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Autoxidation of tartaric acid in air-saturated aqueous solutions in the presence of Fe(II) at low pH, 2.5, shows autocatalytic behavior with distinct initiation, propagation, and termination phases. With increasing pH, the initiation phase speeds up, while the propagation phase shortens and reduces to none. We show that the propagation phase is a chain reaction that occurs via activation of oxygen in the initiation stage with the production of hydrogen peroxide. The subsequent Fenton oxidation that regenerates hydrogen peroxide with a positive feedback is typical of a self-sustained chain reaction. The conditions for such a chain reaction are shown to be similar to those of a dynamical system with critical behavior; namely, the system becomes unstable when the kinetic matrix of pseudo-first-order reaction becomes negatively defined with a negative eigenvalue giving the rate of exponential (chain) growth of the reactive species.

The clinical development of antibody drug conjugates (ADCs) in ovarian cancer began in 2008 with farletuzumab, a humanized monoclonal antibody, and vintafolide, an antigen drug conjugate, both targeting alpha folate receptor. Over the years, this novel class of drugs expanded to agents with a more sophisticated design and structure, targeting tissue factor (TF) in cervical cancer or human epidermal growth factor receptor 2 (HER2) in endometrial cancer. Despite the impressive number of patients included in clinical trials investigating different ADCs across gynecological cancers, it was only recently that the Food and Drug Administration (FDA) granted accelerated approvals to the first ADCs in gynecologic cancer. In September 2021, the FDA approved tisotumab vedotin (TV) in recurrent or metastatic cervical cancer with disease progression on or after chemotherapy. This was followed in November 2022, by the approval of mirvetuximab soravtansine (MIRV) for adult patients with folate receptor alpha (FRα) positive, platinum-resistant epithelial ovarian, fallopian tube, or primary peritoneal cancer, who have received one to three prior systemic treatment regimens. Currently, the field of ADCs is rapidly expanding and more than 20 ADC formulations are in clinical trials for the treatment of ovarian, cervical and endometrial tumors. This review summarizes key evidence supporting their use and therapeutic indications, including results from late-stage development trials investigating MIRV in ovarian cancer and TV in cervical cancer. We also outline new concepts in the field of ADCs, including promising targets such as NaPi2 and novel drug delivery platforms such as dolaflexin with a scaffold-linker. Finally, we briefly present challenges in the clinical management of ADC toxicities and the emerging role of ADC combination therapies, including chemotherapy, anti-angiogenic and immunotherapeutic agents.

Objective. The primary purpose of this study was to determine if farletuzumab, an antifolate receptor-alpha monoclonal antibody, improved progression-free survival (PFS) versus placebo when added to standard chemo-therapy regimens in patients with platinum-sensitive recurrent ovarian cancer (OC) in first relapse (platinum-free interval: 6-36 months) with low cancer antigen 125 (CA-125) levels. Gynecologic Oncology (2023) Methods. Eligibility included CA-125

This article reviews treatments and targets of interest in endometrial cancer by molecular subtype. The Cancer Genome Atlas (TCGA) classifies four molecular subtypes—mismatch repair deficient (dMMR)/microsatellite instability-high (MSI-H); copy number high (CNH)/p53abn; copy number low (CNL)/no specific molecular profile (NSMP); and POLEmut—which are validated and highly prognostic. Treatment consideration by subtype is now recommended. In March and April 2022, respectively, the US Food and Drug Administration (FDA) fully approved and the European Medicines Agency adopted a positive opinion recommending the anti-programmed cell death protein-1 (PD-1) antibody pembrolizumab for advanced/recurrent dMMR/MSI-H endometrial cancer which has progressed on or following a platinum-containing therapy. A second anti-PD-1, dostarlimab, received accelerated approval by the FDA and conditional marketing authorization by the European Medicines Agency in this group. The combination of pembrolizumab/lenvatinib for mismatch repair proficient/microsatellite stable endometrial cancer, including p53abn/CNH and NSMP/CNL, received accelerated FDA approval in conjunction with Australia’s Therapeutic Goods Administration and Health Canada in September 2019. The FDA and European Medicines Agency made full recommendations in July 2021 and October 2021. Trastuzumab is National Comprehensive Cancer Network (NCCN) compendium listed for human epidermal growth factor receptor-2-positive serous endometrial cancer, which is primarily within the p53abn/CNH subtype. In addition to hormonal therapy, maintenance therapy with selinexor (exportin-1 inhibitor) showed potential benefit inp53-wildtype cases in a subset analysis and is being investigated prospectively. Other treatment regimens being evaluated in NSMP/CNL are hormonal combinations with cyclin-dependent kinase 4/6 inhibitors and letrozole. Ongoing trials are evaluating immunotherapy in combination with frontline chemotherapy and other targeted agents. Treatment de-escalation is being evaluated in POLEmut cases given its favorable prognosis with or without adjuvant therapy. Molecular subtyping has important prognostic and therapeutic implications, and should guide patient management and clinical trial design in endometrial cancer, which is a molecularly driven disease.

