Agent Topotecan Research Articles

Abstract 4562: Activity of PARP1-selective inhibitor SNV-001 in models of HRD cancers as monotherapy and in combination

Abstract Current FDA approved poly (ADP-ribose) polymerase inhibitors (PARPi) inhibit both PARP1 and PARP2. Genetic studies indicate that loss of PARP1 is synthetic lethal with homologous recombination deficiency (HRD), including BRCA1/2 mutations, and PARP2 depletion may be linked to hematologic toxicity. To date, no first generation PARPi has been successfully combined with standard of care (SoC) agents in the clinic without significant reduction in dosing intensity because of overlapping toxicities. A selective inhibitor could potentially improve the therapeutic index and provide additional opportunities in combination strategies especially with drugs that are prone to myelosuppression. SNV-001 potently and selectively inhibits PARP1 in biochemical and cellular assays. SNV-001 inhibits PARylation in the low nanomolar range with over 500-fold selectivity against PARP2. SNV-001 did not inhibit any other PARP family member tested up to 10 uM in biochemical assays confirming its selectivity for PARP1. In human erythroid and myeloid colony formation assays, the IC50s of SNV-001 for both lineages were approximately 100-times higher than the target efficacious concentration based on the IC95 for inhibition of DLD1 (BRCA2 -/-) colony formation. In a subcutaneous MDA-MB-436 (BRCA1mut) xenograft model, oral administration of SNV-001 once daily resulted in a dose-dependent tumor regression. SNV-001 showed greater efficacy at lower doses compared to olaparib at 100 mg/kg or at equivalent doses of AZD9574. SNV-001 showed promising synergy with SoC chemotherapies or DDR inhibitors in cell proliferation assays. SNV-001 enhanced cytotoxicity of the DNA-damaging agent topotecan or carboplatin in Capan-1 (BRCA2mut) and HCC1395 (BRCA1mut) cells. Synergy was seen between SNV-001 and cell cycle check point inhibitors camonsertib (ATRi) or adavosertib (WEE1i) in DLD1 (BRCA2 -/-) and UWB1.289 (BRCA1mut) cells, as DNA damage induced by PARP1 trapping relies on cell cycle modulation for repair. Combination of SNV-001 with polymerase theta inhibitor ART558 or USP1 inhibitor KSQ4279 exhibited synergistic effects in DLD1 (BRCA2 -/-) or UWB1.289 (BRCA1mut) cells. These results suggest that inhibition of microhomology-mediated end-joining and translesion synthesis pathways may further exacerbate SNV-001-induced DDR in certain cellular contexts.In summary, the PARP1 selective inhibitor SNV-001 demonstrates potent antitumor activity and shows promising synergy between SoC chemotherapies or DDR inhibitors. Current data highlight a potential strategy for improving treatment efficacy and overcoming resistance. Further clinical investigations are warranted to validate this combination strategy. Citation Format: Zhentian Li, Qipeng Fan, Mingming Gao, Jun Pan, Liangxing Wu, Wenqing Yao, Phillip C. Liu. Activity of PARP1-selective inhibitor SNV-001 in models of HRD cancers as monotherapy and in combination [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4562.

Multilayered Polyurethane/Poly(vinyl alcohol) Nanofibrous Mats for Local Topotecan Delivery as a Potential Retinoblastoma Treatment.

Local chemotherapy using polymer drug delivery systems has the potential to treat some cancers, including intraocular retinoblastoma, which is difficult to treat with systemically delivered drugs. Well-designed carriers can provide the required drug concentration at the target site over a prolonged time, reduce the overall drug dose needed, and suppress severe side effects. Herein, nanofibrous carriers of the anticancer agent topotecan (TPT) with a multilayered structure composed of a TPT-loaded inner layer of poly(vinyl alcohol) (PVA) and outer covering layers of polyurethane (PUR) are proposed. Scanning electron microscopy showed homogeneous incorporation of TPT into the PVA nanofibers. HPLC-FLD proved the good loading efficiency of TPT (≥85%) with a content of the pharmacologically active lactone TPT of more than 97%. In vitro release experiments demonstrated that the PUR cover layers effectively reduced the initial burst release of hydrophilic TPT. In a 3-round experiment with human retinoblastoma cells (Y-79), TPT showed prolonged release from the sandwich-structured nanofibers compared with that from a PVA monolayer, with significantly enhanced cytotoxic effects as a result of an increase in the PUR layer thickness. The presented PUR-PVA/TPT-PUR nanofibers appear to be promising carriers of active TPT lactone that could be useful for local cancer therapy.

