Ovarian Cancer Therapeutics Research Articles

Folate receptor-targeted therapeutics for ovarian cancer

Folate receptor-targeted therapeutics for ovarian cancer

Potential targets for ovarian clear cell carcinoma: a review of updates and future perspectives.

Advances in surgical and medical treatments for ovarian cancer have improved prognoses. Platinum drugs in particular are pivotal for the medical treatment of ovarian cancer. However, previous studies have revealed that some histological subtypes, such as clear cell carcinoma, are resistant to medical treatment, including that with platinum drugs. Consequently, the clinical prognosis of advanced clear cell carcinoma is remarkably inferior, primarily because of its chemoresistant behavior. The prevalence of clear cell carcinoma is approximately 5 % in the West, but in Japan, its prevalence is particularly high, at approximately 25 %. Current medical treatments for advanced clear cell carcinoma are difficult to administer, and they have poor efficacy, warranting the development of novel target-based therapies. In this review, we describe medical treatments for clear cell carcinoma and discuss future prospects for therapy. In particular, we focus on the mechanism of platinum resistance in clear cell carcinoma, including the role of annexin A4, one of the most investigated factors of platinum resistance, as well as the mutant genes and overexpressed proteins such as VEGF, PI3K/AKT/mTOR signaling pathway, ARID1A, hepatocyte nuclear factor-1β, ZNF217. We also review targeted molecular therapeutics for epithelial ovarian cancer and discuss their role in clear cell carcinoma treatment. We review the drugs targeting angiogenesis (bevacizumab, sorafenib, and pazopanib), growth factors (gefitinib, erlotinib, lapatinib, trastuzumab, and AMG479), and signaling pathways (temsirolimus, dasatinib, and imatinib), and other drugs (oregovomab, volociximab, and iniparib). This current review summarizes and discusses the clinical significance of these factors in ovarian clear cell carcinoma as well as their potential mechanisms of action. It may provide new integrative understanding for future studies on their exact role in ovarian clear cell carcinoma.

MiR-30d Blocked Transforming Growth Factor β1-Induced Epithelial-Mesenchymal Transition by Targeting Snail in Ovarian Cancer Cells.

MicroRNAs (miRs) are essential regulators of gene expression by suppressing translation or causing degradation of target mRNA. Growing evidence sheds light on the crucial roles of miR dysregulation in cancer development and progression. In this study, we focused on the role of miR-30d in transforming growth factor β1 (TGF-β1)-initiated epithelial-mesenchymal transition (EMT) in ovarian cancer cells. Transforming growth factor β1 (10 ng/mL) was used to initiate EMT in SKOV3 and 3AO cells. The expression of miR-30 family members was determined by quantitative real-time polymerase chain reaction. Messenger RNA and protein levels of E-cadherin, N-cadherin, vimentin, and Snail were detected by quantitative real-time polymerase chain reaction and Western blot, respectively. Cell migration and invasion capacities were evaluated by Transwell chamber assay. Luciferase activity assay was performed to verify the direct inhibition of Snail by miR-30d. MiR-30b, MiR-30c, and MiR-30d were down-regulated during TGF-β1-induced EMT in SKOV3 and 3AO ovarian cancer cells. Restoration of miR-30d by miR-30d mimic reversed TGF-β1-induced EMT phenotypes including the morphological changes, expression pattern of molecular markers (E-cadherin, N-cadherin), and migratory and invasive capabilities in ovarian cancer cells. Furthermore, Snail was identified as the direct target of miR-30d. Our results revealed that miR-30d functioned as a suppressor of ovarian cancer progression by decreasing Snail expression and thus blocking TGF-β1-induced EMT process, suggesting the potentiality of miR-30d analogs to be used as therapeutics for ovarian cancer.

Targeting Human Poly(ADP-Ribose) Polymerase-1 with Natural Medicines and Its Potential Applications in Ovarian Cancer Therapeutics.

Targeting poly(ADP-ribose) polymerase-1 (PARP-1) has been established as an efficient therapeutics for advanced ovarian cancer. In this study, we describe an integrated procedure that combines virtual computer screening and an experimental enzyme assay to discover novel potent PARP-1 inhibitors from more than 130000 commercially available natural products. The protocol employed a stepwise strategy to fast exclude typical PARP-1 non-binders and then performing rigorous prediction to identify promising candidates with high potency against PARP-1. Consequently, eight natural products were hit and tested to determine their inhibitory activities against the PARP-1 catalytic domain. From these, four compounds, i.e., puerarin, phloretin, chlorogenic acid, and biochanin A, were found to have high or moderate potencies with inhibitory IC50 values of 6, 470, 25, and 86 nM, respectively. The values are comparable to that (IC50 = 1.94 nM) of the FDA-approved agent olaparib. Structural and energetic analyses of the modeled structures of the PARP-1 catalytic domain complexed with the newly identified inhibitors revealed a common binding mode in the complexes: the active site of PARP-1 is composed of a thin polar helix and a flat non-polar pocket; the inhibitors can form a number of hydrogen bonds and electrostatic forces with the helix, while tightly packing against the pocket to define chemical interactions.

