Toxicity Of CPT-11 Research Articles

Role of MRPs transporters in pharmacokinetics and intestinal toxicity of irinotecan

To identify additional genetic markers contributing to variability in CPT-11 disposition and toxicity, we assessed impact of the multiple drug-resistant transporters 1, 2, and 3 (MRP1, MRP2, and MRP3) on the intestinal toxicity, pharmacokinetics, tissue distribution and biliary excretion of CPT-11 using a knockout mouse model. Mrp1/3 knockout had minor impact on intestinal toxicity of CPT-11, tissue distribution, biliary excretion, and PK parameter of its active metabolites SN38. Conversely, Mrp2−/− mice, with low carboxylesterase activity, displayed insensitivity to CPT-11 toxicity due to reduced intestinal exposure to SN38. In PK studies, Mrp1/2 knockout significantly increased the AUC of CPT-11 compared to their AUC in FVB mice. However, the AUC of SN38 in Mrp2 −/− mice was decreased by 3.25-fold. Mrp3 knockout only slightly increased SN38 plasma exposure. Lastly, Mrp2/3 knockout increased biliary excretion amount of CPT-11 by 67.2% and 48.5% compared to wild-type mice, respectively. Consequently, Mrp1/3 deficiency didn't change SN38 tissue distribution. Finally, correlation analysis demonstrated that tissue exposure to SN38 was better correlated with toxicity than plasma AUC of SN38. Mrp1/2/3 deficiency showed a minor impact on PK, biliary excretion, distribution and intestinal exposure of SN38, and as a result, did not affect the intestinal toxicity of CPT-11.

Investigation of the Possible Protective Effect of N-Acetylcysteine (NAC) against Irinotecan (CPT-11)-Induced Toxicity in Rats.

Irinotecan (CPT-11) is a chemotherapeutic agent involved in the treatment regimens for several malignancies such as colorectal cancer. N-acetylcysteine (NAC) is a strong antioxidant and anti-inflammatory agent used in the treatment of several diseases related to oxidative stress and inflammation. This study aimed at investigating whether NAC provides protection against hepatorenal and gastrointestinal tissue damage induced by CPT-11. Thirty-two Wistar albino rats were divided into four groups as control, NAC, CPT-11, and CPT-11+NAC. Following the experimental period, blood, and tissue samples (liver, kidney, stomach, and small intestine) were collected, and biochemical indicators, together with pro-inflammatory cytokines (TNF-α and IL-1β), matrix metalloproteinases (MMPs), malondialdehyde (MDA), glutathione peroxidase (GPx) and superoxide dismutase (SOD) levels were evaluated. Both the biochemical indicators and the pro-inflammatory cytokines, MMP, and MDA levels increased in animals treated with CPT-11, while SOD and GPx activities decreased. Histopathological evaluation revealed structural damage in all examined tissues. With NAC administration, significant improvements were observed, both biochemically and histologically. In conclusion, the results of the present study suggest that NAC treatment together with CPT-11 may have a beneficial effect on reducing CPT-11 toxicity in rats, by modulating inflammation and the oxidant-antioxidant balance. These results strongly promote further investigative studies.

Berberine Improves Irinotecan-Induced Intestinal Mucositis Without Impairing the Anti-colorectal Cancer Efficacy of Irinotecan by Inhibiting Bacterial β-glucuronidase.

Irinotecan (CPT11), a broad-spectrum cytotoxic anticancer agent, induces a series of toxic side-effects. The most conspicuous side-effect is gastrointestinal mucositis, including nausea, vomiting, and diarrhea. A growing body of evidence indicates that bacteria β-glucuronidase (GUS), an enzyme expressed by intestinal microbiota, converts the inactive CPT11 metabolite SN38G to the active metabolite SN38 to ultimately induce intestinal mucositis. We sought to explore the potential efficacy and underlying mechanisms of berberine on CPT11-induced mucositis. Our study showed that berberine (50 mg/kg; i. g.) mitigated the CPT11-induced loss of mucosal architecture, ulceration, and neutrophil infiltration. Meanwhile, berberine improved mucosal barrier function by increasing the number of globlet cells, protecting trans-endothelial electrical resistance (TEER), reducing permeability and increasing tight junction proteins expression. LC-MS analysis showed that berberine decreased the content of SN38 in feces, which correlated with decreases in both GUS activity and GUS-producing bacteria. Further molecular docking and Lineweaver-Burk plots analyses suggested that berberine functions as a potential non-competitive inhibitor against GUS enzyme. Of note, berberine maintained the anti-tumor efficacy of CPT11 in a tumor xenograft model while abrogating the intestinal toxicity of CPT11. Overall, we identified for the first time the remission effects of berberine on intestinal mucositis induced by CPT11 without impairing the anti-colorectal cancer efficacy of CPT11 partially via inhibiting bacterial GUS enzyme.