Influence of Tyrosyl-DNA Phosphodiesterase 1 Inhibitor on the Proapoptotic and Genotoxic Effects of Anticancer Agent Topotecan

To date, various strategies have been proposed to increase the efficiency of cancer therapy. It is known that the action of DNA repair system can determine the resistance of cancer cells to DNA-damaging chemotherapy and radiotherapy, and one of these ways to increase therapeutic efficiency is the search for inhibitors of enzymes of the DNA repair system. Inhibition of the DNA repair enzyme tyrosyl-DNA phosphodiesterase1 (Tdp1) leads to an increase in the effectiveness of the topoisomerase 1 (Top1) inhibitor, the anticancer drug topotecan. Covalent complexes Top1-DNA, which are normally short-lived and are not a threat to the cell, are stabilized under the influence of topotecan and lead to cell death. Tdp1 eliminates such stabilized complexes and thus weaken the effect of topotecan therapy. We have previously shown that the use of the usnic acid hydrazonothiazole derivative OL9-119 in combination with topotecan increased the antitumor and antimetastatic efficacy of the latter in a mouse model of Lewis lung carcinoma. In this work, it was shown that the combined use of topotecan and Tdp1 inhibitor, the hydrazonothiazole derivative of usnic acid OL9-119, leads to an increase in the DNA-damaging effect of topotecan which is used in the clinic for the treatment of cancer. The study of the proapoptotic effect of the compound OL9-119 showed that the compound itself does not induce apoptosis, but increases the proapoptotic effect of topotecan. The results of the study could be used to improve the effectiveness of anticancer therapy and/or to reduce the therapeutic dose of topotecan and, therefore, the severity of side effects.

Elucidation of the Mechanism of Topotecan-induced Antitumor Immune Activation

Danger-associated molecular patterns (DAMPs) derived from damaged or dying cells elicit inflammation and potentiate antitumor immune responses. We found that treatment of cancer cells with the antitumor agent topotecan (TPT), an inhibitor of topoisomerase I (TOP1), induces DAMP secretion that triggers dendritic cell activation and cytokine production. TPT administration inhibits tumor growth in tumor-bearing mice, which is accompanied by infiltration of activated DCs and CD8+ T cells. These effects are abrogated in mice lacking stimulator of interferon genes (STING), an essential molecule in cytosolic DNA-mediated innate immune responses. Furthermore, we identified ribosomal protein L15 (RPL15), a 60S ribosomal protein, as a novel TPT target and showed that TPT inhibited pre-ribosomal subunit formation via its binding to RPL15, resulting in the induction of DAMP-mediated antitumor immune activation independent of TOP1. RPL15 knockdown induced DAMP secretion and increased the cytotoxic T lymphocyte (CTL) population but decreased the T-regulatory cell (Treg) population in a B16-F10 murine melanoma model, which sensitized B16-F10 tumors against programmed death receptor-1(PD-1) blockade. Our study identified a novel TPT target protein and showed that ribosomal stress is a trigger of DAMP secretion, which contributes to antitumor immunotherapy.

A sensitive surfactant-mediated spectrofluorimetric determination of chemotherapeutic agent topotecan in human serum and its investigation of DNA binding mechanism

A sensitive surfactant-mediated spectrofluorimetric determination of chemotherapeutic agent topotecan in human serum and its investigation of DNA binding mechanism

Reshaping Tumor Blood Vessels to Enhance Drug Penetration with a Multistrategy Synergistic Nanosystem

Due to the abnormal tumor vasculature and dense stroma, there is limited tumor perfusion in the immunosuppressive tumor microenvironment (TME). In order to reshape tumor blood vessels and enhance the penetration of anticancer drugs into the tumor tissue, an anionic liposome nanosystem with a "sandwich" structure was prepared. The chemotherapeutic agent topotecan (TPT) was encapsulated in the lipid hydrophilic layer, and the sensitizer indocyanine green (ICG) was loaded into the hydrophobic layer. In addition, the positively charged vascular normalization drug erlotinib (ERL) was adsorbed to the outermost layer of the microenvironment. The nanosystem showed superior tumor permeability in vitro/in vivo experiments compared with the ERL-treated group. The nanosystem entered the tumor through normalization of blood vessels after the action of ERL. Ultrasound treatment improves the vascular permeability, allowing the nanoparticles to penetrate blood vessels and reach tumor cells. Finally, in addition to cytotoxic effects, TPT can also down-regulate the expression of HIF-1α and so prolong the vascular normalization time. These experimental results showed that the nanosystem effectively improves the tumor microenvironment. This work indicates the great potential of vascular normalization combined with sonodynamic therapy and chemotherapy to enhance efficiency.