Editorial introductions

Current Opinion in Oncology was launched in 1989. It is one of a successful series of review journals whose unique format is designed to provide a systematic and critical assessment of the literature as presented in the many primary journals. The field of psychiatry is divided into 15 sections that are reviewed once a year. Each section is assigned a Section Editor, a leading authority in the area, who identifies the most important topics at that time. Here we are pleased to introduce the Journal's Section Editors for this issue. SECTION EDITORS Bertrand Coiffier Bertrand Coiffier is Professor of Hematology at the Department of Hematology, Hospices Civils de Lyon and the University Claude Bernard, Lyon, France. After having specialized in hematology, Professor Coiffier has held a number of positions at the Hematology Department of the Edouard-Herriot Hospital, Lyon and then the Centre Hospitalier Lyon-Sud, Pierre-Benite. He is a member of several international societies, including the American Society of Hematology, the American Society of Clinical Oncology, the International Society for Medical Oncology, the European Haematology Association, and the European Society for Medical Oncology. Professor Coiffier is a founding member of the Group d’Etude des Lymphomes de l’Adulte (GELA) and has been President of the GELA since 2006. He has published over 300 papers in peer-reviewed journals. Anne-Sophie Michallet Anne-Sophie Michallet is a Doctor of Hematology at the Department of Hematology, Hospices Civils de Lyon and the University Claude Bernard, Lyon, France. After having specialized in hematology, she focused on lymphoproliferative disorders. She is a member of the French Group of Chronic Lymphocyic Leukemia (GFCLL and GOELAMS), the Group d’Etude des Lymphomes de l’Adulte (GELA); and a member of the société française de greffe de moelle (SFGM-TC). Martin GoreMartin GoreProfessor Martin Gore is the Medical Director of the Royal Marsden Hospital and Professor of Cancer Medicine at the Institute of Cancer Research. He qualified in medicine at St Bartholomew's Hospital, London and spent time in general practice before training in general internal medicine at University College Hospital, London. He joined the Ludwig Institute of Cancer Research in 1981 where he worked as a clinical scientist and was awarded a PhD for his work on breast cancer. Professor Gore serves on The Commission on Human Medicines, is the founder Co-Patron of the Rarer Cancers Foundation and was a founder Senior Investigator of the UK's National Institute for Health Research. He previously chaired the Department of Health's Gene Therapy Advisory Committee, is a past President of the UK Melanoma Study Group and a past Chair of the National Cancer Research Institute's Melanoma Clinical Studies Group. He has served on the Health and Safety Executive's Scientific Advisory Committee on Genetically Modified Organisms, the Council of the International Gynaecologic Cancer Society, the Programme Committee of the Annual Meeting of the American Society of Clinical Oncology and he was co-founder of the European International Kidney Cancer Symposium. He is a Medical Advisor to the US-based patient advocacy group the Kidney Cancer Association who awarded him the Eugene P. Schonfeld Memorial Lecturership in 2014, the first non-US recipient. He is the editor of the Gynaecological Cancer Section of Current Opinion in Oncology and has served on the editorial board of several journals including the Journal of Clinical Oncology. He has published over 500 articles and edited ten textbooks. Professor Gore's main research interests are the development of novel therapeutics in ovarian cancer, melanoma and renal cell carcinoma, particularly in the area of immunotherapy, gene therapy and targeted agents.

Metformin and epithelial ovarian cancer therapeutics.