Significance of aldo-keto reductase 1C3 and ATP-binding cassette transporter B1 in gain of irinotecan resistance in colon cancer cells

Irinotecan (CPT11) is widely prescribed for treatment of various intractable cancers such as advanced and metastatic colon cancer cells, but its continuous treatment promotes the resistance development. In this study, we established CPT11-resistant variants of three human colon cancer (DLD1, RKO and LoVo) cell lines, and found that gain of the resistance elicited an up-regulation of aldo-keto reductase (AKR) 1C3 in the cells. Additionally, the sensitivity to CPT11 toxicity was decreased and increased by overexpression and knockdown, respectively, of the enzyme. Moreover, the resistant cells suppressed formation of reactive 4-hydroxy-2-nonenal by CPT11 treatment, and the suppressive effect was almost completely abolished by addition of an AKR1C3 inhibitor. These results suggest that up-regulated AKR1C3 contributes to promotion of the chemoresistance by detoxifying the reactive aldehyde. Western blot and real-time polymerase-chain reaction analyses and ATP-binding cassette (ABC) B1-functional assay revealed that, among three ABC transporters, ABCB1 was the most highly up-regulated by development of the CPT11 resistance, inferring a significant contribution of pregnane-X receptor-dependent signaling to the ABCB1 up-regulation. The combined treatment with inhibitors of AKR1C3 and ABCB1 potently sensitized the resistant cells to CPT11 and its active metabolite SN38. Taken together, our results suggest that combination of AKR1C3 and ABCB1 inhibitors is effective as adjuvant therapy to enhance CPT11 sensitivity of intractable colon cancer cells.

PLX038: a PEGylated prodrug of SN-38 independent of UGT1A1 activity.

The purpose of this study was to determine the importance of UGT1A1 activity on the metabolism and pharmacokinetics of a releasable PEG ~ SN-38 conjugate, PLX038A. Irinotecan (CPT-11) is converted to the topoisomerase 1 inhibitor SN-38 by first-pass hepatic metabolism and is converted to its glucuronide SN-38G by UGT1A1. With diminished UGT1A1 activity, the high liver exposure to SN-38 can cause increased toxicity of CPT-11. In contrast, releasable PEG ~ SN-38 conjugates-such as PLX038-release SN-38 in the vascular compartment, and only low levels of SN-38 are expected to enter the liver by transport through the OATP1B1 transporter. We measured CPT-11 and PLX038A metabolites in plasma and bile, and determined pharmacokinetics of PLX038A in UGT1A-deficient and replete rats. Compared to CPT-11, treatment of rats with PLX038A results in very low levels of biliary SN-38 and SN-38G, a low flux through UGT1A, and a low SN-38G/SN-38 ratio in plasma. Further, the pharmacokinetics of plasma PLX038A and SN-38 in rats deficient in UGT1A is unchanged compared to normal rats. The disposition of PEGylated SN-38 is independent of UGT1A activity in rats, and PLX038 may find utility in full-dose treatment of patients who are UGT1A1*28 homozygotes or have metastatic disease with coincidental or incidental liver dysfunction.

A phase II trial of irinotecan in patients with advanced or recurrent endometrial cancer and correlation with biomarker analysis

ObjectiveChemotherapy for advanced or recurrent endometrial cancer requires further development. Irinotecan hydrochloride (CPT-11) suppresses tumor growth in several endometrial cancer strains. The present study evaluated the anti-tumor activity and toxicity of CPT-11 in patients with advanced or recurrent endometrial cancer. MethodsEnrolled patients had advanced endometrial cancer with measurable lesions and received 2 pretreatment regimens. A 90-minute intravenous infusion of CPT-11 (100 mg/m2) was given on days 1, 8, and 15 of a 4-week cycle, aiming for an effect with ≤2 cycles. Treatment was continued until the primary disease worsened or severe toxicity occurred. The primary endpoint was response rate, and the secondary endpoints were progression-free survival, overall survival, and adverse events. Antitumor effect and adverse events were evaluated according to RECIST version 1.1 and NCI-CTC AE version 3.0, respectively. ResultsTwenty-two patients were registered (11 endometrioid carcinomas and 11 serous carcinomas). The median duration of the treatment-free interval (TFI) was 7.5 months, and the median number of administered cycles per patient was 4. Response rate was 36.4% (complete response: 1 patient, partial response: 7 patients). Clinical benefit rate, including stable disease, was 77.3%. Median progression-free and overall survival was 4.4 and 18.4 months, respectively. Observed adverse events included grade 4 hematotoxicity (neutropenia and thrombocytopenia), and grade 2 or 3 non-hematotoxicity (diarrhea). All adverse events were manageable. Biomarker predictors of therapeutic effectiveness were not observed. ConclusionAs a single agent, CPT-11 has anti-tumor activity for advanced or recurrent endometrial cancer and has manageable adverse events.

