An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel administration: evaluating the protective role of dimethyl fumarate

An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel administration: evaluating the protective role of dimethyl fumarate | 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 An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel administration: evaluating the protective role of dimethyl fumarate Sebastian Cubides-Cely, Mario Negrette-Guzmán This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4974358/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 Docetaxel is a taxane antineoplastic widely used against different types of cancer. However, its efficacy is limited mainly by its myelotoxicity and peripheral neuropathy with neutropenia and nociceptive alterations as the main clinical signs, respectively. These adverse effects undermine the quality of life of patients leading them to treatment withdrawal. In this study, we set up a unique preclinical scheme for induction of both effects associated with docetaxel administration, in such a way that we can evaluate them in the same animal and carry out future protection trials. Methods Four docetaxel administration schemes were tested varying dose and dosage. Four days after the last dose, behavioral/sensory paw pressure, tail pressure and hot plate tests were conducted. Next, euthanasia was performed, and blood was obtained for total cell count and other toxicological markers. Once the scheme that better showed significant alterations in both nociception and neutropenia was chosen, joint administration with 100 mg/kg/day oral dimethyl fumarate was carried out to evaluate its protective effect. Results A scheme with six doses (5 mg/kg) of docetaxel administered weekly was chosen for protection trials. Dimethyl fumarate showed protection in nociceptive tests compared to the damage group. However, it did not show protection against neutropenia. Conclusion The confirmed experimental model is clinically representative as it was designed for rats through equivalent data obtained from clinical assays. It was useful to evaluate the protective potential of dimethyl fumarate, showing how it could attenuate docetaxel-induced peripheral neuropathy, but not neutropenia. docetaxel peripheral neuropathy nociception myelotoxicity neutropenia dimethyl fumarate Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 INTRODUCTION Docetaxel is a semi-synthetic taxane, paclitaxel analog, approved by the U.S. Food and Drug Administration (FDA) for breast, ovarian, lung, and prostate cancer, among others[ 1 ], [ 2 ]. Its activity relies on specific binding to beta-tubulin subunits of microtubules, inhibiting tubulin depolymerization and, thus, stabilizing microtubules. Consequently, the cell cycle is arrested in mitosis, leading to cell death [ 3 ]. In addition, docetaxel is also able to lead cancer cells to intrinsic apoptosis by inhibiting the expression of anti-apoptotic genes BCL-2 and BCL-X and to arrest cancer growth by downregulating the androgen receptor signaling pathway [ 4 ]. Cancer cell death depends on drug concentration and exposure time. However, docetaxel can produce adverse reactions such as peripheral neuropathy, febrile neutropenia, infusion reactions, fatigue, fluid retention, pneumonitis, and gastrointestinal complications [ 5 ]. Taxane-associated peripheral neurotoxicity is an acute pain syndrome consisting of diffuse muscle pain, mainly in the legs, hips and lower back, although it may spread, resulting in myalgia and/or arthralgia. Incidences of docetaxel-induced peripheral neuropathy are highly variable. However, studies have reported up to 64% of overall incidence with 7.4% of severe neuropathy [ 6 ]. The exact pathophysiological mechanism of taxane-induced peripheral neurotoxicity is not yet fully described; however, axonal microtubule stability and polar reconfiguration enhanced by taxanes can alter axonal transport of different substances and mitochondria, leading to a loss of axonal integrity or degeneration, which, in turn, results in abnormal nerve physiology [ 6 ]. According to the American Society of Clinical Oncology (ASCO) and the European Society for Medical Oncology (ESMO) guidelines, there are no recommended or effective agents to prevent chemotherapy-induced peripheral neuropathy. Clinical trials have shown that duloxetine treatment reduces pain, numbness, and tingling symptoms, but only to a moderate degree, with a better protective effect in the treatment of peripheral neuropathy due to platinum-based drugs rather than other therapies such as taxanes [ 7 ]. Other options, such as tricyclic antidepressants or anticonvulsant drugs, have been explored, but trials show mixed results [ 8 ], [ 9 ]. On the other hand, neutropenia remains a condition that has no treatment other than prevention against possible infections once acquired. Severe neutropenia often requires modification of the chemotherapy regimen and may compromise anticancer efficacy. Prevention of neutropenia is an important patient benefit for reasons of safety, treatment efficacy, and cost-effectiveness [ 10 ]. There are recent evidence-based guidelines for the prevention of infections associated with chemotherapy-induced neutropenia [ 11 ]. However, antibiotics are the standard treatment for febrile neutropenia and are increasingly used for prophylaxis in patients at low risk of febrile neutropenia[ 12 ]. Granulocyte colony stimulating factors (G-CSF) stimulate the production and development of neutrophils, which are necessary to fight infections. Some G-CSF such as pegfilgrastim and filgrastim have been used for the treatment of febrile neutropenia caused by chemotherapeutics [ 13 ]. On the other hand, plinabulin, a non-G-CSF-selective microtubule-binding immunomodulatory agent that possesses hematopoietic stem cell protective properties and anticancer benefits [ 10 ], has previously been administered in animals, where improvement in neutropenia induced by various chemotherapies, including docetaxel, doxorubicin hydrochloride, and cyclophosphamide, was observed through a mechanism other than G-CSF [ 14 ]. Dimethyl fumarate, a fumaric acid ester (α, β-unsaturated carboxylic acid ester), is an FDA and European Medicines Agency (EMA) approved drug, which is effective in the treatment of relapsing/remitting multiple sclerosis, and is considered a type of indirect antioxidant, as it promotes the activation and stabilization of nuclear factor erythroid 2-related factor 2 (Nrf2) [ 15 ], [ 16 ]. Dimethyl fumarate enters the body orally and dissociates in the intestine, where it subsequently interacts with immune cells in the blood circulation and crosses the blood-brain barrier into the central nervous system [ 17 ]. There are different works involving hepatoprotection, neuroprotection and anti-arthritis protection using daily oral doses between 25 and 100 mg/kg of dimethyl fumarate for several weeks, showing an Nrf2-mediated cytoprotective effect in Wistar rats [ 18 ]. Although peripheral neuropathy and neutropenia are widely reported chemotherapy-induced adverse effects, dose reduction or discontinuation of treatment remains the more effective treatment for their co-management. More preclinical investigation on strategies for inhibiting or attenuating docetaxel-induced peripheral neuropathy and neutropenia without affecting its therapeutic efficacy is required, ensuring the quality of data and reducing the number of research subjects ethically. In this work, we establish a rat model for simultaneous induction of myelotoxicity and peripheral neuropathy by docetaxel, which is useful for studying protective therapeutic alternatives and allows the evaluation of both effects in the same animal. Four different administration schemes were tested based on clinical information and experimental models. Sensory behavioral tests were performed to obtain markers of peripheral neuropathy, and blood was obtained to determine neutropenia. The scheme representing changes better in both nociceptive thresholds (peripheral neuropathy) and neutropenia (myelotoxicity) was chosen. Using this experimental model, we first report that dimethyl fumarate is a potential protective agent against docetaxel-induced peripheral neuropathy, but not against neutropenia. MATERIAL AND METHODS Drugs Docetaxel was purchased from BLAŪ Farmacéutica, (Brazil), and prepared and mixed according to the manufacturer's instructions. Dimethyl fumarate was purchased from Sigma-Aldrich (St Loius, MI, USA) and was resuspended in carboxyl methyl cellulose (CMC, Sigma-Aldrich, St Loius, MI, USA), which was prepared at 1% by diluting in distilled water. Animals Male Wistar rats weighing 240 ± 30 g at the start of the treatments were used. Animals were maintained, cared for and used according to Law 84 of 1989 (Congress of Colombia), Title V of Resolution 8430 of 1993 (Ministry of Health, Colombia) and the guidelines described in the Guide for the Care and Use of Laboratory Animals of the National Academy of Sciences of the United States [ 19 ]. Animals were kept in stainless steel cages 30x40 cm x20 cm high (6 rats per cage) at a temperature of 21°C − 24°C, relative humidity of 55–70%, photoperiod of 12 hours of light and 12 hours of darkness with sterile water and specialized food ad libitum . Protocols were approved by the Comité de Ética en Investigación Científica (CEINCI) of the Universidad Industrial de Santander (Committee Minutes No. 10 of 2021 and No.12 of 2024). Rats were habituated and trained daily, always in the morning hours, to diminish stress levels and improve the reliability of the model. Four days before the start of administration schemes, rats were habituated to the room, caretakers, researchers, and equipment used in experiments. Animals also underwent individual training to teach them to remain still and calm during oral administration, restraining for intravenous administration, and using analgesimeters. At the end of this period, the change in the behavior of rats was observed. Establishment of the docetaxel damage model Four docetaxel administration schemes were tested varying dose and dosage (Fig. 1 ). Scheme 1 consisted of a 15 mg/kg single dose. Scheme 2 included two doses of 15 mg/kg administered three weeks apart. In Scheme 3, a 5 mg/kg dose was administered weekly until 6 doses were completed. Scheme 4 consisted of three 15 mg/kg doses each administered every 3 weeks. In all cases, docetaxel was administered intravenously in the tail. Each scheme evaluated two experimental groups, the control group (CT, n = 8), which was administered normal saline, and the docetaxel-treated group (DT, n = 8). Administration schemes were proposed based on clinical practice and trials from literature [ 20 ], [ 21 ] equivalent for rats [ 22 ]. However, some reports where either peripheral neuropathy or neutropenia was induced in rats, but not simultaneously, were also considered [ 23 ], [ 24 ], [ 25 ]. Behavioral testing, euthanasia and blood cell count Four days after the last administration in each scheme, sensory behavioral testing paw and tail pressure and hot plate were performed using the PANLAB analgesimeter model LE7306 Randall-Selitto and the PANLAB analgesimeter model LE7406 Hot Plate (Panlab, S.L.U., Barcelona, Spain), respectively. Nociceptive testing was based on physical indicators of discomfort (withdrawal reflexes, licking and voicing). To ensure ethical treatment of the animals, maximum cut-off values of 10 seconds of paw and tail pressure latency and a weight of up to 650 g were adjusted. For the case of the hot plate method, 60 seconds of latency at 50 ± 0.1°C was taken as the cut-off. All tests were recorded in video format for later data verification and analysis. Once the nociceptive tests were concluded, the rats were euthanized under anesthesia with ketamine (90 mg/kg) + xylazine (15 mg/kg) intraperitoneally (i.p.) by exsanguination through the left ventricle of the animal. Hemograms and blood biochemistry Complete differential cell counts were performed in blood samples to determine neutropenia using a Genvet VH50 hematology analyzer (Genrui Biotech Inc., Shenzhen, China). Blood biochemistry tests were performed using kits commercially available following the manufacturer's instructions (LabTest Diagnóstica S.A., Minas Gerais, Brazil); creatine kinase activity (CK-MB. Ref. 118), aspartate aminotransferase (AST, Ref. 109), alanine aminotransferase (ALT, Ref. 1008), alkaline phosphatase (Ref. 79), creatinine K concentration (Ref. 96), and urea (Ref. 104) were analyzed. Assessment of dimethyl fumarate protection Once selected, the best scheme that ensured neutropenia and peripheral neurotoxicity was challenged with a protection alternative proposed: dimethyl fumarate, a known Nrf2 inducer and FDA-approved drug. This administration scheme consisted of a control group (CT, n = 8), which received saline i.v. as a vehicle for docetaxel and oral carboxymethylcellulose (CMC) as a vehicle for dimethyl fumarate; a docetaxel group (DT, n = 8), administered with docetaxel i.v. along with CMC; a co-treated group (DT + DF, n = 8) administered with docetaxel i.v. and dimethyl fumarate orally at 100 mg/Kg/day [ 18 ], [ 26 ]; an additional control group was administered with only dimethyl fumarate (DF, n = 8) and saline as vehicle for docetaxel. Docetaxel or its vehicle were administered according to the scheme selected for simultaneous induction of nociceptive alterations and neutropenia. Dimethyl fumarate or CMC were administered one hour before the administration of docetaxel (or its vehicle). As previously described for the establishment of the damage model, four days after the end of the treatment, animals were subjected to sensory behavioral tests and subsequently euthanized, obtaining blood for hemograms and blood biochemistry. In addition, sciatic nerves were obtained and stored for further studies on mechanisms of damage and protection. Statistical analysis All results were expressed as the mean ± standard deviation of data. Data processing was performed by t-test analysis comparing groups DT and CT for each of the four evaluated damage schemes. To compare between groups in the protection model, one-way ANOVA followed by Bonferroni multiple comparison tests were used. Significance was determined at p ≤ 0.05. All data were processed, plotted and analyzed using Prism 8 (GraphPad Software). RESULTS Simultaneous induction of nociceptive alterations and neutropenia using docetaxel Docetaxel toxicity was evident throughout the treatment as animal behavior and feeding and living habits were changing. Rats treated with the drug showed a significant lag in body weight gain compared to the control rats, mainly in schemes 3 and 4 (Fig. 2 ). Schemes 1, 3 and 4 showed differences between the groups in the paw pressure test (Fig. 3 a), although docetaxel induced an increase in the nociceptive threshold due to Scheme 1 (P < 0.05), which is not a canonical outcome. The mean pain threshold of DT rats in Scheme 3 was almost a third of that for group CT (P < 0.0001). Despite the entire dose in Scheme 2 being twice the dose in Scheme 1, no significant differences between groups were observed in Scheme 2. In the tail pressure test (Fig. 3 b), the group DT of schemes 2, 3 and 4 had significant falls in nociceptive thresholds compared to the control group. Scheme 1 did not show differences between groups for this test. On the other hand, of the 4 schemes administered, only Scheme 4 showed an alteration in the hot plate test (Fig. 3 c, P < 0.05). However, this alteration, likewise in the paw pressure test of Scheme 1, had an increasing trend in the nociceptive threshold, which is not frequently reported in this kind of study. None of the remaining treatments with docetaxel (schemes 1–3) gave a significant response compared to the CT group in the hot plate test (Fig. 3 c). Once the pain behavioral tests were completed, euthanasia was performed, and blood was taken for the differential neutrophil count. As shown in Fig. 4 , all the docetaxel administration schemes led to neutropenia, with neutrophil count decreases of 49% for Scheme 1 (P < 0.05), 75% for Scheme 2 (P < 0.0001), 80% for Scheme 3 (P < 0.001), and 100% for Scheme 4 (P < 0.0001). Schemes 2, 3 and 4 showed a feature of febrile neutropenia, counts < 0.5*10 9 /L. However, null or non-detectable neutrophil counts obtained in Scheme 4 may make it few attractive experimentally. Since the results shown so far, Scheme 3 joins together most of the characteristics desirable for the experimental model of simultaneous induction. First, decreases in paw and tail pressure thresholds (two of the three nociceptive tests assessed) and neutrophil counts were observed with good significance, which validates and gives confidence to the model. Although Scheme 4 showed similar features, the undetectable neutrophil counts would make it difficult to evaluate potential myeloprotective agents. In addition, this severe neutropenia with null cell counts in Scheme 4 is not the most observed clinical outcome. Schemes 1 and 2 only showed responses in one of the nociceptive tests executed. Even though Scheme 3 implies the largest number of doses over a long period, its outcomes demonstrated that it is the completer and most reliable scheme for evaluating potential neuro and myeloprotective chemotherapeutics. The following results were obtained by applying Scheme 3 as a model of simultaneous induction of peripheral neurotoxicity and myelotoxicity by docetaxel. Protective role of dimethyl fumarate against docetaxel-induced nociceptive alterations and neutropenia As was seen above, animals treated with docetaxel (group DT) showed a significant decrease in body weight gain throughout the treatment. This trend was not counteracted or attenuated in the group co-treated with dimethyl fumarate and docetaxel (DT + DF). Regarding dimethyl fumarate-treated group (DF), no difference was observed with respect to the control group (CT, data not shown). Nociceptive thresholds were reproduced for the DT group according to the damage model obtained, as shown in Fig. 5 . Treatment with dimethyl fumarate allowed an attenuation of docetaxel-induced decreases in paw (DT + DF, Fig. 5 a, P < 0.05 vs DT) and tail (DT + DF, Fig. 5 b, P < 0.05 vs DT) pressure nociceptive thresholds, indicating a possible recovery of the damage caused to the peripheral nerves. These markers values were similar to those for the CT group when rats were treated with only dimethyl fumarate (DF, Fig. 5 a and 5 b). Neutropenia was reproduced for the DT group in this model of docetaxel-induced peripheral neuropathy and myelotoxicity with protection by dimethyl fumarate as in Scheme 3; however, neutrophil count gave some null or undetectable values that led to more severe neutropenia (Fig. 6 , approx. 0.03*10 9 /L, P < 0.0001 vs CT). Despite some point values of the neutrophil count being higher than in the DT group, there was not a significant improvement in the DT + DF group. Treatment with only dimethyl fumarate (DF group) had no effect on neutrophil count compared to the CT group (Fig. 6 ). Complete blood count and other toxicity plasma markers Table 1 shows the remaining data from hemograms to analyze changes in other cell counts due to the treatment in Scheme 3 and the effect of dimethyl fumarate. DT group had decreased counts in white blood cells (WBC, leukopenia) and red blood cells (RBC, erithropenia) compared to the CT group. Indeed, lower counts were observed in all blood cells except for basophils (#BAS). In addition, hemoglobin (HGB) levels also were lower by docetaxel administration. Dimethyl fumarate treatment did not recover normal values nor attenuate these falls in either case, but neither only dimethyl fumarate (DF group) affected counts or levels compared with the control. Additional tests were performed on blood plasma collected from the CT, DT, DT + DF and DF rats to determine other types of damage in addition to neuro and myelotoxicity and the effects of dimethyl fumarate on those. As shown in Table 1 , plasma creatinine, alkaline phosphatase (ALP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatine kinase activity (CK-MB) determinations produced lower levels in the DT group related to the non-treated group, with creatinine and CK-MB displayed values within normal physiological ranges. The levels of these markers for the DT + DF group did not show a trend towards a recovery of normal levels. Table 1 Complete blood count and other toxicity plasma markers from Wistar rats under Scheme 3 (docetaxel-induced peripheral neuropathy and myelotoxicity) and the effect of dimethyl fumarate. Test Unit GROUP CT DT DT + DF DF Normal Values WBC 10 9 /L 4.809 ± 1.463 1.164 ± 0.324 **** 1.070 ± 0.287 4.489 ± 2.098 1.96–8.25 #NEU 10 9 /L 0.961 ± 0.272 0.031± 0.041 **** 0.116 ± 0.111 0.991 ± 0.466 0.22–1.57 #LYM 10 9 /L 3.420 ± 1.194 0.591 ± 0.646 **** 0.606 ± 0.518 3.230 ± 1.636 1.41–7.11 #MON 10 9 /L 0.313 ± 0.110 0.026 ± 0.035 **** 0.040 ± 0.041 0.325 ± 0.147 0.03–0.18 #EOS 10 9 /L 0.029 ± 0.012 0.006 ± 0.007 *** 0.010 ± 0.009 0.026 ± 0.007 0.01–0.16 #BAS 10 9 /L 0.008± 0.007 0.004 ± 0.005 0.008 ± 0.007 0.006 ± 0.005 0–0.005 RBC 10 12 /L 7.893± 0.326 5.191± 0.443 **** 5.335 ± 0.371 7.825 ± 0.432 7.27–9.65 HGB g/dL 16.313 ± 0.511 