Cytotoxicity Of Irinotecan Research Articles

Local Delivery of Irinotecan to Recurrent GBM Patients at Reoperation Offers a Safe Route of Administration.

Glioblastomas are impossible to completely resect and almost always recur at the borders of the resection margin. There is no established chemotherapy regimen available to patients who recur, while systemic treatment is hampered by the blood-brain barrier. Here, we report on the first evaluation in humans of the intraparenchymal injection of irinotecan into the resection cavity after surgical resection of recurrent glioblastoma patients. The cytotoxicity of irinotecan was compared to SN-38 in primary cells from recurrent glioblastoma patients. Irinotecan was injected at multiple (~30) sites of the resection cavity wall at a depth of 3 to 5 mm. SN-38 was more cytotoxic than irinotecan at concentrations below 1 µM due to enzyme kinetics. The intraparenchymal administration of irinotecan was safe, with good wound healing and an absence of swelling, inflammation, or pseudo-abscess formation. The median survival post irinotecan administration was 32.6 weeks. The median overall survival was 30.5 months, with a two-year survival rate of 56%. This study demonstrates that local delivery of irinotecan into the brain parenchyma offers a safe route of administration over systemic delivery in the treatment of recurrent glioblastoma.

Synergistic cytotoxicity of irinotecan combined with polysaccharide-based nanoparticles for colorectal carcinoma

Functional polymeric nanoparticles (NPs) with antitumor potential were combined with the topoisomerase I inhibitor, irinotecan (IRT), to enhance cytotoxicity against colorectal cancers. The negatively charged γ-polyglutamic acid (γ-PGA) or fucoidan (FCD) was complexed with the positively charged chitosan (CS) to encapsulate IRT. The size of the γ-PGA/CS/IRT NPs and FCD/CS/IRT NPs were 146.0 ± 8.0 nm and 230.8 ± 2.5 nm, respectively, with polydispersity index ≤0.3. The cellular uptake ability of FCD/CS-FITC NPs was better than that of γ-PGA/CS-FITC NPs, especially in p-selectin positive HCT116 colorectal cancer cells (4.8 ± 0.8 μg/mL vs 11.4 ± 2.2 μg/mL). The IC50 of FCD/CS/IRT NPs was 2.4 times lower than that of γ-PGA/CS/IRT NPs in HCT116 cells (4.8 ± 0.8 μg/mL vs 11.4 ± 2.2 μg/mL), indicating its superior antitumor potential. The combination of irinotecan and fucoidan-based NPs exhibited a synergistic effect (CI <1), resulting in better anticancer activity of FCD/CS/IRT NPs than irinotecan alone. The apoptosis-related proteins, caspase 3, caspase 9, and poly(ADP-ribose) polymerase (PARP), were prominently increased in FCD/CS/IRT NPs-treated HCT116 cells by 2.3 folds, 3.5 folds, and 6.3 folds, respectively. All results support that fucoidan-based irinotecan-loaded nanoparticles possess the ability to effectively enhance cellular uptake and induce synergistic apoptosis of colorectal cancer cells.

ATM kinase inhibitor AZD0156 in combination with irinotecan and 5-fluorouracil in preclinical models of colorectal cancer

BackgroundAZD0156 is an oral inhibitor of ATM, a serine threonine kinase that plays a key role in DNA damage response (DDR) associated with double-strand breaks. Topoisomerase-I inhibitor irinotecan is used clinically to treat colorectal cancer (CRC), often in combination with 5-fluorouracil (5FU). AZD0156 in combination with irinotecan and 5FU was evaluated in preclinical models of CRC to determine whether low doses of AZD0156 enhance the cytotoxicity of irinotecan in chemotherapy regimens used in the clinic.MethodsAnti-proliferative effects of single-agent AZD0156, the active metabolite of irinotecan (SN38), and combination therapy were evaluated in 12 CRC cell lines. Additional assessment with clonogenic assay, cell cycle analysis, and immunoblotting were performed in 4 selected cell lines. Four colorectal cancer patient derived xenograft (PDX) models were treated with AZD0156, irinotecan, or 5FU alone and in combination for assessment of tumor growth inhibition (TGI). Immunofluorescence was performed on tumor tissues. The DDR mutation profile was compared across in vitro and in vivo models.ResultsEnhanced effects on cellular proliferation and regrowth were observed with the combination of AZD0156 and SN38 in select models. In cell cycle analysis of these models, increased G2/M arrest was observed with combination treatment over either single agent. Immunoblotting results suggest an increase in DDR associated with irinotecan therapy, with a reduced effect noted when combined with AZD0156, which is more pronounced in some models. Increased TGI was observed with the combination of AZD0156 and irinotecan as compared to single-agent therapy in some PDX models. The DDR mutation profile was variable across models.ConclusionsAZD0156 and irinotecan provide a rational and active combination in preclinical colorectal cancer models. Variability across in vivo and in vitro results may be related to the variable DDR mutation profiles of the models evaluated. Further understanding of the implications of individual DDR mutation profiles may help better identify patients more likely to benefit from treatment with the combination of AZD0156 and irinotecan in the clinical setting.