Curcumin protects heart tissue against irinotecan-induced damage in terms of cytokine level alterations, oxidative stress, and histological damage in rats.

Irinotecan (CPT-11), commonly used in the treatment of many cancer types, may have several side effects that limit the use of CPT-11 in specific tissues such as the heart. In the current study, positive effects of curcumin (CRC) was determined in terms of antioxidant and anti-inflammatory properties against heart damage, caused by CPT-11, in rats. Rats were divided randomly into four equal groups (Control, CPT-11, CRC, and CPT-11 + CRC). CPT-11 10mg/kg/day was administered intraperitoneally and CRC 100mg/kg-1 was given orally. Blood and tissue samples were collected from all groups at day 30 for the detection of oxidative stress, histological changes, and cytokine levels. Results showed that CPT-11 caused dramatic changes in heart tissue for oxidative stress parameters (TBARS, SOD, CAT, GSH, and GPx levels), histological tissue damage, and cytokine levels (TNF and IL-4). CRC therapy reversed the elevated oxidative stress, histological tissue damages, and immunological changes and protected cardiac tissue against CPT-11 toxicity when given together with CPT-11.In conclusion, CPT-11 caused adverse effects on cytokine levels, histological alterations, and oxidative stress in rats. However, CRC treatment eliminated these toxic effects with its antioxidant and anti-inflammatory properties. Thus, these results suggest that CRC may play a protective role against CPT-11 toxicity in heart tissue of rats.

Abstract A052: Predictive wild-type RAS/BRAF and pharmacogenetic biomarkers drive selection of metastatic colorectal cancer patients fit for intensive first-line FIr-C/FOx-C schedule adding cetuximab to triplet chemotherapy

Abstract Background: Intensive-first line bevacizumab (BEV) addition to triplet chemotherapy significantly increased clinical outcome in metastatic colorectal cancer (MCRC) patients. Cetuximab (CET) addition to doublet chemotherapy significantly increased clinical outcome in RAS wild-type MCRC. This phase II study investigated safety and activity of FIr-C/FOx-C schedule of weekly CET added to triplet chemotherapy in first line RAS wild-type MCRC. Methods: The phase II study was planned according to Simon two-step design: delta 15% (p0 70%, p1 85%), power 80%, α 5%, β 20%; projected objective responses (ORR), I step 14/19 patients. FIr-C/FOx-C schedule: 5-fluorouracil (5-FU) 12h-timed-flat-infusion 900 mg/m2, days 1-2, 8-9, 15-16, 22-23; associated to weekly alternating irinotecan (CPT-11) 160 mg/m2, days 1,15 or oxaliplatin (OXP) 80 mg/m2, days 8, 22; CET 400 mg/m2 loading dose, then 250 mg/m2 days 1,8,15,22; every 4 weeks. Toxicity and limiting toxicity syndromes (LTS) were evaluated and compared using chi-square test; activity and efficacy using log-rank test. 5-FU degradation rate (5-FUDR) and single nucleotide polymorphisms (SNP) of 4 genes (DYPD, CYP3A4, ABCB1, UGT1A1) involved in metabolism of 5-FU and CPT-11 were evaluated in individual patients showing LTS and at the recommended doses as predictive biomarkers of toxicity. Results: Twenty-nine consecutive patients <75 years with primary/intermediate Cumulative Illness Rating Scale (CIRS) were enrolled. From April 2014, only KRAS/NRAS wild-type patients were eligible: median age 59; young-elderly (yE) 7 (24%); liver-limited metastases (L-L) 7, 24%. Due to limiting gastrointestinal toxicity, recommended doses of CPT-11 and 5-FU were 120 and 750 mg/m2, respectively. Activity data met the primary endpoint: 14 OR out of 18 evaluable patients as treated (76%). Preliminary phase II data confirm 17 OR out of 23 evaluable patients intent-to-treat (74%). Liver metastasectomies were performed in 14%, 57% L-L. At median follow-up 18 months, median progression-free survival (PFS) was 12 months, median overall survival (OS) 23 months. OS was significantly worse in yE compared to non-elderly patients (p = 0.045). At the recommended doses, received DI was >80% for each drug and cumulative G3-4 toxicities in 13 patients were: diarrhea 23%, asthenia 15%, vomiting 8%, hypertransaminasemy 8%. LTS, consisting of the LT, associated or not to G2 or other LT, were observed overall in 19 patients (65.5%), 83% in yE. LTS were prevalently multiple than single site (59% versus 7%, chi square 7.703, p = 0.006). Reduced FUDR, CYP3A4 and UGT1A1 SNPs, also frequently associated, were particularly observed in patients showing gastrointestinal LTS, while not in patients without LTS. Conclusion: Intensive first-line FIr-C/FOx-C regimen at the modified recommended doses is tolerable and highly effective in RAS wild-type MCRC patients, with significantly increased LTS multiple sites, justifying careful selection of fit patients using a panel of 5 predictive biomarkers of 5-FU and CPT11 toxicity. Citation Format: Gemma Bruera, Silvia Massacese, Antonella Dal Mas, Corrado Ficorella, Eugenio Ciacco, Giuseppe Calvisi, Maurizio Simmaco, Enrico Ricevuto. Predictive wild-type RAS/BRAF and pharmacogenetic biomarkers drive selection of metastatic colorectal cancer patients fit for intensive first-line FIr-C/FOx-C schedule adding cetuximab to triplet chemotherapy [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A052.

