Real-world pharmacokinetics of ramucirumab in combination with erlotinib and docetaxel for advanced non-small cell lung cancer: a prospective cohort study

Real-world pharmacokinetics of ramucirumab in combination with erlotinib and docetaxel for advanced non-small cell lung cancer: a prospective cohort study | 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 Research Article Real-world pharmacokinetics of ramucirumab in combination with erlotinib and docetaxel for advanced non-small cell lung cancer: a prospective cohort study Kazumasa Akagi, Shinnosuke Takemoto, Mayu Ohuchi, Shigehiro Yagishita, and 13 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8482971/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 Purpose In pivotal phase III studies of non-small cell lung cancer (NSCLC), ramucirumab (RAM) serum concentrations were associated with clinical outcomes in docetaxel (DTX) plus RAM treatment (DTX group), but not in erlotinib (ERL) plus RAM treatment (ERL group). This study prospectively examined the relationship between serum RAM concentrations and clinical outcomes in a real-world setting. Methods This study included patients with advanced NSCLC who received RAM in combination with DTX or ERL. Blood samples were obtained at trough levels before RAM administration. The day 1 sample of the second cycle was defined as C trough and measured using liquid chromatography–tandem mass spectrometry. The associations between C trough and clinical outcomes were evaluated. Results C trough ranged from 4.3–36.4 µg/mL in the DTX group and 10.2–44.5 µg/mL in the ERL group. Edema was more commonly observed in the high C trough group than in the DTX group. C trough was not associated with prognosis in the DTX group, while the median progression-free survival (PFS) was significantly longer in high C trough (8.3 vs 16.4 months, p = 0.032) within the ERL group. The median overall survival was significantly longer in low modified Glasgow Prognostic Score (mGPS-low) (32.8 vs 13.6 months, p = 0.042) within the DTX group, and PFS was significantly longer in mGPS-low (18.2 vs 6.1 months, p = 0.004) within the ERL group. Conclusion RAM C trough may influence survival after ERL + RAM treatment. The mGPS appears to be a potential prognostic factor in patients with NSCLC treated with RAM. Ramucirumab Pharmacokinetics Concentration Erlotinib Docetaxel Non-small cell lung cancer Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 1. Introduction Ramucirumab (RAM) is a human IgG1 monoclonal antibody that targets human vascular endothelial growth factor receptor-2 (VEGFR-2). RAM demonstrates anti-angiogenic and anti-tumor activities through the inhibition of vascular endothelial growth factor (VEGF) binding to VEGFR-2[ 1 ]. RAM has been approved for use in combination with docetaxel (DTX) or erlotinib (ERL) in lung cancer. The REVEL trial demonstrated that DTX + RAM improved survival as a second-line treatment for patients with stage IV non-small cell lung cancer (NSCLC) and remains an important therapeutic option beyond second-line therapy[ 2 ]. The RELAY trial showed that ERL + RAM demonstrated superior progression-free survival (PFS) compared to placebo plus ERL in patients with untreated epithelial growth factor receptor (EGFR)-mutated metastatic NSCLC[ 3 ]. Recently, EGFR-mutant lung cancer has undergone a paradigm shift in treatment, with regimens such as FLAURA2[ 4 ] and MARIPOSA [ 5 ] becoming standard therapies. However, survival analysis from the RELAY trial suggested that ERL + RAM may be an effective treatment option, especially for patients with EGFR L858R mutations[ 6 ]. Furthermore, recent positive trials combining RAM with osimertinib[ 7 ] have indicated that RAM remains an important therapeutic option. In the population pharmacokinetics (PK) analysis of the REVEL trial, higher RAM exposure was associated with improved clinical outcomes[ 8 ]. Conversely, PPK analysis of the RELAY trial showed no association between RAM exposure and response[ 9 ]. A system was established to measure serum concentrations of RAM using liquid chromatography–tandem mass spectrometry (LC-MS/MS), and trough levels (C trough ) were analyzed after the first administration of RAM in patients with advanced or recurrent NSCLC, together with clinical information. These results suggest that patients with a higher C trough tended to have a better prognosis. Conversely, patients with lower C trough levels include a greater proportion of cases with cachexia, indicating that accelerated clearance of RAM in cachexia may reduce therapeutic efficacy[ 10 ]. However, since this was a single-center retrospective study, a prospective, multicenter study was initiated to validate these findings in a real-world setting, including not only DTX plus RAM, but also ERL plus RAM, which has not been previously reported. 2. Methods 2.1. Study design This was a multicenter prospective cohort study. Eligible participants were patients with advanced or recurrent NSCLC who received RAM treatment in combination with DTX or ERL at Nagasaki University Hospital or a collaborating research facility between June 22, 2021, and August 31, 2023. Participants were expected to undergo follow-up for at least 90 days from the start of treatment, and were aged ≥ 20 years at the time of consent. Blood sampling was performed at the trough level immediately before RAM administration. Blood samples were collected on day 1 for each of the first four treatment cycles, starting with the initial administration of RAM, designated as day 1. Samples collected on day 1 of the second treatment cycle were defined as the C trough . 2.2. Data collection The following clinical information was prospectively collected: date of birth, sex, performance status at the time closest to the initiation of RAM, height and weight, smoking history, medical history and comorbidities, and prior treatment information for the primary disease. Imaging data, including chest radiography, chest and abdominal computed tomography (CT) scans, brain CT or magnetic resonance imaging, positron emission tomography-CT, and bone scintigraphy, were reviewed. Clinical assessments and laboratory tests were reviewed, including subjective and objective symptoms and any other grade 3 or higher adverse events. Edema and proteinuria were identified when they appeared or worsened, respectively, during the clinical course. Laboratory data included peripheral blood count, blood biochemistry, urinalysis, and oxygen saturation. Pathological data included tumor histology and gene alterations, EGFR mutations, anaplastic lymphoma kinase and c-ros oncogene 1 rearrangements, and programmed death-ligand 1 (PD-L1) tumor proportion score. Efficacy was assessed in accordance with the Response Evaluation Criteria in Solid Tumors version 1.1[ 11 ], and adverse events were assessed using the National Cancer Institute Common Terminology Criteria for Adverse Events version 5.0[ 12 ]. PFS was defined as the time from the start date of the RAM treatment to disease progression or death from any cause, whereas overall survival (OS) was defined as the time from the start date of the RAM treatment to the last day confirmed as alive or dead from any cause. Cachexia was defined as either a weight loss of ≥ 5% within 6 months before treatment initiation, or a weight loss of ≥ 2% within 2 months, accompanied by a body mass index 0.5 mg/dL or albumin 0.5 mg/dL and albumin < 3.5 g/dL[ 14 ]. 