Development and Validation of Reversed Phase HPLC Method for the Simultaneous Detection of Lactone and Carboxylate Forms of Topotecan Along with Thymoquinone: Application to Nanoparticulate Anticancer Formulation System

Nanomedicine combination therapy, a strategy to delivery multiple drugs via a single nanosystem has opened new avenues to treat cancer and prevent side effects and overcome chemoresistance. The present work utilizes a chemotherapeutic agent topotecan hydrochlorideand a natural chemosensitizer (thymoquinone, TQ) incorporated into poly (lactic-co-glycolic acid) nanoparticles (PLGA NPs) to target tumor. The objective of the present work is to develop a reversed phase HPLC method for simultaneous estimation of TQ along with the pH dependent lactone and carboxylate forms of topotecan and validate its applicability. Chromatographic separation was obtained on Agilent 1200 Infinity series C18 analytical column using gradient elution with acetonitrile and 1% triethylamine acetate (TEAA) buffer (pH 5.5) as mobile phase and a flow rate of 0.8 mL/min. Analytical detection was done at 253 nm. Validation was performed as per ICH guidelines and characterization and stability of the optimized nanoparticles were evaluated. The retention time of 5.93, 8.73, and 19.76 min was obtained for lactone, carboxylate and TQ, respectively. Linearity response was established in the range of 5‒45 µg/mL. The limits of detection and quantification were found to be 15 and 50 ng/mL for lactone, 18 and 59 ng/mL for carboxylate, and 5 and 18 ng/mL for TQ, respectively. The method was validated in terms of robustness, precision, accuracy, specificity and percent loading, entrapment efficiency, and in vitro drug release. The stability of the nanoparticles was determined using the above validated method. The method was successfully validated and applied for routine analyses indicating its use as a suitable analytical tool to determine efficiency of the proposed therapy.

PDCT-04. ETMR ONE: AN INTERNATIONAL REGISTRY AND RESEARCH PLATFORM FOR CHILDREN WITH EMBRYONAL TUMOR WITH MULTILAYER ROSETTES

Abstract Embryonal tumor with multilayer rosettes (ETMR) is a rare and highly-aggressive CNS neoplasm which occurs almost exclusively in young children and is associated with an extremely poor prognosis. Due to the rare nature of ETMR and its recent classification, no large clinical investigations have been conducted to determine the optimal therapy for these tumors. Historical data suggests that extensive surgical resection, radiotherapy, and high-dose chemotherapy are not sufficient treatment in the vast majority of cases with a reported 1-year EFS of 36% and 1-year OS of 45% using these approaches. New treatment regimens incorporating the known biology of ETMR must be investigated. Pre-clinical drug screens using the ETMR BT183 cell line and its xenograft have demonstrated responses to the chemotherapy agents topotecan, doxorubicin, and actinomycin D. Based on this data, a modified IRS-III chemotherapy regimen incorporating these active agents was developed and a small cohort of four children with ETMR were then treated. Three patients completed therapy without subsequent relapse and have EFS of >30 months. The ETMR One international registry and its associated consensus treatment protocol represent the first prospective evaluation of treatment outcomes for children with ETMR. The consensus regimen involves maximal feasible surgical resection, modified IRS-III chemotherapy, high-dose chemotherapy with autologous stem cell rescue, and radiotherapy (as indicated). Patients enrolled on the registry will have their cases periodically reviewed by a medical advisory board that will offer management recommendations, particularly regarding the use of radiotherapy or the need for additional surgical interventions. Through the procurement of patient tumor samples and development of additional cell lines and xenografts, the registry aims to serve as a platform for translational research. Pre-clinical findings will be used to modify the consensus protocol therapy as indicated or to generate additional biology-based phase I/II trials for ETMR.

Small-cell lung cancer growth inhibition: synergism between NMDA receptor blockade and chemotherapy.