BackgroundThe predicament of achieving optimal surgical intervention faced by surgeons in treating ovarian cancer has driven research into improving intra-operative detection of cancer using fluorescent materials.ObjectiveTo provide a literature overview on the clinical use of intra-operative fluorescence-guided surgery for ovarian cancer, either for cytoreductive surgery or sentinel lymph node (SLN) biopsy.MethodsThe systematic review included studies from June 2002 until October 2021 from PubMed, Web of Science, and Scopus as well as those from a search of related literature. Studies were included if they investigated the use of fluorescence-guided surgery in patients with a diagnosis of ovarian cancer. Authors charted variables related to study characteristics, patient demographics, baseline clinical characteristics, fluorescence-guided surgery material, and treatment details, and surgical, oncological, and survival outcome variables. After screening 2817 potential studies, 24 studies were included.ResultsStudies investigating the role of fluorescence-guided surgery to visualize tumor deposits or SLN biopsy included the data of 410 and 118 patients, respectively. Six studies used indocyanine green tracer with a mean SLN detection rate of 92.3% with a pelvic and para-aortic detection rate of 94.8% and 96.7%, respectively. The sensitivity, specificity, and positive predictive value for micrometastases detection of OTL38 and 5-aminolevulinc acid at time of cytoreduction were 92.2% vs 79.8%, 67.3% vs 94.8%, and 55.8% vs 95.8%, respectively.ConclusionFluorescence -guided surgery is a technique that may improve the detection rate of micrometastases and SLN identification in ovarian cancer. Further research is needed to establish whether this will lead to improved patient outcomes.

PURPOSE In VELIA trial, veliparib combined with carboplatin-paclitaxel, followed by maintenance (veliparib-throughout) was associated with improved progression-free survival (PFS) compared with carboplatin-paclitaxel alone in patients with high-grade ovarian carcinomas. We explored the prognostic value of the modeled cancer antigen (CA)-125 elimination rate constant K (KELIM), which is known to be an indicator of the intrinsic tumor chemosensitivity (the faster the rate of CA-125 decline, the higher the KELIM and the higher the chemosensitivity), and its association with benefit from veliparib. PATIENTS AND METHODS Individual KELIM values were estimated from longitudinal CA-125 kinetics. Patients were categorized as having favorable (≥ median) or unfavorable (< median) KELIM. The prognostic value of KELIM for veliparib-related PFS benefit was explored in cohorts treated with primary or interval debulking surgery, according to the surgery completeness, the disease progression risk group, and the homologous recombination (HR) status ( BRCA mutation, HR deficiency [HRD], or HR proficiency [HRP]). RESULTS The data from 854 of 1,140 enrolled patients were analyzed (primary debulking surgery, n = 700; interval debulking surgery, n = 154). Increasing KELIM values were associated with higher benefit from veliparib in HRD cancer, as were decreasing KELIM values in HRP cancer. The highest PFS benefit from veliparib was observed in patients with both favorable KELIM and BRCA mutation (hazard ratio, 0.28; 95% CI, 0.13 to 0.61) or BRCA wild-type HRD cancer (hazard ratio, 0.43; 95% CI, 0.26 to 0.70), consistent with the association between poly (adenosine diphosphate-ribose) polymerase inhibitor efficacy and platinum sensitivity. In contrast, seventy-four percent of patients with a BRCA mutation and unfavorable KELIM progressed within 18 months while on veliparib. The patients with HRP cancer and unfavorable KELIM might have benefited from the veliparib chemosensitizing effect. CONCLUSION In addition to HRD/ BRCA status, the tumor primary chemosensitivity observed during the first-line chemotherapy might be another complementary determinant of poly (adenosine diphosphate-ribose) polymerase inhibitor efficacy.