Abstract 6228: APR-246, which restores p53 function, is highly active against alternative lengthening of telomere (ALT) cell lines and PDXs

Abstract Introduction: Most cancers proliferate by activating telomerase (TA+) while 10% of cancers utilize alternate lengthening of telomeres (ALT). ALT has been associated with resistance to DNA damaging agents, p53 loss-of-function (p53LOF), ATRX mutations, and very poor survival. ATM kinase, which activates functional p53, is constitutively activated in ALT cancers (Science Translational Medicine 18:eabd5750, 2021). We hypothesized that the constitutive activation of ATM kinase in ALT cancers would confer high sensitivity to pharmacological reactivation of p53 function. Methods: We used patient-derived ALT (telomeric DNA C-circle positive) and TA+ neuroblastoma, sarcoma, colorectal, and breast cancer cell lines and xenografts treated with the clinical-stage p53 reactivator eprenetapopt (APR-246) and irinotecan. In vitro cytotoxicity was assayed by DIMSCAN digital imaging microscopy, ATM activation by immunofluorescence microscopy, and protein expression by western blotting. Results: ALT p53LOF cell lines were significantly (p&amp;lt;0.05) less sensitive to DNA-damaging agents relative to TA+ p53LOF comparators. Constitutive phosphorylation (activation) of ATM kinase and the DNA damage marker 53BP1 were observed at telomeres in ALT but not TA+ cell lines. APR-246 induced p53 targets p21 and NOXA and was significantly more cytotoxic (p&amp;lt;0.001) for ALT relative to TA+ cell lines, regardless of TP53 status. Overexpression of p53 in a TP53-null ALT cell line increased sensitivity (p&amp;lt;0.0001) to APR-246. Knockdown of p53 or ATM kinase in ALT TP53 mutated and wild-type cell lines antagonized (p&amp;lt;0.0001) APR-246 activity. Induction of telomere dysfunction in a TA+ p53LOF neuroblastoma cell line using dominant-negative TRF2 (a shelterin protein that blocks ATM activation at telomeres) activated ATM and sensitized cells to APR-246 (p&amp;lt;0.01). APR-246 enhanced the cytotoxicity of irinotecan (as SN38) in ALT cell lines in vitro to a higher degree than in TA+ p53LOF cell lines (p&amp;lt;0.05). Single-agent APR-246 significantly (p&amp;lt;0.0001) increased event-free survival (EFS) of mice with ALT xenografts relative to controls. APR-246 enhanced (p&amp;lt;0.0001) the activity of irinotecan in 3 neuroblastoma, 2 rhabdomyosarcoma, 1 colorectal, and 1 triple-negative breast ALT xenograft models with most mice (47/56) achieving complete responses and an EFS of &amp;gt;100 days (45/56) compared to no complete responses (median EFS ~ 34 days) in mice treated with only irinotecan (p&amp;lt;0.0001). APR-246 + irinotecan had no significant effect relative to irinotecan alone (p=0.08) on EFS of mice with TA+ p53LOF xenografts (median EFS ~ 40 days). Conclusion: ALT cancers of a variety of histologies are highly resistant to DNA damaging agents, have p53LOF, and constitutive activation of ATM kinase, which confers high sensitivity to p53 reactivation by APR-246. APR-246 warrants clinical testing in patients with ALT cancers. Citation Format: Shawn J. Macha, Balakrishna Koneru, Trevor Burrow, Charles Zhu, Dzmitry Savitski, Jonas Nance, Kristyn McCoy, Cody Eslinger, C Patrick Reynolds. APR-246, which restores p53 function, is highly active against alternative lengthening of telomere (ALT) cell lines and PDXs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6228.

DFT studies of camptothecins cytotoxicity III: camptothecin, irinotecan and SN-38

Abstract Geometries of camptothecin, irinotecan, SN-38, and of their hypothetical Cu(II) complexes were optimized at the B3LYP/6-311G* level of theory. Their electron structure, evaluated in terms of Mulliken population analysis and Quantum Theory of Atoms-in-Molecule, was subsequently related to in vitro cytotoxicity. Electron density transfer from the relevant active sites to Cu decreases in the sequence irinotecan &gt; SN-38 &gt; camptothecin. The absolute values of their metal-ligand interaction energies exhibit the same trend. Discrepancy with the least relative in vitro cytotoxicity of irinotecan can be explained by differences in its pharmacokinetics.