Clarithromycin co-administration does not increase irinotecan (CPT-11) toxicity in colorectal cancer patients.

Irinotecan (CPT-11) is used to treat advanced colorectal cancer. The drug is activated by carboxylesterases and rendered inactive by CYP3A4. Recently, the efficacy of combining CPT-11 and anti-epidermal growth factor receptor (EGFR) agents was confirmed in patients with KRAS wild-type metastatic colorectal cancer. Clarithromycin (CAM) is a strong CYP3A inhibitor often used to prevent rash associated with anti-EGFR therapy. The objective of this study was to evaluate the risk of increased neutropenia and diarrhea in combining CPT-11 and CAM. Retrospective analyses were conducted at Osaka National Hospital (Osaka, Japan) on the records of colorectal cancer patients treated with a CPT-11-containing regimen between November 2006 and January 2014. The incidence of neutropenia and diarrhea was compared between patients who received CPT-11 and CAM and patients who received CPT-11 without CAM. One-hundred and twenty-eight patients were included in this study, of whom 21 were concomitantly treated with CAM and 107 were not. There was no difference in the incidence of grade 3-4 neutropenia between the CAM co-administration group (10%) and the non-CAM group (16%) [Odds ratio: 0.56 (95% confidence interval: 0.12-2.62), p=0.45]. No difference in the incidence of grade 3-4 diarrhea was found between the CAM co-administration group (0%) and the non-CAM group (4%) (p=0.37). This study did not identify an increase in CPT-11 toxicity by co-administration with CAM.

HuangQin Decoction Attenuates CPT-11-Induced Gastrointestinal Toxicity by Regulating Bile Acids Metabolism Homeostasis.

Irinotecan (CPT-11) is a potent chemotherapeutic agent, however, its clinical usage is often limited by the induction of severe gastrointestinal (GI) toxicity, especially late-onset diarrhea. HuangQin Decoction (HQD), commonly used for the treatment of GI ailments, has been proved could significantly ameliorate the intestinal toxicity of CPT-11. To reveal the mechanisms of CPT-11-induced toxicity and the modulation effects of HQD, a previous untargeted metabolomics study was performed and the results indicated that HQD may protect the GI tract by altering the metabolism of bile acids (BAs). Nevertheless, the untargeted assays are often less sensitive and/or efficient. In order to further confirm our previous findings, here in this paper, serum and tissues metabolic profiles of 17 BAs were analyzed using liquid chromatography-tandem mass spectrometry based targeted metabolomics. The results indicated that serum and tissues levels of most BAs were significantly decreased after CPT-11 administration, except some hydrophobic BAs. Co-treatment with HQD could markedly attenuate CPT-11-induced GI toxicity and reverse the alterations of hydrophobic BAs. Despite the fact that the BAs pool size remained unchanged, the balance of BAs had shifted leading to decreased toxicity after HQD treatment. The present study demonstrated for the first time that the precise interaction between HQD, CPT-11-induced intestinal toxicity and BAs’ homeostasis.