ObjectivesAnetumab ravtansine is an antibody-drug conjugate consisting of a fully human anti-mesothelin monoclonal antibody conjugated to cytotoxic maytansinoid tubulin inhibitor DM4. Mesothelin is highly expressed in ovarian cancer. This phase Ib study determines the safety, pharmacokinetics, and anti-tumor activity of anetumab ravtansine and pegylated liposomal doxorubicin in mesothelin-expressing platinum-resistant ovarian cancer.MethodsAnetumab ravtansine (5.5 or 6.5 mg/kg) and pegylated liposomal doxorubicin (30 mg/m2) were administered intravenously every 3 weeks to 65 patients with platinum-resistant epithelial ovarian cancer. Mesothelin expression was assessed by central immunohistochemistry. Adverse events, tumor response (RECIST 1.1), and progression-free survival were determined. Biomarker samples were assessed by ELISA and next-generation sequencing.ResultsIn dose escalation, nine patients received anetumab ravtansine across two doses (5.5 or 6.5 mg/kg). The maximum tolerated dose of anetumab ravtansine was 6.5 mg/kg every 3 weeks and no dose-limiting toxicities were observed. In dose expansion, 56 patients were treated at the maximum tolerated dose. The most common treatment-emergent adverse events of any grade were nausea (47.7%), decreased appetite (43.1%), fatigue (38.5%), diarrhea (32.3%), and corneal disorder (29.2%). In all treated patients the objective response rate was 27.7% (95% CI 17.3% to 40.2%), including one complete (1.5%) and 17 partial responses (26.2%), with median duration of response of 7.6 (95% CI 3.3 to 10.2) months and median progression-free survival of 5.0 (95% CI 3.2 to 6.0) months. In an exploratory analysis of a sub-set of patients (n=19) with high mesothelin expression who received ≤3 prior lines of systemic therapy, the objective response rate was 42.1% (95% CI 20.3% to 66.5%) with a median duration of response of 8.3 (95% CI 4.1 to 12.0) months and median progression-free survival of 8.5 (95% CI 4.0 to 11.4) months.ConclusionsAnetumab ravtansine and pegylated liposomal doxorubicin showed tolerability and promising clinical activity. These results established the dose schedule and the mesothelin-positive target population of this combination for a phase III study in platinum-resistant ovarian cancer.Trial registration numberNCT02751918.

Background The phase 3 VELIA trial evaluated veliparib with carboplatin/paclitaxel and as maintenance in patients with high-grade serous ovarian carcinoma. Methods Patients with previously untreated stage III–IV high-grade serous ovarian carcinoma were randomized 1:1:1 to control (placebo with carboplatin/paclitaxel and placebo maintenance), veliparib-combination-only (veliparib with carboplatin/paclitaxel and placebo maintenance), or veliparib-throughout (veliparib with carboplatin/paclitaxel and veliparib maintenance). Randomization stratification factors included geographic region (Japan versus North America or rest of the world). Primary end point was investigator-assessed median progression-free survival. Efficacy, safety, and pharmacokinetics were evaluated in a subgroup of Japanese patients. Results Seventy-eight Japanese patients were randomized to control (n = 23), veliparib-combination-only (n = 30), and veliparib-throughout (n = 25) arms. In the Japanese subgroup, median progression-free survival for veliparib-throughout versus control was 27.4 and 19.1 months (hazard ratio, 0.46; 95% confidence interval, 0.18–1.16; p = 0.1 [not significant]). In the veliparib-throughout arm, grade 3/4 leukopenia, neutropenia, and thrombocytopenia rates were higher for Japanese (32%/88%/32%) versus non-Japanese (17%/56%/28%) patients. Grade 3/4 anemia rates were higher in non-Japanese (65%) versus Japanese (48%) patients. Early introduction of olanzapine during veliparib monotherapy maintenance phase may help prevent premature discontinuation of veliparib, via its potent antiemetic efficacy. Conclusions Median progression-free survival was numerically longer in Japanese patients in the veliparib-throughout versus control arm, consistent with results in the overall study population. Pharmacokinetics were comparable between Japanese and non-Japanese patients. Data for the subgroup of Japanese patients were not powered to show statistical significance but to guide further investigation.