2.3. Measurement of serum RAM concentration At the time of blood sampling immediately before RAM administration, 6 mL of blood was collected in a serum separation tube. After incubation at room temperature for 30 min, the sample was centrifuged at 2000 rpm for 10 min at room temperature to separate the serum. One milliliter of serum was transferred into a cryotube and stored at − 80°C. If storage at − 80°C was not feasible at a given facility, − 20°C storage was permitted. The stored serum samples were transported frozen to the National Cancer Center Research Institute, where serum RAM concentrations were measured using LC-MS/MS coupled with the nSMOL method, as previously reported[ 10 ]. If treatment was delayed or discontinued owing to adverse events at the scheduled C trough sampling point, the blood test conducted at that time was used as the trough value, provided that blood sampling had been performed. 2.4. Ethical considerations and registration Written informed consent was obtained from all enrolled patients, and the study was conducted in accordance with the Declaration of Helsinki and Good Clinical Practice Guidelines. The study protocol was reviewed and approved by the Clinical Review Board of Nagasaki University (CRB7180001) on June 22, 2021 (approval no. 21062121). 2.5. Statistical analysis Continuous variables were presented as means and medians for parametric and nonparametric variables, respectively. The Fisher’s exact test was used for group comparisons of categorical variables. Student’s t -test was used to compare parametric continuous variables between groups, whereas the Mann–Whitney U test was used to compare nonparametric continuous variables between groups. Univariate and logistic regression analyses were used to assess continuous and categorical variables, respectively. Cox univariate regression analysis was used to assess survival, and the log-rank test was used for intergroup comparisons. Statistical significance was set at p < 0.05. The proportional hazard assumption was assessed using Schoenfeld residuals, and it was confirmed that no violations were observed. Analyses were performed using R (version 4.5.0) and GraphPad Prism (version 9.5.1; GraphPad Software, USA). 2.6. Data statement Additional data are available upon request from the corresponding author. 3. Results 3.1. Patient characteristics In the DTX group, the median age was 64 years, and half of the patients had EGFR mutations. Eleven patients (68.8%) had received two or more prior treatment regimens, including immune checkpoint inhibitors (ICI) in eight patients (50.0%) and EGFR-tyrosine inhibitors (TKIs) in six patients (37.5%). Pleural effusion and edema were observed in six patients (37.5%). Six patients (37.5%) had an mGPS of 0, while 10 patients (62.5%) had an mGPS of 1 or 2. Cachexia was present in only two patients (12.5%). The median age of patients in the ERL group was 73 years. EGFR mutations included exon 19 deletion (del19) in three patients (37.5%) and L858R in five patients (62.5%). Four (50.0 %) patients had a history of EGFR-TKI treatment. Pleural effusion was observed in six patients (75.0%) and edema in two patients (25.0%). Five patients (62.5%) had an mGPS of 0, and three patients (37.5%) had an mGPS of 1 or 2. Cachexia was present in two patients (25.0%) (Table 1). Divided into Q1 and Q2–4, the DTX group showed six cases (54.5%) with EGFR mutations, including insertion in 20, and five cases (45.5%) with PD‑L1 negativity in Q2–4 (Table S1). In the ERL group, all patients in Q2–4 had an mGPS of 1, whereas in Q1, five patients (83.3%) had an mGPS of 0 (Table S2). 3.2. Pharmacokinetics of RAM Trough blood sampling was performed in 23 patients at the start of cycle 1, 23 at cycle 2, 18 at cycle 3, and 14 at cycle 4 (Fig. 1A). In one case in the DTX group, C trough sampling was missing, and in another case, the pre-dose concentration was missing. Additionally, there was one case in which ERL + RAM was administered 35 days after treatment with DTX + RAM, in which a detectable pre-dose RAM concentration was observed. Under the above conditions, C trough in the DTX group ranged from 4.3–36.4 μg/mL, and the median was 15.3 μg/mL. C trough was sorted from Q1–Q4 by interquartile range (4.3 ≤ Q1 ≤ 10.7, 10.7 < Q2 ≤ 15.3, 15.3 < Q3 ≤ 18.1 and 18.1 < Q4 ≤ 36.4 μg/mL. Meanwhile, C trough in the ERL group ranged from 10.2–44.5 μg/mL, and the median was 26.2 μg/mL. C trough was sorted from Q1–Q4 by interquartile range (10.2 ≤ Q1 ≤ 24.3, 24.3 < Q2 ≤ 26.2, 26.2 < Q3 ≤ 37.4, and 37.4 < Q4 ≤ 44.5 μg/mL (Fig. 1B). C trough was significantly higher in the ERL group than in the DTX group after excluding one overlapping case between the two groups from the analysis ( p =0.0047, t-test) (Fig. 1C). 3.3. RAM C trough and adverse events The relationship between C trough and the presence or absence of RAM-specific adverse events, namely, bleeding, edema, hypertension, febrile neutropenia (FN), and proteinuria, was evaluated. In the DTX group, patients with edema exhibited higher C trough levels, whereas no association was observed between C trough and other adverse events. In the ERL group, no association was found between the C trough and any adverse events (Fig.2). 3.4. RAM C trough, mGPS and efficacy The association between RAM efficacy and C trough (Q1 vs. Q2–4) and mGPS (mGPS=0; mGPS-low vs. mGPS=1 or 2; mGPS-high) was analyzed. In the DTX and ERL groups, patients in the Q2–4 group tended to have a higher objective response rate (ORR) (Fig. 3A). Next, log-rank tests were performed to evaluate the survival outcomes. In the DTX group, no association was observed between C trough and PFS or OS. The median PFS was 6.7 months in the mGPS-low and 4.2 months in the mGPS-high, which was not significantly different. The Kaplan–Meier curve indicated a trend toward longer PFS in the mGPS-low group. The median OS was 32.8 months in mGPS-low and 13.6 months in mGPS-high, respectively, and was significantly longer in mGPS-low ( p = 0.042, log-rank test) (Fig.3B). In the ERL group, the median PFS was 8.3 months in Q1 and 16.4 months in the Q2-4 group, respectively, and was significantly longer in the Q2-4 group ( p = 0.032, log-rank test). The median OS was 16.6 months in the Q1 group and was not reached in the Q2-4 group, which was not significantly different. The Kaplan–Meier curve indicated a trend toward longer OS in Q2-4. The median PFS was 18.2 months in the mGPS-low and 6.1 months in the mGPS-high groups, respectively, and was significantly longer in the mGPS-low ( p = 0.004, log-rank test). The median OS was not reached in the mGPS-low and 16.6 months in the mGPS-high group, which was not significantly different. The Kaplan–Meier curve indicated a trend toward longer PFS in the mGPS-low group (Fig.3C). Subsequently, univariate analysis was performed to identify the factors influencing survival outcomes. In the DTX group, the best response to prior treatment was significantly associated with PFS and OS (HR: 0.27, 95% CI: 0.08–0.91, p = 0.035 for PFS; HR: 0.11, 95% CI: 0.02–0.55, p = 0.007 for OS), whereas C trough , cachexia, and mGPS were not associated with prognosis (Fig. 4A). In the ERL group, the presence of pleural effusion and C trough showed a trend toward association with PFS and OS, although the differences were not statistically significant (HR: 0.11, 95% CI: 0.01–1.24, p = 0.074 for PFS and OS) (Fig. 4B). 3.6. Factors influencing RAM C trough To identify factors influencing C trough , univariate analyses were conducted separately for the DTX and ERL groups. In the DTX group, a history of ICI treatment was associated with lower serum concentrations, whereas edema was associated with higher concentrations. In the ERL group, a high mGPS contributed to lower serum concentrations, whereas a favorable response to prior treatment and the presence of pleural effusion were associated with higher concentrations (Fig.5). 