Extremely Low Frequency Magnetic Field Alters Cytotoxicity of Irinotecan in Glioblastoma: A Preliminary Observation

Objective: Exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) emitted from electric appliances used in daily life may cause various changes at the cellular level. Glioblastoma multiform (GBM) is an aggressive brain cancer resulting in high mortality. Despite advances in treatment methods, GBM has remained an incurable disease. It has become important that necessary precautions are taken to overcome factors that may reduce the efficiency of antineoplastic agents used for GBM patients. Irinotecan, an antineoplastic agent, is one of the second line drugs for GBM patients. The aim of this preliminary research was to examine the impact of ELF-EMF on irinotecan cytotoxicity in GBM cells. Method: GBM cells (U87) in the control group were only treated with irinotecan (0, 1, 2, 10, 25, 50, 75, and 150 µM). ELFEMF (50 Hz, 1 mT) and irinotecan were applied simultaneously for 1 h to similar cells in the study group. Cytotoxicity was determined via XTT assay. Results: While the IC50 (half maximal inhibitory concentration) value was 14.31 µM in the control group, and 20.51 µM in the study group. Conclusion: The results of the study indicated that ELF-EMF reduced the efficiency of irinotecan in the U87 cells. Therefore, patients undergoing chemotherapy with irinotecan should be given more attention so as to avoid ELF-EMFs. The effects of ELF-EMF on irinotecan cytotoxicity in U87 cells were determined through an exactly unknown mechanism. Further studies on ELF-EMFs and irinotecan cytotoxicity are required to illuminate the topic.

A green tea extract and epigallocatechin-3-gallate attenuate the deleterious effects of irinotecan in an oral epithelial cell model

ObjectiveTo investigate the ability of a green tea extract and epigallocatechin-3-gallate (EGCG) to protect oral epithelial cells against the deleterious effects of the chemotherapeutic agent irinotecan, with respect to cytotoxicity; reactive oxygen species (ROS) generation; cytokine and matrix metalloproteinase (MMP) production; and cell proliferation and migration. MethodsThe B11 oral keratinocyte and GMSM-K oral epithelial cell lines were used in this study. Cell viability was determined using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay. A fluorometric assay was used to quantify ROS production. Cell proliferation was assessed using a fluorescent cell tracker dye, while a migration assay kit was used to monitor cell migration. Cytokine and MMP secretion was quantified by an enzyme-linked immunosorbent assay. ResultsThe green tea extract and EGCG reduced the cytotoxicity of irinotecan toward oral keratinocyte and epithelial cell lines. Irinotecan-induced intracellular ROS generation by oral keratinocytes was reduced by the green tea extract and EGCG. Irinotecan negatively affected the proliferation and migration of oral keratinocytes in a dose-dependent manner. However, these effects were not neutralized by the green tea extract, while EGCG showed a trend to attenuate the irinotecan-induced decrease in cell migration. The green tea extract and EGCG also had a dose-dependent inhibitory effect on irinotecan-induced secretion of interleukin-6 and interleukin-8 by oral epithelial cells. Lastly, the irinotecan-induced decrease in the secretion of MMP-2 and MMP-9 by oral epithelial cells was partially restored by the green tea extract and EGCG. ConclusionsThe green tea extract and EGCG, through anti-cytotoxic, anti-oxidative, and anti-inflammatory properties, may protect the oral mucosa against the deleterious effects of the chemotherapeutic agent irinotecan and may be of interest for treating oral mucositis.

A mathematical model of the circadian clock and drug pharmacology to optimize irinotecan administration timing in colorectal cancer

Scheduling anticancer drug administration over 24 h may critically impact treatment success in a patient-specific manner. Here, we address personalization of treatment timing using a novel mathematical model of irinotecan cellular pharmacokinetics and –dynamics linked to a representation of the core clock and predict treatment toxicity in a colorectal cancer (CRC) cellular model. The mathematical model is fitted to three different scenarios: mouse liver, where the drug metabolism mainly occurs, and two human colorectal cancer cell lines representing an in vitro experimental system for human colorectal cancer progression. Our model successfully recapitulates quantitative circadian datasets of mRNA and protein expression together with timing-dependent irinotecan cytotoxicity data. The model also discriminates time-dependent toxicity between the different cells, suggesting that treatment can be optimized according to their cellular clock. Our results show that the time-dependent degradation of the protein mediating irinotecan activation, as well as an oscillation in the death rate may play an important role in the circadian variations of drug toxicity. In the future, this model can be used to support personalized treatment scheduling by predicting optimal drug timing based on the patient’s gene expression profile.

