 162:213–227. https://doi.org/10.1016/bs.pmbts.2018.12.011 Holmer R, Wätzig GH, Tiwari S, Rose-John S, Kalthoff H (2015) Interleukin-6 trans-signaling increases the expression of carcinoembryonic antigen-related cell adhesion molecules 5 and 6 in colorectal cancer cells. BMC Cancer 15:975. https://doi.org/10.1186/s12885-015-1950-1 Honneffer JB, Minamoto Y, Suchodolski JS (2014) Microbiota alterations in acute and chronic gastrointestinal inflammation of cats and dogs. World J Gastroenterol 20(44):16489–16497. https://doi.org/10.3748/wjg.v20.i44.16489 Jain M, Ingole SD, Deshmukh RS, Bharucha SV, Nagvekar AS, Gaikwad RV, Kharde SD (2021) CEA, CA 15 – 3, and miRNA expression as potential biomarkers in canine mammary tumors. Chromosome Res 29(2):175–188. https://doi.org/10.1007/s10577-021-09652-7 Kelleher M, Singh R, O'Driscoll CM, Melgar S (2019) Carcinoembryonic antigen (CEACAM) family members and Inflammatory Bowel Disease. Cytokine Growth Factor Rev 47:21–31. https://doi.org/10.1016/j.cytogfr.2019.05.008 Kim S-H, Yoo DS, Park D-H, Cho S-S, Park S, Jung B-G, Park S-I, Bae C-S (2024) Combined Application of CEA, CA 15 – 3, and CA 27–29 for the Evaluation of Diagnostic Performance in Canine Mammary Gland Tumors. J Veterinary Clin 41:359–369. https://doi.org/10.17555/jvc.2024.41.6.359 Kwon YJ, Lee HS, Shim JY, Lee YJ (2018) Serum carcinoembryonic antigen is positively associated with leukocyte count in Korean adults. J Clin Lab Anal 32(3):e22291. https://doi.org/10.1002/jcla.22291 Ledecky V, Valencakova-Agyagosova A, Lepej J, Frischova Z, Hornak S, Nagy V (2013) Determination of carcinoembryonic antigen and cancer antigen values with the radioimmunoassay method in healthy females dogs. Vet Med 58(5):277–283. https://doi.org/10.17221/6810-VETMED Loewenstein MS, Zamcheck N (1978) Carcinoembryonic antigen (CEA) levels in benign gastrointestinal disease states. Cancer 42(3 Suppl):1412–1418. https://doi.org/10.1002/1097-0142(197809)42:3+3.0.co;2-8 Nakagoe T, Tsuji T, Sawai T, Tanaka K, Hidaka S, Shibasaki S, Nanashima A, Yamaguchi H, Yasutake T, Sugawara K, Inokuchi N, Kamihira S (2003) The relationship between circulating interleukin-6 and carcinoembryonic antigen in patients with colorectal cancer. Anticancer Res 23(4):3561–3564 Paniz C, Grotto D, Schmitt GC et al (2005) Physiopathology of vitamin B12 deficiency and its laboratorial diagnosis. J Bras Patol Med Lab 41:323–334. https://doi.org/10.1590/S1676-24442005000500007 Pinheiro BQ, da Silva IN, Faustino AM, Machado da Silva LD (2022) The prognostic value of serum neoplastic biomarkers CA 15 – 3 and CEA in canine mammary neoplasms: a review. Revista Brasileira de Reprodução Anim 46(3):290–297. https://doi.org/10.21451/1809-3000.RBRA2022.021 Reiter W, Stieber P, Reuter C, Nagel D, Lau-Werner U, Lamerz R (2000) Multivariate analysis of the prognostic value of CEA and CA 19 – 9 serum levels in colorectal cancer. Anticancer Res 20(6D):5195–5198 Rueda JR, Porto CD, Franco RP, da Costa IB, Bueno LMC, Girio RJS, Manhoso FFR, Bueno PCDS, Repetti CSF (2024) Mammary neoplasms in female dogs: Clinical, diagnostic and therapeutic aspects. Vet Med 69(4):99–114. https://doi.org/10.17221/4/2024-VETMED Rule AH, Goleski-Reilly C, Sachar DB, Vandevoorde J, Janowitz HD (1973) Circulating carcinoembryonic antigen (CEA): relationship to clinical status of patients with inflammatory bowel disease. Gut 14(11):880–884. https://doi.org/10.1136/gut.14.11.880 Senhorello ILS, Terra EM, Sueiro FAR, Firmo BF, Anai LA, Goloni C, Canavari IC, Ampuero RAN, Pereira RS, Tinucci-Costa M (2020) Clinical value of carcinoembryonic antigen in mammary neoplasms of bitches. Vet Comp Oncol 18(3):315–323. https://doi.org/10.1111/vco.12552 Senhorello ILS, Terra EM, Sueiro FAR, Pereira RS, Firmo BF, Martinelli C, Tinucci-Costa M (2024) Clinical value of alpha-fetoprotein in the detection of mammary carcinoma in female dogs. Arq Bras Med Vet Zootec 76(5):e13231. https://doi.org/10.1590/1678-4162-13231 Singh D, Rokade PP, Gangwar NK, Gabhane MG, Malik S, Gangwar K, Prabhu SN, Singh R, Singh DD, Kumari S, Chaudhary S, Choudhary JK (2025) Diagnostic value of carcinoembryonic antigen, cancer antigen 15 – 3, and cell-free DNA as blood biomarkers in early detection of canine mammary tumor. J Circulating Biomarkers 14:30–38. https://doi.org/10.33393/jcb.2025.3564 Soares EDS, Valente FL, Rocha CC, Real Pereira CE, Sarandy TB, de Oliveira FLD, de Morais Calado SL, Borges APB (2023) Prognostic Factors for Cancer-Specific Survival and Disease-Free Interval of Dogs with Mammary Carcinomas. Vet Med Int 2023:6890707. https://doi.org/10.1155/2023/6890707 Tchoupa AK, Schuhmacher T, Hauck CR (2014) Signaling by epithelial members of the CEACAM family - mucosal docking sites for pathogenic bacteria. Cell Commun Signal 12:27. https://doi.org/10.1186/1478-811X-12-27 Trapé J, Filella X, Alsina-Donadeu M, Juan-Pereira L, Bosch-Ferrer Á, Rigo-Bonnin R, Oncology Section of the Catalan Association of Clinical Laboratory Science (2011) Increased plasma concentrations of tumour markers in the absence of neoplasia. Clin Chem Lab Med 49(10):1605–1620. https://doi.org/10.1515/CCLM.2011.694 Vieira FT, Acosta ICL, Martins TF, Filho JM, Krawczak FDS, Barbieri ARM, Egert L, Fernandes DR, Braga FR, Labruna MB, Dietze R (2018) Tick-borne infections in dogs and horses in the state of Espírito Santo, Southeast Brazil. Vet Parasitol 249:43–48. https://doi.org/10.1016/j.vetpar.2017.11.005 Wang G, Qin Y, Zhang J, Zhao J, Liang Y, Zhang Z, Qin M, Sun Y (2014) Nipple discharge of CA15-3, CA125, CEA and TSGF as a new biomarker panel for breast cancer. Int J Mol Sci 15(6):9546–9565. https://doi.org/10.3390/ijms15069546 Woolcock AD, Keenan A, Cheung C, Christian JA, Moore GE (2017) Thrombocytosis in 715 Dogs (2011–2015). J Vet Intern Med 31(6):1691–1699. https://doi.org/10.1111/jvim.14831 Zhou H, Fuks A, Alcaraz G, Bolling TJ, Stanners CP (1993) Homophilic adhesion between Ig superfamily carcinoembryonic antigen molecules involves double reciprocal bonds. J Cell Biol 122(4):951–960. https://doi.org/10.1083/jcb.122.4.951 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7481550","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Short Report","associatedPublications":[],"authors":[{"id":507001466,"identity":"90ae1d28-652d-4062-92b9-a303edee3902","order_by":0,"name":"Leonardo Willian Amite Lababrín","email":"","orcid":"","institution":"Universidade Vila Velha","correspondingAuthor":false,"prefix":"","firstName":"Leonardo","middleName":"Willian Amite","lastName":"Lababrín","suffix":""},{"id":507001467,"identity":"344fc1d1-b94d-414f-9524-0d99e8d44338","order_by":1,"name":"Lara Coslop Comércio","email":"","orcid":"","institution":"Universidade Vila Velha","correspondingAuthor":false,"prefix":"","firstName":"Lara","middleName":"Coslop","lastName":"Comércio","suffix":""},{"id":507001468,"identity":"0df70c90-467e-4c7d-9380-56160c7054b1","order_by":2,"name":"Ana Carolina Teixeira Brandão Fagundes","email":"","orcid":"","institution":"Universidade Vila Velha","correspondingAuthor":false,"prefix":"","firstName":"Ana","middleName":"Carolina Teixeira Brandão","lastName":"Fagundes","suffix":""},{"id":507001469,"identity":"42174073-2e6a-4b73-89d8-a43a2a46a239","order_by":3,"name":"Gabriela Cirino Varnier","email":"","orcid":"","institution":"Universidade Vila Velha","correspondingAuthor":false,"prefix":"","firstName":"Gabriela","middleName":"Cirino","lastName":"Varnier","suffix":""},{"id":507001470,"identity":"39cbe4ca-d096-4823-aeb7-94172e39d74e","order_by":4,"name":"Camila Goloni","email":"","orcid":"","institution":"Faculdade de Medicina Veterinária, UNESP","correspondingAuthor":false,"prefix":"","firstName":"Camila","middleName":"","lastName":"Goloni","suffix":""},{"id":507001471,"identity":"51f849ca-c007-47a9-8165-4e0995edff9e","order_by":5,"name":"Felipe Augusto Ruiz Sueiro","email":"","orcid":"","institution":"Médico-veterinário","correspondingAuthor":false,"prefix":"","firstName":"Felipe","middleName":"Augusto Ruiz","lastName":"Sueiro","suffix":""},{"id":507001472,"identity":"87df0f5d-1931-4264-a281-7ade1c343f7c","order_by":6,"name":"Rodrigo Storti Pereira","email":"","orcid":"","institution":"Médico-veterinário","correspondingAuthor":false,"prefix":"","firstName":"Rodrigo","middleName":"Storti","lastName":"Pereira","suffix":""},{"id":507001473,"identity":"0bd87daa-07f3-432e-a42e-c3ada66d138c","order_by":7,"name":"Mirela Tinucci-Costa","email":"","orcid":"","institution":"UNESP","correspondingAuthor":false,"prefix":"","firstName":"Mirela","middleName":"","lastName":"Tinucci-Costa","suffix":""},{"id":507001474,"identity":"850c2e4f-9141-4b9f-ba2f-86a62eed0e88","order_by":8,"name":"Igor Luiz Salardani Senhorello","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA+klEQVRIiWNgGAWjYBACNigtA8QGQGQD4TI2ENbCA9WSRlgLDEC1MBwmrIWPf/GxDwx/7Hj4Zx/e+Lmg4Hzidvb2Zw8Yd9zD7TCJZ8kzGNuSeSTOpRVLzzC4nbiz54y5AeOZYjxazhgDncHMw3CGx0CaB6hlw40cNgnGtgT8Whj+1PPIn+Ex/s1jcC5xw/3nz/Br4e8BamE7zGNwhscMaMsBoC0MZgRsYUtmSGw7zmN4hq3Mmscg2XjDmRxzg8QzuLXI9x8+zPDhT7Wc3Bnmzbd5/tjJbjh+/NmDjztwa2GQAMqhS7NhiKAA/gNYXItPwygYBaNgFIw8AADRKU5ptuGSgAAAAABJRU5ErkJggg==","orcid":"","institution":"Faculdade de Medicina Veterinária, UNESP","correspondingAuthor":true,"prefix":"","firstName":"Igor","middleName":"Luiz Salardani","lastName":"Senhorello","suffix":""}],"badges":[],"createdAt":"2025-08-28 15:08:32","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7481550/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7481550/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":90320026,"identity":"7007e67d-72e0-4896-afff-17dd481d3969","added_by":"auto","created_at":"2025-09-01 10:42:30","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":8884,"visible":true,"origin":"","legend":"\u003cp\u003eGraphical representation of cea values (ng/ml) in the experimental groups.