BackgroundUterine clear cell and serous carcinomas have a high propensity for locoregional and distant spread, tend to be more advanced at presentation, and carry a higher risk of recurrence and death than endometrioid cancers. Limited prospective data exist to guide evidence-based management of these rare malignancies.ObjectiveThe American Radium Society sought to summarize evidence-based guidelines developed by a multidisciplinary expert panel that help to guide the management of uterine clear cell and serous carcinomas.MethodsThe American Radium Society Appropriate Use Criteria presented in this manuscript were developed by a multidisciplinary expert panel using an extensive analysis of current published literature from peer-reviewed journals. A well-established methodology (modified Delphi) was used to rate the appropriate use of diagnostic and therapeutic procedures for the management of uterine clear cell and serous carcinomas.ResultsThe primary treatment for non-metastatic uterine clear cell and serous carcinomas is complete surgical staging, with total hysterectomy, salpingo-oophorectomy, omentectomy, and lymph node staging. Even in early-stage disease, patients with uterine clear cell and serous carcinomas have a worse prognosis than those with type I endometrial cancers, warranting consideration for adjuvant therapy regardless of the stage. Given the aggressive nature of these malignancies, and until further research determines the most appropriate adjuvant therapy, it may be reasonable to counsel patients about combined-modality treatment with systemic chemotherapy and radiotherapy.ConclusionPatients diagnosed with uterine clear cell and serous carcinomas should undergo complete surgical staging. Multimodal adjuvant therapies should be considered in the treatment of both early-stage and advanced-stage disease. Further prospective studies or multi-institutional retrospective studies are warranted to determine optimal sequencing of therapy and appropriate management of patients based on their unique risk factors. Long-term surveillance is indicated due to the high risk of locoregional and distant recurrence.

Ovarian cancer is the leading cause of gynecologic cancer-related death in the United States. Historically, studies have demonstrated that ovarian cancer is a heterogeneous disease with several patient and oncologic characteristics, including BRCA status and residual disease at surgery, known to be predictive of clinical outcomes. However, during the last decade, the discovery and approval of bevacizumab and poly(adenosine diphosphate-ribose) polymerase inhibitors have moved the frontline treatment paradigm beyond platinum-doublet therapy for women with advanced ovarian cancer. Subsequently, investigators have sought to assess the therapeutic efficacy of these agents in women who are considered "high" risk and "low" risk to determine which patients may benefit the most from aggressive therapy and in whom additional treatment may be avoided. We reviewed historic and contemporary definitions of "high-risk" and "low-risk" ovarian cancer and how this has been incorporated into the subset analyses of randomized, clinical trials of therapeutic agents, including bevacizumab and poly(adenosine diphosphate-ribose) polymerase inhibitors. Next, we provided an in-depth discussion of landmark trials for frontline maintenance therapy with bevacizumab and/or poly(adenosine diphosphate-ribose) polymerase inhibitors, focusing on the impact of treatment efficacy according to a "high-risk" and "low-risk" paradigm. Furthermore, we highlighted that recent data have challenged this dichotomous classification, notably from the Gynecologic Oncology Group-0218, ICON7, SOLO-1, and PAOLA-1 trials. Although some studies have suggested that certain populations of women with advanced ovarian cancer may have a more favorable prognosis and be considered "low risk," the risk of progression and death remains unacceptably high in all women. Furthermore, in many cases, those considered the lowest risk have the most treatment benefit from maintenance therapy with poly(adenosine diphosphate-ribose) polymerase inhibitors and/or bevacizumab. From these data, we have advocated that virtually all women with advanced ovarian cancer are high risk and that the use of our most effective therapies in the frontline setting holds promise for potentially curing more patients. Lastly, we critically discuss the practice of using subanalyses in clinical trials, with emphasis that although this practice is important for hypothesis generation, caution must be taken before accepting findings from subanalyses as actual treatment effects.