0.387 11.888± 0.942 **** 12.163 ± 0.602 16.200 ± 0.886 13,7–17.6 Crea mg/dL 0.733 ± 0.142 0.568 ± 0.041* 0.528 ± 0.030 0.757 ± 0.101 0.46–0.88 ALP U/L 78.670 ± 19.512 28.711 ± 2.669*** 29.592 ± 4.751 55.211 ± 8.620** 85.4–311.7 AST U/L 75.613 ± 18.870 50.940 ± 9.319* 62.496 ± 14.107 69.851 ± 32.688 65.8–266.2 ALT U/L 29.682 ± 6.569 19.337 ± 3928* 20.679 ± 5.096 20.341 ± 5.369* 26.3–68.5 Urea mg/dL 31.899 ± 7.722 30.411 ± 6.424 29.620 ± 6.996 36.139 ± 8.608 19,62 − 23,02 CK-MB U/L 2473.161 ± 815.85 1989.088 ± 784.76 1741.884 ± 435.27 1394.350 ± 449.37** 233–4367 WBC: white blood cell count; #NEU: neutrophil count; #LYM: lymphocyte count; #MON: monocyte count; #EOS: eosinophil count; #BAS: basophil count; RBC: red blood cell count; HGB: hemoglobin; Crea: creatinine; ALP: alkaline phosphatase; AST: aspartate aminotransferase; ALT: alanine aminotransferase; CK-MB: creatine kinase; CT: control group; DT: docetaxel group; DT + DF: co-administered group; DF: dimethyl fumarate group. Data are expressed as mean ± SD and analyzed with one-way ANOVA followed by Bonferroni multiple comparison tests, n = 8. *P < 0.05, **P < 0.02, ***P < 0.001, ****P < 0.0001 vs CT. Normal values were obtained from [ 27 ], [ 28 ]. DISCUSSION Because of its toxicity profile, docetaxel treatments have been associated with 24–35% of patients that require infusion delay, 12–22% requiring dose reduction, and 11–20% discontinuing the treatment. In addition to that docetaxel remains a standard and often-used agent to improve overall survival in advanced cancer [ 29 ], these facts make it imperative to find alternatives that prevent or at least attenuate adverse effects and thus widen the therapeutic index of the treatment. As expected, treatment with docetaxel showed associated general toxicity, which was initially reflected in the weight control of the DT group in all treatment schemes (Fig. 2 ). This outcome correlates with the feeding behavior observed in the animals of this group during the treatments, which has been reported before [ 30 ][ 23 ], even in clinical studies [ 31 ]. Low levels of hemoglobin, creatinine and the plasma enzymes ALP, AST, ALT, and CK-MB (Table 1 ) in the DT group of Scheme 3 could be associated with this nutritional deficit. Zhao and co-workers [ 30 ] also observed reduced levels of ALP, AST and ALT, unlike other authors who reported the more regular chemotherapeutic-induced increase in the plasma levels of these enzymes [ 32 ]. However, cancer patients who receive docetaxel have experienced decreases in colloid osmotic plasma pressure because of a capillary protein leak syndrome seemingly induced by docetaxel [ 33 ]. An increase in plasma volume could also explain this phenomenon. The systematic decrease observed in plasma markers in the DT group of Scheme 3 (Table 1 ) could also be explained by any of these reasons. Docetaxel-induced nociceptive alterations have been studied in rats before [ 23 ], although not simultaneously along myelotoxicity markers. Increases in nociceptive thresholds of DT groups, such as paw pressure in Scheme 1 (Fig. 3 a) and hot plate in Scheme 4 (Fig. 3 c), were not considered because they are not standard outcomes in preclinical studies of chemotherapeutic-induced peripheral neuropathy. Hot plate tests did not shed significant differences in any other scheme (Fig. 3 c) and only mechanical nociceptive tests were considered reliable in this study. In this way, only schemes 3 and 4 showed significant decreases in both paw and tail pressure nociceptive thresholds (Fig. 3 a and 3 b, respectively), signals of mechanical hyperalgesia in rodents. As nociception describes the peripheral neuronal response to noxious stimuli potentially damaging tissues, we can consider those results supporting a peripheral neuropathy model [ 34 ]. Neutropenia is a well-known adverse effect induced by docetaxel [ 35 ] and rat models have been proposed for studying it [ 14 ], [ 24 ]. All the schemes set up in this work had significant decreases in the number of neutrophils due to the treatment with docetaxel compared to the control (Fig. 4 ). However, schemes 2, 3 and 4 gave counts featured as febrile neutropenia (< 0.5 x 10 9 /L). Scheme 4 was discarded as eligible for neutropenia assessment since the neutrophil count was zero or undetectable, making it challenging to propose protective agents against docetaxel-induced myelotoxicity. Taking together these results and those obtained in pain behavioral tests (paragraph above), Scheme 3 showed better attributes for being used as the definitive experimental model for simultaneous induction of peripheral neuropathy and myelotoxicity. Table 1 shows other characteristics found commonly in clinical practice with docetaxel that are modelled in Scheme 3, such as leukopenia, anemia, and lymphopenia [ 31 ], as well as other not often reported, such as low counts of monocytes and eosinophils. Using Scheme 3 as a joint model for docetaxel-induced peripheral neuropathy and myelotoxicity allowed us first to evaluate dimethyl fumarate as a neuro and myeloprotective agent and a combination alternative for cancer chemotherapy with docetaxel in the same experiment. At least, at a 100 mg/Kg dose co-administered with docetaxel, dimethyl fumarate attenuated mechanical nociceptive alterations suggesting a protective role in peripheral nerve exposure to docetaxel (Fig. 5 ). Previously, it has been shown that dimethyl fumarate exerts a protective role in peripheral neuropathy experimental rat models, as induced surgically [ 16 ] as by chemotherapeutics, such as oxaliplatin [ 36 ] and paclitaxel [ 37 ]. Even though some individual neutrophil counts were higher in the DT + DF group, no significant difference was observed concerning the DT group (Fig. 6 ). Neither was it observed that dimethyl fumarate could improve other blood cell counts or markers of general toxicity shown in Table 1 (i.e. RBC, ALP, AST). Further experiments must be done with higher doses or with a pre-treatment administration scheme of dimethyl fumarate so that a Nrf2-mediated protective response is induced before docetaxel exposure. CONCLUSIONS An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel, a very used cancer drug, was established. The model is highly reliable and allows the evaluation of promissory protective agents against both adverse effects using fewer animals. Besides, there is equivalence with a reported clinical scheme where docetaxel is administered weekly at smaller doses. By using this model, it was assessed preliminary the protective aptitude of dimethyl fumarate against docetaxel toxicity. Dimethyl fumarate showed attenuation of nociceptive alterations and no effect against docetaxel-induced neutropenia. However, the model may be yet used in a dimethyl fumarate pre-treatment mode or with higher doses. Declarations Conflict of Interest. The authors declare no conflict of interest. MN-G participates as researcher in other works funded by MINCIENCIAS. Welfare of animals. Animals were maintained, cared for and used according to Law 84 of 1989 (Congress of Colombia), Title V of Resolution 8430 of 1993 (Ministry of Health, Colombia) and the guidelines described in the Guide for the Care and Use of Laboratory Animals of the National Academy of Sciences of the United States. Protocols were approved by the Comité de Ética en Investigación Científica (CEINCI) of the Universidad Industrial de Santander (Committee Minutes No. 10 of 2021 and No.12 of 2024). Funding. This work was supported by the Ministerio de Ciencia, Tecnología e Innovación (MINCIENCIAS), Colombia, grant #722–2021. Author Contribution S.C.: methodologies setting up and performance; data acquisition and analysis; manuscript draft. M.N.: work conception and design; data interpretation; manuscript revision and correction; manuscript submitted version approval. Data Availability Some raw data supporting the findings of this study are available within the paper and its Supplementary Information. Some values of pressure and reaction time from nociceptive tests were corrected after revising video recordings. Blood cell count data was directly recorded from the equipment display and exists as Excel sheets. References New PZ, Jackson CE, Rinaldi D, Burris H, Barohn RJ (1996) Peripheral neuropathy secondary to docetaxel (Taxotere). 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Ann Oncol 15(9):1358–1365. 10.1093/annonc/mdh349 Bria E et al (2006) Weekly docetaxel as second line chemotherapy for advanced non-small-cell lung cancer: Meta-analysis of randomized trials. Dec. 10.1016/j.ctrv.2006.07.003 Center for Drug Evaluation and Research, Food, Administration D (2005) Guidance for Industry Estimating the Maximum Safe Starting Dose in Initial Clinical Trials for Therapeutics in Adult Healthy Volunteers Healthy Volunteers, [Online]. Available: http://www.fda.gov/cder/guidance/index.htm Kim ST et al (2018) Oct., Phosphatidylcholine attenuated docetaxel-induced peripheral neurotoxicity in rats, Drug Chem Toxicol, vol. 41, no. 4, pp. 476–485, 10.1080/01480545.2017.1390580 Branda RF, Powden C, Brooks EM, Yildirim Z, Naud SJ, McCormack JJ (2006) Vitamin E but not St. John’s wort mitigates leukopenia caused by cancer chemotherapy in rats. Translational Res 148(6):315–324. 10.1016/j.trsl.2006.05.007 Zhu WX, Zhao K, Chu SS, Liu ZL (2012) Evaluation of essential oil and its three main active ingredients of Chinese chenopodium ambrosioides (family: Chenopodiaceae) against blattella germanica. J Arthropod Borne Dis 6(2):90–97 Lal R, Dhaliwal J, Dhaliwal N, Dharavath RN, Chopra K (May 2021) Activation of the Nrf2/HO-1 signaling pathway by dimethyl fumarate ameliorates complete Freund’s adjuvant-induced arthritis in rats. Eur J Pharmacol 899. 10.1016/j.ejphar.2021.174044 León AC et al (2011) Goñi., Hematological and biochemical parameters in Sprague Dawley laboratory rats breed in CENPALAB, Cenp: SPRD, Revista electrónica de Veterinaria, vol. 12 Mary LA, Giknis, Clifford CB (2008) Clinical Laboratory Parameters for CrI. WI (Han) Martini A et al (2021) Sep., Predicting toxicity-related docetaxel discontinuation and overall survival in metastatic castration-resistant prostate cancer: a pooled analysis of open phase 3 clinical trial data, Prostate Cancer Prostatic Dis, vol. 24, no. 3, pp. 743–749, 10.1038/s41391-021-00326-3 Zhao M et al (2010) Aug., Evaluation of docetaxel-loaded intravenous lipid emulsion: Pharmacokinetics, tissue distribution, antitumor activity, safety and toxicity, Pharm Res, vol. 27, no. 8, pp. 1687–1702, 10.1007/s11095-010-0180-0 Hirata T et al (2021) A multicenter study of docetaxel at a dose of 100 mg/m2 in japanese patients with advanced or recurrent breast cancer. Intern Med 60(8):1183–1190. 