4. Discussion This study demonstrated that the RAM C trough may serve as a potential prognostic factor, particularly in the ERL group. To the best of our knowledge, this is the first report of a PK analysis of ERL in a real-world setting. Additionally, mGPS emerged as a strong prognostic factor in DTX and ERL groups. The PK results were consistent with those of previous reports[15], confirming the reproducibility of our LC-MS/MS method. This approach is regarded as useful because it eliminates the need for anti‑drug antibody design and yields results comparable to those of the ligand-binding assay in a shorter time frame[16]. C trough was significantly higher in the ERL group than in the DTX group, which was primarily attributable to the difference in dosing intervals (q2w vs. q3w). Previous studies reported that steady-state serum RAM concentrations are associated with FN and hypertension in the DTX group[8], whereas no significant correlation was observed in the ERL group[9]. However, in our study, no consistent association was found between C trough and toxicity in either group, except for edema in the DTX group. Toxicity should be evaluated by including parameters such as the maximum serum concentration and the area under the curve of RAM. The tumor response tended to be higher in the Q2–4 group. Notably, in the ERL group, additional therapeutic benefits were observed despite half of the patients having a history of TKI treatment. In the DTX group, unlike our previous report[10], C trough was not associated with survival. One possible explanation is that the Q2–4 group included a higher proportion of EGFR‑mutant cases with ins20 following TKI failure and cases with low PD‑L1 expression, together with a lack of correlation between cachexia and prognosis. In contrast to the ERL group, a significant prolongation of PFS was observed in the Q2–4 group. In the RELAY PPK analysis, no exposure–efficacy relationship was observed. One proposed explanation is VEGFR-2 saturation at low RAM concentrations. Moreover, the study population generally had a low disease burden and minimal cachexia, which reduced the likelihood that RAM clearance was influenced by disease status [9]. In this study, 75% of patients had received prior treatment, with half of them having a history of EGFR-TKI treatment. Previous reports have suggested that VEGFR-2 expression in tumors increases after EGFR-TKI treatment[17], raising the possibility that VEGFR-2 may no longer become saturated. Additionally, 25% of the patients in the ERL group had cachexia, indicating a higher disease burden than in the RELAY group and reflecting a real-world population. In cachexia, weight loss occurs along with enhanced protein catabolism, which is thought to accelerate the clearance of antibody drugs; similar phenomena have been reported for various antibody treatments [18-20]. Overall, these findings suggest that an exposure–efficacy relationship may be observed in a real-world setting. In the DTX group, edema was associated with an increased C trough , whereas a history of ICI treatment was associated with a decreased C trough . In the ERL group, pleural effusion was associated with increased C trough . Edema induced by DTX is thought to result primarily from increased vascular permeability, leading to the leakage of water and proteins into the interstitium[21], and the accompanying intravascular dehydration may have contributed to the elevated C trough . Previous reports have shown that RAM achieved favorable control rates of malignant pleural effusion in patients with NSCLC [22], suggesting that a reduction in third-space fluid may have led to increased C trough . With respect to prior ICI treatment, it has been reported that the affinity of anti-drug antibodies (ADAs) matures over time, resulting in increased drug clearance through interactions with more monoclonal antibodies; therefore, prior exposure to monoclonal antibodies such as ICIs may have facilitated the development of ADAs against RAM[23]. Reports have indicated that high VEGFR-2 expression is a poor prognostic factor for pembrolizumab treatment in NSCLC[24]. If VEGFR2‑high tumor cells are selected after ICI treatment, target-mediated drug disposition may accelerate RAM clearance. Regarding the combination of ICIs and anti‑angiogenic agents, a phase II trial comparing pembrolizumab plus RAM with the standard of care in patients with NSCLC who failed ICI monotherapy was positive[25], suggesting that the addition of an anti‑angiogenic agent may help overcome ICI resistance. However, several subsequent phase III trials have failed, highlighting the complexity of the relationship between ICIs and the VEGF/VEGFR axis. Thus, the clinical utility of these combinations remains controversial and requires careful evaluation[26]. The mGPS is a simple prognostic factor, and its usefulness was inadvertently validated in this study. As one of the scores reflecting systemic inflammation in cachexia, mGPS has demonstrated strong prognostic ability across various stages of NSCLC, regardless of the treatment modality, including surgery, radiotherapy, chemotherapy, and immunotherapy[14]. Consistent with previous findings, mGPS was identified as a factor influencing C trough in the ERL group in our univariate analysis. However, C trough may be influenced by multiple factors beyond cachexia, as discussed above. In the DTX group, C trough did not impact prognosis, whereas mGPS still tended to serve as a prognostic factor. Although mGPS is composed of high inflammation and hypoalbuminemia and biologically represents a rather nonspecific index without capturing a particular condition, it may nonetheless be a simple and clinically useful prognostic factor, even in patients with NSCLC treated with RAM. This study had several limitations. First, the number of cases in each group was extremely small. Increasing the sample size might have provided greater statistical power and more convincing results. Interpretation was further complicated by overlapping cases between the DTX and ERL groups; blood sampling was performed only at the trough point, and ADAs were not measured. Based on these findings, strategies such as therapeutic drug monitoring to maintain serum concentrations in patients with cachexia receiving RAM should be considered. However, toxicity has been insufficiently evaluated and requires further validation. Additionally, a comprehensive assessment of RAM concentrations in the tumor tissue or pleural effusion, as well as serum concentrations of concomitant drugs, will be necessary. In summary, RAM C trough may contribute to survival outcomes after ERL plus RAM treatment. Moreover, the mGPS appears to be a simple and useful prognostic factor for patients with NSCLC treated with RAM. Declarations Conflict of Interest Kazuto Ashizawa and Hirokazu Taniguchi have received honorarium from Chugai Pharmaceutical Co.,Ltd.. Funding sources This study did not receive any specific grants from funding agencies in the public, commercial, or not-for-profit sectors. Author Contribution Kazumasa Akagi; Conceptualization, Methodology, Formal analysis, Data Curation, Writing-Original Draft, VisualizationShinnosuke Takemoto; Investigation, Writing-Review & Editing, Project administrationMayu Ohuchi; Formal analysis, Writing-Review & EditingShigehiro Yagishita; Resources, Writing-Review & EditingRyosuke Ogata, Hiromi Tomono, Noritaka Honda, Yasuhiro Umeyama, Yosuke Dotsu, Midori Matsuo, Hirokazu Taniguchi, Hiroshi Gyotoku, Minoru Fukuda, Hiroshi Soda, and Kazuto Ashizawa; Writing-Review & EditingAkinobu Hamada; Resources, Writing-Review & EditingHiroshi Mukae; Supervision Acknowledgement We thank Editage [http://www.editage.com] for editing and reviewing the manuscript for English language. The authors thank Erika Imamura (Department of Respiratory Medicine, Nagasaki University Graduate School of Biomedical Sciences) for her technical assistance with the experiments. 