OBJECTIVE: Progression-free survival (PFS) is an important early efficacy endpoint in ovarian cancer (OC) and its relevance to patients should be assessed. PRIMA, a phase III trial, assessed niraparib in patients with OC; this post hoc analysis examined the relationship between disease progression in OC and health-related quality of life (HRQoL). METHODS: The PRIMA trial randomized patients with advanced OC responsive to first-line platinum-based chemotherapy to once daily maintenance oral niraparib or placebo. This post hoc analysis evaluated the impact of disease progression on HRQoL by comparing HRQoL at the last visit pre-progression to end of treatment (EoT), and after 4, 8, 12, and 24 weeks. Assessments included the Functional Assessment of Cancer Therapy-Ovarian Symptom Index (FOSI), the European Quality of Life Five Dimension Five Level questionnaire (EQ-5D-5L) and EQ Visual Analogue Scale (EQ-VAS), the European Organization for Research and Treatment of Cancer (EORTC) Quality of Life Questionnaire (EORTC-QLQ-C30), and the EORTC Quality of Life Questionnaire Ovarian Cancer module (EORTC-QLQ-OV28). RESULTS: This post hoc analysis included 733 patients. Mean FOSI, EQ-5D-5L, and EQ-VAS scores deteriorated from last visit pre-progression to EoT and remained low up to 24-week follow-up. Least squares mean changes from last visit pre-progression to EoT were -2.1 (95% confidence interval -2.4, -1.7) for FOSI, -4.6 (-5.6, -3.5) for the EQ-5D-5L index, and -7.9 (-9.6, -6.3) for EQ-VAS. CONCLUSIONS: Disease progression negatively impacted HRQoL in patients with OC. PFS is clinically relevant, and prolonging PFS may preserve HRQoL. ispartof: GYNECOLOGIC ONCOLOGY vol:166 issue:3 pages:494-502 ispartof: location:United States status: published

Understanding real-world treatment patterns and proportions of eligible patients in each line of treatment is imperative to inform future clinical trial designs and multi-line treatment algorithm development.We conducted a retrospective observational cohort study of adult women who received first-line (1 L) therapy for r/mCC between 01 September 2014 and 31 December 2019, using The US Oncology Network electronic health records and chart review data. Patients were followed to 31 December 2020. Patient demographic and clinical characteristics, treatment patterns, and clinical outcomes were assessed descriptively.A total of 262 patients with r/mCC met study inclusion criteria (mean age = 53 years). The majority of patients in 1 L received platinum-based chemotherapy doublet plus bevacizumab (66%) or chemotherapy doublet alone (24%). Nearly half the patients (48%) completing 1 L received 2 L therapy. Among these patients, there was no consistent 2 L treatment of choice. Overall median time to treatment discontinuation was 3.5 months from 1 L treatment initiation, and median overall treatment-free interval was 2.1 months from 1 L discontinuation. Besides elevated serum creatinine, abnormal BMI indicated a directional trend for lower likelihood of receiving 2 L. Other predictors may include no prior bevacizumab, worse ECOG, and earlier disease prevention.50% of the patients who initiated 1 L treatment did not receive 2 L therapy, highlighting the need for novel and effective treatment options. As the treatment landscape continues to evolve, we anticipate that more patients will live longer with more treatment options across multiple lines of therapies in the r/mCC setting.

PURPOSE Single-agent chemotherapies have limited activity and considerable toxicity in patients with platinum-resistant epithelial ovarian cancer (PROC). Mirvetuximab soravtansine (MIRV) is an antibody-drug conjugate targeting folate receptor α (FRα). SORAYA is a single-arm, phase II study evaluating efficacy and safety of MIRV in patients with PROC. METHODS SORAYA enrolled FRα-high patients with PROC who had received one to three prior therapies, including required bevacizumab. The primary end point was confirmed objective response rate (ORR) by investigator; duration of response was the key secondary end point. RESULTS One hundred six patients were enrolled; 105 were evaluable for efficacy. All patients had received prior bevacizumab, 51% had three prior lines of therapy, and 48% received a prior poly ADP-ribose polymerase inhibitor. Median follow-up was 13.4 months. ORR was 32.4% (95% CI, 23.6 to 42.2), including five complete and 29 partial responses. The median duration of response was 6.9 months (95% CI, 5.6 to 9.7). In patients with one to two priors, the ORR by investigator was 35.3% (95% CI, 22.4 to 49.9) and in patients with three priors was 30.2% (95% CI, 18.3 to 44.3). The ORR by investigator was 38.0% (95% CI, 24.7 to 52.8) in patients with prior poly ADP-ribose polymerase inhibitor exposure and 27.5% (95% CI, 15.9 to 41.7) in those without. The most common treatment-related adverse events (all grade and grade 3-4) were blurred vision (41% and 6%), keratopathy (29% and 9%), and nausea (29% and 0%). Treatment-related adverse events led to dose delays, reductions, and discontinuations in 33%, 20%, and 9% of patients, respectively. CONCLUSION MIRV demonstrated consistent clinically meaningful antitumor activity and favorable tolerability and safety in patients with FRα-high PROC who had received up to three prior therapies, including bevacizumab, representing an important advance for this biomarker-selected population.