10.2169/internalmedicine.5089-20 Feng S-Q et al (2018) Combined treatment with apatinib and docetaxel in A549 xenograft mice and its cellular pharmacokinetic basis, [Online]. Available: www.chinaphar.com Semb KA, Aamdal S, Oian P (1998) Capillary protein leak syndrome appears to explain fluid retention in cancer patients who receive docetaxel treatment., Journal of Clinical Oncology, vol. 16, no. 10, pp. 3426–3432, Oct. 10.1200/JCO.1998.16.10.3426 Deuis JR, Dvorakova LS, Vetter I (2017) Methods used to evaluate pain behaviors in rodents, Sep. 06, Frontiers Media S.A. 10.3389/fnmol.2017.00284 Frederiks CN, Lam SW, Guchelaar HJ, Boven E (2015) Genetic polymorphisms and paclitaxel- or docetaxel-induced toxicities: A systematic review. W.B. Saunders Ltd. 10.1016/j.ctrv.2015.10.010 Miyagi A, Kawashiri T, Shimizu S, Shigematsu N, Kobayashi D, Shimazoe T (2019) Dimethyl Fumarate Attenuates Oxaliplatin-Induced Peripheral Neuropathy without Affecting the Anti-tumor Activity of Oxaliplatin in Rodents Singh J et al (2022) Dimethyl Fumarate Ameliorates Paclitaxel-Induced Neuropathic Pain in Rats. Cureus Sep. 10.7759/cureus.28818 Additional Declarations No competing interests reported. Supplementary Files Rawdata.zip 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-4974358","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":358796587,"identity":"dde89c25-2b2e-48dd-992b-c83c31aa6e26","order_by":0,"name":"Sebastian Cubides-Cely","email":"","orcid":"","institution":"Universidad Industrial de Santander","correspondingAuthor":false,"prefix":"","firstName":"Sebastian","middleName":"","lastName":"Cubides-Cely","suffix":""},{"id":358796588,"identity":"5763e987-eff8-40b4-9009-6076d6e29251","order_by":1,"name":"Mario Negrette-Guzmán","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAxklEQVRIiWNgGAWjYBADOQZmEMVGghZj0rUkNjAQq0W3/ezBh1/b6tI3HGd/wPCh7DADf3sDfi1mZ/KSjWXb2HI3HOYxYJxx7jCDxJkDBLQcyDGTlmzjAWlhYOZtO8xgIJFAQMv5N+a/Jdsk0g0Osz9g/kuUlhs5Zowf2wwSDICKmRmJ0/LGWJrhXILhTKBfDvacS+ch7JfzOYYff5TVyfOdP/7wwY8yazmCIQYCzDxQBsh4HjwKEYDxB1HKRsEoGAWjYMQCALoEQVC7WHXCAAAAAElFTkSuQmCC","orcid":"","institution":"Universidad Industrial de Santander","correspondingAuthor":true,"prefix":"","firstName":"Mario","middleName":"","lastName":"Negrette-Guzmán","suffix":""}],"badges":[],"createdAt":"2024-08-25 23:49:14","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4974358/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4974358/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":66863394,"identity":"b1bb90ed-f52e-4dea-b72d-1389447c799f","added_by":"auto","created_at":"2024-10-17 08:45:22","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":784987,"visible":true,"origin":"","legend":"\u003cp\u003eDocetaxel administration schemes assessed for simultaneous induction of peripheral neuropathy and neutropenia. i.v.: intravenously.\u003c/p\u003e","description":"","filename":"Fig.1.png","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/88df33d97b0bf7f607eea245.png"},{"id":66864069,"identity":"33349e20-395e-41e0-bc1f-8a18b3609488","added_by":"auto","created_at":"2024-10-17 08:53:22","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":428436,"visible":true,"origin":"","legend":"\u003cp\u003eBody weight gain in rats of four administration schemes of docetaxel for simultaneous induction of peripheral neuropathy and neutropenia. Body weight was measured on days of docetaxel administration for schemes 1 to 4 (S1, S2, S3, and S4) and the day of euthanasia. DT: docetaxel group; CT: control group. Data are expressed as mean ±SD and analyzed by t-test, n = 8. **P\u0026lt;0.02, ***P\u0026lt;0.001 vs CT.\u003c/p\u003e","description":"","filename":"Fig.2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/a9a98f5436c926615926542b.jpg"},{"id":66863392,"identity":"e5dfefdf-d9e5-4419-b087-2998b35fd406","added_by":"auto","created_at":"2024-10-17 08:45:22","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":210801,"visible":true,"origin":"","legend":"\u003cp\u003eMechanical and thermal nociceptive thresholds in rats of four administration schemes of docetaxel for simultaneous induction of peripheral neuropathy and neutropenia. All tests were conducted four days after the last docetaxel dose administration. Values for pressure in paw (\u003cem\u003ea\u003c/em\u003e) and tail (\u003cem\u003eb\u003c/em\u003e) are given in grams (\u003cem\u003eg\u003c/em\u003e). Times of latency at 50±0.1 °C for the hot plate test (c) are in seconds (s). DT: docetaxel group; CT: control group. Data are expressed as mean ±SD and analyzed by t-test, n = 8. *P\u0026lt;0.05, **P\u0026lt;0.02, ***P\u0026lt;0.001, ****P\u0026lt;0.0001 vs. CT.\u003c/p\u003e","description":"","filename":"Fig.3.png","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/b9e3fd2df8af508f08fb6884.png"},{"id":66864067,"identity":"ac07f14d-6f05-4d1a-93f2-29ea3946a933","added_by":"auto","created_at":"2024-10-17 08:53:22","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1084796,"visible":true,"origin":"","legend":"\u003cp\u003eNeutrophil counts in rats of four administration schemes of docetaxel for simultaneous induction of peripheral neuropathy and neutropenia. DT: docetaxel group; CT: control group. Data are expressed as mean ±SD and analyzed by t-test, n = 8. *P\u0026lt;0.05, ***P\u0026lt;0.001, ****P\u0026lt;0.0001 vs CT.\u003c/p\u003e","description":"","filename":"Fig.4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/93c31761b5cecadd968b3ac1.jpg"},{"id":66863395,"identity":"129e4bb3-885c-4773-a46a-d2549ab14326","added_by":"auto","created_at":"2024-10-17 08:45:22","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":100442,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of dimethyl fumarate treatment on docetaxel-induced nociceptive alterations in Wistar rats under Scheme 3. Values for pressure in paw (a) and tail (b) are given in grams (g). CT: control group; DT: docetaxel group; DT+DF: co-administered group; DF: dimethyl fumarate group. Data are expressed as mean ±SD and analyzed with one-way ANOVA followed by Bonferroni multiple comparison tests, n = 8. ****P\u0026lt;0.0001 vs CT; *P\u0026lt;0.05 vs DT.\u003c/p\u003e","description":"","filename":"Fig.5.png","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/188f17131f831470e2978af6.png"},{"id":66863398,"identity":"ed476e2c-cde9-448e-870e-8a7f1406caaa","added_by":"auto","created_at":"2024-10-17 08:45:22","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":461479,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of dimethyl fumarate treatment on docetaxel-induced neutropenia in Wistar rats under Scheme 3.\u003cstrong\u003e \u003c/strong\u003eCT: control group; DT: docetaxel group; DT+DF: co-administered group; DF: dimethyl fumarate group. Data are expressed as mean ±SD and analyzed with one-way ANOVA followed by Bonferroni multiple comparison tests, n = 8. ****P\u0026lt;0.0001 vs CT; ns: not significant at P\u0026lt;0.05 vs DT.\u003c/p\u003e","description":"","filename":"Fig.6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/9595b12b1eae39850c3a5f88.jpg"},{"id":66866566,"identity":"c085b16d-816c-4f7c-a798-10acc09cadcd","added_by":"auto","created_at":"2024-10-17 09:09:26","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3654854,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/58f713cd-6427-4f5d-9878-7778f5f06e4a.pdf"},{"id":66864071,"identity":"0f97a2a8-fa61-4bb4-885f-3424b20cfe4f","added_by":"auto","created_at":"2024-10-17 08:53:22","extension":"zip","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":1148387,"visible":true,"origin":"","legend":"","description":"","filename":"Rawdata.zip","url":"https://assets-eu.researchsquare.com/files/rs-4974358/v1/564908f9773822c12d6ca81c.zip"}],"financialInterests":"No competing interests reported.","formattedTitle":"An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel administration: evaluating the protective role of dimethyl fumarate","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eDocetaxel is a semi-synthetic taxane, paclitaxel analog, approved by the U.S. Food and Drug Administration (FDA) for breast, ovarian, lung, and prostate cancer, among others[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Its activity relies on specific binding to beta-tubulin subunits of microtubules, inhibiting tubulin depolymerization and, thus, stabilizing microtubules. Consequently, the cell cycle is arrested in mitosis, leading to cell death [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. In addition, docetaxel is also able to lead cancer cells to intrinsic apoptosis by inhibiting the expression of anti-apoptotic genes BCL-2 and BCL-X and to arrest cancer growth by downregulating the androgen receptor signaling pathway [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Cancer cell death depends on drug concentration and exposure time. However, docetaxel can produce adverse reactions such as peripheral neuropathy, febrile neutropenia, infusion reactions, fatigue, fluid retention, pneumonitis, and gastrointestinal complications [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eTaxane-associated peripheral neurotoxicity is an acute pain syndrome consisting of diffuse muscle pain, mainly in the legs, hips and lower back, although it may spread, resulting in myalgia and/or arthralgia. Incidences of docetaxel-induced peripheral neuropathy are highly variable. However, studies have reported up to 64% of overall incidence with 7.4% of severe neuropathy [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. The exact pathophysiological mechanism of taxane-induced peripheral neurotoxicity is not yet fully described; however, axonal microtubule stability and polar reconfiguration enhanced by taxanes can alter axonal transport of different substances and mitochondria, leading to a loss of axonal integrity or degeneration, which, in turn, results in abnormal nerve physiology [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eAccording to the American Society of Clinical Oncology (ASCO) and the European Society for Medical Oncology (ESMO) guidelines, there are no recommended or effective agents to prevent chemotherapy-induced peripheral neuropathy. Clinical trials have shown that duloxetine treatment reduces pain, numbness, and tingling symptoms, but only to a moderate degree, with a better protective effect in the treatment of peripheral neuropathy due to platinum-based drugs rather than other therapies such as taxanes [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Other options, such as tricyclic antidepressants or anticonvulsant drugs, have been explored, but trials show mixed results [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e], [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eOn the other hand, neutropenia remains a condition that has no treatment other than prevention against possible infections once acquired. Severe neutropenia often requires modification of the chemotherapy regimen and may compromise anticancer efficacy. Prevention of neutropenia is an important patient benefit for reasons of safety, treatment efficacy, and cost-effectiveness [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. There are recent evidence-based guidelines for the prevention of infections associated with chemotherapy-induced neutropenia [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. However, antibiotics are the standard treatment for febrile neutropenia and are increasingly used for prophylaxis in patients at low risk of febrile neutropenia[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Granulocyte colony stimulating factors (G-CSF) stimulate the production and development of neutrophils, which are necessary to fight infections. Some G-CSF such as pegfilgrastim and filgrastim have been used for the treatment of febrile neutropenia caused by chemotherapeutics [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. On the other hand, plinabulin, a non-G-CSF-selective microtubule-binding immunomodulatory agent that possesses hematopoietic stem cell protective properties and anticancer benefits [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e], has previously been administered in animals, where improvement in neutropenia induced by various chemotherapies, including docetaxel, doxorubicin hydrochloride, and cyclophosphamide, was observed through a mechanism other than G-CSF [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eDimethyl fumarate, a fumaric acid ester (α, β-unsaturated carboxylic acid ester), is an FDA and European Medicines Agency (EMA) approved drug, which is effective in the treatment of relapsing/remitting multiple sclerosis, and is considered a type of indirect antioxidant, as it promotes the activation and stabilization of nuclear factor erythroid 2-related factor 2 (Nrf2) [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e], [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Dimethyl fumarate enters the body orally and dissociates in the intestine, where it subsequently interacts with immune cells in the blood circulation and crosses the blood-brain barrier into the central nervous system [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. There are different works involving hepatoprotection, neuroprotection and anti-arthritis protection using daily oral doses between 25 and 100 mg/kg of dimethyl fumarate for several weeks, showing an Nrf2-mediated cytoprotective effect in Wistar rats [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eAlthough peripheral neuropathy and neutropenia are widely reported chemotherapy-induced adverse effects, dose reduction or discontinuation of treatment remains the more effective treatment for their co-management. More preclinical investigation on strategies for inhibiting or attenuating docetaxel-induced peripheral neuropathy and neutropenia without affecting its therapeutic efficacy is required, ensuring the quality of data and reducing the number of research subjects ethically. In this work, we establish a rat model for simultaneous induction of myelotoxicity and peripheral neuropathy by docetaxel, which is useful for studying protective therapeutic alternatives and allows the evaluation of both effects in the same animal. Four different administration schemes were tested based on clinical information and experimental models. Sensory behavioral tests were performed to obtain markers of peripheral neuropathy, and blood was obtained to determine neutropenia. The scheme representing changes better in both nociceptive thresholds (peripheral neuropathy) and neutropenia (myelotoxicity) was chosen. Using this experimental model, we first report that dimethyl fumarate is a potential protective agent against docetaxel-induced peripheral neuropathy, but not against neutropenia.\u003c/p\u003e"},{"header":"MATERIAL AND METHODS","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eDrugs\u003c/h2\u003e \u003cp\u003eDocetaxel was purchased from BLAŪ Farmac\u0026eacute;utica, (Brazil), and prepared and mixed according to the manufacturer's instructions. Dimethyl fumarate was purchased from Sigma-Aldrich (St Loius, MI, USA) and was resuspended in carboxyl methyl cellulose (CMC, Sigma-Aldrich, St Loius, MI, USA), which was prepared at 1% by diluting in distilled water.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eAnimals\u003c/h2\u003e \u003cp\u003eMale Wistar rats weighing 240\u0026thinsp;\u0026plusmn;\u0026thinsp;30 g at the start of the treatments were used. Animals were maintained, cared for and used according to Law 84 of 1989 (Congress of Colombia), Title V of Resolution 8430 of 1993 (Ministry of Health, Colombia) and the guidelines described in the Guide for the Care and Use of Laboratory Animals of the National Academy of Sciences of the United States [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Animals were kept in stainless steel cages 30x40 cm x20 cm high (6 rats per cage) at a temperature of 21\u0026deg;C \u0026minus;\u0026thinsp;24\u0026deg;C, relative humidity of 55\u0026ndash;70%, photoperiod of 12 hours of light and 12 hours of darkness with sterile water and specialized food \u003cem\u003ead libitum\u003c/em\u003e. Protocols were approved by the Comit\u0026eacute; de \u0026Eacute;tica en Investigaci\u0026oacute;n Cient\u0026iacute;fica (CEINCI) of the Universidad Industrial de Santander (Committee Minutes No. 10 of 2021 and No.12 of 2024).\u003c/p\u003e \u003cp\u003eRats were habituated and trained daily, always in the morning hours, to diminish stress levels and improve the reliability of the model. Four days before the start of administration schemes, rats were habituated to the room, caretakers, researchers, and equipment used in experiments. Animals also underwent individual training to teach them to remain still and calm during oral administration, restraining for intravenous administration, and using analgesimeters. At the end of this period, the change in the behavior of rats was observed.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eEstablishment of the docetaxel damage model\u003c/h2\u003e \u003cp\u003eFour docetaxel administration schemes were tested varying dose and dosage (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Scheme 1 consisted of a 15 mg/kg single dose. Scheme 2 included two doses of 15 mg/kg administered three weeks apart. In Scheme 3, a 5 mg/kg dose was administered weekly until 6 doses were completed. Scheme 4 consisted of three 15 mg/kg doses each administered every 3 weeks. In all cases, docetaxel was administered intravenously in the tail. Each scheme evaluated two experimental groups, the control group (CT, n\u0026thinsp;=\u0026thinsp;8), which was administered normal saline, and the docetaxel-treated group (DT, n\u0026thinsp;=\u0026thinsp;8). Administration schemes were proposed based on clinical practice and trials from literature [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e], [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e] equivalent for rats [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. However, some reports where either peripheral neuropathy or neutropenia was induced in rats, but not simultaneously, were also considered [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e], [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e], [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eBehavioral testing, euthanasia and blood cell count\u003c/h2\u003e \u003cp\u003eFour days after the last administration in each scheme, sensory behavioral testing paw and tail pressure and hot plate were performed using the PANLAB analgesimeter model LE7306 Randall-Selitto and the PANLAB analgesimeter model LE7406 Hot Plate (Panlab, S.L.U., Barcelona, Spain), respectively. Nociceptive testing was based on physical indicators of discomfort (withdrawal reflexes, licking and voicing). To ensure ethical treatment of the animals, maximum cut-off values of 10 seconds of paw and tail pressure latency and a weight of up to 650 g were adjusted. For the case of the hot plate method, 60 seconds of latency at 50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1\u0026deg;C was taken as the cut-off. All tests were recorded in video format for later data verification and analysis. Once the nociceptive tests were concluded, the rats were euthanized under anesthesia with ketamine (90 mg/kg)\u0026thinsp;+\u0026thinsp;xylazine (15 mg/kg) intraperitoneally (i.p.) by exsanguination through the left ventricle of the animal.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eHemograms and blood biochemistry\u003c/h2\u003e \u003cp\u003eComplete differential cell counts were performed in blood samples to determine neutropenia using a Genvet VH50 hematology analyzer (Genrui Biotech Inc., Shenzhen, China). Blood biochemistry tests were performed using kits commercially available following the manufacturer's instructions (LabTest Diagn\u0026oacute;stica S.A., Minas Gerais, Brazil); creatine kinase activity (CK-MB. Ref. 118), aspartate aminotransferase (AST, Ref. 109), alanine aminotransferase (ALT, Ref. 1008), alkaline phosphatase (Ref. 79), creatinine K concentration (Ref. 96), and urea (Ref. 104) were analyzed.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eAssessment of dimethyl fumarate protection\u003c/h2\u003e \u003cp\u003eOnce selected, the best scheme that ensured neutropenia and peripheral neurotoxicity was challenged with a protection alternative proposed: dimethyl fumarate, a known Nrf2 inducer and FDA-approved drug. This administration scheme consisted of a control group (CT, n\u0026thinsp;=\u0026thinsp;8), which received saline i.v. as a vehicle for docetaxel and oral carboxymethylcellulose (CMC) as a vehicle for dimethyl fumarate; a docetaxel group (DT, n\u0026thinsp;=\u0026thinsp;8), administered with docetaxel i.v. along with CMC; a co-treated group (DT\u0026thinsp;+\u0026thinsp;DF, n\u0026thinsp;=\u0026thinsp;8) administered with docetaxel i.v. and dimethyl fumarate orally at 100 mg/Kg/day [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]; an additional control group was administered with only dimethyl fumarate (DF, n\u0026thinsp;=\u0026thinsp;8) and saline as vehicle for docetaxel. Docetaxel or its vehicle were administered according to the scheme selected for simultaneous induction of nociceptive alterations and neutropenia. Dimethyl fumarate or CMC were administered one hour before the administration of docetaxel (or its vehicle).