Ovarian cancer is one of the most common lethal gynecological malignancies world-wide. Despite an initial 70-80% response rate, most patients relapse within 1-2 years and develop chemo-resistance. Hence, the identification of novel drugs or the repositioning of known drugs to re-sensitize ovarian cancer cells to existing chemotherapy regimens is needed. Here, we evaluated the effect of metformin (an anti-diabetic drug) on ovarian cancer cells, based on its putative effect on other solid tumors. Primary cultures of epithelial ovarian cancer cells established from ascitic fluids of untreated ovarian cancer patients and the SKOV-3 ovarian cancer-derived cell line were used. The respective cells were treated with metformin, carboplatin and paclitaxel alone and its various combinations and their effects, including the ability to induce apoptosis, were examined. Concomitantly, the cells were assessed for the expression of several apoptosis-related mRNAs and proteins using quantitative real time PCR, flowcytometry and Western blotting. We found that metformin induced apoptosis in the ovarian cancer cells tested, and provoked a cell cycle arrest in the G0/G1 and S-phase. Metformin induced apoptosis by down-regulating Bcl-2 and Bcl-xL expression, and up-regulating Bax and Cytochrome c expression. We also found that the apoptosis induction by metformin could be enhanced by a combinatorial use of carboplatin and/or paclitaxel. Our data indicate that metformin can induce apoptosis in both primary ovarian cancer cells and in SKOV-3 cells. When metformin was combined with carboplatin or paclitaxel, an increased apoptotic activity was observed, implicating a chemo-adjuvant potential.

Highly effective antiangiogenesis via magnetic mesoporous silica-based siRNA vehicle targeting the VEGF gene for orthotopic ovarian cancer therapy.

Therapeutic antiangiogenesis strategies have demonstrated significant antitumor efficacy in ovarian cancer. Recently, RNA interference (RNAi) has come to be regarded as a promising technology for treatment of disease, especially cancer. In this study, vascular endothelial growth factor (VEGF)-small interfering RNA (siRNA) was encapsulated into a magnetic mesoporous silica nanoparticle (M-MSN)-based, polyethylenimine (PEI)-capped, polyethylene glycol (PEG)-grafted, fusogenic peptide (KALA)-functionalized siRNA delivery system, termed M-MSN_VEGF siRNA@PEI-PEG-KALA, which showed significant effectiveness with regard to VEGF gene silencing in vitro and in vivo. The prepared siRNA delivery system readily exhibited cellular internalization and ease of endosomal escape, resulting in excellent RNAi efficacy without associated cytotoxicity in SKOV3 cells. In in vivo experiments, notable retardation of tumor growth was observed in orthotopic ovarian tumor-bearing mice, which was attributed to significant inhibition of angiogenesis by systemic administration of this nanocarrier. No obvious toxic drug responses were detected in major organs. Further, the magnetic core of M-MSN_VEGF siRNA@PEI-PEG-KALA proved capable of probing the site and size of the ovarian cancer in mice on magnetic resonance imaging. Collectively, the results demonstrate that an M-MSN-based delivery system has potential to serve as a carrier of siRNA therapeutics in ovarian cancer.

TXNDC17 promotes paclitaxel resistance via inducing autophagy in ovarian cancer

Paclitaxel is recommended as a first-line chemotherapeutic agent against ovarian cancer, but drug resistance becomes a major limitation of its success clinically. The key molecule or mechanism associated with paclitaxel resistance in ovarian cancer still remains unclear. Here, we showed that TXNDC17 screened from 356 differentially expressed proteins by LC-MS/MS label-free quantitative proteomics was more highly expressed in paclitaxel-resistant ovarian cancer cells and tissues, and the high expression of TXNDC17 was associated with poorer prognostic factors and exhibited shortened survival in 157 ovarian cancer patients. Moreover, paclitaxel exposure induced upregulation of TXNDC17 and BECN1 expression, increase of autophagosome formation, and autophagic flux that conferred cytoprotection for ovarian cancer cells from paclitaxel. TXNDC17 inhibition by siRNA or enforced overexpression by a pcDNA3.1(+)-TXNDC17 plasmid correspondingly decreased or increased the autophagy response and paclitaxel resistance. Additionally, the downregulation of BECN1 by siRNA attenuated the activation of autophagy and cytoprotection from paclitaxel induced by TXNDC17 overexpression in ovarian cancer cells. Thus, our findings suggest that TXNDC17, through participation of BECN1, induces autophagy and consequently results in paclitaxel resistance in ovarian cancer. TXNDC17 may be a potential predictor or target in ovarian cancer therapeutics.