To assess the impact of frailty in patients with ovarian cancer on surgical procedures and outcomes.A retrospective review of patients with stage II-IV ovarian cancer from April 2013 to September 2017 was performed. Patients were triaged by laparoscopy to determine primary resectability. The adjusted modified frailty index score (amFI) was calculated and amFI ≥2 classified as high frailty. Clinical outcomes, progression free survival (PFS) and overall survival (OS) were estimated.592 patients met inclusion criteria; amFI of 0, 1 and ≥ 2 was noted in 57%, 29%, and 14%, respectively. Patients with high frailty were less likely to be offered laparoscopic assessment for primary surgery (49% v. 43% v. 28% for amFI = 0, 1, and ≥ 2, p = 0.004), and more likely to have a Fagotti score ≥ 8 (58%, 48%, and 34%, p = 0.04). Only 17% of the high frailty cohort had primary tumor reductive surgery compared to 26% and 34% in patients with amFI = 1 and amFI = 0 (p = 0.02). Furthermore, patients with higher amFI were less likely to undergo any tumor reductive surgery (85% v. 74% v. 59%, p0.001). Postoperative complications were more frequent in patients with higher amFI (44% v. 56% v. 64%, p = 0.01). Death within thirty days of treatment initiation was significantly higher in patients with high frailty (0.4% v. 2% v. 9%, p = 0.005). In multivariate analysis, high frailty was associated with worse PFS (p = 0.02) and OS (p0.05).Postoperative morbidity, PFS, and OS were worse in patients with high frailty scores. Quantification of frailty may be useful for clinical decision making in patients with newly diagnosed advanced ovarian cancer.

Supplementary Data Word Document

Hematologic effects of daily metronomic (MET) nab-paclitaxel.

Delta-like ligand 4 (Dll4), one of the Notch ligands, is overexpressed in ovarian cancer, especially in tumors resistant to anti-VEGF therapy. Here, we examined the biologic effects of dual anti-Dll4 and anti-VEGF therapy in ovarian cancer models. Using Dll4-Fc blockade and anti-Dll4 antibodies (murine REGN1035 and human REGN421), we evaluated the biologic effects of Dll4 inhibition combined with aflibercept or chemotherapy in orthotopic mouse models of ovarian cancer. We also examined potential mechanisms by which dual Dll4 and VEGF targeting inhibit tumor growth using immunohistochemical staining for apoptosis and proliferation markers. Reverse-phase protein arrays were used to identify potential downstream targets of Dll4 blockade. Dual targeting of VEGF and Dll4 with murine REGN1035 showed superior antitumor effects in ovarian cancer models compared with either monotherapy. In the A2780 model, REGN1035 (targets murine Dll4) or REGN421 (targets human Dll4) reduced tumor weights by 62% and 82%, respectively; aflibercept alone reduced tumor weights by 90%. Greater therapeutic effects were observed for Dll4 blockade (REGN1035) combined with either aflibercept or docetaxel (P < 0.05 for the combination vs. aflibercept). The superior antitumor effects of REGN1035 and aflibercept were related to increased apoptosis in tumor cells compared with the monotherapy. We also found that GATA3 expression was significantly increased in tumor stroma from the mice treated with REGN1035 combined with docetaxel or aflibercept, suggesting an indirect effect of these combination treatments on the tumor stroma. These findings identify that dual targeting of Dll4 and VEGF is an attractive therapeutic approach. Mol Cancer Ther; 15(6); 1344–52. ©2016 AACR.