BackgroundSmall-cell lung cancer (SCLC) has a poor prognosis since there is currently no effective therapy for commonly recurring disease. In our previous study, both primary and recurrent human tumors have been shown to express functional N-methyl-D-aspartate (NMDA) receptors, and blockade of these receptors with GluN1 and GluN2B antagonists decreased tumor cell viability in vitro, and growth of tumor xenografts in nu/nu mice.Materials and methodsIn this study, we examine the influence of the GluN2B antagonist ifenprodil and the channel-blocker antagonist memantine, on cell viability and growth of tumor xenografts of recurrent SCLC (rSCLC) in mice.ResultsBoth antagonists significantly reduced cell viability and levels of components of the ERK1/2 pathway, increased apoptosis, and at very safe levels significantly reduced the growth of tumors in mice. Each antagonist and topotecan had additive effects to reduce cell viability with significant synergy demonstrated for the case of memantine. More significantly, combination treatments of xenografts in mice with ifenprodil and the chemotherapeutic agent topotecan produced clear additive effects that completely stopped tumor growth. Moreover, the ifenprodil and topotecan combination showed excellent supra-addition or synergy of inhibition for tumors ≤300 mm in size (P=4.7E−4). Combination treatment of memantine with topotecan also showed clear addition but, unlike ifenprodil, no synergy for the doses chosen.ConclusionSince topotecan is a drug of choice for treatment of rSCLC, our findings suggest that combining this agent with NMDA receptor blockade using the GluN2B antagonist, ifenprodil, will significantly improve patient outcomes.

Increased delivery of chemotherapy to the vitreous by inhibition of the blood-retinal barrier

Treatment of retinoblastoma -a pediatric cancer of the developing retina- might benefit from strategies to inhibit the blood-retinal barrier (BRB). The potent anticancer agent topotecan is a substrate of efflux transporters BCRP and P-gp, which are expressed at the BRB to restrict vitreous and retinal distribution of xenobiotics. In this work we have studied vitreous and retinal distribution, tumor accumulation and antitumor activity of topotecan, using pantoprazole as inhibitor of BCRP and P-gp. We used rabbit and mouse eyes as BRB models and patient-derived xenografts as retinoblastoma models. To validate the rabbit BRB model we stained BCRP and P-gp in the retinal vessels. Using intravitreous microdialysis we showed that the penetration of the rabbit vitreous by lactone topotecan increased significantly upon concomitant administration of pantoprazole (P=0.0285). Pantoprazole also increased topotecan penetration of the mouse vitreous, measured as the vitreous-to-plasma topotecan concentration ratio at the steady state (P=0.0246). Pantoprazole increased topotecan antitumor efficacy and intracellular penetration in retinoblastoma in vitro, but did not enhance intratumor drug distribution and survival in mice bearing the intraocular human tumor HSJD-RBT-2. Anatomical differences with the clinical setting likely limited our in vivo study, since xenografts were poorly vascularized masses that loaded most of the vitreous compartment. We conclude that pharmacological modulation of the BRB is feasible, enhances anticancer drug distribution into the vitreous and might have clinical implications in retinoblastoma. Chemical compounds included in this manuscriptTopotecan (PubChem CID: 60700)Pantoprazole sodium (PubChem CID: 15008962)

Abstract 1557: Longitudinal expression response of glycosylation related genes, regulators, and targets in cancer cell lines treated with eleven anti-tumor agents

Abstract Cancer cells have profound alterations in glycosylation processes, which affects the functioning of malignant cells and their interaction with surrounding environment. We examined the effects of treatment of cancer cell lines with eleven antitumor agents (bortezomib, cisplatin, dasatinib, doxorubicin, erlotinib, gemcitabine, paclitaxel, sirolimus, sorafenib, sunitinib, and topotecan) on transcriptional changes in 179 candidate genes involved in biochemical glycosylation pathways or encoding glycosylation targets, regulators of glycosylation processes, and components of cancer pathways affected by glycosylation. In order to achieve this goal, we analyzed time course gene expression information for the NCI-60 cancer cell line panel using the data from the NCI Transcriptional Pharmacodynamic Workbench (NCI TPW), a resource developed at the National Cancer Institute. Our analysis identified 145 glycosylation related genes that responded to drug treatment in a concerted manner. Many of these genes had a concerted response to multiple agents; however, the direction and the magnitude of concerted response were specific to individual drugs. Expression of multiple genes prior to treatment and/or transcriptional changes in response to treatment were significantly correlated with drug chemosensitivity, measured as log(GI50). Treatment with dasatinib resulted in the largest number of significant correlations of chemosensitivity with gene expression prior to treatment and with gene expression changes in response to treatment. Significant correlations of transcriptional changes after treatment and/or of pre-treatment gene expression levels of glycosylation related genes and glycosylation targets were also observed for other antitumor agents. Chemosensitivity to kinase inhibitors and DNA damaging agents was significantly correlated with expression of genes involved in N- and O-glycosylation, fucosylation, biosynthesis of poly-N-acetyllactosamine, removal of misfolded proteins, binding to hyaluronic acid and other glycans, and cell adhesion. In particular, tumor cell sensitivity or resistance to multiple agents were significantly correlated with changes in expression of C1GALT1C1 (COSMC), FUCA1, SDC1, MUC1, members of the MGAT, GALNT, B4GALT, B3GNT, MAN, and EDEM gene families, and other genes involved in glycosylation processes or encoding glycosylation targets and signaling regulators. We present a catalog of dynamic concerted changes in expression of glycosylation related genes, their ligands, regulators, and targets in response to chemotherapy, and provide a list of genes that influence the strength of response of cancer cells to treatment. These glycosylation related genes may be considered as potential candidates for drug targeting in combination therapy to enhance the effects of treatment. Funded by NCI Contract No. HHSN261200800001E. Citation Format: Julia Krushkal, Yingdong Zhao, Curtis Hose, Anne Monks, James H. Doroshow, Richard Simon. Longitudinal expression response of glycosylation related genes, regulators, and targets in cancer cell lines treated with eleven anti-tumor agents [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1557. doi:10.1158/1538-7445.AM2017-1557