UCN-01 enhances cytotoxicity of irinotecan in colorectal cancer stem-like cells by impairing DNA damage response.

Colorectal cancer (CRC) is one of the most common and lethal cancers worldwide. Despite recent progress, the prognosis of advanced stage CRC remains poor, mainly because of cancer recurrence and metastasis. The high morbidity and mortality of CRC has been recently ascribed to a small population of tumor cells that hold the potential of tumor initiation, i.e. cancer stem cells (CSCs), which play a pivotal role in cancer recurrence and metastasis and are not eradicated by current therapy. We screened CRC-SCs in vitro with a library of protein kinase inhibitors and showed that CRC-SCs are resistant to specific inhibition of the major signaling pathways involved in cell survival and proliferation. Nonetheless, broad-spectrum inhibition by the staurosporin derivative UCN-01 blocks CRC-SC growth and potentiates the activity of irinotecan in vitro and in vivo CRC-SC-derived models. Reverse-Phase Protein Microarrays (RPPA) revealed that, albeit CRC-SCs display individual phospho-proteomic profiles, sensitivity of CRC-SCs to UCN-01 relies on the interference with the DNA damage response mediated by Chk1. Combination of LY2603618, a specific Chk1/2 inhibitor, with irinotecan resulted in a significant reduction of CRC-SC growth in vivo, confirming that irinotecan treatment coupled to inhibition of Chk1 represents a potentially effective therapeutic approach for CRC treatment.

Identification of Circadian Determinants of Cancer Chronotherapy through In Vitro Chronopharmacology and Mathematical Modeling.

Cancer chronotherapy aims at enhancing tolerability and efficacy of anticancer drugs through their delivery according to circadian clocks. However, mouse and patient data show that lifestyle, sex, genetics, drugs, and cancer can modify both host circadian clocks and metabolism pathways dynamics, and thus the optimal timing of drug administration. The mathematical modeling of chronopharmacology could indeed help moderate optimal timing according to patient-specific determinants. Here, we combine in vitro and in silico methods, in order to characterize the critical molecular pathways that drive the chronopharmacology of irinotecan, a topoisomerase I inhibitor with complex metabolism and known activity against colorectal cancer. Large transcription rhythms moderated drug bioactivation, detoxification, transport, and target in synchronized colorectal cancer cell cultures. These molecular rhythms translated into statistically significant changes in pharmacokinetics and pharmacodynamics according to in vitro circadian drug timing. The top-up of the multiple coordinated chronopharmacology pathways resulted in a four-fold difference in irinotecan-induced apoptosis according to drug timing. Irinotecan cytotoxicity was directly linked to clock gene BMAL1 expression: The least apoptosis resulted from drug exposure near BMAL1 mRNA nadir (P < 0.001), whereas clock silencing through siBMAL1 exposure ablated all the chronopharmacology mechanisms. Mathematical modeling highlighted circadian bioactivation and detoxification as the most critical determinants of irinotecan chronopharmacology. In vitro-in silico systems chronopharmacology is a new powerful methodology for identifying the main mechanisms at work in order to optimize circadian drug delivery.

PTH-272 Modulation of NRF2 alters the responsiveness of colorectal cancer cells to irinotecan