\u003c/p\u003e","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7481550/v1/55d674d8f3c74c12e5a74f00.png"},{"id":96253200,"identity":"854d3177-64d9-4959-9539-69589f022683","added_by":"auto","created_at":"2025-11-19 07:42:08","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":674095,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7481550/v1/27df09fe-d269-40f9-bf91-fc5832fd4892.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Diagnostic Specificity of Serum Carcinoembryonic Antigen for Canine Mammary Neoplasms: The Impact of Non-Neoplastic Diseases","fulltext":[{"header":"Introduction","content":"\u003cp\u003eThe ongoing search for methods and strategies to improve patient diagnosis, monitoring, and prognosis is essential in clinical practice (Senhorello et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Soares et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Chen et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). In human medicine, serological biomarkers have been introduced to assist in patient management from diagnosis through the completion of treatment. These biomarkers are invaluable for the early detection of cancer recurrence, monitoring inflammatory diseases, and determining patient prognosis (Wang et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Hall et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Chen et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Anjan et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eCarcinoembryonic antigen (CEA) is one of the most commonly used serological biomarkers in human medicine. It is a glycoprotein involved in inflammatory and neoplastic processes (Egawa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1996\u003c/span\u003e; Kelleher et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2019\u003c/span\u003e) and is primarily produced by the gastrointestinal mucosa under normal physiological conditions (Zhou et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e1993\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn humans, excessive production of this glycoprotein occurs in cases of colorectal carcinoma and is also expressed in other malignant and benign tumors within normal mucosal epithelium (Egawa et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1996\u003c/span\u003e). Consequently, its most common clinical application is as a tumor biomarker, particularly for monitoring various organs, especially colorectal and breast tumors (Hall et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eAdditionally, elevated CEA levels have been noted in patients with hepatic, pancreatic, and gastrointestinal diseases, often associated with inflammation (Loewenstein et al. 1978; Kelleher et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Hence, CEA is not exclusively elevated in malignant conditions (Pe\u0026ntilde;a et al. 2021).\u003c/p\u003e\u003cp\u003eIn veterinary medicine, the interest in CEA as a tumor biomarker, particularly for canine mammary neoplasms, has been growing. Studies have shown that CEA can be detected in healthy female dogs, and its levels are elevated in those with mammary neoplasms (Senhorello et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Fan et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Furthermore, CEA can be detected in serum via ELISA and in mammary tissue using Western blot and PCR, utilizing both canine and human antibodies (Campos et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Senhorello et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Fan et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Jain et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). However, despite its potential, CEA is still considered poorly studied in veterinary medicine and cannot be reliably used for primary diagnosis of mammary neoplasms (Pinheiro et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Recent literature also highlights that CEA levels may be elevated in other non-mammary neoplastic conditions, such as ovarian cancer (Kim et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2024\u003c/span\u003e), reinforcing its lack of absolute specificity in canine oncology. The broader consensus indicates that no single ideal biomarker that meets all diagnostic and prognostic criteria has been identified in veterinary medicine, particularly due to the persistent lack of specificity (\u0026Ccedil;olakoğlu et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2025\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eAlthough some studies have evaluated CEA levels in healthy female dogs and those with mammary neoplasms, demonstrating its sensitivity and specificity for monitoring mammary tumors (Senhorello et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), no research has yet assessed CEA in male dogs or compared CEA levels with other non-neoplastic diseases in dogs. Given that infectious and neoplastic diseases comprise a significant portion of veterinary clinical cases, identifying potential coexisting conditions that may influence biomarker levels is crucial for accurately interpreting results (Vieira et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Fan et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In human medicine, the impact of various diseases on CEA levels is well understood, enabling a more precise evaluation of this biomarker in clinical practice (Pe\u0026ntilde;a et al. 2021). This highlights a critical gap in veterinary literature, as the behavior of CEA in the presence of common non-neoplastic conditions or across sexes in healthy animals remains largely unexplored, potentially impacting diagnostic accuracy and clinical interpretation.\u003c/p\u003e\u003cp\u003eThis study aimed to evaluate serum CEA levels in healthy dogs, dogs with mammary neoplasms, dogs with gastroenteritis of unknown origin, and dogs with ehrlichiosis. In addition to comparing mean values, we aimed to evaluate the diagnostic value of CEA across all disease groups, assess the impact of non-neoplastic diseases on its diagnostic accuracy, and investigate the correlation between CEA levels and hematological and biochemical variables in dogs with non-neoplastic diseases.\u003c/p\u003e\u003cp\u003eWe hypothesize that CEA levels will be elevated in both neoplastic and non-neoplastic diseases in dogs, mirroring observations in humans. If confirmed, non-neoplastic diseases may interfere with the specificity of CEA in dogs with mammary neoplasms.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cem\u003eEthical Approval\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThis study was submitted to and approved by the Animal Experimentation Ethics Committee (protocol no. 589-2021) at Vila Velha University, Esp\u0026iacute;rito Santo, Brazil, as well as the Animal Experimentation Ethics Committee of the Faculty of Agricultural and Veterinary Sciences at UNESP-Jaboticabal (protocol no. 10801/15). Owners who participated in the study received detailed guidance and signed an informed consent form. Additionally, a declaration verifying the origin of the samples was obtained to confirm their provenance when the clinical laboratories of the institutions provided serum samples. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eSample Collection \u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eSamples were obtained from animals treated at the Veterinary Hospital of UVV (Vila Velha, ES), Seres Veterinary Hospital (Vila Velha, ES), and VetSa\u0026uacute;de Veterinary Hospital (Vit\u0026oacute;ria, ES). These animals underwent clinical examinations at the request of the attending veterinarians. After confirming the diagnoses and reviewing the medical records, blood serum samples were selected and stored in the clinical pathology laboratory\u0026apos;s sample bank at the UVV Veterinary Hospital. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAll blood samples were centrifuged following standard procedures to separate the serum. The serum samples were individually aliquoted and stored at -20\u0026deg;C for further analysis of the CEA biomarker. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eAdditionally, some control group samples and samples from dogs with mammary neoplasms were obtained from a separate research project. The CEA measurements from these samples were incorporated into this study, as the same methodology was employed. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003eA total of 118 dogs were selected, with the mammary neoplasm group consisting solely of female dogs. \u0026nbsp;Animals were included regardless of breed or age. The collected biological material was categorized into five experimental groups:\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eGroup 1 (G1):\u003c/strong\u003e 29 serum aliquots from healthy dogs.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eGroup 2 (G2):\u003c/strong\u003e 56 serum aliquots from female dogs diagnosed with mammary carcinomas.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eGroup 3 (G3):\u003c/strong\u003e 17 serum aliquots from dogs diagnosed with ehrlichiosis.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eGroup 4 (G4):\u003c/strong\u003e 16 serum aliquots from dogs with gastroenteritis of unknown origin.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eGroup 5 (G5):\u003c/strong\u003e 33 serum aliquots from dogs with non-neoplastic diseases, comprising samples from G3 and G4. This group was specifically created to evaluate the combined impact of distinct non-neoplastic inflammatory and infectious conditions on CEA levels, allowing for a broader assessment of biomarker specificity.\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eAll animals underwent a comprehensive clinical evaluation, which included a physical examination, a complete blood count, and a serum biochemistry analysis. Dogs with mammary carcinoma (group 2) additionally had imaging studies (thoracic radiography and abdominal ultrasonography) for clinical staging based on the TNM classification according to the World Health Organization (WHO) guidelines. Definitive diagnosis was confirmed through histopathological analysis of tissue samples obtained following total, unilateral, or bilateral mastectomy, adhering to the established consensus guidelines for canine mammary tumors (Cassali et al. 2020).\u003c/p\u003e\n\u003cp\u003eDogs in Group 3 (ehrlichiosis) were diagnosed based on compatible clinical signs and laboratory findings, confirmed by the SNAP 4Dx test (IDEXX\u0026reg; Laboratories) for antibody detection and/or RT-PCR for detection of \u003cem\u003eEhrlichia canis\u003c/em\u003e or \u003cem\u003eEhrlichia ewingii\u003c/em\u003e antigens. For the purpose of this study, ehrlichiosis was considered as a systemic inflammatory non-neoplastic condition impacting CEA levels, without focusing on species-specific clinical or biochemical differentiations, which were beyond the scope of this investigation. Patients with gastroenteritis of unknown origin (group 3) were diagnosed based on clinical symptoms, exclusion of other possible causes, and ultrasonographic findings.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eHealthy dogs (group 1) were selected from routine consultations at the \u0026quot;Professor Ricardo Alexandre Rippler\u0026quot; Veterinary Hospital, UVV, Esp\u0026iacute;rito Santo, Brazil, and the \u0026quot;Governador Laudo Natel\u0026quot; Veterinary Hospital, UNESP, Jaboticabal, S\u0026atilde;o Paulo, Brazil. These animals were evaluated as part of elective neutering procedures and underwent a thorough physical examination, complete blood count, and serum biochemistry to confirm their health status.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eExclusion criteria included the presence of concurrent diseases unrelated to those listed in this study, incomplete clinical records, absence of diagnostic tests confirming disease status, and serum samples that were hemolyzed, lipemic, or icteric after centrifugation.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eSerological Marker\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eA human ELISA kit (AccuBind ELISA Kits, MONOBIND INC, catalog #1825-300) was used to determine serum CEA levels. The use of human-based assays for canine CEA determination is supported by previous studies, which demonstrate sufficient cross-reactivity and clinical utility for this biomarker in dogs (Campos et al. 2012; Pinheiro et al. 2022). The readings were performed using an ELISYS UNO Human analyzer, following the manufacturer\u0026apos;s recommended protocol and previously established methodology (Senhorello et al. 2020)\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eStatistical Analysis\u0026nbsp;\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eEach dog was considered an experimental unit for analyzing CEA levels and laboratory variables. The data were subjected to analysis of variance, and since they did not follow a normal distribution, the CEA values were transformed by raising them to the power of 0.25. When significant differences were detected in the F-test, means were compared using Tukey\u0026rsquo;s test. Specifically, hematological variables, including leukocytes, neutrophils, bands, lymphocytes, monocytes, eosinophils, and platelet counts, along with biochemical parameters such as creatinine, alanine aminotransferase (ALT), urea, and alkaline phosphatase, were correlated with CEA levels in the non-neoplastic disease groups using Pearson\u0026rsquo;s correlation coefficient.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe area under the curve (AUC) of the receiver operating characteristic (ROC) curve was utilized to assess the diagnostic value of the serum marker for all diseases. The cutoff point was determined by comparing the control group with dogs diagnosed with mammary carcinoma. Sensitivity and specificity were calculated for all groups. Crucially, to evaluate the impact of non-neoplastic diseases on CEA specificity, an additional sensitivity and specificity analysis was performed by comparing the mammary carcinoma group (G2) against the non-neoplastic disease groups (G3, G4, and the combined G5). This comparative approach aimed to determine the potential for false-positive results resulting from these confounding conditions. Analyses were conducted using GraphPad Prism v. 10.4 software with a significance level of 5%.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eA total of 118 animals participated in the study, consisting of 17 males (14.4%) and 101 females (85.6%), with ages ranging from 3 months to 15 years. No significant difference was found in CEA levels between males (0.89 \u0026plusmn; 0.84 ng/mL) and females (1.00 \u0026plusmn; 0.75 ng/mL) without mammary neoplasia (P = 0.6224).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMean CEA levels were notably highest in the mammary carcinoma group (G2; 1.94 ng/mL) and lowest in the control group (G1; 0.65 ng/mL). Significant elevations in CEA were observed in both the mammary carcinoma group (G2) compared to healthy controls (G1) (P \u0026lt; 0.0001), and importantly, in the non-neoplastic gastroenteritis group (G4) (p = 0.0237) and the combined non-neoplastic diseases group (G5) (P = 0.0120) when compared to healthy controls (G1). Furthermore, a significant difference was detected between the mammary carcinoma group (G2) and the combined non-neoplastic diseases group (G5) (P = 0.0466), indicating a distinct profile of CEA elevation in these conditions. No significant differences were noted among the other group comparisons (P \u0026gt; 0.05) (Table 1, Figure 1).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1 \u0026ndash;\u0026nbsp;\u003c/strong\u003eMean and standard deviation of CEA levels in experimental groups.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"3\" cellpadding=\"0\" width=\"564\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroup (n)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eCEA (ng/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003eG1 (29)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e0.65 (\u0026plusmn;0.32) a\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003eG2 (56)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e1.94 (\u0026plusmn;0.89) b\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003eG3 (17)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e1.29 (\u0026plusmn;1.06) abc\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003eG4 (16)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e1.50 (\u0026plusmn;0.98) bc\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 309px;\"\u003e\n \u003cp\u003eG5 (33)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 249px;\"\u003e\n \u003cp\u003e1.39 (\u0026plusmn;1.02) c\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cem\u003eCEA: Carcinoembryonic Antigen. Differing letters signify statistically significant differences between groups (P \u0026lt; 0.05); identical letters indicate no significant difference (P \u0026gt; 0.05).