The Role of Intestinal Microbiota in Development of Irinotecan Toxicity and in Toxicity Reduction through Dietary Fibres in Rats

CPT-11 is a drug used as chemotherapy for colorectal cancer. CPT-11 causes toxic side-effects in patients. CPT-11 toxicity has been attributed to the activity of intestinal microbiota, however, intestinal microbiota may also have protective effects in CP!-11 chemotherapy. This study aimed to elucidate mechanisms through which microbiota and dietary fibres could modify host health. Rats bearing a Ward colon carcinoma were treated with a two-cycle CPT-11/5-fluorouracil therapy recapitulating clinical therapy of colorectal cancer. Animals were fed with a semi-purified diet or a semi-purified diet was supplemented with non-digestible carbohydrates (isomalto-oligosaccharides, resistant starch, fructo-oligosaccharides, or inulin) in 3 independent experiments. Changes in intestinal microbiota, bacteria translocating to mesenteric lymphnodes, cecal GUD activity, and cecal SCFA production, and the intestinal concentration of CPT-11 and its metabolites were analysed. Non-digestible carbohydrates significantly influenced feed intake, body weight and other indicators of animal health. The identification of translocating bacteria and their quantification in cecal microbiota indicated that overgrowth of the intestine by opportunistic pathogens was not a major contributor to CPT-11 toxicity. Remarkably, fecal GUD activity positively correlated to body weight and feed intake but negatively correlated to cecal SN-38 concentrations and IL1-β. The reduction in CPT-11 toxicity by non-digestible carbohydrates did not correlate to stimulation of specific bacterial taxa. However, cecal butyrate concentrations and feed intake were highly correlated. The protective role of intestinal butyrate production was substantiated by a positive correlation of the host expression of MCT1 (monocarboxylate transporter 1) with body weight as well as a positive correlation of the abundance of bacterial butyryl-CoA gene with cecal butyrate concentrations. These correlations support the interpretation that the influence of dietary fibre on CPT-11 toxicity is partially mediated by an increased cecal production of butyrate.

Intestinal glucuronidation protects against chemotherapy-induced toxicity by irinotecan (CPT-11)

Camptothecin (CPT)-11 (irinotecan) has been used widely for cancer treatment, particularly metastatic colorectal cancer. However, up to 40% of treated patients suffer from severe late diarrhea, which prevents CPT-11 dose intensification and efficacy. CPT-11 is a prodrug that is hydrolyzed by hepatic and intestinal carboxylesterase to form SN-38, which in turn is detoxified primarily through UDP-glucuronosyltransferase 1A1 (UGT1A1)-catalyzed glucuronidation. To better understand the mechanism associated with toxicity, we generated tissue-specific Ugt1 locus conditional knockout mouse models and examined the role of glucuronidation in protecting against irinotecan-induced toxicity. We targeted the deletion of the Ugt1 locus and the Ugt1a1 gene specifically in the liver (Ugt1(ΔHep)) and the intestine (Ugt1(ΔGI)). Control (Ugt1(F/F)), Ugt1(ΔHep), and Ugt1(ΔGI) adult male mice were treated with different concentrations of CPT-11 daily for four consecutive days. Toxicities were evaluated with regard to tissue glucuronidation potential. CPT-11-treated Ugt1(ΔHep) mice showed a similar lethality rate to the CPT-11-treated Ugt1(F/F) mice. However, Ugt1(ΔGI) mice were highly susceptible to CPT-11-induced diarrhea, developing severe and lethal mucositis at much lower CPT-11 doses, a result of the proliferative cell loss and inflammation in the intestinal tract. Comparative expression levels of UGT1A1 in intestinal tumors and normal surrounding tissue are dramatically different, providing for the opportunity to improve therapy by differential gene regulation. Intestinal expression of the UGT1A proteins is critical toward the detoxification of SN-38, whereas induction of the UGT1A1 gene may serve to limit toxicity and improve the efficacy associated with CPT-11 treatment.

Molecular insights into microbial β-glucuronidase inhibition to abrogate CPT-11 toxicity.

Bacterial β-glucuronidases expressed by the symbiotic intestinal microbiota appear to play important roles in drug-induced epithelial cell toxicity in the gastrointestinal (GI) tract. For the anticancer drug CPT-11 (irinotecan) and the nonsteroidal anti-inflammatory drug diclofenac, it has been shown that removal of the glucuronide moieties from drug metabolites by bacterial β-glucuronidases in the GI lumen can significantly damage the intestinal epithelium. Furthermore, selective disruption of bacterial β-glucuronidases by small molecule inhibitors alleviates these side effects, which, for CPT-11 {7-ethyl-10-[4-(1-piperidino)-1-piperidino]}, can be dose limiting. Here we characterize novel microbial β-glucuronidase inhibitors that inhibit Escherichia coli β-glucuronidase in vitro with Ki values between 180 nM and 2 μM, and disrupt the enzyme in E. coli cells, with EC50 values as low as 300 nM. All compounds are selective for E. coli β-glucuronidase without inhibiting purified mammalian β-glucuronidase, and they do not impact the survival of either bacterial or mammalian cells. The 2.8 Å resolution crystal structure of one inhibitor bound to E. coli β-glucuronidase demonstrates that it contacts and orders only a portion of the "bacterial loop" present in microbial, but not mammalian, β-glucuronidases. The most potent compound examined in this group was found to protect mice against CPT-11-induced diarrhea. Taken together, these data advance our understanding of the chemical and structural basis of selective microbial β-glucuronidase inhibition, which may improve human drug efficacy and toxicity.