Abstract LB-13: Hyaluronic acid-based CD44 targeted nanoparticle delivery of combination MDR1 siRNA/paclitaxel to overcome drug resistance in ovarian cancer

Abstract A major obstacle in the success of chemotherapy in ovarian cancer is the emergence of multidrug resistance (MDR). Overexpression of the MDR1 gene and corresponding P-glycoprotein (Pgp) efflux pumps is one of the best characterized MDR mechanisms. Although MDR1 siRNA based strategies are emerging as highly promising approaches to reverse MDR, the systemic delivery still remains a great challenge. In the present study, CD44 targeting hyaluronic acid (HA) based self-assembling nanoparticle systems were designed with MDR1 siRNA to evaluate its delivery efficiency and combination anticancer therapeutic efficacy with paclitaxel in vitro and in vivo. The results showed that HA-poly(ethyleneimine)/HA-poly(ethylene glycol) (HA-PEI/HA-PEG) nanoparticle successfully delivered MDR1 siRNA into MDR ovarian cancer cell lines, SKOV-3TR and OVCAR8TR. Western blot analysis, calcein AM retention assay, and MTT assay further demonstrated that HA-PEI/HA-PEG nanoparticles loaded with MDR1 siRNA efficiently down-regulated the expression of MDR1 and inhibited the functional activity of Pgp, and subsequently increase cell sensitivity to paclitaxel. HA-PEI/HA-PEG/MDR1 siRNA nanoparticle therapy followed by paclitaxel treatment produced a significant inhibitory effect on the drug-resistant tumor growth as compared with control groups in MDR ovarian cancer xenograft mouse models. These results indicated CD44-targeted HA-PEI/HA-PEG nanoparticle platform is an effective siRNA delivery system for potential siRNA-based anticancer therapeutics in ovarian cancer. Citation Format: Xiaoqian Yang, Arun lyer, Francis hornicek, Mansoor Amiji, Zhenfeng Duan. Hyaluronic acid-based CD44 targeted nanoparticle delivery of combination MDR1 siRNA/paclitaxel to overcome drug resistance in ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-13. doi:10.1158/1538-7445.AM2014-LB-13

Abstract 1543: Systematic functional assessment of driver mutations for individual ovarian cancers

Abstract Although epithelial ovarian cancer is an intractable cancer and the overall five-year survival rate is approximately 40 to 50%, clinically effective molecular-targeted therapeutics to treat ovarian cancer has not been available thus far. Recent progress of massively parallel sequencing technologies has been enabling us to identify substantial number of genomic aberrations including somatic mutations, indels and translocations in variable cancer types exemplified by epithelial ovarian cancer and non-small cell lung cancer. However, it is still very difficult to identify a ‘driver’ gene, which is functionally relevant for cancer growth, from hundreds of mutations that are usually found from a single cancer sample of an individual patient. An in vitro cultured cell line, which is originated from a clinical sample, can be utilized to assess functions of the genes that are identified to have somatic aberrations in the paired clinical tumor. We have been establishing multiple ovarian cancer cell lines derived from clinical samples and analyzing pairs of the cell lines and clinical samples with exon-capture sequencing. The genes, which are somatically mutated and shared in a pair of cell line and clinical sample, are targeted with RNA interference in the cell line, and subsequently the effect on cell proliferation and survival is examined. As such, a driver mutation for an individual cancer can be identified by a set of experimental perturbations. Identified driver mutations would be then validated with publicly available databases and with clinical archive samples. As a proof-of-principle, somatic mutations of PIK3CA and KRAS were detected as drivers to promote cell proliferation of a carcinosarcoma. We believe this approach can increase an opportunity to identify molecular targets in ovarian cancer therapeutics. Citation Format: Seiichi Mori, Takako Yokomizo, Takeshi Fujiwara, Osamu Goto, Haruko Iwase, Kazuyoshi Kato, Tokuichi Kawaguchi, Tetsuo Noda. Systematic functional assessment of driver mutations for individual ovarian cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1543. doi:10.1158/1538-7445.AM2014-1543

