Ovarian Cancer Patient-derived Xenografts Research Articles

Ovarian carcinoma patient derived xenografts reproduce their tumor of origin and preserve an oligoclonal structure.

Advanced Epithelial Ovarian Cancer (EOC) patients frequently relapse by 24 months and develop resistant disease. Research on EOC therapies relies on cancer cell lines established decades ago making Patient Derived Xenografts (PDX) attractive models, because they are faithful representations of the original tumor. We established 35 ovarian cancer PDXs resulting from the original graft of 77 EOC samples onto immuno-compromised mice. PDXs covered the diversity of EOC histotypes and graft take was correlated with early patient death. Fourteen PDXs were characterized at the genetic and histological levels. PDXs reproduced phenotypic features of the ovarian tumors of origin and conserved the principal characteristics of the original copy number change (CNC) profiles over several passages. However, CNC fluctuations in specific subregions comparing the original tumor and the PDXs indicated the oligoclonal nature of the original tumors. Detailed analysis by CGH, FISH and exome sequencing of one case, for which several tumor nodules were sampled and grafted, revealed that PDXs globally maintained an oligoclonal structure. No overgrowth of a particular subclone present in the original tumor was observed in the PDXs. This suggested that xenotransplantation of ovarian tumors and growth as PDX preserved at least in part the clonal diversity of the original tumor. We believe our data reinforce the potential of PDX as exquisite tools in pre-clinical assays.

AAV9 delivering a modified human Mullerian inhibiting substance as a gene therapy in patient-derived xenografts of ovarian cancer

To improve ovarian cancer patient survival, effective treatments addressing chemoresistant recurrences are particularly needed. Mullerian inhibiting substance (MIS) has been shown to inhibit the growth of a stem-like population of ovarian cancer cells. We have recently engineered peptide modifications to human MIS [albumin leader Q425R MIS (LRMIS)] that increase production and potency in vitro and in vivo. To test this novel therapeutic peptide, serous malignant ascites from highly resistant recurrent ovarian cancer patients were isolated and amplified to create low-passage primary cell lines. Purified recombinant LRMIS protein successfully inhibited the growth of cancer spheroids in vitro in a panel of primary cell lines in four of six patients tested. Adeno-associated virus (AAV) -delivered gene therapy has undergone a clinical resurgence with a good safety profile and sustained gene expression. Therefore, AAV9 was used as a single i.p. injection to deliver LRMIS to test its efficacy in inhibiting growth of palpable tumors in patient-derived ovarian cancer xenografts from ascites (PDXa). AAV9-LRMIS monotherapy resulted in elevated and sustained blood concentrations of MIS, which significantly inhibited the growth of three of five lethal chemoresistant serous adenocarcinoma PDXa models without signs of measurable or overt toxicity. Finally, we tested the frequency of MIS type II receptor expression in a tissue microarray of serous ovarian tumors by immunohistochemistry and found that 88% of patients bear tumors that express the receptor. Taken together, these preclinical data suggest that AAV9-LRMIS provides a potentially well-tolerated and effective treatment strategy poised for testing in patients with chemoresistant serous ovarian cancer.

Molecular and clinical implementations of ovarian cancer mouse avatar models.

Innovation in oncology drug development has been hindered by lack of preclinical models that reliably predict clinical activity of novel therapies in cancer patients. Increasing desire for individualize treatment of patients with cancer has led to an increase in the use of patient-derived xenografts (PDX) engrafted into immune-compromised mice for preclinical modeling. Large numbers of tumor-specific PDX models have been established and proved to be powerful tools in pre-clinical testing. A subset of PDXs, referred to as Avatars, establish tumors in an orthotopic and treatment naïve fashion that may represent the most clinical relevant model of individual human cancers. This review will discuss ovarian cancer (OC) PDX models demonstrating the opportunities and limitations of these models in cancer drug development, and describe concepts of clinical trials design in Avatar guided therapy.

Cediranib combined with chemotherapy reduces tumor dissemination and prolongs the survival of mice bearing patient-derived ovarian cancer xenografts with different responsiveness to cisplatin.