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Nat Rev Clin Oncol 21(6):468–482. 10.1038/s41571-024-00886-y Table 1 Table 1 is available in the Supplementary Files section. Additional Declarations No competing interests reported. Supplementary Files 20251127supplementarytable.docx Table1Characteristicsofpatientsbytreatmentregimen.docx 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-8482971","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":569869398,"identity":"815025c2-7b1c-4c98-a038-b662daa67216","order_by":0,"name":"Kazumasa Akagi","email":"","orcid":"","institution":"Nagasaki University Graduate School of Biomedical Sciences","correspondingAuthor":false,"prefix":"","firstName":"Kazumasa","middleName":"","lastName":"Akagi","suffix":""},{"id":569869400,"identity":"4b931070-bbdb-4c4d-8ab1-e554426ce95e","order_by":1,"name":"Shinnosuke 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Samples were collected on day 1 of each treatment cycle and defined as Pre (cycle 1), C\u003csub\u003etrough\u003c/sub\u003e (cycle 2), C2\u003csub\u003etrough\u003c/sub\u003e (cycle 3), and C3 t\u003csub\u003erough\u003c/sub\u003e (cycle 4). \u003cstrong\u003e(B) \u003c/strong\u003eRamucirumab pharmacokinetics in the docetaxel and erlotinib group. Error bars show interquartile range. C\u003csub\u003etrough\u003c/sub\u003e was sorted from Q1–Q4 by interquartile range in each regimen.\u003cstrong\u003e (C) \u003c/strong\u003eComparison of C\u003csub\u003etrough\u003c/sub\u003e between the docetaxel and erlotinib group using Student t-test. Error bars represent 95% confidence intervals, and ** indicates \u003cem\u003ep \u003c/em\u003e≤ 0.01. Abbreviations: RAM; ramucirumab, DTX; docetaxel, ERL; erlotinib\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/c27cd95c4d30395bd25e5407.jpeg"},{"id":99813709,"identity":"5003cfb7-ffdf-4552-bc56-d371b408b8f4","added_by":"auto","created_at":"2026-01-08 14:39:32","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":233104,"visible":true,"origin":"","legend":"\u003cp\u003eRamucirumab C\u003csub\u003etrough\u003c/sub\u003e and ramucirumab-specific adverse events in each regimen. Error bars represent 95% confidence intervals, and ns indicates\u003cem\u003e p\u003c/em\u003e \u0026gt; 0.05; * indicates \u003cem\u003ep\u003c/em\u003e ≤ 0.05. Abbreviations: RAM; ramucirumab, DTX; docetaxel, ERL; erlotinib, FN; febrile neutropenia\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/0645d0b1468a84053866e912.jpeg"},{"id":99813289,"identity":"874425f1-84fe-4e1a-92ed-0f4676b5aeab","added_by":"auto","created_at":"2026-01-08 14:38:48","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":327993,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003e(A) \u003c/strong\u003eTumor response by ramucirumab C\u003csub\u003etrough\u003c/sub\u003e in each regimen. The bar in red represents the overall response rate.\u003cstrong\u003e (B)\u003c/strong\u003e Kaplan-Meier curve by C\u003csub\u003etrough\u003c/sub\u003e and modified Glasgow Prognostic Score in the docetaxel group.\u003cstrong\u003e (C)\u003c/strong\u003e Kaplan-Meier curve by C\u003csub\u003etrough\u003c/sub\u003e and modified Glasgow Prognostic Score in the erlotinib group. Abbreviations: RAM; ramucirumab, DTX; docetaxel, ERL; erlotinib, ORR: overall response rate, SD; stable disease, PD; progressive disease, NE; not evaluable, CR; complete remission, PR; partial response, PFS; progression-free survival, OS; overall survival, mGPS; modified Glasgow Prognostic Score\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/e2cdc332f998d40a6493c392.jpeg"},{"id":99813329,"identity":"b47a24cb-7d60-4ee9-9341-328e63c4c243","added_by":"auto","created_at":"2026-01-08 14:38:51","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":558030,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003e(A) \u003c/strong\u003eForest plot of Cox univariate regression analysis for survival in the docetaxel group. \u003cstrong\u003e(B) \u003c/strong\u003eForest plot of Cox univariate regression analysis for survival in the erlotinib group. Abbreviations: RAM; ramucirumab, DTX; docetaxel, ERL; erlotinib, PFS; progression-free survival, OS; overall survival, HR; hazard ratio, 95%CI; 95% confidence intervals, PD-L1; programmed death-ligand 1, ICI; immune checkpoint inhibitor, VEGF/VEGFR; vascular endothelial growth factor/ vascular endothelial growth factor/ receptor, EGFR-TKI; epidermal growth factor receptor-tyrosine kinase inhibitor, PR; partial response, SD; stable disease\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/4cc85f2a5158a9227def2faa.jpeg"},{"id":99813413,"identity":"de573633-b3d5-4727-8450-640daf6f7137","added_by":"auto","created_at":"2026-01-08 14:39:10","extension":"jpeg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":374035,"visible":true,"origin":"","legend":"\u003cp\u003eForest plot of univariate linear regression analysis for ramucirumab C\u003csub\u003etrough\u003c/sub\u003e in each regimen. Abbreviations: RAM; ramucirumab, DTX; docetaxel, ERL; erlotinib, 95%CI; 95% confidence intervals, PD-L1; programmed death-ligand 1, ICI; immune checkpoint inhibitor, VEGF/VEGFR; vascular endothelial growth factor/ vascular endothelial growth factor/ receptor, EGFR-TKI; epidermal growth factor receptor-tyrosine kinase inhibitor, PR; partial response, SD; stable disease\u003c/p\u003e","description":"","filename":"floatimage5.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/d14d0fed85d9003f1debdcf9.jpeg"},{"id":100955882,"identity":"faed28f3-2698-460c-8aa0-9248981b5aee","added_by":"auto","created_at":"2026-01-23 07:29:20","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2444694,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/f8473e50-c834-40ce-bf35-8d056eca118c.pdf"},{"id":99813343,"identity":"399cc468-385d-49c5-9278-9c2071134faf","added_by":"auto","created_at":"2026-01-08 14:38:53","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":26491,"visible":true,"origin":"","legend":"","description":"","filename":"20251127supplementarytable.docx","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/571fcf3e99bc67302ca0b6e7.docx"},{"id":99813389,"identity":"c503f2f4-8584-4963-99de-5fff57ff767b","added_by":"auto","created_at":"2026-01-08 14:38:59","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":21304,"visible":true,"origin":"","legend":"","description":"","filename":"Table1Characteristicsofpatientsbytreatmentregimen.docx","url":"https://assets-eu.researchsquare.com/files/rs-8482971/v1/e2c0e986de087f3a388bd1e5.