<h3>Introduction</h3> Chemotherapy is essential to improve outcomes in colorectal cancer (CRC). Irinotecan is a pro-drug commonly used in CRC which requires conversion to the active metabolite Sn38. The cytoprotective protein NRF2 has been associated with chemo-resistance and a worse prognosis in a number of cancers.¹It also regulates the activity of enzymes (including CES1 and UGT1A1) that coordinate irinotecan metabolism and efflux. ²NRF2 remains bound to the regulatory protein KEAP1 in its quiescent state. The aim of this study is to establish the value of modulating NRF2 as a novel strategy for enhancing the efficacy of irinotecan-based chemotherapy either through increased conversion of irinotecan to Sn38 or by sensitisation of CRC cells to its effect. <h3>Method</h3> NRF2 was modulated in the human CRC cell line HCT116 using small inhibitory RNA (siRNA) targeting <i>NRF2</i>and <i>KEAP1 </i>or the chemical inhibitor brusatol and inducer CDDO-Me. Cells were then exposed to irinotecan over a range of concentrations for 48 h. Cell viability was measured by MTS assay and expressed as a percentage of control for the calculation of IC₅₀values. GraphPad Prism 6^®was used for all statistical analysis and the generation of dose-response curves. <h3>Results</h3> Successful NRF2 modulation was confirmed by western blotting for Nrf2 and downstream targets NQO1 and HO-1. The IC₅₀value of irinotecan in untreated control cells was 90µM. Induction of NRF2 by <i>KEAP1</i>siRNA (IC₅₀220µM) or CDDO-Me (IC₅₀325µM) significantly reduced irinotecan cytotoxicity (p &lt; 0.0001). In contrast, inhibition of NRF2 by <i>NRF2</i>siRNA (IC₅₀11µM) or brusatol (IC₅₀55µM) had the opposite effect (p &lt; 0.0001). <h3>Conclusion</h3> Inhibition of NRF2 sensitises CRC cells to irinotecan cytotoxicity and may allow improved response to treatment, or permit lower doses with fewer side effects. We are currently validating these findings <i><i>in vivo</i></i>and examining the contribution of processes that mediate the disposition of irinotecan. The availability of NRF2 inhibiting and inducing compounds may allow for successful modulation in the clinical setting. <h3>Disclosure of interest</h3> J. Evans Grant/ Research Support from: Cancer Research UK, B. Winiarski: None Declared, P. Sutton: None Declared, D. Palmer: None Declared, N. Kitteringham: None Declared. <h3>References</h3> Kawasaki Y, Ishigami S, Arigami T, Uenosono Y, Yanagita S, Uchikado Y, <i>et al</i>. Clinicopathological significance of nuclear factor (erythroid-2)-related factor 2 (Nrf2) expression in gastric cancer. BMC Cancer 2015;<b>15</b>(1):5 Wang M, Zhu JY, Chen S, Qing Y, Wu D, Lin YM, <i>et al</i>. Effects of co-treatment with sulforaphane and autophagy modulators on uridine 5’-diphospho-glucuronosyltransferase 1A isoforms and cytochrome P450 3A4 expression in Caco-2 human colon cancer cells. Oncol Lett. 2014;8(6):2407–16

Image-based evaluations of distribution and cytotoxicity of Irinotecan (CPT-11) in a multi-compartment micro-cell coculture device

We recently developed a polydimethylsiloxane (PDMS)-based three-compartment microfluidic cocultivation device enabling real-time interactions of different cell populations as an advanced physiologically-relevant cell-based assay. This device had valves and small magnetic stirrer-based internal pumps for easy and flexible perfusion operations. In this study, we applied this device for the evaluation of Irinotecan (CPT-11) toxicity to the lung, because it is detoxified by the liver and accumulated in the fat in humans. We successfully cultured representative three different tissue model cells in each compartment under the individual culture conditions and also in entire perfusion. Growth inhibition of rat lung epithelial cell line L-2, was measured when administered with 50μM CPT-11 under various cocultivation conditions with respect to the presences and absence of primary rat hepatocytes (liver tissue model) and adipocyte-like cells (fat tissue model) induced from a mouse fibroblast cell line, 3T3-L1. Although CPT-11 showed moderate toxicity to the pure culture of L-2 cells in the device after 72h of perfusion culture, this was lowered mainly in the presence of the liver tissue. Inhibition of the L-2 cell growth agreed with the area under curve (AUC) values obtained from fluorescent image-based analyses in each compartment. These results demonstrate that developed simple and flexible microfluidic cocultivation device, with appropriate image-based analyses, can be used in evaluating toxicokinetic behaviors of drug candidates in systemic levels.

Synergistic Cytotoxicity of Irinotecan and Cisplatin in Dual-Drug Targeted Polymeric Nanoparticles

Two unexplored aspects for irinotecan and cisplatin (I&C) combination chemotherapy are: actively targeting both drugs to a specific diseased cell type, and delivering both drugs on the same vehicle to ensure their synchronized entry into the cell at a well-defined ratio. In this work, the authors report the use of targeted polymeric nanoparticles (NPs) to coencapsulate and deliver I&C to cancer cells expressing the prostate-specific membrane antigen. Targeted NPs were prepared in a single step by mixing four different precursors inside microfluidic devices. I&C were encapsulated in 55-nm NPs and showed an eightfold increase in internalization by prostate-specific membrane antigen-expressing LNCaP cells compared with nontargeted NPs. NPs coencapsulating both drugs exhibited strong synergism in LNCaP cells with a combination index of 0.2. The strategy of coencapsulating both I&C in a single NP targeted to a specific cell type could potentially be used to treat different types of cancer.