\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eThe diagnostic performance of serum CEA was evaluated using ROC curve analysis (Table 2). Employing an optimal cutoff value of 1.08 ng/mL, derived from the comparison between healthy controls (G1) and mammary carcinoma cases (G2), CEA demonstrated high sensitivity (82.14%) and specificity (93.10%) for detecting mammary carcinomas. However, when this established cutoff was applied to assess sensitivity across other groups, CEA\u0026apos;s performance was markedly lower for G4 (50.00%), G5 (48.48%), and G3 (47.06%). Crucially, to elucidate the interference of non-neoplastic diseases on CEA\u0026apos;s diagnostic utility for mammary carcinomas, we conducted a comparative analysis between the mammary carcinoma group (G2) and the non-neoplastic disease groups (G3, G4, and G5). While sensitivity for mammary carcinomas remained constant at 82.14% across these comparisons, specificity decreased dramatically to 52.94% for G3, 50.00% for G4, and 51.52% for G5. This stark reduction unequivocally demonstrates a significant compromise in CEA\u0026apos;s specificity in the presence of common non-neoplastic conditions, highlighting its limitations as a standalone biomarker in a broader clinical context.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2 \u0026ndash;\u0026nbsp;\u003c/strong\u003eSensitivity and specificity values for diseased animals compared to the control group and non-neoplastic disease groups compared to mammary carcinoma cases.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"3\" cellpadding=\"0\" width=\"567\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eGroup\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSensitivity (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eIC (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eSpecificity (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eIC (%)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAUC\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG1 vs. G2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e82.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e70.16-90.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e93.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e78.04-98.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e\u0026lt;0.0001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG1 vs. G3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e47.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e26.17-69.04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e93.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e78.04-98.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0298\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG1 vs. G4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e50.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e28.00-72.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e93.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e78.04-98.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0028\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG1 vs. G5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e48.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e32.50-64.78\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e93.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e78,04-98,77\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0018\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG3 vs. G2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e82.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e70.16-90.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e52.94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e30.96-73.83\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.71\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0065\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG4 vs. G2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e82.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e70.16-90.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e50.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e28.00-72.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0415\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003eG5 vs. G2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 102px;\"\u003e\n \u003cp\u003e82.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 74px;\"\u003e\n \u003cp\u003e70.16-90.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e51.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e35.22-67.50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 45px;\"\u003e\n \u003cp\u003e0.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 63px;\"\u003e\n \u003cp\u003e0.0023\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cem\u003eAUC: Area under the curve; IC: Confidence interval\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eAmong the hematological parameters evaluated, a significant moderate positive correlation was observed between platelet counts and CEA levels in both the ehrlichiosis group (G3; r = 0.68, P = 0.01) and the combined non-neoplastic diseases group (G5; r = 0.37, P = 0.01). No other significant correlations were found for leukocyte differentials (leukocytes, neutrophils, bands, lymphocytes, monocytes, or eosinophils) (P \u0026gt; 0.05). Furthermore, evaluation of correlations between CEA levels and serum biochemical parameters (creatinine, ALT, urea, and alkaline phosphatase) in animals with non-neoplastic diseases (n = 28) yielded no significant associations (P \u0026gt; 0.05) (Table 3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3 \u0026ndash;\u0026nbsp;\u003c/strong\u003ePearson correlation coefficients between CEA levels and hematological and biochemical parameters.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"3\" cellpadding=\"0\" width=\"564\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eG3\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eG4\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eG5\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eVariable\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003er\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003er\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003er\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eLeukocytes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.125\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.41\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.06\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eBands\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eNeutrophils\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.84\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.27\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eLymphocytes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.43\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eMonocytes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.06\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.82\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.39\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eEosinophils\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.66\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.65\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003ePlatelets\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e0.68\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e0.01\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e0.37\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e\u003cstrong\u003e0.01\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eRBCs\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eHemoglobin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eHematocrit\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.12\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eCreatinine\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.70\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eALT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.08\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.68\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eUrea\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.39\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd\u003e\n \u003cp\u003eAlkaline Phosphatase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e-0.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd\u003e\n \u003cp\u003e0.26\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cem\u003eALT: alanine aminotransferase\u0026nbsp;\u003c/em\u003e\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eCurrent studies on CEA in veterinary medicine primarily focus on its association with mammary neoplasms in female dogs (Senhorello et al. 2020; Fan et al. 2021; Jain et al. 2021). However, no studies have compared CEA levels between male and female dogs; existing research has only evaluated this biomarker in healthy females and those with mammary neoplasia (Ledecky et al. 2013; Senhorello et al. 2020; Fan et al. 2021).