Cediranib is a pan-vascular endothelial growth factor receptor tyrosine kinase inhibitor that affects tumor angiogenesis and is under investigation in clinical studies on ovarian cancer. Using a panel of eleven patient-derived ovarian cancer xenografts (EOC-PDX) growing orthotopically in the peritoneal cavity of nude mice we investigated the effect of cediranib as monotherapy or in combination with chemotherapy on overall survival (primary endpoint, at euthanasia), and tumor dissemination and metastasis in the peritoneal cavity (secondary endpoint, interim analysis). The response of EOC-PDX to cediranib varied (increment of lifespan, ILS between 12 and 85 %) in the different EOC-PDX, independently from tumor responsiveness to cisplatin (DDP). Cediranib combined with DDP and in maintenance regimen prolonged the survival of mice bearing EOC-PDX with different responsiveness to DDP (ILS between 34 and 224 % with only DDP and between 135 and 337 % with DDP plus Cediranib); survival was extended with the addition of paclitaxel to chemotherapy (50-77 % complete remissions). Cediranib reduced ascites of advanced EOC-PDX, but had limited effect on tumor dissemination; only combined with chemotherapy, ascites and metastases were both reduced. The reduction of tumor dissemination was associated to the increase of overall survival. In conclusion, the response to cediranib differs in the various EOC-PDX, reproducing the heterogeneous response of cancer patients to angiogenesis inhibitors. Cediranib potentiated chemotherapy, significantly inhibiting tumor progression and dissemination to metastatic organs, even in tumors poorly responsive to DDP. EOC-PDX preclinical models with different responsiveness to Cediranib may help in identifying determinants of response to cediranib and mechanisms of adaptation to antiangiogenic treatments.

P7.01 - Characterization of 148 Ovarian Cancer tumografts (Avatars) using BROCA-HR deep sequencing

ABSTRACT Ovarian cancer is the most lethal gynecologic malignancy. Current standard of care post surgical cytoreduction is combination platinum/taxane chemotherapy, with initial response varying widely; subsets of carcinomas demonstrate resistance or sensitivity from the onset. The underlying cause of this response heterogeneity remains unknown. Patient-derived xenografts (PDX) serve as useful in vivo models to study molecular response markers and test the efficacy of targeted therapies. Our group has demonstrated a high engraftment rate (>70%) of ovarian cancer PDXs (Avatars) by injecting treatment naive patient tumor directly into the peritoneal cavity of immunocompromised mice, in an effort to better mimic the anatomic context by which ovarian cancer naturally develops. We now have 328 engrafted Avatar models representing the wide range of ovarian cancer subtypes, including serous (65%), mixed epithelial (10%), endometrioid (7%), clear cell (4%), mucinous (3%), carcinosarcoma (1%) and other (10%). Using these models, we are performing a massively parallel sequencing strategy referred to as BROCA-HR to detect all mutation classes (e.g. gene rearrangements, copy number variations, etc.) and gene aberrations within the Fanconi Anemia-BRCA homologous recombination (HR), non-homologous end joining (NHEJ), PTEN, and DNA Mismatch repair pathways. To date, 148 Avatar models have undergone BROCA sequencing. Observed deleterious mutations included 12 BRCA1 (8%), 6 BRCA2 (4%), and 4 PIK3CA (3%); additional loss of function mutations were also evident in the following genes: ATM, RAD51C, FANCM, FANCD2, FANCA, CHEK2, PALB2, MHS6, CDK12 and GEN1. These models are representative of mutations seen in the TCGA mutation set. Models harboring said mutations are currently being tested with platinum agents and/or niraparib. Taken together, the Avatars represent a high throughput model system for pre-clinical testing in vivo that effectively recapitulates ovarian cancer (e.g. mutations in BRCA1, BRCA2 and other DNA damage response proteins). We have been able to establish avatars at a high rate, including those with mutations in BRCA1, BRCA2 and other DNA damage response genes. These models will be a unique resource for future studies of biomarkers and novel therapeutic approaches.

P4.01 - In vivo synergism between chemotherapy and the 3C23K monoclonal antibody targeting the AMH-type II receptor in ovarian cancer PDX models

ABSTRACT Background: Mullerian inhibiting substance, also called anti-Mullerian hormone (AMH), belongs to the TGF-alpha family and plays a key role during sexual development. Various reports have underlined expression of AMHRII in human gynecologic cancers including ovarian cancers (OC). Based on these considerations, a humanized glyco-engineered monoclonal antibody (3C23K) specific to AMHRII has been developed. In this study, we have assessed AMHRII expression in a panel of OC patient-derived xenografts (PDXs) and evaluated in vivo efficacy of 3C23K. Materials and Methods: AMHRII expression was studied in 14 OC PDXs, using flow cytometry (FC), immunohistochemistry (IHC) and immunofluorescence (IF). In vivo efficacy of 3C23K alone (20 mg/kg twice a week, IP) or in combination with standard chemotherapy (CT) (carboplatin 66 mg/kg IP days 1-22 + paclitaxel 30 mg/kg IP days 1-22) has been evaluated in 4 AMHRII-positive undifferentiated, mucinous or serous adenocarcinoma OC PDXs. On these 4 models, expression of AMHRII was studied before and after treatments, as well as tumor-associated macrophages (TAMs) infiltrate. Results: Using FC analyses, 6 over the 14 PDX tested (43%) were positive with at least 25% of tumor cells expressing AMHR2. Data obtained with IHC and IF analyses were in concordance with FC results. In vivo experiments showed an intermediate efficacy of the 3C23K when administered alone and a synergistic effect when associated with CT. In particular, 3C23K was found to consistently increase the proportion of complete response (CR) on any of the models tested, namely OV54 (188 cumulated days of CR vs 47), OV25 (30 vs 10), OV21 (12 vs 0) and OV16 (67 vs 37). 3C23K administration did not modify tumor AMHRII expression. Stromal FC analyses showed a high TAMs infiltrate of M2 type compatible with induction of Antibody Dependent Cell Phagocytosis by the glyco-engineered 3C23K antibody. Conclusions: Using a panel of OC PDXs, we confirmed the expression of AMHRII in human OC. Targeting this receptor by 3C23K, dramatically increases the efficacy of a standard CT. Our results therefore suggest that such combination could be further developed in OC patients whose prognosis remains dismal. 3C23K is planned to enter in early clinical phase this year.