Treatment of Colorectal Cancer Using a Combination of Liposomal Irinotecan (Irinophore C™) and 5-Fluorouracil

PurposeTo investigate the use of liposomal irinotecan (Irinophore C™) plus or minus 5-fluorouracil (5-FU) for the treatment of colorectal cancer.Experimental DesignThe effect of irinotecan (IRI) and/or 5-FU exposure times on cytotoxicity was assessed in vitro against HT-29 or LS174T human colon carcinoma cells. The pharmacokinetics and biodistribution of Irinophore C™ (IrC™) and 5-FU, administered alone or in combination, were compared in vivo. A subcutaneous model of HT-29 human colorectal cancer in Rag2-M mice was utilized to assess the efficacy of IrC™ alone, and in combination with 5-FU.ResultsThe cytotoxicity of IRI and 5-FU were strongly dependent on exposure time. Synergistic interactions were observed following prolonged exposure to IRI/5-FU combinations. Pharmacokinetics/biodistribution studies demonstrated that the 5-FU elimination rate was decreased significantly when 5-FU was co-administered intravenously with IrC™, versus alone. Significant decreases in 5-FU elimination were also observed in plasma, with an associated increase of 5-FU in some tissues when 5-FU was given by intraperitoneal injection and IrC™ was given intravenously. The elimination of IrC™ was not significantly different when administered alone or in combination with 5-FU. Therapeutic studies demonstrated that single agent IrC™ was significantly more effective than the combination of IRI/5-FU; surprisingly, IrC™/5-FU combinations were no more effective than IrC™ alone. The administration of combinations of 5-FU (16 mg/kg) and IrC™ (60 mg IRI/kg) showed increased toxicity when compared to IrC™ alone. Treatment with IrC™ alone (60 mg IRI/kg) delayed the time required for a 5-fold increase in initial tumor volume to day 49, compared to day 23 for controls. When IrC™ (40 mg IRI/kg) was used in combination with 5-FU (16 mg/kg), the time to increase tumor volume 5-fold was 43 days, which was comparable to that achieved when using IrC™ alone (40 mg IRI/kg).ConclusionsSingle agent IrC™ was well tolerated and has significant therapeutic potential. IrC™ may be a suitable replacement for IRI treatment, but its use with free 5-FU is complicated by IrC™-engendered changes in 5-FU pharmacokinetics/biodistribution which are associated with increased toxicity when using the combination.

Abstract 3353: Effect of erlotinib on the pharmacokinetics of irinotecan and its metabolites: A drug-drug interaction study.

Abstract Purpose: Since irinotecan and erlotinib are used in treatment for non-small cell lung cancer (NSCLC), co-administration of these drugs might be an effective treatment for NSCLC. However, drug-drug interaction should be concerned because these drugs are metabolized by CYP3A4 and excreted into bile by ABC transporters. To clarify the drug-drug interaction when these drugs are co-administered, we examined the effect of this co-administration on the pharmacokinetics of irinotecan, its active metabolite 7-ethyl-10-hydroxycamptothecin (SN-38), SN-38 glucuronide (SN-38G) in animal model. Methods: Cytotoxicity of irinotecan and SN-38 with erlotinib against NSCLC cell line (A549) was assessed. To elucidate the effect of drug-drug interaction between irinotecan and erlotinib, male Sprague-Dawley rats were treated with 10 mg/kg irinotecan and 25 mg/kg erlotinib. The concentrations of irinotecan, SN-38, SN-38G and erlotinib in plasma, bile, and tissues were analyzed by high-performance liquid chromatography. Pharmacokinetic parameters were calculated by non-compartmental model methods. Results: Co-exposured erlotinib enhanced the cytotoxicity of SN-38 in a dose-dependent manner, however it did not significantly affect the cytotoxicity of irinotecan against A549 that is resistant to erlotinib. Additionally, co-administration of erlotinib resulted in a significant increase of plasma concentration of irinotecan, SN-38 and SN-38G compared to irinotecan alone, but decrease in the biliary excretion. Erlotinib significantly increased the concentration of irinotecan and SN-38G in kidney and that of SN-38 in lung and jejunum. On the other hand, co-administration of irinotecan intended to decrease the plasma concentration of erlotinib but significant differences of pharmacokinetic parameters were not observed. Also, levels of erlotinib in tissues were not affected by co-administration of irinotecan. Conclusions: We proposed the possibility of combination therapy of irinotecan with erlotinib because the plasma concentration of irinotecan and its active metabolite SN-38 significantly increased in vivo and the cytotoxicity of SN-38 against NSCLC cell line enhanced. New effective salvage chemotherapy regimens are needed in patients with NSCLC. We conducted a phase I/II clinical study of combination chemotherapy with irinotecan and erlotinib for recurrent NSCLC because our data have shown synergistic/additive effects between irinotecan and epidermal growth factor receptor-tyrosine kinase inhibitor. Citation Format: Emiko Sanematsu, Issei Toyooka, Yuki Takashima, Tamio Okimoto, Yukari Tsubata, Hideyuki Saito, Takeshi Isobe, Akinobu Hamada. Effect of erlotinib on the pharmacokinetics of irinotecan and its metabolites: A drug-drug interaction study. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3353. doi:10.1158/1538-7445.AM2013-3353