PTEN is known to be frequently mutated in uterine cancer and also dephosphorylates FAK. Here, we examined the impact of PTEN alterations on the response to treatment with a FAK inhibitor (GSK2256098). In vitro and in vivo therapeutic experiments were carried out using PTEN-mutated and PTEN-wild-type models of uterine cancer alone and in combination with chemotherapy. Treatment with GSK2256098 resulted in greater inhibition of pFAKY397 in PTEN-mutated (Ishikawa) than in PTEN-wild-type (Hec1A) cells. Ishikawa cells were more sensitive to GSK2256098 than the treated Hec1A cells. Ishikawa cells were transfected with a wild-type PTEN construct and pFAKY397 expression was unchanged after treatment with GSK2256098. Decreased cell viability and enhanced sensitivity to chemotherapy (paclitaxel and topotecan) in combination with GSK2256098 was observed in Ishikawa cells as compared with Hec1a cells. In the Ishikawa orthoptopic murine model, treatment with GSK2256098 resulted in lower tumor weights and fewer metastases than mice inoculated with Hec1A cells. Tumors treated with GSK2256098 had lower microvessel density (CD31), less cellular proliferation (Ki67), and higher apoptosis (TUNEL) rates in the Ishikawa model when compared with the Hec1a model. From a large cohort of evaluable patients, increased FAK and pFAKY397 expression levels were significantly related to poor overall survival. Moreover, PTEN levels were inversely related to pFAKY397 expression. These preclinical data demonstrate that PTEN-mutated uterine cancer responds better to FAK inhibition than does PTEN wild-type cancer. Therefore, PTEN could be a biomarker for predicting response to FAK-targeted therapy during clinical development. Mol Cancer Ther; 14(6); 1466–75. ©2015 AACR.

Mean mouse weights in orthotopic mouse models of uterine cancer.

Primate Hematology

A list of copy number alterations, rearrangements and short variants detected by Foundation Medicine NGS

Supplementary data

Table S2 shows data concerning the association between baseline characteristics and presence of BRCA reversion mutations in pretreatment circulating cell-free DNA

Supplementary Tables, Figures and Video legends, Tables 2,3,5 and all Supplementary Figures. Supplementary Table 2. Confirmation of cis configuration of BRCA1 primary and secondary mutations in case 4 by colony PCR. Supplementary Table 3. IC50 (mircoM) values of the PARPi and platinum drugs in parental OVCAR8 cell line and OVCAR8 RAD51C KO clone, and the fold change in IC50 values. Supplementary Table 5. Sequences of primers used for site-directed mutagenesis. Supplementary Figure 1. Foundation Medicine NGS analysis of the 12 cases with archival tissue and/or pre-treatment and post-progression biopsies. Supplementary Figure 2. Sanger sequencing trace of the primary and secondary BRCA1 mutations in cis configuration in case 4 post-progression biopsy sample. Supplementary Figure 3. In vitro response to PARP inhibitor therapy and platinum agents in RAD51C deficient cell lines, with primary or secondary mutations in RAD51C. Supplementary Figure 4. RAD51 foci formation in geminin positive cells deficient for RAD51C, complemented with primary or secondary mutations in RAD51C. Supplementary Figure 5. Diagram of HR reporter assay. Supplementary Figure 6. RAD51C expression in MCF10A cells and in yeast. Supplementary Figure 7. Analysis of serial sections by direct PCR sequencing approach of a post-progression biopsy containing multiple secondary mutations in RAD51C. Supplementary Figure 8. Molecular Dynamics Modeling of WT RAD51D protein and RAD51D protein with secondary mutation c.770_776delinsA, p.S257_R259delinsK. Supplementary Figure 9. In vitro response to PARP inhibitor therapy and cisplatin in RAD51D deficient CHO cell line, with primary or secondary mutation in RAD51D. Supplementary Figure 10. Examination of the parental PEO4 cell line, PEO4 cells with the homozygous frameshift RAD51D mutation (c.762_763del, D254E*fs72) in the same exon as the primary mutation and PEO4 cells with the homozygous secondary RAD51D mutation (c.770_776delinsA, S257_R259delinsK). Supplementary Figure 11. Modeling of tumor clonal fractions in the post-progression biopsy sample with germline RAD51C mutation and multiple secondary mutations.