\u003c/p\u003e \u003cp\u003eAs previously described for the establishment of the damage model, four days after the end of the treatment, animals were subjected to sensory behavioral tests and subsequently euthanized, obtaining blood for hemograms and blood biochemistry. In addition, sciatic nerves were obtained and stored for further studies on mechanisms of damage and protection.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eAll results were expressed as the mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation of data. Data processing was performed by t-test analysis comparing groups DT and CT for each of the four evaluated damage schemes. To compare between groups in the protection model, one-way ANOVA followed by Bonferroni multiple comparison tests were used. Significance was determined at p\u0026thinsp;\u0026le;\u0026thinsp;0.05. All data were processed, plotted and analyzed using Prism 8 (GraphPad Software).\u003c/p\u003e \u003c/div\u003e"},{"header":"RESULTS","content":"\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eSimultaneous induction of nociceptive alterations and neutropenia using docetaxel\u003c/h2\u003e \u003cp\u003eDocetaxel toxicity was evident throughout the treatment as animal behavior and feeding and living habits were changing. Rats treated with the drug showed a significant lag in body weight gain compared to the control rats, mainly in schemes 3 and 4 (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSchemes 1, 3 and 4 showed differences between the groups in the paw pressure test (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea), although docetaxel induced an increase in the nociceptive threshold due to Scheme 1 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05), which is not a canonical outcome. The mean pain threshold of DT rats in Scheme 3 was almost a third of that for group CT (P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). Despite the entire dose in Scheme 2 being twice the dose in Scheme 1, no significant differences between groups were observed in Scheme 2.\u003c/p\u003e \u003cp\u003eIn the tail pressure test (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb), the group DT of schemes 2, 3 and 4 had significant falls in nociceptive thresholds compared to the control group. Scheme 1 did not show differences between groups for this test. On the other hand, of the 4 schemes administered, only Scheme 4 showed an alteration in the hot plate test (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec, P\u0026thinsp;\u0026lt;\u0026thinsp;0.05). However, this alteration, likewise in the paw pressure test of Scheme 1, had an increasing trend in the nociceptive threshold, which is not frequently reported in this kind of study. None of the remaining treatments with docetaxel (schemes 1\u0026ndash;3) gave a significant response compared to the CT group in the hot plate test (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eOnce the pain behavioral tests were completed, euthanasia was performed, and blood was taken for the differential neutrophil count. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e, all the docetaxel administration schemes led to neutropenia, with neutrophil count decreases of 49% for Scheme 1 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05), 75% for Scheme 2 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001), 80% for Scheme 3 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001), and 100% for Scheme 4 (P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). Schemes 2, 3 and 4 showed a feature of febrile neutropenia, counts\u0026thinsp;\u0026lt;\u0026thinsp;0.5*10\u003csup\u003e9\u003c/sup\u003e/L. However, null or non-detectable neutrophil counts obtained in Scheme 4 may make it few attractive experimentally.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSince the results shown so far, Scheme 3 joins together most of the characteristics desirable for the experimental model of simultaneous induction. First, decreases in paw and tail pressure thresholds (two of the three nociceptive tests assessed) and neutrophil counts were observed with good significance, which validates and gives confidence to the model. Although Scheme 4 showed similar features, the undetectable neutrophil counts would make it difficult to evaluate potential myeloprotective agents. In addition, this severe neutropenia with null cell counts in Scheme 4 is not the most observed clinical outcome. Schemes 1 and 2 only showed responses in one of the nociceptive tests executed. Even though Scheme 3 implies the largest number of doses over a long period, its outcomes demonstrated that it is the completer and most reliable scheme for evaluating potential neuro and myeloprotective chemotherapeutics. The following results were obtained by applying Scheme 3 as a model of simultaneous induction of peripheral neurotoxicity and myelotoxicity by docetaxel.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eProtective role of dimethyl fumarate against docetaxel-induced nociceptive alterations and neutropenia\u003c/h2\u003e \u003cp\u003eAs was seen above, animals treated with docetaxel (group DT) showed a significant decrease in body weight gain throughout the treatment. This trend was not counteracted or attenuated in the group co-treated with dimethyl fumarate and docetaxel (DT\u0026thinsp;+\u0026thinsp;DF). Regarding dimethyl fumarate-treated group (DF), no difference was observed with respect to the control group (CT, data not shown). Nociceptive thresholds were reproduced for the DT group according to the damage model obtained, as shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e. Treatment with dimethyl fumarate allowed an attenuation of docetaxel-induced decreases in paw (DT\u0026thinsp;+\u0026thinsp;DF, Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea, P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 vs DT) and tail (DT\u0026thinsp;+\u0026thinsp;DF, Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb, P\u0026thinsp;\u0026lt;\u0026thinsp;0.05 vs DT) pressure nociceptive thresholds, indicating a possible recovery of the damage caused to the peripheral nerves. These markers values were similar to those for the CT group when rats were treated with only dimethyl fumarate (DF, Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eNeutropenia was reproduced for the DT group in this model of docetaxel-induced peripheral neuropathy and myelotoxicity with protection by dimethyl fumarate as in Scheme 3; however, neutrophil count gave some null or undetectable values that led to more severe neutropenia (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e, approx. 0.03*10\u003csup\u003e9\u003c/sup\u003e/L, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001 vs CT). Despite some point values of the neutrophil count being higher than in the DT group, there was not a significant improvement in the DT\u0026thinsp;+\u0026thinsp;DF group. Treatment with only dimethyl fumarate (DF group) had no effect on neutrophil count compared to the CT group (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eComplete blood count and other toxicity plasma markers\u003c/h2\u003e \u003cp\u003eTable\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e shows the remaining data from hemograms to analyze changes in other cell counts due to the treatment in Scheme 3 and the effect of dimethyl fumarate. DT group had decreased counts in white blood cells (WBC, leukopenia) and red blood cells (RBC, erithropenia) compared to the CT group. Indeed, lower counts were observed in all blood cells except for basophils (#BAS). In addition, hemoglobin (HGB) levels also were lower by docetaxel administration. Dimethyl fumarate treatment did not recover normal values nor attenuate these falls in either case, but neither only dimethyl fumarate (DF group) affected counts or levels compared with the control.\u003c/p\u003e \u003cp\u003eAdditional tests were performed on blood plasma collected from the CT, DT, DT\u0026thinsp;+\u0026thinsp;DF and DF rats to determine other types of damage in addition to neuro and myelotoxicity and the effects of dimethyl fumarate on those. As shown in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, plasma creatinine, alkaline phosphatase (ALP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatine kinase activity (CK-MB) determinations produced lower levels in the DT group related to the non-treated group, with creatinine and CK-MB displayed values within normal physiological ranges. The levels of these markers for the DT\u0026thinsp;+\u0026thinsp;DF group did not show a trend towards a recovery of normal levels.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComplete blood count and other toxicity plasma markers from Wistar rats under Scheme 3 (docetaxel-induced peripheral neuropathy and myelotoxicity) and the effect of dimethyl fumarate.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"7\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eTest\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\" morerows=\"1\" rowspan=\"2\"\u003e \u003cp\u003eUnit\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c6\" namest=\"c3\"\u003e \u003cp\u003eGROUP\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eCT\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eDT\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eDT\u0026thinsp;+\u0026thinsp;DF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eDF\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eNormal Values\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eWBC\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e4.809\u0026thinsp;\u0026plusmn;\u0026thinsp;1.463\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1.164\u0026thinsp;\u0026plusmn;\u0026thinsp;0.324 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.070\u0026thinsp;\u0026plusmn;\u0026thinsp;0.287\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e4.489\u0026thinsp;\u0026plusmn;\u0026thinsp;2.098\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e1.96\u0026ndash;8.25\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e#NEU\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.961\u0026thinsp;\u0026plusmn;\u0026thinsp;0.272\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.031\u0026plusmn;\u003c/p\u003e \u003cp\u003e0.041 