Modulation of Esterified Drug Metabolism by Tanshinones from Salvia miltiorrhiza (“Danshen”)

The roots of Salvia miltiorrhiza ("Danshen") are used in traditional Chinese medicine for the treatment of numerous ailments including cardiovascular disease, hypertension, and ischemic stroke. Extracts of S. miltiorrhiza roots in the formulation "Compound Danshen Dripping Pill" are undergoing clinical trials in the United States. To date, the active components of this material have not been conclusively identified. We have determined that S. miltiorrhiza roots contain potent human carboxylesterase (CE) inhibitors, due to the presence of tanshinones. K(i) values in the nM range were determined for inhibition of both the liver and intestinal CEs. As CEs hydrolyze clinically used drugs, the ability of tanshinones and S. miltiorrhiza root extracts to modulate the metabolism of the anticancer prodrug irinotecan (CPT-11) was assessed. Our results indicate that marked inhibition of human CEs occurs following incubation with both pure compounds and crude material and that drug hydrolysis is significantly reduced. Consequently, a reduction in the cytotoxicity of irinotecan is observed following dosing with either purified tanshinones or S. miltiorrhiza root extracts. It is concluded that remedies containing tanshinones should be avoided when individuals are taking esterified agents and that patients should be warned of the potential drug-drug interaction that may occur with this material.

Phase II study of bevacizumab plus irinotecan and carboplatin for recurrent WHO grade 3 malignant gliomas with no prior bevacizumab failure.

2095^ Background: No standard treatment exists for recurrent WHO grade 3 malignant glioma patients. Bevacizumab (BV) with irinotecan has significant anti-tumor activity for recurrent glioblastoma. Carboplatin has anti-glioma activity and can potentiate the cytotoxicity of irinotecan. We evaluated the progression-free survival (PFS) of BV in combination to irinotecan and carboplatin in recurrent WHO grade 3 malignant gliomas, as well as its safety. Methods: Adult patients with measurable recurrent WHO grade 3 malignant glioma, ≥12 weeks after radiation therapy, ≥4 weeks after chemotherapy, with adequate organ function, KPS ≥70%, no prior failure or grade ≥3 toxicity to the agents, and no contraindications to BV were eligible for study. Patients received BV at 10 mg/kg with irinotecan on days 1 and 15 of a 28-day cycle. The dose of irinotecan was 125 mg/m2 for patients not on enzyme inducing anti-epileptics (EIAEDs) and 340 mg/m2 for patients on EIAEDs. All patients received carboplatin at an AUC of 4 on day 1 of each cycle. MRIs were obtained every 8 weeks. Results: As planned, 39 WHO grade III malignant glioma patients were enrolled on study. Median age was 47 (range, 26-71). At a median follow up of 14 months, the 6-month PFS is 69% and the median PFS is 11 months. A median of 8 cycles were given. Seven patients completed the planned course of treatment (12 cycles) with hypometabolic PET scan and nine patients remain on study. Fifteen patients came off study due to disease progression and eight due to toxicity. As of 1/25/2012, 22 patients are still alive and 17 have died. Grade 3-4 toxicities included: neutropenia (grade 3, n=12; grade 4, n= 1), thrombocytopenia (grade 3, n=6; grade 4, n=4), nausea (grade 3, n=7), diarrhea (grade 3, n=2), deep venous thrombosis (grade 3, n=2), febrile neutropenia (grade 3, n=1; grade 4, n=1), fatigue (grade 3, n=8; grade 4, n=1), cerebral infarction (grade 4, n=3), intracranial hemorrhage (grade 4, n=1), posterior reversible encephalopathy syndrome (grade 3, n=1). Conclusions: The combination of bevacizumab, irinotecan and carboplatin for WHO grade 3 malignant glioma patients is effective and with no more than expected toxicity.