Abstract PR01: Adaptive resistance of patient-derived ovarian cancer cells to PI3K/mTOR inhibition

Abstract Relapse due to drug resistance remains a major challenge for successful cancer treatment despite advances in chemotherapy efficacy and the development of targeted therapies. We have previously reported a therapy-induced adaptive response that is associated with treatment resistance and are investigating whether this adaptive response program could be targeted to increase treatment efficacy (Muranen et al, Cancer Cell 2012). The PI3K pathway is amplified in high-grade serous ovarian tumor and represents an attractive target in this cancer subtype. We found that both in vitro and in vivo exposure of patient-derived ovarian cancer cells to the PI3K/mTOR dual inhibitor GNE-493 increases the levels of the anti-apoptotic Bcl-2 protein, and multiple tyrosine kinase receptors, such as EGFR and HER2. Concurrent treatment of these cancer cells with GNE-493 and ABT737 (a Bcl-2 and Bcl-xl dual inhibitor) causes a reduction in cell number and increased cell death. Ongoing studies are underway to comprehensively characterize the adaptive response signature of a panel of patient-derived ovarian cancer cells and study whether targeting multiple different upregulated proteins (e.g. EGFR) may increase PI3K/mTOR treatment efficacy. Therapy resistance has also been associated with exposure of tumor cells to pro-survival factors secreted by supporting cells in the tumor microenvironment (Straussman et al, Nature 2012 and Wilson et al, Nature 2012). We found that the growth-factors EGF, inflammatory cytokine IL-6 and ascites fluid from ovarian cancer patients induces up-regulation of Bcl-2 as well as IGFR, which masked the adaptive response to PI3K/mTOR inhibitors. We are performing ongoing studies using orthotopic patient-derived ovarian cancer xenografts to investigate the patterns of residual disease in vivo with the ultimate goal to identify whether Bcl-2 targeted drugs will enhance the efficacy of PI3K/mTOR inhibition and whether there is differential sensitivity at distinct anatomical sites (pancreas, diaphragm, peritoneal wall). Our studies provide a framework to identify adaptive response programs and tumor microenvironment factors in patient-derived ovarian cancer cells that limit treatment efficacy of PI3K/mTOR pathway inhibition and to develop therapies to abrogate this resistance. Citation Format: Ioannis Zervantonakis, Hsing-Yu Chen, Taru Muranen, Joyce Liu, Ronny Drapkin, Ursula Matulonis, Joan Brugge. Adaptive resistance of patient-derived ovarian cancer cells to PI3K/mTOR inhibition. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Drug Sensitivity and Resistance: Improving Cancer Therapy; Jun 18-21, 2014; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(4 Suppl): Abstract nr PR01.

732: Heterogeneous response to bevacizumab combined with chemotherapy in patient-derived ovarian cancer xenografts

732: Heterogeneous response to bevacizumab combined with chemotherapy in patient-derived ovarian cancer xenografts

78: Proffered Paper: Dual wavelength near-infrared fluorescence imaging of VEGF and IGF-1R in ovarian cancer patient-derived xenografts

78: Proffered Paper: Dual wavelength near-infrared fluorescence imaging of VEGF and IGF-1R in ovarian cancer patient-derived xenografts

Molecular correlates of platinum response in human high-grade serous ovarian cancer patient-derived xenografts