Molecular Insights into Microbial -Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

Molecular Insights into Microbial -Glucuronidase Inhibition to Abrogate CPT-11 Toxicity

Irinotecan (CPT-11) Chemotherapy Alters Intestinal Microbiota in Tumour Bearing Rats

Intestinal microbiota mediate toxicity of irinotecan (CPT-11) cancer therapies and cause systemic infection after CPT-11-induced loss of barrier function. The intestinal microbiota and their functions are thus potential targets for treatment to mitigate CPT-11 toxicity. However, microbiota changes during CPT-11 therapy remain poorly described. This study analysed changes in intestinal microbiota induced by CPT-11 chemotherapy. Qualitative and quantitative taxonomic analyses, and functional analyses were combined to characterize intestinal microbiota during CPT-11-based chemotherapy, and in presence or absence of oral glutamine, a treatment known to reduce CPT-11 toxicity. In the first set of experiments tumour-bearing rats received a dose-intensive CPT-11 regimen (125 mg kg−1×3 days), with or without oral glutamine bolus (0.75 g kg−1). In a subsequent more clinically-oriented chemotherapy regimen, rats received two cycles of CPT-11 (50 mg kg−1) followed by 5-flurouracil (50 mg kg−1). The analysis of fecal samples over time demonstrated that tumours changed the composition of intestinal microbiota, increasing the abundance of clostrridial clusters I, XI, and Enterobacteriaceae. CPT-11 chemotherapy increased cecal Clostridium cluster XI and Enterobacteriaceae, particularly after the dose-intensive therapy. Glutamine treatment prevented the reduced abundance of major bacterial groups after CPT-11 administration; i.e. total bacteria, Clostridium cluster VI, and the Bacteroides-group. Virulence factor/toxin genes of pathogenic Escherichia coli and Clostridium difficile were not detected in the cecal microbiota. In conclusion, both colon cancer implantation and CPT-11-based chemotherapies disrupted the intestinal microbiota. Oral glutamine partially mitigated CPT-11 toxicity and induced temporary changes of the intestinal microbiota.

Prevalence of topoisomerase I genetic mutations and UGT1A1 polymorphisms associated with irinotecan in individuals of Asian descent

Topoisomerase I (TOP-I) mutations have been shown to be correlated to irinotecan resistance in vitro. However, the prevalence of TOP-I germline mutations has yet to be systematically elucidated. On the other hand, polymorphisms of UGT1A1 have been shown to be associated with CPT-11 toxicity in clinical situations. The primary aim of this study was to investigate the prevalence of mutations in the TOP-I exons associated with CPT-11 resistance, including untreated cancer tissue. A secondary aim was to confirm the less frequent UGT1A1*28 and more frequent UGT1A1*6 in individuals of Asian descent compared to Caucasians and individuals of African descent. The prevalence of 5 reported TOP-I mutations in exons was investigated in volunteers (n=236) using DNA sequencing of the PCR products. The prevalence of TOP-I mutations in untreated lung cancer tissues (n=16) was also investigated. Additionally, 3 UGT1A1 polymorphisms, UGT1A1*6, *27 and *28, were investigated in volunteers (n=126). There were no mutations of TOP-I in any of the 236subjects or in the untreated lung tissues. Among 128subjects, the distribution of homozygous polymorphisms of UGT1A1 was: UGT1A1*28 in 3 (2.4%) and UGT1A1*6 in 4 (3.2%) subjects, and co-occurrence of heterozygous polymorphisms for both UGT1A1*6 and UGT1A1*28 in 4 (3.2%) subjects, and for UGT1A1*27 and UGT1A1*28 in 1 subject (0.8%). The Hardy-Weinberg deviation test showed there was no significant deviation from the equilibrium, and the association analysis indicated no significant linkage between UGT1A1*6 and UGT1A1*28. In conclusion, TOP-I genetic mutations correlated to CPT-11 resistance were not detected in any of the subjects and untreated lung cancer tissues. Less frequent UGT1A1*28 and more frequent UGT1A1*6 were confirmed in East Asian individuals compared to Caucasians and individuals of African descent. Linkage disequilibrium was not detected between UGT1A1*6 and UGT1A1*28.