Abstract 3784: Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells

Abstract The field of ovarian cancer therapeutics has made a substantial leap forward with the discovery that ovarian cancers with a BRCA mutation, can be exploited with “synthetic lethality.” In patients who have a BRCA germline mutation, this phenomenon is achieved by blocking PARP (poly-ADP-ribosyl-polymerase), which has a role in compensating for the missing or low levels of the BRCA protein. PARP activity is essential for DNA repair in patients with BRCA mutations as their cells attempt to maintain a normal genotype; however, PARP is not enough to compensate for the BRCA mutation, and these patients eventually develop ovarian cancer. Out of the five major mechanisms of DNA repair, PARP compensates for missing BRCA1 and/or BRCA2 proteins required for homologous repair, base excision repair and nuclear excision repair. If PARP is inhibited in BRCA-mutant patients, DNA mutations accumulate and are not repaired because PARP is not available to ribosylate the areas of cut or damaged DNA, which would signal that repair is needed. The BRCA-mutant genotype, in which there is a germline-inactivating mutation in one allele and loss of heterozygosity or somatic mutation in the other allele, only occurs in 5-10% of the cancer population. We wondered, “is there a way to apply the concept of “synthetic lethality” to the 90-95% of the ovarian cancer patients who do not have BRCA mutations?” We tried to answer that query by testing different gene-expression, modulating agents. We found that low-concentration hydroxyurea can sometimes potentiate the cytotoxicity of alkylating agents, suggesting a mechanism that involves DNA repair. We found synergistic apoptosis in 4 (ovcar-8, TOV21G, PA-1 and UACC1598) ovarian cancer cell lines treated with hydroxyurea and the PARP inhibitor, Olaparib, and in cells treated with curcumin with Olaparib. When the Ovcar-8 cancer cell line was treated with a single agent, the percentage of apoptosis was ≤ 17%, and when treated with synergistic agents together, relative apoptosis increased to ≤ 45%. In these cell lines, both curcumin and hydroxyurea significantly decreased BRCA1 and BRCA2 mRNA and protein levels. We also found that curcumin triggered BRCA1,2 promoter methylation, which led to cessation of gene expression. Whereas, hydroxyurea, not only caused BRCA1,2 promoter methylation, but also elicited an increase in miRNA hsa-miR-17-5p and hsa-miR-182-5p, causing BRCA1 mRNA degradation. This finding suggests greater than 5-10% of wild-type, BRCA ovarian cancer patients may benefit from PARP inhibitors and DNA cross-linking agents, such as mitomycin C. if they can be induced into “BRCAness,” Citation Format: Yuehua Mao, Richard D Dinnen, Robert L. Fine. Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3784. doi:10.1158/1538-7445.AM2014-3784

Epithelial–mesenchymal transition-associated miRNAs in ovarian carcinoma, with highlight on the miR-200 family: Prognostic value and prospective role in ovarian cancer therapeutics

MicroRNAs (miRNAs) are a family of short ribonucleic acids found to play a pivotal role in cancer pathogenesis. MiRNAs are crucial in cellular differentiation, growth, stress response, cell death and other fundamental cellular processes, and their involvement in ovarian cancer has been recently shown. They can repress the expression of important cancer-related genes and they can also function both as oncogenes and tumour suppressor genes. During epithelial–mesenchymal transition (EMT), epithelial cells lose their cell polarity and cell–cell adhesion and gain migratory and invasive properties. In the ovarian surface epithelium, EMT is considered the key regulator of the post-ovulatory repair process and it can be triggered by a range of environmental stimuli. The aberrant expression of the miR-200 family (miR-200a, miR-200b, miR-200c, miR-141 and miR-429) in ovarian carcinoma and its involvement in ovarian cancer initiation and progression has been well-demonstrated. The miR-200 family members seem to be strongly associated with a pathologic EMT and to have a metastasis suppressive role. MiRNA signatures can accurately distinguish ovarian cancer from the normal ovary and can be used as diagnostic tools to predict the clinical response to chemotherapy. Recent evidence suggests a growing list of new miRNAs (miR-187, miR-34a, miR-506, miRNA-138, miR-30c, miR-30d, miR-30e-3p, miR-370 and miR-106a, among others) that are also implicated in ovarian carcinoma-associated EMT, either enhancing or suppressing it. MiRNA-based gene therapy provides a prospective anti-tumour approach for integrated cancer therapy. The aim of nanotechnology-based delivery approach for miRNA therapy is to overcome challenges in miRNA delivery and to effectively encourage the reprogramming of miRNA networks in cancer cells, which may lead to a clinically translatable miRNA-based therapy to benefit ovarian cancer patients.

Development of nanoscale approaches for ovarian cancer therapeutics and diagnostics.

Ovarian cancer is the deadliest of all gynecological cancers and the fifth leading cause of death due to cancer in women. This is largely due to late-stage diagnosis, poor prognosis related to advanced-stage disease, and the high recurrence rate associated with development of chemoresistance. Survival statistics have not improved significantly over the last three decades, highlighting the fact that improved therapeutic strategies and early detection require substantial improvements. Here, we review and highlight nanotechnology-based approaches that seek to address this need. The success of Doxil, a PEGylated liposomal nanoencapsulation of doxorubicin, which was approved by the FDA for use on recurrent ovarian cancer, has paved the way for the current wave of nanoparticle formulations in drug discovery and clinical trials. We discuss and summarize new nanoformulations that are currently moving into clinical trials and highlight novel nanotherapeutic strategies that have shown promising results in preclinical in vivo studies. Further, the potential for nanomaterials in diagnostic imaging techniques and the ability to leverage nanotechnology for early detection of ovarian cancer are also discussed.