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Real-world pharmacokinetics of ramucirumab in combination with erlotinib and docetaxel for advanced non-small cell lung cancer: a prospective cohort study","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eRamucirumab (RAM) is a human IgG1 monoclonal antibody that targets human vascular endothelial growth factor receptor-2 (VEGFR-2). RAM demonstrates anti-angiogenic and anti-tumor activities through the inhibition of vascular endothelial growth factor (VEGF) binding to VEGFR-2[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. RAM has been approved for use in combination with docetaxel (DTX) or erlotinib (ERL) in lung cancer. The REVEL trial demonstrated that DTX\u0026thinsp;+\u0026thinsp;RAM improved survival as a second-line treatment for patients with stage IV non-small cell lung cancer (NSCLC) and remains an important therapeutic option beyond second-line therapy[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. The RELAY trial showed that ERL\u0026thinsp;+\u0026thinsp;RAM demonstrated superior progression-free survival (PFS) compared to placebo plus ERL in patients with untreated epithelial growth factor receptor (EGFR)-mutated metastatic NSCLC[\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Recently, EGFR-mutant lung cancer has undergone a paradigm shift in treatment, with regimens such as FLAURA2[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e] and MARIPOSA [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e] becoming standard therapies. However, survival analysis from the RELAY trial suggested that ERL\u0026thinsp;+\u0026thinsp;RAM may be an effective treatment option, especially for patients with EGFR L858R mutations[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Furthermore, recent positive trials combining RAM with osimertinib[\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e] have indicated that RAM remains an important therapeutic option.\u003c/p\u003e \u003cp\u003eIn the population pharmacokinetics (PK) analysis of the REVEL trial, higher RAM exposure was associated with improved clinical outcomes[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Conversely, PPK analysis of the RELAY trial showed no association between RAM exposure and response[\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. A system was established to measure serum concentrations of RAM using liquid chromatography\u0026ndash;tandem mass spectrometry (LC-MS/MS), and trough levels (C\u003csub\u003etrough\u003c/sub\u003e) were analyzed after the first administration of RAM in patients with advanced or recurrent NSCLC, together with clinical information. These results suggest that patients with a higher C\u003csub\u003etrough\u003c/sub\u003e tended to have a better prognosis. Conversely, patients with lower C\u003csub\u003etrough\u003c/sub\u003e levels include a greater proportion of cases with cachexia, indicating that accelerated clearance of RAM in cachexia may reduce therapeutic efficacy[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. However, since this was a single-center retrospective study, a prospective, multicenter study was initiated to validate these findings in a real-world setting, including not only DTX plus RAM, but also ERL plus RAM, which has not been previously reported.\u003c/p\u003e"},{"header":"2. Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Study design\u003c/h2\u003e \u003cp\u003eThis was a multicenter prospective cohort study. Eligible participants were patients with advanced or recurrent NSCLC who received RAM treatment in combination with DTX or ERL at Nagasaki University Hospital or a collaborating research facility between June 22, 2021, and August 31, 2023. Participants were expected to undergo follow-up for at least 90 days from the start of treatment, and were aged\u0026thinsp;\u0026ge;\u0026thinsp;20 years at the time of consent. Blood sampling was performed at the trough level immediately before RAM administration. Blood samples were collected on day 1 for each of the first four treatment cycles, starting with the initial administration of RAM, designated as day 1. Samples collected on day 1 of the second treatment cycle were defined as the C\u003csub\u003etrough\u003c/sub\u003e.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Data collection\u003c/h2\u003e \u003cp\u003eThe following clinical information was prospectively collected: date of birth, sex, performance status at the time closest to the initiation of RAM, height and weight, smoking history, medical history and comorbidities, and prior treatment information for the primary disease. Imaging data, including chest radiography, chest and abdominal computed tomography (CT) scans, brain CT or magnetic resonance imaging, positron emission tomography-CT, and bone scintigraphy, were reviewed. Clinical assessments and laboratory tests were reviewed, including subjective and objective symptoms and any other grade 3 or higher adverse events. Edema and proteinuria were identified when they appeared or worsened, respectively, during the clinical course. Laboratory data included peripheral blood count, blood biochemistry, urinalysis, and oxygen saturation. Pathological data included tumor histology and gene alterations, EGFR mutations, anaplastic lymphoma kinase and c-ros oncogene 1 rearrangements, and programmed death-ligand 1 (PD-L1) tumor proportion score. Efficacy was assessed in accordance with the Response Evaluation Criteria in Solid Tumors version 1.1[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], and adverse events were assessed using the National Cancer Institute Common Terminology Criteria for Adverse Events version 5.0[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. PFS was defined as the time from the start date of the RAM treatment to disease progression or death from any cause, whereas overall survival (OS) was defined as the time from the start date of the RAM treatment to the last day confirmed as alive or dead from any cause. Cachexia was defined as either a weight loss of \u0026ge;\u0026thinsp;5% within 6 months before treatment initiation, or a weight loss of \u0026ge;\u0026thinsp;2% within 2 months, accompanied by a body mass index\u0026thinsp;\u0026lt;\u0026thinsp;20[\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. The modified Glasgow Prognostic Score (mGPS) was defined as follows: score 0 for C-reactive protein (CRP)\u0026thinsp;\u0026le;\u0026thinsp;0.5 mg/dL and albumin\u0026thinsp;\u0026ge;\u0026thinsp;3.5 g/dL; score 1 for CRP\u0026thinsp;\u0026gt;\u0026thinsp;0.5 mg/dL or albumin\u0026thinsp;\u0026lt;\u0026thinsp;3.5 g/dL; and score 2 for CRP\u0026thinsp;\u0026gt;\u0026thinsp;0.5 mg/dL and albumin\u0026thinsp;\u0026lt;\u0026thinsp;3.5 g/dL[\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3. Measurement of serum RAM concentration\u003c/h2\u003e \u003cp\u003eAt the time of blood sampling immediately before RAM administration, 6 mL of blood was collected in a serum separation tube. After incubation at room temperature for 30 min, the sample was centrifuged at 2000 rpm for 10 min at room temperature to separate the serum. One milliliter of serum was transferred into a cryotube and stored at \u0026minus;\u0026thinsp;80\u0026deg;C. If storage at \u0026minus;\u0026thinsp;80\u0026deg;C was not feasible at a given facility, \u0026minus;\u0026thinsp;20\u0026deg;C storage was permitted. The stored serum samples were transported frozen to the National Cancer Center Research Institute, where serum RAM concentrations were measured using LC-MS/MS coupled with the nSMOL method, as previously reported[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. If treatment was delayed or discontinued owing to adverse events at the scheduled C\u003csub\u003etrough\u003c/sub\u003e sampling point, the blood test conducted at that time was used as the trough value, provided that blood sampling had been performed.