Are we turning to more than a first line treatment of metastatic colorectal cancer with high dose irinotecan ?: A monocentric institution safety analysis of 46 patients

Irinotecan (CPT11) at 180 mg/m(2) with LV5FU2 for metastatic colorectal cancer (MCRC) has response rates (RRs) of 56 and 4% as first- and second-line treatments, respectively [1-2], and higher doses of CPT11 result in higher RRs. The present cohort analysis aimed to evaluate the effect of increasing doses of this combination treatment in clinical practice. Chemo-naive and pretreated patients with MCRC received CPT11 and LV5FU2 (5FU 48-h CI 2400 mg/m(2), D1 bolus leucovorin 200 mg/m(2)), followed by 5FU 400 mg/m(2) (cycles d1-d15). CPT11 dose was increased by 20 mg/m(2) at each cycle, from 180 mg/m(2) up to 260 mg/m(2), unless grade 3 toxicities other than alopecia arose. Between March 2002 and September 2005, 46 patients were recruited (median age: 62.3 years). A total of 512 cycles of chemotherapy were administered (median: 9 cycles/patient; range: 3-41). Median follow-up was 16.2 months. Altogether, 27 patients had received prior chemotherapy: 24 with an oxaliplatin-based regimen; seven with CPT11; and five with LV5FU2 or oral 5FU. Doses of 260 mg/m(2) were used in 17 patients, 240 mg/m(2) in seven, 220 mg/m(2) in six and 200 mg/m(2) in five, while 11 remained at 180 mg/m(2); 121 cycles used 260 mg/m(2) (24%), with 76 cycles at 240 mg/m(2) (14%), 78 cycles at 220 mg/m(2) and 58 cycles at 200mg/m(2). The objective response (OR) was 40%, with stable disease (SD) in 45% and disease progression (DP) in 11%. In the first-line therapy group, partial/complete responses were 55%, with SD in 30% and DP in 15%. In pretreated patients, OR was 30.5%, SD was 58.5% and DP was 11%. Nine patients (20%) had a therapeutic break (median: 5.1 months; range: 3-10). Overall median survival was 17 months, with 16.5 months in pretreated patients and 19.6 months in the first-line group. Toxicity grades 3-4 and overall incidence per cycle were: neutropenia, 3-22%; diarrhea, 4-22%; vomiting, 2-20%; alopecia, 20-26%; anemia, 0.2-2%; thrombocytopenia, 0-0%; and mucositis, 0.4-2.2%. The toxicity of high-dose CPT11+LV5FU2 chemotherapy was well tolerated when the dose was progressively increased according to individual tolerability, with 37% of patients receiving CPT11 at 260 mg/m(2). Progression-free survival (PFS) increased with higher doses of CPT11. In the chemo-naive and pretreated subgroups, the median PFS was 10.9 and 8.8 months, respectively (P=0.698, NS). Optimization of CPT11 doses in pretreated patients appears to pave the way for new treatment options.

Effect of poly (ethylene oxide)-poly (propylene oxide)-poly (ethylene oxide) micelles on pharmacokinetics and intestinal toxicity of irinotecan hydrochloride: potential involvement of breast cancer resistance protein (ABCG2)

Intestinal toxicity and low levels of systemic drug exposure are among the major problems associated with tumour therapy. We have developed poly (ethylene oxide)-poly (propylene oxide)-poly (ethylene oxide) (PEO-PPO-PEO) micelles loaded with irinotecan hydrochloride (CPT-11) hoping to decrease CPT-11-induced intestinal toxicity while increasing its systemic exposure. In addition, we have investigated the potential involvement of breast cancer resistance protein (BCRP) in biliary excretion, pharmacokinetics, and intestinal toxicity of CPT-11. PEO-PPO-PEO micelles were prepared using PEO(20)-PPO(70)-PEO(20) and lecithin. The effect of PEO-PPO-PEO micelles on BCRP-mediated cellular accumulation and transport efflux of CPT-11 was evaluated in MDCKII/BCRP cells. The biliary excretion, intestinal damage, and pharmacokinetic study of CPT-11-loaded PEO-PPO-PEO micelles were investigated in rats. The obtained micelles could effectively inhibit BCRP-mediated CPT-11 efflux in MDCKII/BCRP cells, and significantly decrease the drug biliary excretion in rats. Moreover, intestinal toxicity, assessed by microscopic examination of pathological damage, was ameliorated in rats injected with PEO-PPO-PEO micelles compared with rats injected with CPT-11 alone. Treatment with PEO-PPO-PEO micelles resulted in prolonged circulation time in blood and increased bioavailability of CPT-11 and SN-38 (7-ethyl-10-hydroxycamptothecin). PEO-PPO-PEO micelles were identified as promising carriers able to reduce intestinal toxicity and increase antitumour therapeutic effect of CPT-11. The study indicated a potential involvement of BCRP in CPT-11 pharmacokinetics and CPT-11-induced intestinal toxicity.