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.116\u0026thinsp;\u0026plusmn;\u0026thinsp;0.111\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.991\u0026thinsp;\u0026plusmn;\u0026thinsp;0.466\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.22\u0026ndash;1.57\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e#LYM\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e3.420\u0026thinsp;\u0026plusmn;\u0026thinsp;1.194\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.591\u0026thinsp;\u0026plusmn;\u0026thinsp;0.646 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.606\u0026thinsp;\u0026plusmn;\u0026thinsp;0.518\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e3.230\u0026thinsp;\u0026plusmn;\u0026thinsp;1.636\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e1.41\u0026ndash;7.11\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e#MON\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.313\u0026thinsp;\u0026plusmn;\u0026thinsp;0.110\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.026\u0026thinsp;\u0026plusmn;\u0026thinsp;0.035 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.040\u0026thinsp;\u0026plusmn;\u0026thinsp;0.041\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.325\u0026thinsp;\u0026plusmn;\u0026thinsp;0.147\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.03\u0026ndash;0.18\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e#EOS\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.029\u0026thinsp;\u0026plusmn;\u0026thinsp;0.012\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.006\u0026thinsp;\u0026plusmn;\u0026thinsp;0.007 ***\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.010\u0026thinsp;\u0026plusmn;\u0026thinsp;0.009\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.026\u0026thinsp;\u0026plusmn;\u0026thinsp;0.007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.01\u0026ndash;0.16\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003e#BAS\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e9\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.008\u0026plusmn;\u003c/p\u003e \u003cp\u003e0.007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.004\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.008\u003c/p\u003e \u003cp\u003e\u0026plusmn;\u0026thinsp;0.007\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.006\u0026thinsp;\u0026plusmn;\u0026thinsp;0.005\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0\u0026ndash;0.005\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eRBC\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10\u003csup\u003e12\u003c/sup\u003e/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e7.893\u0026plusmn;\u003c/p\u003e \u003cp\u003e0.326\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e5.191\u0026plusmn;\u003c/p\u003e \u003cp\u003e0.443 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5.335\u003c/p\u003e \u003cp\u003e\u0026plusmn;\u0026thinsp;0.371\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e7.825\u0026thinsp;\u0026plusmn;\u0026thinsp;0.432\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e7.27\u0026ndash;9.65\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eHGB\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eg/dL\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e16.313\u0026thinsp;\u0026plusmn;\u0026thinsp;0.511\u003c/p\u003e \u003cp\u003e0.387\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e11.888\u0026plusmn;\u003c/p\u003e \u003cp\u003e0.942 ****\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e12.163\u003c/p\u003e \u003cp\u003e\u0026plusmn;\u0026thinsp;0.602\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e16.200\u0026thinsp;\u0026plusmn;\u0026thinsp;0.886\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e13,7\u0026ndash;17.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eCrea\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg/dL\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e0.733\u0026thinsp;\u0026plusmn;\u0026thinsp;0.142\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e0.568\u0026thinsp;\u0026plusmn;\u0026thinsp;0.041*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.528\u0026thinsp;\u0026plusmn;\u0026thinsp;0.030\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e0.757\u0026thinsp;\u0026plusmn;\u0026thinsp;0.101\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e0.46\u0026ndash;0.88\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eALP\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eU/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e78.670\u0026thinsp;\u0026plusmn;\u0026thinsp;19.512\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e28.711\u0026thinsp;\u0026plusmn;\u0026thinsp;2.669***\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e29.592\u0026thinsp;\u0026plusmn;\u0026thinsp;4.751\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e55.211\u0026thinsp;\u0026plusmn;\u0026thinsp;8.620**\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e85.4\u0026ndash;311.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eAST\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eU/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e75.613\u0026thinsp;\u0026plusmn;\u0026thinsp;18.870\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e50.940\u0026thinsp;\u0026plusmn;\u0026thinsp;9.319*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e62.496\u0026thinsp;\u0026plusmn;\u0026thinsp;14.107\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e69.851\u0026thinsp;\u0026plusmn;\u0026thinsp;32.688\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e65.8\u0026ndash;266.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eALT\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eU/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e29.682\u0026thinsp;\u0026plusmn;\u0026thinsp;6.569\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e19.337\u0026thinsp;\u0026plusmn;\u0026thinsp;3928*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e20.679\u0026thinsp;\u0026plusmn;\u0026thinsp;5.096\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e20.341\u0026thinsp;\u0026plusmn;\u0026thinsp;5.369*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e26.3\u0026ndash;68.5\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eUrea\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003emg/dL\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e31.899\u0026thinsp;\u0026plusmn;\u0026thinsp;7.722\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e30.411\u0026thinsp;\u0026plusmn;\u0026thinsp;6.424\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e29.620\u0026thinsp;\u0026plusmn;\u0026thinsp;6.996\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e36.139\u0026thinsp;\u0026plusmn;\u0026thinsp;8.608\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e19,62\u0026thinsp;\u0026minus;\u0026thinsp;23,02\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eCK-MB\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eU/L\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e \u003cp\u003e2473.161\u0026thinsp;\u0026plusmn;\u0026thinsp;815.85\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c4\"\u003e \u003cp\u003e1989.088\u0026thinsp;\u0026plusmn;\u0026thinsp;784.76\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1741.884\u0026thinsp;\u0026plusmn;\u0026thinsp;435.27\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c6\"\u003e \u003cp\u003e1394.350\u0026thinsp;\u0026plusmn;\u0026thinsp;449.37**\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e233\u0026ndash;4367\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eWBC: white blood cell count; #NEU: neutrophil count; #LYM: lymphocyte count; #MON: monocyte count; #EOS: eosinophil count; #BAS: basophil count; RBC: red blood cell count; HGB: hemoglobin; Crea: creatinine; ALP: alkaline phosphatase; AST: aspartate aminotransferase; ALT: alanine aminotransferase; CK-MB: creatine kinase; CT: control group; DT: docetaxel group; DT\u0026thinsp;+\u0026thinsp;DF: co-administered group; DF: dimethyl fumarate group. Data are expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD and analyzed with one-way ANOVA followed by Bonferroni multiple comparison tests, n\u0026thinsp;=\u0026thinsp;8. *P\u0026thinsp;\u0026lt;\u0026thinsp;0.05, **P\u0026thinsp;\u0026lt;\u0026thinsp;0.02, ***P\u0026thinsp;\u0026lt;\u0026thinsp;0.001, ****P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001 vs CT. Normal values were obtained from [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e], [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eBecause of its toxicity profile, docetaxel treatments have been associated with 24\u0026ndash;35% of patients that require infusion delay, 12\u0026ndash;22% requiring dose reduction, and 11\u0026ndash;20% discontinuing the treatment. In addition to that docetaxel remains a standard and often-used agent to improve overall survival in advanced cancer [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e], these facts make it imperative to find alternatives that prevent or at least attenuate adverse effects and thus widen the therapeutic index of the treatment.\u003c/p\u003e \u003cp\u003eAs expected, treatment with docetaxel showed associated general toxicity, which was initially reflected in the weight control of the DT group in all treatment schemes (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). This outcome correlates with the feeding behavior observed in the animals of this group during the treatments, which has been reported before [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e][\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e], even in clinical studies [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. Low levels of hemoglobin, creatinine and the plasma enzymes ALP, AST, ALT, and CK-MB (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) in the DT group of Scheme 3 could be associated with this nutritional deficit. Zhao and co-workers [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e] also observed reduced levels of ALP, AST and ALT, unlike other authors who reported the more regular chemotherapeutic-induced increase in the plasma levels of these enzymes [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. However, cancer patients who receive docetaxel have experienced decreases in colloid osmotic plasma pressure because of a capillary protein leak syndrome seemingly induced by docetaxel [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. An increase in plasma volume could also explain this phenomenon. The systematic decrease observed in plasma markers in the DT group of Scheme 3 (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) could also be explained by any of these reasons.