\u003c/p\u003e\n\u003cp\u003eIn the present study, serum CEA levels were assessed in both males and females, revealing no significant differences between the sexes. Similarly, human studies have reported no correlation between CEA levels and sex but rather with underlying diseases. These include neoplastic conditions such as gastric, colorectal, and breast carcinomas (Deng et al. 2015; Reiter et al. 2000; Wang et al. 2014), as well as non-neoplastic conditions such as inflammatory bowel disease, hepatitis, pancreatitis, and COVID-19 (Hao et al. 2019; Chen et al. 2021; Anjan et al. 2024).\u003c/p\u003e\n\u003cp\u003eThe highest mean CEA concentrations were found in female dogs with mammary neoplasms (G2) in comparison to healthy females (G1) and dogs with non-neoplastic diseases (G5). These findings confirm the potential of this biomarker for monitoring female dogs with mammary neoplasms, supporting the studies by Senhorello et al. (2020) and Fan et al. (2021), which noted elevated CEA levels in patients with mammary tumors. This observation is consistent with recent findings by Singh et al. (2025), who reported significantly higher serum CEA levels in canines with malignant mammary tumors compared to healthy controls. Furthermore, following mastectomy, the serum concentration of this biomarker significantly dropped and remained low in patients evaluated in follow-up assessments (Senhorello et al. 2020). However, it is crucial to recognize that despite its prognostic potential in mammary neoplasms, CEA alone exhibits limited specificity and may not be a standalone indicator for early diagnosis or definitive assessment, as also highlighted by Singh et al. (2025), who found that CEA alone was not highly specific and showed no significant difference between benign tumors and healthy controls\u003c/p\u003e\n\u003cp\u003eIn humans, elevated serum CEA concentrations are associated with both neoplastic and non-neoplastic diseases (Hao et al. 2019; Pe\u0026ntilde;a et al. 2021). Addressing a critical gap in veterinary literature, this study is the first to comprehensively investigate CEA levels in dogs affected by common non-neoplastic conditions. While no significant difference was observed between the control group (G1) and animals diagnosed solely with ehrlichiosis (G3), a significant elevation in mean CEA levels was noted in the gastroenteritis group (G4) and, notably, in the combined group of non-neoplastic diseases (G5) when compared to healthy controls (G1). This pivotal finding strongly suggests that CEA levels in dogs are not exclusively indicative of neoplastic processes and can be significantly influenced by inflammatory and infectious non-neoplastic conditions, mirroring observations in human medicine and contrasting with the absence of significant CEA elevation in benign mammary tumors reported by Singh et al. (2025). This difference highlights the diverse etiologies that can influence CEA concentrations in canines, underscoring the need for careful differential diagnosis.\u003c/p\u003e\n\u003cp\u003eOur results align with findings in human medicine, where elevated serum CEA levels have been reported in patients diagnosed with ulcerative colitis, inflammatory bowel disease, and other inflammation-associated conditions (Rule et al. 1973; Loewenstein et al. 1978; Hao et al. 2019; Kelleher et al. 2019). According to Tchoupa et al. (2014), carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) are part of a subfamily of immunoglobulin-related proteins that possess a high affinity for cellular adhesion to specific pathogens, such as Escherichia coli (Kelleher et al. 2019). Since this bacterium is one of the principal agents involved in the increased secretion of pro-inflammatory cytokines in dogs (Honneffer et al. 2014), it may explain the rise in CEA levels noted in gastroenteritis cases.\u003c/p\u003e\n\u003cp\u003eFurthermore, \u003cem\u003eEscherichia coli\u003c/em\u003e is the primary microorganism isolated from human patients with inflammatory bowel disease, further reinforcing this similarity (Tchoupa et al. 2014). Cellular adhesion can stimulate an innate immune response, activate specific kinases, promote the stimulation of small G proteins, and induce new gene expression events (Tchoupa et al. 2014). These processes can lead to cellular damage and inflammation, ultimately resulting in epithelial injury, which may contribute to tumorigenesis, tissue fibrosis, and increased serum CEA levels (Hao et al. 2019).\u003c/p\u003e\n\u003cp\u003eSimilarly, ehrlichiosis, a common hemoparasitic infection in dogs, can induce the gene expression of various cytokines, including interleukin-6 (IL-6), a key mediator of acute-phase responses (Gr\u0026ouml;ne et al. 1998; Brachelente et al. 2005). The known correlation between circulating CEA levels and IL-6 in human patients with certain cancers (Nakagoe and Sawai, 2003; Holmer et al. 2015) could provide a plausible explanation for the observed elevations in CEA in some dogs with ehrlichiosis. These specific findings, alongside those from gastroenteritis, highlight that common inflammatory and infectious conditions can indeed elevate CEA levels in canines, thus complicating the interpretation of this biomarker for cancer screening or monitoring. This highlights the importance of a comprehensive diagnostic approach, rather than relying solely on CEA as an indicator. Further studies are indeed necessary to fully elucidate these specific pathophysiological mechanisms.\u003c/p\u003e\n\u003cp\u003eIt is important to note that, in addition to evaluating whether non-neoplastic diseases can elevate CEA levels in dogs, this study plays a crucial role in the clinical application of CEA for animals diagnosed with mammary cancer, as previous research supports this use (Senhorello et al. 2020; Fan et al. 2021; Jain et al. 2021). However, we still do not fully comprehend which diseases may affect marker values during patient follow-up.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe optimal CEA cutoff point (1.08 ng/mL), established through a comparison between healthy controls and patients with mammary carcinoma, served as a crucial reference for evaluating the biomarker\u0026rsquo;s diagnostic utility. Our findings, consistent with previous research (Senhorello et al. 2020), demonstrated good sensitivity (82.14%) and specificity (93.10%) for CEA in detecting mammary carcinomas. However, it is noteworthy that even in studies confirming CEA elevation in malignant mammary tumors, the individual specificity of CEA has been reported as relatively low (Singh et al. 2025). This further underscores the importance of our investigation into confounding factors. Moreover, when this established cutoff was applied to assess sensitivity across other non-neoplastic groups, CEA\u0026apos;s performance was considerably lower, aligning with human studies on CEA in non-malignant conditions (Hao et al. 2019). This disparity in performance highlights the inherent challenge of biomarker interpretation in diverse clinical presentations, a concern further amplified by recent discussions on the general inspecificity of tumor biomarkers in veterinary medicine (\u0026Ccedil;olakoğlu et al. 2025).\u003c/p\u003e\n\u003cp\u003eCrucially, our study demonstrated that common non-neoplastic conditions significantly compromise CEA\u0026apos;s specificity for mammary carcinomas, to a greater extent than the inherent limitations observed in other studies (Singh et al. 2025). When CEA measurement was evaluated against non-neoplastic diseases (G3, G4, and G5), the specificity for mammary carcinomas decreased dramatically (to 52.94% for G3, 50.00% for G4, and 51.52% for G5). This stark reduction unequivocally indicates that non-neoplastic diseases, such as gastroenteritis and ehrlichiosis, significantly heighten the risk of false-positive CEA results for mammary carcinomas. Consequently, a patient with mammary tumors undergoing CEA-based follow-up, who subsequently develops one of these common non-neoplastic conditions, may exhibit altered CEA values directly attributable to the intercurrent disease, thereby severely undermining the reliability of the biomarker for neoplastic monitoring. This finding provides direct evidence supporting the necessity of considering confounding non-neoplastic conditions when interpreting CEA levels in canine patients, a previously unaddressed clinical challenge.