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4. Ethical considerations and registration\u003c/h2\u003e \u003cp\u003e Written informed consent was obtained from all enrolled patients, and the study was conducted in accordance with the Declaration of Helsinki and Good Clinical Practice Guidelines. The study protocol was reviewed and approved by the Clinical Review Board of Nagasaki University (CRB7180001) on June 22, 2021 (approval no. 21062121).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003e2.5. Statistical analysis\u003c/h2\u003e \u003cp\u003eContinuous variables were presented as means and medians for parametric and nonparametric variables, respectively. The Fisher\u0026rsquo;s exact test was used for group comparisons of categorical variables. Student\u0026rsquo;s \u003cem\u003et\u003c/em\u003e-test was used to compare parametric continuous variables between groups, whereas the Mann\u0026ndash;Whitney \u003cem\u003eU\u003c/em\u003e test was used to compare nonparametric continuous variables between groups. Univariate and logistic regression analyses were used to assess continuous and categorical variables, respectively. Cox univariate regression analysis was used to assess survival, and the log-rank test was used for intergroup comparisons. Statistical significance was set at \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05. The proportional hazard assumption was assessed using Schoenfeld residuals, and it was confirmed that no violations were observed. Analyses were performed using R (version 4.5.0) and GraphPad Prism (version 9.5.1; GraphPad Software, USA).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e2.6. Data statement\u003c/h2\u003e \u003cp\u003eAdditional data are available upon request from the corresponding author.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results","content":"\u003cp\u003e\u003cstrong\u003e3.1. Patient characteristics\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn the DTX group, the median age was 64 years, and half of the patients had EGFR mutations. Eleven patients (68.8%) had received two or more prior treatment regimens, including immune checkpoint inhibitors (ICI) in eight patients (50.0%) and EGFR-tyrosine inhibitors (TKIs) in six patients (37.5%). Pleural effusion and edema were observed in six patients (37.5%). Six patients (37.5%) had an mGPS of 0, while 10 patients (62.5%) had an mGPS of 1 or 2. Cachexia was present in only two patients (12.5%).\u003c/p\u003e\n\u003cp\u003eThe median age of patients in the ERL group was 73 years. EGFR mutations included exon 19 deletion (del19) in three patients (37.5%) and L858R in five patients (62.5%). Four (50.0 %) patients had a history of EGFR-TKI treatment. Pleural effusion was observed in six patients (75.0%) and edema in two patients (25.0%). Five patients (62.5%) had an mGPS of 0, and three patients (37.5%) had an mGPS of 1 or 2. Cachexia was present in two patients (25.0%) (Table 1). Divided into Q1 and Q2\u0026ndash;4, the DTX group showed six cases (54.5%) with EGFR mutations, including insertion in 20, and five cases (45.5%) with PD‑L1 negativity in Q2\u0026ndash;4 (Table S1). In the ERL group, all patients in Q2\u0026ndash;4 had an mGPS of 1, whereas in Q1, five patients (83.3%) had an mGPS of 0 (Table S2).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.2. Pharmacokinetics of RAM\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTrough blood sampling was performed in 23 patients at the start of cycle 1, 23 at cycle 2, 18 at cycle 3, and 14 at cycle 4 (Fig. 1A). In one case in the DTX group, C\u003csub\u003etrough\u003c/sub\u003e sampling was missing, and in another case, the pre-dose concentration was missing. Additionally, there was one case in which ERL + RAM was administered 35 days after treatment with DTX + RAM, in which a detectable pre-dose RAM concentration was observed. Under the above conditions, C\u003csub\u003etrough\u003c/sub\u003e in the DTX group ranged from 4.3\u0026ndash;36.4 \u0026mu;g/mL, and the median was 15.3 \u0026mu;g/mL. C\u003csub\u003etrough\u003c/sub\u003e was sorted from Q1\u0026ndash;Q4 by interquartile range (4.3 \u0026le; Q1 \u0026le; 10.7, 10.7 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ2 \u0026le; 15.3, 15.3 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ3 \u0026le; 18.1 and 18.1 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ4 \u0026le; 36.4 \u0026mu;g/mL. Meanwhile, C\u003csub\u003etrough\u003c/sub\u003e in the ERL group ranged from 10.2\u0026ndash;44.5 \u0026mu;g/mL, and the median was 26.2 \u0026mu;g/mL. C\u003csub\u003etrough\u003c/sub\u003e was sorted from Q1\u0026ndash;Q4 by interquartile range (10.2 \u0026le; Q1 \u0026le; 24.3, 24.3 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ2 \u0026le; 26.2, 26.2 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ3 \u0026le; 37.4, and 37.4 \u003cem\u003e\u0026lt;\u0026nbsp;\u003c/em\u003eQ4 \u0026le; 44.5 \u0026mu;g/mL (Fig. 1B). C\u003csub\u003etrough\u003c/sub\u003e was significantly higher in the ERL group than in the DTX group after excluding one overlapping case between the two groups from the analysis (\u003cem\u003ep\u003c/em\u003e=0.0047, t-test) (Fig. 1C).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.3. RAM\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eC\u003csub\u003etrough\u003c/sub\u003e and adverse events\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe relationship between C\u003csub\u003etrough\u003c/sub\u003e and the presence or absence of RAM-specific adverse events, namely, bleeding, edema, hypertension, febrile neutropenia (FN), and proteinuria, was evaluated. In the DTX group, patients with edema exhibited higher C\u003csub\u003etrough\u003c/sub\u003e levels, whereas no association was observed between C\u003csub\u003etrough\u003c/sub\u003e and other adverse events. In the ERL group, no association was found between the C\u003csub\u003etrough\u003c/sub\u003e and any adverse events (Fig.2).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.4. RAM\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eC\u003csub\u003etrough,\u0026nbsp;\u003c/sub\u003emGPS and efficacy\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe association between RAM efficacy and C\u003csub\u003etrough\u003c/sub\u003e (Q1 vs. Q2\u0026ndash;4) and mGPS (mGPS=0; mGPS-low vs. mGPS=1 or 2; mGPS-high) was analyzed. In the DTX and ERL groups, patients in the Q2\u0026ndash;4 group tended to have a higher objective response rate (ORR) (Fig. 3A).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eNext, log-rank tests were performed to evaluate the survival outcomes. In the DTX group, no association was observed between C\u003csub\u003etrough\u003c/sub\u003e and PFS or OS. The median PFS was 6.7 months in the mGPS-low and 4.2 months in the mGPS-high, which was not significantly different. The Kaplan\u0026ndash;Meier curve indicated a trend toward longer PFS in the mGPS-low group. The median OS was 32.8 months in mGPS-low and 13.6 months in mGPS-high, respectively, and was significantly longer in mGPS-low (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e= 0.042, log-rank test) (Fig.3B).\u003c/p\u003e\n\u003cp\u003eIn the ERL group, the median PFS was 8.3 months in Q1 and 16.4 months in the Q2-4 group, respectively, and was significantly longer in the Q2-4 group (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e= 0.032, log-rank test). The median OS was 16.6 months in the Q1 group and was not reached in the Q2-4 group, which was not significantly different. The Kaplan\u0026ndash;Meier curve indicated a trend toward longer OS in Q2-4. The median PFS was 18.2 months in the mGPS-low and 6.1 months in the mGPS-high groups, respectively, and was significantly longer in the mGPS-low (\u003cem\u003ep\u0026nbsp;\u003c/em\u003e= 0.004, log-rank test). The median OS was not reached in the mGPS-low and 16.6 months in the mGPS-high group, which was not significantly different. The Kaplan\u0026ndash;Meier curve indicated a trend toward longer PFS in the mGPS-low group (Fig.3C).