DNA-Containing Exosomes Derived from Cancer Cells Treated with Topotecan Activate a STING-Dependent Pathway and Reinforce Antitumor Immunity.

Danger-associated molecular patterns derived from damaged or dying cells elicit inflammation and potentiate antitumor immune responses. In this article, we show that treatment of breast cancer cells with the antitumor agent topotecan (TPT), an inhibitor of topoisomerase I, induces danger-associated molecular pattern secretion that triggers dendritic cell (DC) activation and cytokine production. TPT administration inhibits tumor growth in tumor-bearing mice, which is accompanied by infiltration of activated DCs and CD8+ T cells. These effects are abrogated in mice lacking STING, an essential molecule in cytosolic DNA-mediated innate immune responses. Furthermore, TPT-treated cancer cells release exosomes that contain DNA that activate DCs via STING signaling. These findings suggest that a STING-dependent pathway drives antitumor immunity by responding to tumor cell-derived DNA.

Microwave assisted process intensification and kinetic modelling: Extraction of camptothecin from Nothapodytes nimmoniana plant

Process intensification of natural products extraction is possible through assistance of microwaves, ultrasound, high pressure, enzyme pretreatments and supercritical fluids. In the present paper microwave-assisted process intensification is investigated for the extraction of camptothecin (CPT) from Nothapodytes nimmoniana plant. CPT is an industrial raw material for the production of anticancer agents topotecan and irinotecan and semisynthetic analogs of CPT. Nothapodytes nimmoniana is found wild in India and is a prime source for CPT. Effects of various process factors such as stirring, solid to liquid ratio, microwave power, irradiation time and particle size on extraction kinetics have been studied. The results showed that stirring, microwave power and the temperature had a significant effect on extraction rate and yields of camptothecin. Overall process intensification has been achieved with extraction yield of 0.41% w/w of CPT, an efficiency of about 97% in very short time of just 2min as against 6h by classical string extraction method. Scanning electron microscopy (SEM) analysis of the spent plant material was performed to assess the effect of microwaves and found complete cell disruption has happened. This cell disruption and associated enhancement in mass transfer could be the prime reason for the process intensification. An attempt has been made to fit the extraction kinetics data with different kinetic models such as second order rate model, power law model and two site kinetic models. The second order rate model appears to be the best fit to explain the microwave assisted kinetics.

Ion-Pairing Contribution to the Liposomal Transport of Topotecan as Revealed by Mechanistic Modeling

Actively loaded liposomal formulations of anticancer agents have been widely explored due to their high drug encapsulation efficiencies and prolonged drug retention. Mathematical models to predict and optimize drug loading and release kinetics from these nanoparticle formulations would be useful in their development and may allow researchers to tune release profiles. Such models must account for the driving forces as influenced by the physicochemical properties of the drug and the microenvironment, and the liposomal barrier properties. This study employed mechanistic modeling to describe the active liposomal loading and release kinetics of the anticancer agent topotecan (TPT). The model incorporates ammonia transport resulting in generation of a pH gradient, TPT dimerization, TPT lactone ring-opening and -closing interconversion kinetics, chloride transport, and transport of TPT-chloride ion-pairs to describe the active loading and release kinetics of TPT in the presence of varying chloride concentrations. Model-based predictions of the kinetics of active loading at varying loading concentrations of TPT and release under dynamic dialysis conditions were in reasonable agreement with experiments. These findings identify key attributes to consider in optimizing and predicting loading and release of liposomal TPT that may also be applicable to liposomal formulations of other weakly basic pharmaceuticals.