Abstract 3660: Bicarbonate attenuates irinotecan-induced cytotoxicity by altering intracellular pH in HT29 cells

Abstract [Background/Aim] The therapeutic efficacy of irinotecan (CPT-11) is often limited by the induction of severe late-onset diarrhea. CPT-11 and its active metabolite (SN-38) have a labile α-hydroxy-lactone ring that undergoes pH-dependent reversible hydrolysis, and the toxicity is more potent in the lactone (LA) form at acidic pH compared to that in the carboxylate (CA) form in physiological and higher pH. We have previously demonstrated that sodium bicarbonate (Bicarb) reduced the intestinal toxicity of CPT-11/SN-38 in in vivo, in a mouse model, and in a cultured intestinal cancer (HT29) cell line. The hypothesis was that protective action of Bicarb might be due, at least in part, to reduced CPT-11/SN-38-induced intracellular (pHi) acidification. [Method] HT29 cells were exposed to ∼1µM SN-38 LA or CA form for 2h and further incubated in fresh medium at two different extracellular pH (6.2 and 7.4) adjusted by HCl with 44 mM Bicarb for an additional 96h. In addition, the HT29 cells exposed to SN-38 LA were also incubated at 5-set points of extracellular pH at 6.8, 7.0, 7.4, 7.8, 8.2 adjusted by either increasing concentrations of Bicarb (5, 10, 20, 44, 100mM, respectively) or HCl/NaOH in combination with Trice, Tris, and NaH2PO4 to maintain extracellualr pH during culture. The cell proliferation and apoptosis were examined by MTT assay and Flow Cytometry, respectively. The pHi following SN-38 exposure was also analyzed using Snarf-1 with two concentrations (10 and 30mM) of Bicarb. [Result] The cell proliferation was significantly decreased on exposure to SN-38 LA form compared to the CA form, while there was no difference in the lower cell growth on exposure to LA form between pH 6.2 and 7.4. In the SN-38 exposed cells, the cell proliferation and apoptosis were significantly higher and lower, respectively, dependent on the concentration of Bicarb. However, the decreased cell viabilities were not influenced by extracellular pH without Bicarb. The SN-38 LA exposure induced a significant decrease of pHi, and the incubation with 30mM Bicarb significantly prevented from the acidification of pHi compared to that in 10mM Bicarb. [Conclusion] The present study showed that cell cytotoxicity of SN-38 was independent of extracellular pH but dependent on pHi that converts the SN-38 form, while the acidification of pHi was reduced by Bicarb. The novel protective mechanism of Bicarb action against intestinal cell injury by CPT-11/SN-38 would be beneficial for anti-cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3660.

Relations between strain and gender dependencies of irinotecan toxicity and UGT1A1, CES2 and TOP1 expressions in mice

Irinotecan hydrochloride (CPT-11) can display severe toxicities in individual cancer patients. CPT-11 is bio-activated through CES, detoxified through UGT1A1 and inhibits TOP1. CPT-11 toxicity and UGT1A1, CES2 and TOP1 mRNAs and UGT1A1 protein were determined in male and female C57BL/6, B6D2F1 and B6CBAF1, as potential models for tailoring CPT-11 delivery. CPT-11 was administered intravenously (40–90 mg/kg/day for 4 days at 7 h after light onset). The relations between dose and lethal toxicity or body weight loss were steep and similar in C57BL/6 (lethality, p = 0.001; weight loss, p = 0.002) and B6D2F1 ( p = 0.01; p = 0.03, respectively), but weak in B6CBAF1. Females displayed less toxicity than males ( p < 0.001). Mean mRNA expression of UGT1A1 was highest in B6CBAF1 ( p = 0.039) and in females ( p < 0.001). Both CES2 and TOP1 varied according to strain and gender ( p < 0.001). The three gene expression data explained the most severe toxicity of CPT-11 in male B6D2F1, but displayed inconsistent relations with toxicity in the other groups. Mean UGT1A1 protein expression was highest in males as compared to females, and so by ∼8-fold in C57BL/6 as compared to B6D2F1 ( p < 0.0001). Genetic background and gender significantly altered the molecular prediction of irinotecan toxicity by UGT1A1, CES2 and TOP1 mRNA expressions.