\u003c/p\u003e\n\u003cp\u003eSubsequently, univariate analysis was performed to identify the factors influencing survival outcomes. In the DTX group, the best response to prior treatment was significantly associated with PFS and OS (HR: 0.27, 95% CI: 0.08\u0026ndash;0.91, \u003cem\u003ep\u003c/em\u003e = 0.035 for PFS; HR: 0.11, 95% CI: 0.02\u0026ndash;0.55, \u003cem\u003ep\u003c/em\u003e = 0.007 for OS), whereas C\u003csub\u003etrough\u003c/sub\u003e, cachexia, and mGPS were not associated with prognosis (Fig. 4A). In the ERL group, the presence of pleural effusion and C\u003csub\u003etrough\u003c/sub\u003e showed a trend toward association with PFS and OS, although the differences were not statistically significant (HR: 0.11, 95% CI: 0.01\u0026ndash;1.24, \u003cem\u003ep\u003c/em\u003e = 0.074 for PFS and OS) (Fig. 4B).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e3.6. Factors influencing RAM\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003eC\u003csub\u003etrough\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo identify factors influencing C\u003csub\u003etrough\u003c/sub\u003e, univariate analyses were conducted separately for the DTX and ERL groups. In the DTX group, a history of ICI treatment was associated with lower serum concentrations, whereas edema was associated with higher concentrations. In the ERL group, a high mGPS contributed to lower serum concentrations, whereas a favorable response to prior treatment and the presence of pleural effusion were associated with higher concentrations (Fig.5).\u003c/p\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eThis study demonstrated that the RAM C\u003csub\u003etrough\u003c/sub\u003e may serve as a potential prognostic factor, particularly in the ERL group. To the best of our knowledge, this is the first report of a PK analysis of ERL in a real-world setting. Additionally, mGPS emerged as a strong prognostic factor in DTX and ERL groups.\u003c/p\u003e\n\u003cp\u003eThe PK results were consistent with those of previous reports[15], confirming the reproducibility of our LC-MS/MS method.\u0026nbsp;This approach is regarded as useful because it eliminates the need for anti‑drug antibody design and yields results comparable to those of the ligand-binding assay in a shorter time frame[16].\u0026nbsp;C\u003csub\u003etrough\u003c/sub\u003e was significantly higher in the ERL group than in the DTX group, which was primarily attributable to the difference in dosing intervals (q2w vs. q3w). Previous studies reported that steady-state serum RAM concentrations are associated with FN and hypertension in the DTX group[8], whereas no significant correlation was observed in the ERL group[9]. However, in our study, no consistent association was found between\u0026nbsp;C\u003csub\u003etrough\u003c/sub\u003e and toxicity in either group, except for edema in the DTX group. Toxicity should be evaluated by including parameters such as the maximum serum concentration and the area under the curve of RAM.\u003c/p\u003e\n\u003cp\u003eThe tumor response tended to be higher in the Q2–4 group. Notably, in the ERL group, additional therapeutic benefits were observed despite half of the patients having a history of TKI treatment. In the DTX group, unlike our previous report[10], C\u003csub\u003etrough\u003c/sub\u003e was not associated with survival. One possible explanation is that the Q2–4 group included a higher proportion of EGFR‑mutant cases with ins20 following TKI failure and cases with low PD‑L1 expression, together with a lack of correlation between cachexia and prognosis. In contrast to the ERL group, a significant prolongation of PFS was observed in the Q2–4 group. In the RELAY PPK analysis, no exposure–efficacy relationship was observed. One proposed explanation is VEGFR-2 saturation at low RAM concentrations. Moreover, the study population generally had a low disease burden and minimal cachexia, which reduced the likelihood that RAM clearance was influenced by disease status\u0026nbsp;[9]. In this study, 75% of patients had received prior treatment, with half of them having a history of EGFR-TKI treatment. Previous reports have suggested that VEGFR-2 expression in tumors increases after EGFR-TKI treatment[17], raising the possibility that VEGFR-2 may no longer become saturated. Additionally, 25% of the patients in the ERL group had cachexia, indicating a higher disease burden than in the RELAY group and reflecting a real-world population. In cachexia, weight loss occurs along with enhanced protein catabolism, which is thought to accelerate the clearance of antibody drugs; similar phenomena have been reported for various antibody treatments\u0026nbsp;[18-20]. Overall, these findings suggest that an exposure–efficacy relationship may be observed in a real-world setting.\u003c/p\u003e\n\u003cp\u003eIn the DTX group, edema was associated with an increased C\u003csub\u003etrough\u003c/sub\u003e, whereas a history of ICI treatment was associated with a decreased C\u003csub\u003etrough\u003c/sub\u003e. In the ERL group, pleural effusion was associated with increased C\u003csub\u003etrough\u003c/sub\u003e. Edema induced by DTX is thought to result primarily from increased vascular permeability, leading to the leakage of water and proteins into the interstitium[21], and the accompanying intravascular dehydration may have contributed to the elevated C\u003csub\u003etrough\u003c/sub\u003e. Previous reports have shown that RAM achieved favorable control rates of malignant pleural effusion in patients with NSCLC [22], suggesting that a reduction in third-space fluid may have led to increased C\u003csub\u003etrough\u003c/sub\u003e. With respect to prior ICI treatment, it has been reported that the affinity of anti-drug antibodies (ADAs) matures over time, resulting in increased drug clearance through interactions with more monoclonal antibodies; therefore, prior exposure to monoclonal antibodies such as ICIs may have facilitated the development of ADAs against RAM[23]. Reports have indicated that high VEGFR-2 expression is a poor prognostic factor for pembrolizumab treatment in NSCLC[24]. If VEGFR2‑high tumor cells are selected after ICI treatment, target-mediated drug disposition may accelerate RAM clearance. Regarding the combination of ICIs and anti‑angiogenic agents, a phase II trial comparing pembrolizumab plus RAM with the standard of care in patients with NSCLC who failed ICI monotherapy was positive[25], suggesting that the addition of an anti‑angiogenic agent may help overcome ICI resistance. However, several subsequent phase III trials have failed, highlighting the complexity of the relationship between ICIs and the VEGF/VEGFR axis. Thus, the clinical utility of these combinations remains controversial and requires careful evaluation[26].