Urokinase Plasminogen Activator System–Targeted Delivery of Nanobins as a Novel Ovarian Cancer Therapy

The urokinase system is overexpressed in epithelial ovarian cancer cells and is expressed at low levels in normal cells. To develop a platform for intracellular and targeted delivery of therapeutics in ovarian cancer, we conjugated urokinase plasminogen activator (uPA) antibodies to liposomal nanobins. The arsenic trioxide-loaded nanobins had favorable physicochemical properties and the ability to bind specifically to uPA. Confocal microscopy showed that the uPA-targeted nanobins were internalized by ovarian cancer cells, whereas both inductively coupled plasma optical mass spectrometry (ICP-MS) and fluorescence-activated cell sorting (FACS) analyses confirmed more than four-fold higher uptake of targeted nanobins when compared with untargeted nanobins. In a coculture assay, the targeted nanobins showed efficient uptake in ovarian cancer cells but not in the normal primary omental mesothelial cells. Moreover, this uptake could be blocked by either downregulating uPA receptor expression in the ovarian cancer cells using short-hairpin RNA (shRNA) or by competition with free uPA or uPA antibody. In proof-of-concept experiments, mice bearing orthotopic ovarian tumors showed a greater reduction in tumor burden when treated with targeted nanobins than with untargeted nanobins (47% vs. 27%; P < 0.001). The targeted nanobins more effectively inhibited tumor cell growth both in vitro and in vivo compared with untargeted nanobins, inducing caspase-mediated apoptosis and impairing stem cell marker, aldehyde dehydrogenase-1A1 (ALDH1A1), expression. Ex vivo fluorescence imaging of tumors and organs corroborated these results, showing preferential localization of the targeted nanobins to the tumor. These findings suggest that uPA-targeted nanobins capable of specifically and efficiently delivering payloads to cancer cells could serve as the foundation for a new targeted cancer therapy using protease receptors.

Ovarian cancer clinical trial endpoints: Society of Gynecologic Oncology white paper

ObjectiveTo explore the value of multiple clinical endpoints in the unique setting of ovarian cancer. MethodsA clinical trial workgroup was established by the Society of Gynecologic Oncology to develop a consensus statement via multiple conference calls, meetings and white paper drafts. ResultsClinical trial endpoints have profound effects on late phase clinical trial design, result interpretation, drug development, and regulatory approval of therapeutics. Selection of the optimal clinical trial endpoint is particularly provocative in ovarian cancer where long overall survival (OS) is observed. The lack of new regulatory approvals and the lack of harmony between regulatory bodies globally for ovarian cancer therapeutics are of concern. The advantages and disadvantages of the numerous endpoints available are herein discussed within the unique context of ovarian cancer where both crossover and post-progression therapies potentially uncouple surrogacy between progression-free survival (PFS) and OS, the two most widely supported and utilized endpoints. The roles of patient reported outcomes (PRO) and health related quality of life (HRQoL) are discussed, but even these widely supported parameters are affected by the unique characteristics of ovarian cancer where a significant percentage of patients may be asymptomatic. Original data regarding the endpoint preferences of ovarian cancer advocates is presented. ConclusionsEndpoint selection in ovarian cancer clinical trials should reflect the impact on disease burden and unique characteristics of the treatment cohort while reflecting true patient benefit. Both OS and PFS have led to regulatory approvals and are clinically important. OS remains the most objective and accepted endpoint because it is least vulnerable to bias; however, the feasibility of OS in ovarian cancer is compromised by the requirement for large trial size, prolonged time-line for final analysis, and potential for unintended loss of treatment effect from active post-progression therapies. A large magnitude of effect in PFS improvement should establish benefit, and further communication with regulatory authorities to clarify acceptable endpoints should be undertaken.