\u003c/p\u003e \u003cp\u003eDocetaxel-induced nociceptive alterations have been studied in rats before [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e], although not simultaneously along myelotoxicity markers. Increases in nociceptive thresholds of DT groups, such as paw pressure in Scheme 1 (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea) and hot plate in Scheme 4 (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec), were not considered because they are not standard outcomes in preclinical studies of chemotherapeutic-induced peripheral neuropathy. Hot plate tests did not shed significant differences in any other scheme (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec) and only mechanical nociceptive tests were considered reliable in this study. In this way, only schemes 3 and 4 showed significant decreases in both paw and tail pressure nociceptive thresholds (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb, respectively), signals of mechanical hyperalgesia in rodents. As nociception describes the peripheral neuronal response to noxious stimuli potentially damaging tissues, we can consider those results supporting a peripheral neuropathy model [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eNeutropenia is a well-known adverse effect induced by docetaxel [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e] and rat models have been proposed for studying it [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e], [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. All the schemes set up in this work had significant decreases in the number of neutrophils due to the treatment with docetaxel compared to the control (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). However, schemes 2, 3 and 4 gave counts featured as febrile neutropenia (\u0026lt;\u0026thinsp;0.5 x 10\u003csup\u003e9\u003c/sup\u003e/L). Scheme 4 was discarded as eligible for neutropenia assessment since the neutrophil count was zero or undetectable, making it challenging to propose protective agents against docetaxel-induced myelotoxicity. Taking together these results and those obtained in pain behavioral tests (paragraph above), Scheme 3 showed better attributes for being used as the definitive experimental model for simultaneous induction of peripheral neuropathy and myelotoxicity. Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e shows other characteristics found commonly in clinical practice with docetaxel that are modelled in Scheme 3, such as leukopenia, anemia, and lymphopenia [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e], as well as other not often reported, such as low counts of monocytes and eosinophils.\u003c/p\u003e \u003cp\u003eUsing Scheme 3 as a joint model for docetaxel-induced peripheral neuropathy and myelotoxicity allowed us first to evaluate dimethyl fumarate as a neuro and myeloprotective agent and a combination alternative for cancer chemotherapy with docetaxel in the same experiment. At least, at a 100 mg/Kg dose co-administered with docetaxel, dimethyl fumarate attenuated mechanical nociceptive alterations suggesting a protective role in peripheral nerve exposure to docetaxel (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). Previously, it has been shown that dimethyl fumarate exerts a protective role in peripheral neuropathy experimental rat models, as induced surgically [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e] as by chemotherapeutics, such as oxaliplatin [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e] and paclitaxel [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Even though some individual neutrophil counts were higher in the DT\u0026thinsp;+\u0026thinsp;DF group, no significant difference was observed concerning the DT group (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). Neither was it observed that dimethyl fumarate could improve other blood cell counts or markers of general toxicity shown in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e (i.e. RBC, ALP, AST). Further experiments must be done with higher doses or with a pre-treatment administration scheme of dimethyl fumarate so that a Nrf2-mediated protective response is induced before docetaxel exposure.\u003c/p\u003e"},{"header":"CONCLUSIONS","content":"\u003cp\u003eAn experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel, a very used cancer drug, was established. The model is highly reliable and allows the evaluation of promissory protective agents against both adverse effects using fewer animals. Besides, there is equivalence with a reported clinical scheme where docetaxel is administered weekly at smaller doses. By using this model, it was assessed preliminary the protective aptitude of dimethyl fumarate against docetaxel toxicity. Dimethyl fumarate showed attenuation of nociceptive alterations and no effect against docetaxel-induced neutropenia. However, the model may be yet used in a dimethyl fumarate pre-treatment mode or with higher doses.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eConflict of Interest.\u003c/h2\u003e \u003cp\u003eThe authors declare no conflict of interest. MN-G participates as researcher in other works funded by MINCIENCIAS.\u003c/p\u003e \u003ch2\u003eWelfare of animals.\u003c/h2\u003e \u003cp\u003e Animals were maintained, cared for and used according to Law 84 of 1989 (Congress of Colombia), Title V of Resolution 8430 of 1993 (Ministry of Health, Colombia) and the guidelines described in the Guide for the Care and Use of Laboratory Animals of the National Academy of Sciences of the United States. Protocols were approved by the Comit\u0026eacute; de \u0026Eacute;tica en Investigaci\u0026oacute;n Cient\u0026iacute;fica (CEINCI) of the Universidad Industrial de Santander (Committee Minutes No. 10 of 2021 and No.12 of 2024).\u003c/p\u003e\u003ch2\u003eFunding.\u003c/h2\u003e \u003cp\u003eThis work was supported by the Ministerio de Ciencia, Tecnolog\u0026iacute;a e Innovaci\u0026oacute;n (MINCIENCIAS), Colombia, grant #722\u0026ndash;2021.\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eS.C.: methodologies setting up and performance; data acquisition and analysis; manuscript draft. M.N.: work conception and design; data interpretation; manuscript revision and correction; manuscript submitted version approval.\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eSome raw data supporting the findings of this study are available within the paper and its Supplementary Information. Some values of pressure and reaction time from nociceptive tests were corrected after revising video recordings. Blood cell count data was directly recorded from the equipment display and exists as Excel sheets.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eNew PZ, Jackson CE, Rinaldi D, Burris H, Barohn RJ (1996) Peripheral neuropathy secondary to docetaxel (Taxotere). Am Acad Neurol 46:108\u0026ndash;111\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZehua M et al (2022) Docetaxel remodels prostate cancer immune microenvironment and enhances checkpoint inhibitor-based immunotherapy. 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W.B. Saunders Ltd. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1016/j.ctrv.2015.10.010\u003c/span\u003e\u003cspan address=\"10.1016/j.ctrv.2015.10.010\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMiyagi A, Kawashiri T, Shimizu S, Shigematsu N, Kobayashi D, Shimazoe T (2019) Dimethyl Fumarate Attenuates Oxaliplatin-Induced Peripheral Neuropathy without Affecting the Anti-tumor Activity of Oxaliplatin in Rodents\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSingh J et al (2022) Dimethyl Fumarate Ameliorates Paclitaxel-Induced Neuropathic Pain in Rats. Cureus Sep. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.7759/cureus.28818\u003c/span\u003e\u003cspan address=\"10.7759/cureus.28818\" 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":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":"docetaxel, peripheral neuropathy, nociception, myelotoxicity, neutropenia, dimethyl fumarate","lastPublishedDoi":"10.21203/rs.3.rs-4974358/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4974358/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003ePurpose\u003c/h2\u003e \u003cp\u003eDocetaxel is a taxane antineoplastic widely used against different types of cancer. However, its efficacy is limited mainly by its myelotoxicity and peripheral neuropathy with neutropenia and nociceptive alterations as the main clinical signs, respectively. These adverse effects undermine the quality of life of patients leading them to treatment withdrawal. In this study, we set up a unique preclinical scheme for induction of both effects associated with docetaxel administration, in such a way that we can evaluate them in the same animal and carry out future protection trials.\u003c/p\u003e\u003ch2\u003eMethods\u003c/h2\u003e \u003cp\u003eFour docetaxel administration schemes were tested varying dose and dosage. Four days after the last dose, behavioral/sensory paw pressure, tail pressure and hot plate tests were conducted. Next, euthanasia was performed, and blood was obtained for total cell count and other toxicological markers. Once the scheme that better showed significant alterations in both nociception and neutropenia was chosen, joint administration with 100 mg/kg/day oral dimethyl fumarate was carried out to evaluate its protective effect.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eA scheme with six doses (5 mg/kg) of docetaxel administered weekly was chosen for protection trials. Dimethyl fumarate showed protection in nociceptive tests compared to the damage group. However, it did not show protection against neutropenia.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eThe confirmed experimental model is clinically representative as it was designed for rats through equivalent data obtained from clinical assays. It was useful to evaluate the protective potential of dimethyl fumarate, showing how it could attenuate docetaxel-induced peripheral neuropathy, but not neutropenia.\u003c/p\u003e","manuscriptTitle":"An experimental rat model for simultaneous induction of peripheral neuropathy and myelotoxicity by docetaxel administration: evaluating the protective role of dimethyl fumarate","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-10-17 08:45:17","doi":"10.21203/rs.3.rs-4974358/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":"d26e720c-2608-46a3-8b40-4b3a3679f74c","owner":[],"postedDate":"October 17th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-10-17T08:45:20+00:00","versionOfRecord":[],"versionCreatedAt":"2024-10-17 08:45:17","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-4974358","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4974358","identity":"rs-4974358","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}