\u0026quot;\u003c/p\u003e\n\u003cp\u003eThese findings align with those reported in human medicine, as CEA is a non-specific biomarker that can be elevated in non-cancerous conditions, including gastrointestinal diseases, inflammatory processes, and other disorders like renal and hepatic diseases, which may increase CEA levels even without neoplasia. As a result, CEA is not a reliable general tool for cancer detection, and the interpretation of results should take into account the patient\u0026apos;s comorbidities (Trape et al. 2011; Hao et al. 2019; Pe\u0026ntilde;a et al. 2021). Our study extends this crucial understanding to veterinary medicine, emphasizing that clinical context and thorough differential diagnostics are paramount for accurate CEA interpretation in dogs, particularly given the inherent limitations of single biomarkers as noted by Singh et al. (2025).\u003c/p\u003e\n\u003cp\u003eOur correlation analyses aimed to identify potential influences of hematological and biochemical changes, commonly associated with non-neoplastic diseases, on CEA levels. While most variables did not show significant correlations, a notable moderate positive correlation was identified between platelet counts and CEA levels in both the ehrlichiosis group (G4) and the combined non-neoplastic diseases group (G5). This finding provides indirect evidence of the intricate relationship between inflammatory processes and CEA elevation, supporting the broader understanding that CEA can be influenced by systemic inflammation. In inflammatory conditions, thrombocytosis can occur due to IL-6 production, which stimulates thrombopoietin production (Ceresa et al. 2007; Athanasiou et al. 2017). As IL-6 has been linked to increased CEA levels in humans (Nakagoe and Sawai, 2003; Holmer et al. 2015), our results suggest a similar underlying mechanism in dogs, where inflammation, mediated by cytokines like IL-6, could contribute to the observed CEA elevations. These correlations, particularly with platelets, further support the notion that CEA levels are not solely dictated by neoplastic disease, highlighting a crucial area for future mechanistic investigations in veterinary species. The findings are also consistent with the call from Singh et al. (2025) for more comprehensive studies on CEA\u0026apos;s diagnostic value, particularly given its lower individual specificity.\u003c/p\u003e\n\u003cp\u003eRegarding serum biochemical parameters (creatinine, ALT, urea, and alkaline phosphatase), no significant correlations were observed with CEA levels. This assessment was conducted in light of reports in human medicine linking elevated CEA levels to hepatic and renal diseases (Rule et al. 1973; Hao et al. 2019). While ehrlichiosis and gastroenteritis can induce systemic effects, they are not primarily defined by direct hepatic or renal dysfunction leading to consistent enzyme elevations. Thus, the limited number of animals exhibiting significant alterations in these specific parameters within our study groups might explain the lack of observed correlations. This finding suggests that CEA elevation in these specific non-neoplastic conditions may be more strongly associated with inflammatory processes (as indicated by the platelet correlation) rather than direct organ damage, as reflected by these biochemical markers. Future investigations involving a larger cohort of animals with confirmed hepatic or renal pathologies would be beneficial for fully elucidating these relationships.\u003c/p\u003e\n\u003cp\u003eDespite the significant contributions of this study, certain limitations should be acknowledged. Firstly, while novel data on CEA levels in ehrlichiosis and gastroenteritis were provided, the range of non-neoplastic conditions investigated was limited. Further research encompassing a broader spectrum of inflammatory, infectious, and systemic diseases would provide a more comprehensive understanding of CEA\u0026apos;s behavior in canine health and disease. Secondly, although our mechanistic discussion suggests the role of inflammatory mediators, such as interleukin-6 (IL-6), in CEA elevation, these were not directly measured in the study; thus, our explanations remain inferential. Future studies could incorporate direct cytokine quantification to solidify these proposed pathways. Lastly, although the overall sample size was robust, the individual sample sizes for specific non-neoplastic subgroups (G3 and G4) were relatively modest, and the inherent clinical heterogeneity within these conditions (e.g., varying severity or specific etiologies of gastroenteritis) could introduce variability. These considerations highlight avenues for future research aimed at further refining the diagnostic and prognostic utility of CEA in veterinary medicine.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study provides crucial insights into the behavior of serum Carcinoembryonic Antigen (CEA) in canine patients. CEA levels were confirmed to be significantly elevated in dogs with mammary carcinomas, highlighting their potential for disease monitoring. Importantly, no significant difference was observed in CEA levels between healthy male and female dogs, thereby expanding the scope for CEA\u0026apos;s clinical utility across both sexes. However, a key finding demonstrates that common non-neoplastic conditions, such as gastroenteritis and ehrlichiosis, also lead to a marked increase in CEA concentrations. This elevation significantly compromises CEA\u0026apos;s specificity for mammary neoplasms, thereby increasing the risk of false-positive results. Furthermore, a positive correlation was noted between CEA levels and platelet count, suggesting that inflammatory processes significantly influence the variability of this biomarker. Therefore, while CEA holds value in canine mammary tumor assessment, its accurate interpretation requires a comprehensive clinical evaluation of the patient, taking into account concurrent non-neoplastic conditions. Future research should expand this knowledge by investigating other confounding variables that may affect CEA values and prioritizing the development of multi-biomarker panels to enhance diagnostic accuracy and specificity in veterinary oncology.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was funded by the Funda\u0026ccedil;\u0026atilde;o de Amparo \u0026agrave; Pesquisa do Estado de S\u0026atilde;o Paulo (FAPESP), process number 2016/00128-5.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data supporting the conclusions of this study are available from the author upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of Interest Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that there is no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor Contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eL.W.A.L: Conceptualization, investigation, methodology, writing \u0026ndash; original draft. L.C.C: Investigation, methodology. A.C.T.B.F: Investigation, methodology. G.C.V: \u0026nbsp;Investigation, methodology. C.G: methodology, data analysis. F.A.R.S: Investigation, methodology, data analysis. R.P.S: Investigation, methodology, data analysis. M.T.C: Conceptualization, methodology, resources, funding acquisition, supervision, writing \u0026ndash; original draft. I.L.S.S: Conceptualization, data curation, investigation, methodology, data analysis, supervision, writing \u0026ndash; original draft. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eApproval was granted by the Animal Experimentation Ethics Committee at the Faculty of Agricultural and Veterinary Sciences of UNESP-Jaboticabal (protocol no. 10801/15) and the Committee on Animal Experimentation Ethics (protocol no. 589-2021) at Vila Velha University (UVV).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAnjan MAH, Ahmed QMU, Masum AA, Sami CA, Matin MA, Islam MS, Chowdhury FR, Arafat SM, Rahman M, Hasan MN (2024) Role of Carcinoembryonic Antigen in Severity Assessment and Mortality Prediction in COVID-19 Patients. Cureus 16(9):e69894. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.7759/cureus.69894\u003c/span\u003e\u003cspan address=\"10.7759/cureus.69894\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAthanasiou LV, Polizopoulou ZS, Papavasileiou EG, Mpairamoglou EL, Kantere MC, Rousou XA (2017) Magnitude of reactive thrombocytosis and associated clinical conditions in dogs. Vet Rec 181(10):267. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1136/vr.104042\u003c/span\u003e\u003cspan address=\"10.1136/vr.104042\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ede la Ayala F, Ortiz-Mu\u0026ntilde;oz B, Quintanar-Verd\u0026uacute;guez T et al (2021) Consensus of the Spanish society of laboratory medicine and the Spanish society of medical oncology on the methodology and criteria for evaluation of circulating tumour markers in breast cancer. Clin Transl Oncol 23(7):1272\u0026ndash;1280. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s12094-020-02529-x\u003c/span\u003e\u003cspan address=\"10.1007/s12094-020-02529-x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBrachelente C, M\u0026uuml;ller N, Doherr MG, Sattler U, Welle M (2005) Cutaneous leishmaniasis in naturally infected dogs is associated with a T helper-2-biased immune response. Vet pathol 42(2):166\u0026ndash;175. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1354/vp.42-2-166\u003c/span\u003e\u003cspan address=\"10.1354/vp.42-2-166\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCampos LC, Lavalle GE, Estrela-Lima A, Melga\u0026ccedil;o de Faria JC, Guimar\u0026atilde;es JE, Dutra \u0026Aacute;P, Ferreira E, de Sousa LP, Rabelo \u0026Eacute;M, Vieira da, Costa AF, Cassali GD (2012) CA15.3, CEA and LDH in dogs with malignant mammary tumors. J Vet Intern Med 26(6):1383\u0026ndash;1388. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1111/j.1939-1676.2012.01014.x\u003c/span\u003e\u003cspan address=\"10.1111/j.1939-1676.2012.01014.x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCarlos RS, Muniz Neta ES, Spagnol FH, Oliveira LL, de Brito RL, Albuquerque GR, Almosny NR (2007) Frequency of antibodies anti-Ehrlichia canis, Borrelia burgdorferi and Dirofilaria immitis antigens in dogs from microrregion Ilh\u0026eacute;us-Itabuna, State of Bahia. Brazil Rev Bras Parasitol Vet 16(3):117\u0026ndash;120. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1590/s1984-29612007000300001\u003c/span\u003e\u003cspan address=\"10.1590/s1984-29612007000300001\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCassali GD, Jark PC, Gamba C et al (2020) Consensus Regarding the Diagnosis, Prognosis and Treatment of Canine and Feline Mammary Tumors \u0026ndash; 2019. Braz J Vet Pathol 13(3):555\u0026ndash;574. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.24070/bjvp.1983-0246.v13i3p555-574\u003c/span\u003e\u003cspan address=\"10.24070/bjvp.1983-0246.v13i3p555-574\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCeresa IF, Noris P, Ambaglio C, Pecci A, Balduini CL (2007) Thrombopoietin is not uniquely responsible for thrombocytosis in inflammatory disorders. Platelets 18(8):579\u0026ndash;582. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1080/09537100701593601\u003c/span\u003e\u003cspan address=\"10.1080/09537100701593601\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eChen Q, Kong H, Qi X, Ding W, Ji N, Wu C, Huang C, Wu W, Huang M, Xie W, Liu Y, Tang J (2020) Carcinoembryonic Antigen: A Potential Biomarker to Evaluate the Severity and Prognosis of COVID-19. Front Med 7:579543. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3389/fmed.2020.579543\u003c/span\u003e\u003cspan address=\"10.3389/fmed.2020.579543\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003e\u0026Ccedil;olakoğlu HE, Fidan \u0026Ccedil;, Al\u0026ccedil;ığır ME, G\u0026ouml;bek\u0026ccedil;i AE, Devrim E, Anteplioğlu T, Yazlık MO, \u0026Ouml;zmen \u0026Ouml;, Re\u0026ccedil;ber T, Nemutlu E, Kaya U (2025) Evaluation of serum CA15-3, CEA, Ki-67, PDL-1 and VEGF as a potential diagnostic and prognostic biomarkers in canine mammary tumors. Theriogenology 247:117579. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.theriogenology.2025.117579\u003c/span\u003e\u003cspan address=\"10.1016/j.theriogenology.2025.117579\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eDeng K, Yang L, Hu B, Wu H, Zhu H, Tang C (2015) The prognostic significance of pretreatment serum CEA levels in gastric cancer: a meta-analysis including 14651 patients. PLoS ONE 10(4):e0124151. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1371/journal.pone.0124151\u003c/span\u003e\u003cspan address=\"10.1371/journal.pone.0124151\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eEgawa K, Honda Y, Kuroki M, Inaba Y, Ono T (1996) Carcinoembryonic antigen and related antigens expressed on keratinocytes in inflammatory dermatoses. Br J Dermatol 134(3):451\u0026ndash;459\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFan Y, Ren X, Liu X, Shi D, Xu E, Wang S, Liu Y (2021) Combined detection of CA15-3, CEA, and SF in serum and tissue of canine mammary gland tumor patients. Sci Rep 11(1):6651. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1038/s41598-021-85029-4\u003c/span\u003e\u003cspan address=\"10.1038/s41598-021-85029-4\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eGr\u0026ouml;ne A, Frisk AL, Baumg\u0026auml;rtner W (1998) Cytokine mRNA expression in whole blood samples from dogs with natural canine distemper virus infection. Vet Immunol Immunopathol 65(1):11\u0026ndash;27. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/s0165-2427(98)00170-6\u003c/span\u003e\u003cspan address=\"10.1016/s0165-2427(98)00170-6\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHall C, Clarke L, Pal A, Buchwald P, Eglinton T, Wakeman C, Frizelle F (2019) A Review of the Role of Carcinoembryonic Antigen in Clinical Practice. Ann Coloproctol 35(6):294\u0026ndash;305. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3393/ac.2019.11.13\u003c/span\u003e\u003cspan address=\"10.3393/ac.2019.11.13\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHao C, Zhang G, Zhang L (2019) Serum CEA levels in 49 different types of cancer and noncancer diseases. Prog Mol Biol Transl Sci 162:213\u0026ndash;227. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/bs.pmbts.2018.12.011\u003c/span\u003e\u003cspan address=\"10.1016/bs.pmbts.2018.12.011\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHolmer R, W\u0026auml;tzig GH, Tiwari S, Rose-John S, Kalthoff H (2015) Interleukin-6 trans-signaling increases the expression of carcinoembryonic antigen-related cell adhesion molecules 5 and 6 in colorectal cancer cells. BMC Cancer 15:975. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1186/s12885-015-1950-1\u003c/span\u003e\u003cspan address=\"10.1186/s12885-015-1950-1\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHonneffer JB, Minamoto Y, Suchodolski JS (2014) Microbiota alterations in acute and chronic gastrointestinal inflammation of cats and dogs. World J Gastroenterol 20(44):16489\u0026ndash;16497. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3748/wjg.v20.i44.16489\u003c/span\u003e\u003cspan address=\"10.3748/wjg.v20.i44.16489\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eJain M, Ingole SD, Deshmukh RS, Bharucha SV, Nagvekar AS, Gaikwad RV, Kharde SD (2021) CEA, CA 15\u0026thinsp;\u0026ndash;\u0026thinsp;3, and miRNA expression as potential biomarkers in canine mammary tumors. Chromosome Res 29(2):175\u0026ndash;188. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1007/s10577-021-09652-7\u003c/span\u003e\u003cspan address=\"10.1007/s10577-021-09652-7\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKelleher M, Singh R, O'Driscoll CM, Melgar S (2019) Carcinoembryonic antigen (CEACAM) family members and Inflammatory Bowel Disease. Cytokine Growth Factor Rev 47:21\u0026ndash;31. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.cytogfr.2019.05.008\u003c/span\u003e\u003cspan address=\"10.1016/j.cytogfr.2019.05.008\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKim S-H, Yoo DS, Park D-H, Cho S-S, Park S, Jung B-G, Park S-I, Bae C-S (2024) Combined Application of CEA, CA 15\u0026thinsp;\u0026ndash;\u0026thinsp;3, and CA 27\u0026ndash;29 for the Evaluation of Diagnostic Performance in Canine Mammary Gland Tumors. J Veterinary Clin 41:359\u0026ndash;369. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.17555/jvc.2024.41.6.359\u003c/span\u003e\u003cspan address=\"10.17555/jvc.2024.41.6.359\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKwon YJ, Lee HS, Shim JY, Lee YJ (2018) Serum carcinoembryonic antigen is positively associated with leukocyte count in Korean adults. J Clin Lab Anal 32(3):e22291. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1002/jcla.22291\u003c/span\u003e\u003cspan address=\"10.1002/jcla.22291\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLedecky V, Valencakova-Agyagosova A, Lepej J, Frischova Z, Hornak S, Nagy V (2013) Determination of carcinoembryonic antigen and cancer antigen values with the radioimmunoassay method in healthy females dogs. Vet Med 58(5):277\u0026ndash;283. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.17221/6810-VETMED\u003c/span\u003e\u003cspan address=\"10.17221/6810-VETMED\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLoewenstein MS, Zamcheck N (1978) Carcinoembryonic antigen (CEA) levels in benign gastrointestinal disease states. Cancer 42(3 Suppl):1412\u0026ndash;1418. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1002/1097-0142(197809)42:3+\u0026lt;1412::aid-cncr2820420805\u0026gt;3.0.co;2-8\u003c/span\u003e\u003cspan address=\"10.1002/1097-0142(197809)42:3+%3C1412::aid-cncr2820420805%3E3.0.co;2-8\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eNakagoe T, Tsuji T, Sawai T, Tanaka K, Hidaka S, Shibasaki S, Nanashima A, Yamaguchi H, Yasutake T, Sugawara K, Inokuchi N, Kamihira S (2003) The relationship between circulating interleukin-6 and carcinoembryonic antigen in patients with colorectal cancer. Anticancer Res 23(4):3561\u0026ndash;3564\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePaniz C, Grotto D, Schmitt GC et al (2005) Physiopathology of vitamin B12 deficiency and its laboratorial diagnosis. J Bras Patol Med Lab 41:323\u0026ndash;334. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1590/S1676-24442005000500007\u003c/span\u003e\u003cspan address=\"10.1590/S1676-24442005000500007\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePinheiro BQ, da Silva IN, Faustino AM, Machado da Silva LD (2022) The prognostic value of serum neoplastic biomarkers CA 15\u0026thinsp;\u0026ndash;\u0026thinsp;3 and CEA in canine mammary neoplasms: a review. Revista Brasileira de Reprodu\u0026ccedil;\u0026atilde;o Anim 46(3):290\u0026ndash;297. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.21451/1809-3000.RBRA2022.021\u003c/span\u003e\u003cspan address=\"10.21451/1809-3000.RBRA2022.021\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eReiter W, Stieber P, Reuter C, Nagel D, Lau-Werner U, Lamerz R (2000) Multivariate analysis of the prognostic value of CEA and CA 19\u0026thinsp;\u0026ndash;\u0026thinsp;9 serum levels in colorectal cancer. Anticancer Res 20(6D):5195\u0026ndash;5198\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRueda JR, Porto CD, Franco RP, da Costa IB, Bueno LMC, Girio RJS, Manhoso FFR, Bueno PCDS, Repetti CSF (2024) Mammary neoplasms in female dogs: Clinical, diagnostic and therapeutic aspects. Vet Med 69(4):99\u0026ndash;114. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.17221/4/2024-VETMED\u003c/span\u003e\u003cspan address=\"10.17221/4/2024-VETMED\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRule AH, Goleski-Reilly C, Sachar DB, Vandevoorde J, Janowitz HD (1973) Circulating carcinoembryonic antigen (CEA): relationship to clinical status of patients with inflammatory bowel disease. Gut 14(11):880\u0026ndash;884. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1136/gut.14.11.880\u003c/span\u003e\u003cspan address=\"10.1136/gut.14.11.880\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSenhorello ILS, Terra EM, Sueiro FAR, Firmo BF, Anai LA, Goloni C, Canavari IC, Ampuero RAN, Pereira RS, Tinucci-Costa M (2020) Clinical value of carcinoembryonic antigen in mammary neoplasms of bitches. Vet Comp Oncol 18(3):315\u0026ndash;323. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1111/vco.12552\u003c/span\u003e\u003cspan address=\"10.1111/vco.12552\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSenhorello ILS, Terra EM, Sueiro FAR, Pereira RS, Firmo BF, Martinelli C, Tinucci-Costa M (2024) Clinical value of alpha-fetoprotein in the detection of mammary carcinoma in female dogs. Arq Bras Med Vet Zootec 76(5):e13231. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1590/1678-4162-13231\u003c/span\u003e\u003cspan address=\"10.1590/1678-4162-13231\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSingh D, Rokade PP, Gangwar NK, Gabhane MG, Malik S, Gangwar K, Prabhu SN, Singh R, Singh DD, Kumari S, Chaudhary S, Choudhary JK (2025) Diagnostic value of carcinoembryonic antigen, cancer antigen 15\u0026thinsp;\u0026ndash;\u0026thinsp;3, and cell-free DNA as blood biomarkers in early detection of canine mammary tumor. J Circulating Biomarkers 14:30\u0026ndash;38. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.33393/jcb.2025.3564\u003c/span\u003e\u003cspan address=\"10.33393/jcb.2025.3564\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSoares EDS, Valente FL, Rocha CC, Real Pereira CE, Sarandy TB, de Oliveira FLD, de Morais Calado SL, Borges APB (2023) Prognostic Factors for Cancer-Specific Survival and Disease-Free Interval of Dogs with Mammary Carcinomas. Vet Med Int 2023:6890707. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1155/2023/6890707\u003c/span\u003e\u003cspan address=\"10.1155/2023/6890707\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTchoupa AK, Schuhmacher T, Hauck CR (2014) Signaling by epithelial members of the CEACAM family - mucosal docking sites for pathogenic bacteria. Cell Commun Signal 12:27. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1186/1478-811X-12-27\u003c/span\u003e\u003cspan address=\"10.1186/1478-811X-12-27\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTrap\u0026eacute; J, Filella X, Alsina-Donadeu M, Juan-Pereira L, Bosch-Ferrer \u0026Aacute;, Rigo-Bonnin R, Oncology Section of the Catalan Association of Clinical Laboratory Science (2011) Increased plasma concentrations of tumour markers in the absence of neoplasia. Clin Chem Lab Med 49(10):1605\u0026ndash;1620. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1515/CCLM.2011.694\u003c/span\u003e\u003cspan address=\"10.1515/CCLM.2011.694\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eVieira FT, Acosta ICL, Martins TF, Filho JM, Krawczak FDS, Barbieri ARM, Egert L, Fernandes DR, Braga FR, Labruna MB, Dietze R (2018) Tick-borne infections in dogs and horses in the state of Esp\u0026iacute;rito Santo, Southeast Brazil. Vet Parasitol 249:43\u0026ndash;48. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.vetpar.2017.11.005\u003c/span\u003e\u003cspan address=\"10.1016/j.vetpar.2017.11.005\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWang G, Qin Y, Zhang J, Zhao J, Liang Y, Zhang Z, Qin M, Sun Y (2014) Nipple discharge of CA15-3, CA125, CEA and TSGF as a new biomarker panel for breast cancer. Int J Mol Sci 15(6):9546\u0026ndash;9565. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.3390/ijms15069546\u003c/span\u003e\u003cspan address=\"10.3390/ijms15069546\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eWoolcock AD, Keenan A, Cheung C, Christian JA, Moore GE (2017) Thrombocytosis in 715 Dogs (2011\u0026ndash;2015). J Vet Intern Med 31(6):1691\u0026ndash;1699. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1111/jvim.14831\u003c/span\u003e\u003cspan address=\"10.1111/jvim.14831\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eZhou H, Fuks A, Alcaraz G, Bolling TJ, Stanners CP (1993) Homophilic adhesion between Ig superfamily carcinoembryonic antigen molecules involves double reciprocal bonds. J Cell Biol 122(4):951\u0026ndash;960. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1083/jcb.122.4.951\u003c/span\u003e\u003cspan address=\"10.1083/jcb.122.4.951\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"mammary tumor, ehrlichiosis, gastroenteritis, CEA, biomarker","lastPublishedDoi":"10.21203/rs.3.rs-7481550/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7481550/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eCarcinoembryonic antigen (CEA) is a glycoprotein produced by normal mucosal cells of the gastrointestinal tract and has been utilized as a tumor biomarker in humans. In veterinary medicine, canine patients with mammary neoplasms present elevated CEA levels. However, CEA is also known to increase in other pathological conditions. This study compared serum CEA levels in dogs with mammary carcinoma and those with non-neoplastic diseases. Blood serum samples were collected from 118 dogs (101 females and 17 males) and were divided into five experimental groups: control (G1), mammary neoplasia (G2), ehrlichiosis (G3), gastroenteritis (G4), and non-neoplastic diseases (G5). CEA levels were measured using a human ELISA kit, and the results were analyzed and compared across the experimental groups. Statistical analyses included Tukey's test, ROC curve analysis, and Pearson\u0026rsquo;s correlation, with a significance level of 5%. The results demonstrated significant differences in mean CEA levels between groups G1 and G2 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001), G1 and G4 (P\u0026thinsp;=\u0026thinsp;0.0267), and G1 and G5 (P\u0026thinsp;=\u0026thinsp;0.0070), with elevated values in mammary carcinomas and non-neoplastic diseases. Greater sensitivity and specificity were noted in G2 compared to the other groups. However, specificity significantly dropped when non-neoplastic diseases were compared to mammary carcinomas. Furthermore, a moderate positive correlation was found between CEA levels and platelet count, indicating a possible influence of inflammation on the variation of this biomarker. The results demonstrated that CEA has good diagnostic value for dogs with mammary carcinomas; however, this was not observed for non-neoplastic diseases. However, non-neoplastic diseases can drastically affect CEA specificity. Future studies evaluating additional diseases will be necessary for a better understanding and clinical application of this biomarker in veterinary practice.\u003c/p\u003e","manuscriptTitle":"Diagnostic Specificity of Serum Carcinoembryonic Antigen for Canine Mammary Neoplasms: The Impact of Non-Neoplastic Diseases","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-01 10:42:26","doi":"10.21203/rs.3.rs-7481550/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"f87f09a8-e005-4ef8-a073-3df1a7cf9a7e","owner":[],"postedDate":"September 1st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-18T20:23:09+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-01 10:42:26","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7481550","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7481550","identity":"rs-7481550","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}