IntroductionImprovement in the ability to target underlying drivers and vulnerabilities of high-grade serous ovarian cancer (HG-SOC) requires the development of molecularly annotated pre-clinical models reflective of clinical responses. MethodsWe generated patient-derived xenografts (PDXs) from consecutive, chemotherapy-naïve, human HG-SOC by transplanting fresh human HG-SOC fragments into subcutaneous and intra-ovarian bursal sites of NOD/SCID IL2Rγnull recipient mice, completed molecular annotation and assessed platinum sensitivity. ResultsThe success rate of xenografting was 83%. Of ten HG-SOC PDXs, all contained mutations in TP53, two were mutated for BRCA1, three for BRCA2, and in two, BRCA1 was methylated. In vivo cisplatin response, determined as platinum sensitive (progression-free interval ≥100 d, n = 4), resistant (progression-free interval <100 d, n = 3) or refractory (n = 3), was largely consistent with patient outcome. Three of four platinum sensitive HG-SOC PDXs contained DNA repair gene mutations, and the fourth was methylated for BRCA1. In contrast, all three platinum refractory PDXs overexpressed dominant oncogenes (CCNE1, LIN28B and/or BCL2). ConclusionsBecause PDX platinum response reflected clinical outcome, these annotated PDXs will provide a unique model system for preclinical testing of novel therapies for HG-SOC.

Abstract 3725: Anti-DLL4 (demcizumab) inhibits tumor growth and reduces cancer stem cell frequency in patient-derived ovarian cancer xenografts.

Abstract Ovarian cancer in a major cause cancer related death in women and this disease is often characterized by resistance to chemotherapy and tumor recurrence. Cancer stem cells (CSCs) have been shown to be relatively insensitive to chemotherapeutic agents. Several lines of evidence indicate that aberrant Notch signaling plays a major role in ovarian cancer progression and resistance to standard therapies. DLL4 is a key ligand that activates the Notch pathway and has been shown to regulate both CSC function and tumor angiogenesis. DLL4 expression in tumor and endothelial cells has also been associated with resistance to bevacizumab treatment. In these studies, we investigated the role of anti-DLL4 in a panel of patient-derived ovarian cancer xenograft models. We utilized anti-human DLL4 (OMP-21M18) and anti-murine DLL4 to block Notch signaling in both the tumor and stromal/vascular cells in the xenografts. We found that anti-DLL4 treatment was broadly efficacious in these ovarian cancer models, significantly inhibiting tumor growth both as a single agent and in combination with paclitaxel. We carried out serial transplantation studies to investigate the effect on cancer stem cells and found that anti-DLL4 in combination with paclitaxel profoundly reduced the tumor initiating cell frequency. In contrast, treatment with paclitaxel alone had the opposite effect and increased ovarian CSC frequency. These data suggest that anti-DLL4 treatment sensitizes chemoresistant ovarian tumorigenic cells to therapy and provide compelling evidence that anti-DLL4 (demcizumab) is a promising strategy for the clinical treatment of ovarian cancer. Citation Format: Wan-Ching Yen, Marcus M. Fischer, Jalpa Shah, Jie Wei, Jennifer Cain, Pete Yeung, Lucia Beviglia, Belinda Cancilla, Ann Kapoun, John Lewicki, Austin Gurney, Timothy Hoey. Anti-DLL4 (demcizumab) inhibits tumor growth and reduces cancer stem cell frequency in patient-derived ovarian cancer xenografts. [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 3725. doi:10.1158/1538-7445.AM2013-3725

Patient-Derived Xenograft Models to Improve Targeted Therapy in Epithelial Ovarian Cancer Treatment

Despite increasing evidence that precision therapy targeted to the molecular drivers of a cancer has the potential to improve clinical outcomes, high-grade epithelial ovarian cancer (OC) patients are currently treated without consideration of molecular phenotype, and predictive biomarkers that could better inform treatment remain unknown. Delivery of precision therapy requires improved integration of laboratory-based models and cutting-edge clinical research, with pre-clinical models predicting patient subsets that will benefit from a particular targeted therapeutic. Patient-derived xenografts (PDXs) are renewable tumor models engrafted in mice, generated from fresh human tumors without prior in vitro exposure. PDX models allow an invaluable assessment of tumor evolution and adaptive response to therapy. PDX models have been applied to pre-clinical drug testing and biomarker identification in a number of cancers including ovarian, pancreatic, breast, and prostate cancers. These models have been shown to be biologically stable and accurately reflect the patient tumor with regards to histopathology, gene expression, genetic mutations, and therapeutic response. However, pre-clinical analyses of molecularly annotated PDX models derived from high-grade serous ovarian cancer (HG-SOC) remain limited. In vivo response to conventional and/or targeted therapeutics has only been described for very small numbers of individual HG-SOC PDX in conjunction with sparse molecular annotation and patient outcome data. Recently, two consecutive panels of epithelial OC PDX correlate in vivo platinum response with molecular aberrations and source patient clinical outcomes. These studies underpin the value of PDX models to better direct chemotherapy and predict response to targeted therapy. Tumor heterogeneity, before and following treatment, as well as the importance of multiple molecular aberrations per individual tumor underscore some of the important issues addressed in PDX models.