Protective effects of propolis and related polyphenolic/flavonoid compounds against toxicity induced by irinotecan

Despite the excellent chemotherapeutic effect of irinotecan, its cytotoxicity and genotoxicity in normal cells remains a major problem in chemotherapy. This study was carried out to find whether propolis preparations and related flavonoids (quercetin, naringin) might enhance irinotecan-induced cytotoxicity to tumor cells in mice bearing Ehrlich ascites tumors (EAT) while protecting normal blood, liver, and kidney cells. The preparation of propolis and their flavonoids were given to mice intraperitoneally at a dose of 100 mg kg(-1) body weight for three consecutive days before the ip injection of EAT cells (2×10(6)). Irinotecan was administered ip at dose of 50 mg kg(-1) on days 3, 4, and 5 after tumor cell inoculation. The combination treatment resulted in substantial inhibition of the growth of EAT cells as well as treatment with quercetin or irinotecan alone, whereas other treatment by itself showed little effect. However, when mice were pre-treated with test components prior to irinotecan, the frequencies of irinotecan-induced micronuclei (MN) was decreased but in mice bearing tumor QU and EEP increased number of micronucleated cells. Propolis preparation and related flavonoids were found to exhibit an important immunomodulatory effect and could decrease irinotecan-induced toxic and genotoxic effects to normal cells without effecting irinotecan cytotoxicity in EAT cells.

Epigenetic mechanisms of irinotecan sensitivity in colorectal cancer cell lines

Irinotecan is a topoisomerase-I (Top-I) inhibitor used for the treatment of colorectal cancer. DNA demethylating agents, including 5-azacytidine (5-aza), display synergistic antitumor activity with several chemotherapy drugs. 5-Aza may enhance irinotecan cytotoxicity by at least one of the following mechanisms: (a) Top-I promoter demethylation, (b) activation of genes involved in Top-I transcriptional regulation (p16 or Sp1), and (c) modulation of the cell cycle and apoptosis after DNA damage. The growth-inhibitory effects of SN38, the active metabolite of irinotecan, 5-aza, and their combinations, were studied in four colorectal cancer cell lines. The effects of treatments on cell cycle were analyzed by flow cytometry, and apoptosis was measured by fluorescence microscopy. Top-I, Sp1, and p53 expression modulated by 5-aza were measured by real-time PCR. Methylation of Top-I, p16, 14-3-3sigma, and hMLH1 promoters before and after 5-aza treatment were measured by MethyLight PCR and DNA bisulfite sequencing. Low-dose 5-aza significantly enhanced the apoptotic effect of irinotecan in all colorectal cancer cells, whereas a synergistic cytotoxic effect was observed only in p53-mutated cells (HT29, SW620, and WiDr). This synergistic effect was significantly correlated with Top-I up-regulation by 5-aza, and coupled to p16 demethylation and Sp1 up-regulation. p16 demethylation was also associated with enhanced cell cycle arrest after irinotecan treatment. In contrast, 5-aza down-regulated Top-I expression in the p53 wild-type LS174T cells in a p53-dependent manner, thereby reducing SN38 cytotoxicity. In conclusion, 5-aza modulates Top-I expression by several mechanisms involving Sp1, p16, and p53. If confirmed in other models, these results suggest that p16 and p53 status affects the 5-aza-irinotecan interaction.

GSTP1 Ile105Val polymorphism correlates with progression-free survival in MCRC patients treated with or without irinotecan: a study of the Dutch Colorectal Cancer Group

A Valine residue at position 105 of the GSTP1 protein results in decreased enzyme activity. As nuclear GSTP1 activity decreases irinotecan cytotoxicity, Val-allele carriers may benefit more from irinotecan chemotherapy. Our aim was to investigate the association of GSTP1 genotype with treatment outcome of irinotecan. Progression-free survival (PFS) and toxicity were determined in 267 metastatic colorectal cancer (MCRC) patients who were treated with first-line capecitabine (CAP) plus irinotecan (CAPIRI), or CAP single agent in a prospective randomised phase III trial (CAIRO). GSTP1 genotype was determined by Pyrosequencing. Patients receiving CAP showed a PFS of 6.6 (Ile/Ile), 6.0 (Ile/Val) and 6.5 months (Val/Val); compared to 7.0 (Ile/Ile), 8.8 (Ile/Val) and 9.2 months (Val/Val) with CAPIRI. Median PFS was 2.7 months longer in Val-allele carriers treated with CAPIRI compared to CAP (P=0.005). Patients with the Ile/Ile genotype showed similar PFS with CAPIRI and CAP (7.0 compared to 6.6 months, P=0.972). Toxicity did not differ significantly among genotypes. GSTP1 codon 105 polymorphism may be predictive for the response to irinotecan-based chemotherapy in patients with MCRC, with the Val-allele being associated with a better outcome. Ile/Ile genotype patients do not appear to benefit from the addition of irinotecan to CAP.