Abstract B30: Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells

Abstract The field of ovarian cancer therapeutics has recently made a substantial leap forward with the discovery that ovarian cancers with a BRCA mutation can be exploited with “synthetic lethality”. This phenomenon takes advantage of the role of PARP (poly-ADP ribosyl-polymerase), which compensates for the missing or low BRCA protein in these patients with a BRCA germline mutation. PARP activity is essential for these patients to maintain a normal genotype, albeit not completely normal since they developed ovarian cancer mainly because of their BRCA mutation. Thus, in the 5 major mechanisms of DNA repair PARP compensates for missing BRCA1 or BRCA2 in homologous repair, base excision repair and nuclear excision repair. In these patients, if PARP is inhibited, then massive DNA mutations accumulate and are not repaired because PARP ribosylates the areas of cut or damaged DNA to mark the areas needed for repair to occur. The concept of inhibiting PARP in these BRCA mutated tumors is an elegant idea. The caveat here is the BRCA mutant genotype where there is a germline, inactivating mutation in one allele and LOH or somatic mutation of the other allele only occurs in 5-10% of the cancer population. Thus, could there be a way to apply this concept of synthetic lethality to the other 90-95% of the cancer population without BRCA mutations in ovarian cancer patients? We tried to answer that question by testing different agent which could modulate gene expression, and found that curcumin and hydroxyurea, at low concentrations, could sometimes increase the cytotoxicity of alkylating agents which suggests a mechanism involving DNA repair. Four (ovcar-8, TOV21G, PA-1 and UACC1598) out of nine pretreated ovarian cancer cell lines with hydroxyurea or curcumin has synergistically increase apoptosis induced by PARP inhibitor - Olaparid. In the tested Ovcar-8 cancer cell line, Curcumin (5 μM) alone induced 10% apoptosis, hydroxyurea (100 μM) alone induced 17% apoptosis, Olaparid (2.5 μM) alone induced 7% cell death. Curcumin and Olaparid together induce 45% cell death (165% increase), hydroxurea and Olaparid together induced 40% (67% increase). Both curcumin and hydroxyurea decrease BRCA1 and BRCA2 mRNA and protein level in the tested sensitive cell lines. Further study, we found that curcumin prompt epigenetically BRCA1,2 promoter methylation and lead to gene expression shutting down whereas hydroxyurea induced miRNA hsa-miR-17-5p and hsa-miR-182-5p and lead BRCA1 mRNA degradation. This finding suggest If wt BRCA patients with ovarian cancer could be induced into BRCAness, then perhaps a larger percentage than 5-10% could benefit from PARP inhibitors and DNA crosslinkers like mitomycin C. Citation Format: Yuehua Mao, Richard D Dinnen, Robert L Fine. Induction of BRCAness phenotype by curcumin and hydroxyurea in WT BRCA ovarian cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: From Concept to Clinic; Sep 18-21, 2013; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2013;19(19 Suppl):Abstract nr B30.

Abstract 2996: Systematic functional identification of driver mutations for individual ovarian cancers.

Abstract Although epithelial ovarian cancer is an intractable cancer and the overall five-year survival rate is approximately 40 to 50%, clinically effective molecular-targeted therapeutics to treat ovarian cancer has not been available thus far. Recent progress of massively parallel sequencing technologies has been enabling us to identify substantial number of genomic aberrations including somatic mutations, indels and translocations in variable cancer types exemplified by epithelial ovarian cancer and non-small cell lung cancer. However, it is still very difficult to identify a ‘driver’ gene, which is functionally relevant for cancer growth, from hundreds of mutations that are usually found from a single cancer sample of an individual patient. An in vitro cultured cell line, which is originated from a clinical sample, can be utilized to assess functions of the genes that are identified to have somatic aberrations in the paired clinical tumor. We have been establishing multiple ovarian cancer cell lines derived from clinical samples and analyzing pairs of the cell lines and clinical samples with exon-capture sequencing. The genes, which are somatically mutated and shared in a pair of cell line and clinical sample, are targeted with RNA interference in the cell line, and subsequently the effect on cell proliferation and survival is examined. As such, a driver mutation for an individual cancer can be identified by a set of experimental perturbations. Identified driver mutations would be then validated with publicly available databases and with clinical archive samples. We believe this approach can increase an opportunity to identify molecular targets in ovarian cancer therapeutics. Citation Format: Seiichi Mori, Takeshi Fujiwara, Takako Yokomizo, Haruko Iwase, Kazuyoshi Kato, Yukie Nakashima, Tokuichi Kawaguchi, Tetsuo Noda. Systematic functional identification of driver mutations for individual ovarian cancers. [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 2996. doi:10.1158/1538-7445.AM2013-2996

Abstract 5113: Carboxyl terminal region of MUC16 is critical for tumor cell invasion and drug resistance.