\u003c/p\u003e\n\u003cp\u003eThe mGPS is a simple prognostic factor, and its usefulness was inadvertently validated in this study. As one of the scores reflecting systemic inflammation in cachexia, mGPS has demonstrated strong prognostic ability across various stages of NSCLC, regardless of the treatment modality, including surgery, radiotherapy, chemotherapy, and immunotherapy[14]. Consistent with previous findings, mGPS was identified as a factor influencing C\u003csub\u003etrough\u003c/sub\u003e in the ERL group in our univariate analysis. However, C\u003csub\u003etrough\u003c/sub\u003e may be influenced by multiple factors beyond cachexia, as discussed above. In the DTX group, C\u003csub\u003etrough\u003c/sub\u003e did not impact prognosis, whereas mGPS still tended to serve as a prognostic factor. Although mGPS is composed of high inflammation and hypoalbuminemia and biologically represents a rather nonspecific index without capturing a particular condition, it may nonetheless be a simple and clinically useful prognostic factor, even in patients with NSCLC treated with RAM.\u003c/p\u003e\n\u003cp\u003eThis study had several limitations. First, the number of cases in each group was extremely small. Increasing the sample size might have provided greater statistical power and more convincing results. Interpretation was further complicated by overlapping cases between the DTX and ERL groups; blood sampling was performed only at the trough point, and ADAs were not measured. Based on these findings, strategies such as therapeutic drug monitoring to maintain serum concentrations in patients with cachexia receiving RAM should be considered. However, toxicity has been insufficiently evaluated and requires further validation. Additionally, a comprehensive assessment of RAM concentrations in the tumor tissue or pleural effusion, as well as serum concentrations of concomitant drugs, will be necessary.\u003c/p\u003e\n\u003cp\u003eIn summary, RAM C\u003csub\u003etrough\u003c/sub\u003e may contribute to survival outcomes after ERL plus RAM treatment. Moreover, the mGPS appears to be a simple and useful prognostic factor for patients with NSCLC treated with RAM.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eConflict of Interest\u003c/h2\u003e \u003cp\u003eKazuto Ashizawa and Hirokazu Taniguchi have received honorarium from Chugai Pharmaceutical Co.,Ltd..\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eFunding sources\u003c/h2\u003e \u003cp\u003eThis study did not receive any specific grants from funding agencies in the public, commercial, or not-for-profit sectors.\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eKazumasa Akagi; Conceptualization, Methodology, Formal analysis, Data Curation, Writing-Original Draft, VisualizationShinnosuke Takemoto; Investigation, Writing-Review \u0026amp; Editing, Project administrationMayu Ohuchi; Formal analysis, Writing-Review \u0026amp; EditingShigehiro Yagishita; Resources, Writing-Review \u0026amp; EditingRyosuke Ogata, Hiromi Tomono, Noritaka Honda, Yasuhiro Umeyama, Yosuke Dotsu, Midori Matsuo, Hirokazu Taniguchi, Hiroshi Gyotoku, Minoru Fukuda, Hiroshi Soda, and Kazuto Ashizawa; Writing-Review \u0026amp; EditingAkinobu Hamada; Resources, Writing-Review \u0026amp; EditingHiroshi Mukae; Supervision\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eWe thank Editage [http://www.editage.com] for editing and reviewing the manuscript for English language. 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Oncol Rep 48(6). \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.3892/or.2022.8429\u003c/span\u003e\u003cspan address=\"10.3892/or.2022.8429\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eReckamp KL, Redman MW, Dragnev KH, Minichiello K, Villaruz LC, Faller B, Al Baghdadi T, Hines S, Everhart L, Highleyman L, Papadimitrakopoulou V, Neal JW, Waqar SN, Patel JD, Gray JE, Gandara DR, Kelly K, Herbst RS (2022) Phase II Randomized Study of Ramucirumab and Pembrolizumab Versus Standard of Care in Advanced Non-Small-Cell Lung Cancer Previously Treated With Immunotherapy-Lung-MAP S1800A. J Clin Oncol 40(21):2295\u0026ndash;2306. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1200/JCO.22.00912\u003c/span\u003e\u003cspan address=\"10.1200/JCO.22.00912\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKuo HY, Khan KA, Kerbel RS (2024) Antiangiogenic-immune-checkpoint inhibitor combinations: lessons from phase III clinical trials. Nat Rev Clin Oncol 21(6):468\u0026ndash;482. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1038/s41571-024-00886-y\u003c/span\u003e\u003cspan address=\"10.1038/s41571-024-00886-y\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Table 1","content":"\u003cp\u003eTable 1 is available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"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":"Ramucirumab, Pharmacokinetics, Concentration, Erlotinib, Docetaxel, Non-small cell lung cancer","lastPublishedDoi":"10.21203/rs.3.rs-8482971/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8482971/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePurpose\u003c/h2\u003e \u003cp\u003eIn pivotal phase III studies of non-small cell lung cancer (NSCLC), ramucirumab (RAM) serum concentrations were associated with clinical outcomes in docetaxel (DTX) plus RAM treatment (DTX group), but not in erlotinib (ERL) plus RAM treatment (ERL group). This study prospectively examined the relationship between serum RAM concentrations and clinical outcomes in a real-world setting.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eThis study included patients with advanced NSCLC who received RAM in combination with DTX or ERL. Blood samples were obtained at trough levels before RAM administration. The day 1 sample of the second cycle was defined as C\u003csub\u003etrough\u003c/sub\u003e and measured using liquid chromatography\u0026ndash;tandem mass spectrometry. The associations between C\u003csub\u003etrough\u003c/sub\u003e and clinical outcomes were evaluated.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eC\u003csub\u003etrough\u003c/sub\u003e ranged from 4.3\u0026ndash;36.4 \u0026micro;g/mL in the DTX group and 10.2\u0026ndash;44.5 \u0026micro;g/mL in the ERL group. Edema was more commonly observed in the high C\u003csub\u003etrough\u003c/sub\u003e group than in the DTX group. C\u003csub\u003etrough\u003c/sub\u003e was not associated with prognosis in the DTX group, while the median progression-free survival (PFS) was significantly longer in high C\u003csub\u003etrough\u003c/sub\u003e (8.3 vs 16.4 months, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.032) within the ERL group. The median overall survival was significantly longer in low modified Glasgow Prognostic Score (mGPS-low) (32.8 vs 13.6 months, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.042) within the DTX group, and PFS was significantly longer in mGPS-low (18.2 vs 6.1 months, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.004) within the ERL group.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eRAM C\u003csub\u003etrough\u003c/sub\u003e may influence survival after ERL\u0026thinsp;+\u0026thinsp;RAM treatment. The mGPS appears to be a potential prognostic factor in patients with NSCLC treated with RAM.\u003c/p\u003e","manuscriptTitle":"Real-world pharmacokinetics of ramucirumab in combination with erlotinib and docetaxel for advanced non-small cell lung cancer: a prospective cohort study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-08 14:02:52","doi":"10.21203/rs.3.rs-8482971/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":"8b44010b-ac29-402b-a0f6-88d8adf0765d","owner":[],"postedDate":"January 8th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-02-23T14:24:09+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-08 14:02:52","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8482971","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8482971","identity":"rs-8482971","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}