Thymoquinone from Nigella sativa Seeds Promotes the Antitumor Activity of Noncytotoxic Doses of Topotecan in Human Colorectal Cancer Cells in Vitro.

Topotecan, a topoisomerase I inhibitor, is an anticancer drug widely used in the therapy of lung, ovarian, colorectal, and breast adenocarcinoma. Due to the primary dose-limiting toxicity of topotecan, which is myelosuppressive, it is necessary to identify other chemotherapeutic agents that can work synergistically with topotecan to increase its efficacy and limit its toxicity. Many studies have shown synergism upon the combination of topotecan with other chemotherapeutic agents such as gemcitabine. Other studies have demonstrated that pre-exposing cells to naturally occurring compounds such as thymoquinone, followed by gemcitabine or oxaliplatin, resulted in higher growth inhibition compared to treatment with gemcitabine or oxaliplatin alone. Our aim was to elucidate the underlying mechanism of action of topotecan in the survival and apoptotic pathways in human colon cancer cell lines in comparison to thymoquinone, to study the proapoptotic and antiproliferative effects of thymoquinone on the effectiveness of the chemotherapeutic agent topotecan, and to investigate the potential synergistic effect of thymoquinone with topotecan. Cells were incubated with different topotecan and thymoquinone concentrations for 24 and 48 hours in order to determine the IC50 for each drug. Combined therapy was then tested with ± 2 values for the IC50 of each drug. The reduction in proliferation was significantly dose- and time-dependent. After determining the best combination (40 µM thymoquinone and 0.6 µM topotecan), cell proteins were extracted after treatment, and the expression levels of B-cell lymphoma 2 and of its associated X protein, proteins p53 and p21, and caspase-9, caspase-3, and caspase-8 were studied by Western blot. In addition, cell cycle analysis and annexin/propidium iodide staining were performed. Both drugs induced apoptosis through a p53-independent mechanism, whereas the expression of p21 was only seen in thymoquinone treatment. Cell cycle arrest in the S phase was detected with each compound separately, while combined treatment only increased the production of fragmented DNA. Both compounds induced apoptosis through the extrinsic pathway after 24 hours; however, after 48 hours, the intrinsic pathway was activated by topotecan treatment only. In conclusion, thymoquinone increased the effectiveness of the chemotherapeutic reagent topotecan by inhibiting proliferation and lowering toxicity through p53- and Bax/Bcl2-independent mechanisms.

Topotecan for Relapsed Small-cell Lung Cancer: Systematic Review and Meta-Analysis of 1347 Patients.

Topotecan is the most reliable chemotherapy regimen for relapsed small-cell lung carcinoma (SCLC). The efficacy and adverse effects of topotecan as reported by previous studies varied greatly. The inclusion criterion was a prospective study that was able to provide data for 6-month over-all survival (OS) rate, 1-year OS rate, objective responses, and/or adverse effects of single agent topotecan as a second line chemotherapy for SCLC, written in English language as a full article. Any topotecan regimen were allowed. Binary data were meta-analyzed with the random-model generic inverse variance method. We included 14 articles consisted of 1347 patients. Pooled values were estimated as follows. <Refractory relapse> Six-month OS rate: 37% (95% CI: 28–46%). One-year OS rate: 9% (95% CI: 5–13%). Response rate: 5% (95% CI: 1–8%). <Sensitive relapse> Six-month OS rate: 57% (95% CI: 50–64%). One-year OS rate: 27% (95% CI: 22–32%). Response rate: 17% (95% CI: 11–23%). <Adverse effect> Grade III/IV neutropenia 69% (95% CI: 58–80%). Grade III/IV thrombopenia 41% (95% CI: 34–48%). Grade III/IV anemia 24% (95% CI: 17–30%). Non-hematorogical events were rare. Chemotherapy-related death 2% (95% CI: 1–3%). In conclusion, Topotecan provided a possibly promising outcome for sensitive-relapse SCLC and poor outcome for refractory relapse SCLC. Adverse events were mainly hematological.

Sensitivity of neuroblastoma to the novel kinase inhibitor cabozantinib is mediated by ERK inhibition.