Abstract MUC16, also known as CA125, is routinely used to monitor the progression (as a biomarker) and response to therapeutics (as a prognostic marker) for epithelial ovarian cancer (EOC), but the deregulated expression is not restricted only to EOC. Recent studies from our lab have shown that normal breast and pancreas (ducts) do not express MUC16; however its expression progressively increases in primary and metastatic breast and pancreatic cancers. In addition, we have observed the involvement of MUC16 in enhancing the tumorigenic potential by shRNA studies. In the present study, to understand the mechanistic involvement of MUC16, we have focused on its C-terminal region. MUC16 is a type-I membrane-bound mucin with a heavily glycosylated N-terminal domain, a transmembrane region (TM) and a 32-residue cytoplasmic tail (CT) domain. Though it has been predicted that MUC16 undergoes cleavage/shedding in the extracellular SEA domain(s) by proteases such as MMP7 and neutrophil elastase, the exact nature and site of cleavage and the fate of the membrane bound portion (C-terminal) after release of extracellular portion is not entirely understood. The post cleavage C-ter region is interesting due to presence of (a) stretch of polybasic aminoacids, the site of interaction for cytoskeletal proteins like Ezrin/Radixin/Moesin, (b) potential nuclear localization signal (bioinformatics prediction) which might facilitate its nuclear localization with associated transcription factors modulating the transcription of various target genes and (c) several potential serine, threonine and tyrosine phosphorylation sites facilitating its interaction(s) with proteins influencing oncogenic signaling pathways. Therefore, we hypothesized that the oncogenic potential of MUC16 is in part, mediated by the potential involvement of C-terminal domain of MUC16. Thusly, we have made several truncated versions of MUC16 from the C-terminal end with N-terminal FLAG tag and C-terminal HA-tag and are in the process of characterization. To understand the functional significance of this C-terminal region, we have stably transfected MCF7 (breast cancer), MiaPaCa and T3M4 (pancreatic cancer) cells with an empty vector, FLAG-MUC16-114AA-HA and FLAG-MUC16-114AA-dCT (32 AA in the CT is deleted). Our results from in vitro functional assays indicate that C-terminal region of MUC16 is responsible for enhancing the invasive potential of cancer cells and confers resistance to chemotherapeutic agents such as Cisplatin and Gemcitabine. Moreover, we see that the function of the carboxyl terminal MUC16 is dependent on the stability of the protein, which is a function of N-glycosylation and ubiquitylation. Overall, our studies indicate that C-terminal region of MUC16 is an important mediator of cancer cell invasion and drug resistance. (This work is supported from the Department of Defense BC101014). Citation Format: Srustidhar Das, Maria P. Torres-Gonzalez, Prabin D. Majhi, Imayavaramban Lakshmanan, Moorthy P. Ponnusamy, Satyanarayana Rachagani, Eric Cruz, Dhanya Haridas, Surinder K. Batra. Carboxyl terminal region of MUC16 is critical for tumor cell invasion and drug resistance. [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 5113. doi:10.1158/1538-7445.AM2013-5113

Role of EZH2 in Epithelial Ovarian Cancer: From Biological Insights to Therapeutic Target

EZH2 is the catalytic subunit of polycomb repressive complex 2 (PRC2), which generates a methylation epigenetic mark at lysine 27 residue of histone H3 (H3K27me3) to silence gene expression. EZH2 target genes are involved in a variety of biological processes such as stem cell pluripotency, cell proliferation, and oncogenic transformation. EZH2 is often over-expressed in epithelial ovarian cancer (EOC) cells and in ovarian cancer-associated stromal endothelial cells. Notably, EZH2 promotes cell proliferation, inhibits apoptosis and enhances angiogenesis in EOCs. In contrast to genetic alterations, which are typically non-reversible, epigenetic alterations are reversible. Thus, inhibiting EZH2/PRC2 activity represents an attractive strategy for developing ovarian cancer therapeutics by targeting both ovarian cancer cells and ovarian tumor microenvironment. Here we discuss the progress recently obtained in understanding how EZH2/PRC2 promotes malignant phenotypes of EOC. In addition, we focus on strategies for targeting EZH2/PRC2 to develop novel EOC epigenetic therapeutics.

Oncolytic virotherapy for ovarian cancer.

In the past two decades, more than 20 viruses with selective tropism for tumor cells have been developed as oncolytic viruses (OVs) for treatments of a variety of malignancies. Of these viruses, eleven have been tested in human ovarian cancer models in preclinical studies. So far, nine phase I or II clinical trials have been conducted or initiated using four different types of OVs in patients with recurrent ovarian cancers. In this article, we summarize the different OVs that are being assessed as therapeutics for ovarian cancer. We also present an overview of recent advances in identification of key genetic or immune-response pathways involved in tumorigenesis of ovarian cancer, which provides a better understanding of the tumor specificities and oncolytic properties of OVs. In addition, we discuss how next-generation OVs could be genetically modified or integrated into multimodality regimens to improve clinical outcomes based on recent advances in ovarian cancer biology.