Children with high-risk neuroblastoma have poor survival rates, and novel therapies are needed. We hypothesized that cabozantinib would be effective against neuroblastoma tumor cells and tumors in preclinical models via inhibition of receptor tyrosine kinase signaling pathways. We determined neuroblastoma cell viability after treatment with cabozantinib alone and in combination with 13-cis-retinoic acid, topotecan, and temozolomide using MTT assays. Inhibition of RET and intracellular signaling was measured by Western blot analysis of treated and untreated cells. To investigate the efficacy of cabozantinib against neuroblastoma tumors in vivo, neuroblastoma cells were injected orthotopically into immunocompromised mice, and mice were treated with oral cabozantinib. Tumors were evaluated for growth by determination of in vivo luminescence and final tumor weights. All neuroblastoma cell lines were sensitive to cabozantinib, and IC50 values ranged from 1.6 to 16.2 μM. Cabozantinib treatment was synergistic with 13-cis-retinoic acid and chemotherapy agents topotecan and temozolomide. Cabozantinib treatment inhibited RET phosphorylation in all cell lines and ERK phosphorylation in more sensitive neuroblastoma cell lines. In mice with neuroblastoma xenograft tumors, cabozantinib treatment significantly reduced tumor growth. Treatment of neuroblastoma tumor cells with cabozantinib inhibits RET and ERK phosphorylation and is effective against neuroblastoma tumor cell lines alone and in combination with 13-cis-retinoic acid, topotecan, and temozolomide. Cabozantinib treatment is also effective in reducing tumor growth in vivo. Cabozantinib therefore represents a novel therapeutic agent for neuroblastoma, and further preclinical and clinical studies are warranted.

Abstract 5333: TH-302 potentiates the antitumor activity of topotecan in neuroblastoma and rhabdomyosarcoma preclinical models

Abstract Background: There is strong evidence that tumor cells in the hypoxic regions cause drug resistance and tumor relapse. Hypoxia targeting agents may make cytotoxic therapy more effective. In this study, we investigated the efficacy of TH-302, a hypoxia-activated prodrug of bromo-isophosphoramide mustard, both alone and in combination with the cytotoxic agent topotecan, in preclinical models of neuroblastoma (NBL) and rhabdomyosarcoma (RMS). Materials and methods: A panel of four NBL and three RMS cell lines was tested in vitro under normoxic or hypoxic conditions to assess the effect of TH-302 on cell proliferation, both as a single agent and in combination with topotecan. In vivo activity was also evaluated in different xenograft models. TH-302 (50 mg/kg) and topotecan (1 mg/kg) were administered daily for 2 consecutive weeks, as single agents or in combination. Animal survival was studied with an NBL (SK-N-BE(2)) metastatic tumor model. Cleaved caspase-3 and the hypoxia staining agent pimonidazole were used as apoptotic and hypoxic markers in vivo. Results: Under normoxic conditions, all tested lines showed an anti-proliferative response to TH-302 with the IC50s ranging from 4.6 to 52μM. When exposed to hypoxia (1% O2) overnight, a dramatic decrease of IC50 was observed, ranging from 0.07 to 2.4μM. Adding topotecan to the TH-302 treatment significantly increased cytotoxicity in all tested tumor cell lines. In both NBL models (SK-N-BE(2) and CHLA-20), single-agent TH-302 treatment delayed tumor growth. Complete tumor regression was observed in the combined treatment group after a two-week treatment period, but all the regressed tumors relapsed after treatment ended. In the CHLA-20 xenograft model, single-agent topotecan also induced complete tumor regression. However, median time to relapse in the combination group was significantly longer than in the topotecan group (43 days vs. 16 days; p&amp;lt;0.01). In the combination group, all relapsed tumors remained responsive to the drug treatment when re-challenged with the original treatment regimen. In the neuroblastoma metastatic model, all three regimens prolonged animal survival, but the combined treatment group showed better survival than single-agent treatments (p&amp;lt;0.01). In a RMS xenograft model, combined treatment was also more effective than single-agent TH-302 or topotecan. Double immunostaining of apoptotic and hypoxic markers showed that TH-302 mostly targeted tumor cells residing at hypoxic regions while topotecan was more effective on tumor cells outside the hypoxic zones. The combined treatment induced tumor cell apoptosis in both normoxic and hypoxic regions. Conclusions: TH-302 showed antitumor effects in neuroblastoma and rhabdomyosarcoma xenograft models. Compared to single-agent TH-302 or topotecan, the combination treatment significantly improved tumor response, delayed tumor relapse, and enhanced animal survival. Citation Format: Libo Zhang, Bing Wu, Paula Marrano, Paul Thorner, Sylvain Baruchel. TH-302 potentiates the antitumor activity of topotecan in neuroblastoma and rhabdomyosarcoma preclinical models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5333. doi:10.1158/1538-7445.AM2015-5333

Insights into accelerated liposomal release of topotecan in plasma monitored by a non-invasive fluorescence spectroscopic method

A non-invasive fluorescence method was developed to monitor liposomal release kinetics of the anticancer agent topotecan (TPT) in physiological fluids and subsequently used to explore the cause of accelerated release in plasma. Analyses of fluorescence excitation spectra confirmed that unencapsulated TPT exhibits a red shift in its spectrum as pH is increased. This property was used to monitor TPT release from actively loaded liposomal formulations having a low intravesicular pH. Mathematical release models were developed to extract reliable rate constants for TPT release in aqueous solutions monitored by fluorescence and release kinetics obtained by HPLC. Using the fluorescence method, accelerated TPT release was observed in plasma as previously reported in the literature. Simulations to estimate the intravesicular pH were conducted to demonstrate that accelerated release correlated with alterations in the low intravesicular pH. This was attributed to the presence of ammonia in plasma samples rather than proteins and other plasma components generally believed to alter release kinetics in physiological samples. These findings shed light on the critical role that ammonia may play in contributing to the preclinical/clinical variability and performance seen with actively-loaded liposomal formulations of TPT and other weakly-basic anticancer agents.

Abstract B36: Inhibition of Exportin 1 (XPO1) potently suppresses growth of human neuroblastoma cell lines

Abstract Background: Most children with high-risk neuroblastoma are not cured, and new therapies are needed. Exportin 1 (XPO1; Chromosome Region Maintenance 1 Protein Homolog, CRM1) is the sole nuclear exporter of TP53, FOXO1, RB1, CDKN1A, and other critical tumor suppressor proteins (TSPs). This suggests that inhibition of XPO1 is a rational anti-cancer strategy. KPT-330 is a Selective Inhibitor of Nuclear Export (SINE) that irreversibly binds XPO1 and is currently in Phase 1 clinical trials. SINE inhibits XPO1-mediated nuclear export across multiple tumor types leading to activation of TSPs and cancer cell death. Methods: A panel of 14 genomically-characterized neuroblastoma cell lines was treated with KPT-330 across a 5-log range. Growth inhibition was measured using CellTiter-Glo (Promega) viability assays and a real time growth monitoring system (xCELLigence, Roche). Drug combinations were tested in vitro, and the combination index was calculated using CalcySyn (Biosoft). NOD/SCID mice with neuroblastoma xenografts (both flank and tail-vein injected) were treated orally with KPT-330 (15 mg/kg thrice weekly or 20-25 mg/kg twice weekly), and tumor size was monitored by direct measurement and luciferase luminescence. Results: KPT-330 showed cytotoxicity against 14 neuroblastoma cell lines with a median IC50 of 236 nM (range 51-568). Sensitivity to KPT-330 was independent of MYCN amplification status. Combination of KPT-330 with commonly used cytotoxic agents (irinotecan, topotecan, cisplatin, and doxorubicin) showed additive effects. Flank xenografts using the neuroblastoma cell line SH-SY5Y showed significantly decreased tumor growth after oral administration of KPT-330. After allowing the tumors to grow following 28 days of drug withdrawal, re-exposure to the drug again showed growth suppression. In a model of metastatic neuroblastoma after tail-vein injection of luciferase-transfected neuroblastoma cell lines (SH-SY5Y, IMR-5, and Be2c), KPT-330 showed significant tumor growth inhibition (p&amp;lt;0.001). KPT-330 treatment resulted in decreased Myc family protein levels as well as activation of apoptotic pathways. Major toxicity was weight loss which was overcome with nutritional supplementation. Conclusions: Neuroblastomas show sensitivity to XPO1 inhibition both in vitro and in vivo. Ongoing work is focused on discovering the cellular and genomic factors responsible for increased sensitivity to nuclear export inhibition and potential synergistic combinations. With the expected completion of the first in human phase I trials this year, treatment with KPT-330 has the potential to be rapidly translated into a clinical trial for children with neuroblastoma. Citation Format: Edward F. Attiyeh, Aaron McKeon-Fish, Rebecca Trillo, Yosef Landesman, William Senapedis, Dilara McCauley, Trinayan Kashyap, Sharon Shacham, Michael Kauffman, John M. Maris. Inhibition of Exportin 1 (XPO1) potently suppresses growth of human neuroblastoma cell lines. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr B36.