Although progesterone receptor (PR)–targeted therapies are modestly active in patients with uterine cancer, their underlying molecular mechanisms are not well understood. The clinical use of such therapies is limited because of the lack of biomarkers that predict response to PR agonists (progestins) or PR antagonists (onapristone). Thus, understanding the underlying molecular mechanisms of action will provide an advance in developing novel combination therapies for cancer patients. Nuclear translocation of PR has been reported to be ligand-dependent or -independent. Here, we identified that onapristone, a PR antagonist, inhibited nuclear translocation of ligand-dependent or -independent (EGF) phospho-PR (S294), whereas trametinib inhibited nuclear translocation of EGF-induced phospho-PR (S294). Using orthotopic mouse models of uterine cancer, we demonstrated that the combination of onapristone and trametinib results in superior antitumor effects in uterine cancer models compared with either monotherapy. These synergistic effects are, in part, mediated through inhibiting the nuclear translocation of EGF-induced PR phosphorylation in uterine cancer cells. Targeting MAPK-dependent PR activation with onapristone and trametinib significantly inhibited tumor growth in preclinical uterine cancer models and is worthy of further clinical investigation. Mol Cancer Ther; 17(2); 464–73. ©2017 AACR.

EP-100 is a synthetic lytic peptide that specifically targets the gonadotropin-releasing hormone receptor on cancer cells. To extend the utility of EP-100, we aimed to identify effective combination therapies with EP-100 for ovarian cancer and explore potential mechanisms of this combination. A series of in vitro (MTT assay, immunoblot analysis, reverse-phase protein array, comet assay, and immunofluorescence staining) and in vivo experiments were carried out to determine the biological effects of EP-100 alone and in combination with standard-of-care drugs. EP-100 decreased the viability of ovarian cancer cells and reduced tumor growth in orthotopic mouse models. Of five standard drugs tested (cisplatin, paclitaxel, doxorubicin, topotecan, and olaparib), we found that the combination of EP-100 and olaparib was synergistic in ovarian cancer cell lines. Further experiments revealed that combined treatment of EP-100 and olaparib significantly increased the number of nuclear foci of phosphorylated histone H2AX. In addition, the extent of DNA damage was significantly increased after treatment with EP-100 and olaparib in comet assay. We performed reverse-phase protein array analyses and identified that the PI3K/AKT pathway was inhibited by EP-100, which we validated with in vitro experiments. In vivo experiment using the HeyA8 mouse model demonstrated that mice treated with EP-100 and olaparib had lower tumor weights (0.06 ± 0.13 g) than those treated with a vehicle (1.19 ± 1.09 g), EP-100 alone (0.62 ± 0.78 g), or olaparib alone (0.50 ± 0.63 g). Our findings indicate that combining EP-100 with olaparib is a promising therapeutic strategy for ovarian cancer.

Primate CK and LDH

There is growing recognition of the important role of metronomic chemotherapy in cancer treatment. On the basis of their unique antiangiogenic effects, we tested the efficacy of nab-paclitaxel, which stimulates thrombospondin-1, and topotecan, which inhibits hypoxia-inducible factor 1-α, at metronomic dosing for the treatment of ovarian carcinoma. In vitro and in vivo SKOV3ip1, HeyA8, and HeyA8-MDR (taxane-resistant) orthotopic models were used to examine the effects of metronomic nab-paclitaxel and metronomic topotecan. We examined cell proliferation (Ki-67), apoptosis (cleaved caspase-3), and angiogenesis (microvessel density, MVD) in tumors obtained at necropsy. In vivo therapy experiments demonstrated treatment with metronomic nab-paclitaxel alone and in combination with metronomic topotecan resulted in significant reductions in tumor weight (62% in the SKOV3ip1 model, P < 0.01 and 96% in the HeyA8 model, P < 0.03) compared with vehicle (P < 0.01). In the HeyA8-MDR model, metronomic monotherapy with either cytotoxic agent had modest effects on tumor growth, but combination therapy decreased tumor burden by 61% compared with vehicle (P < 0.03). The greatest reduction in MVD (P < 0.05) and proliferation was seen in combination metronomic therapy groups. Combination metronomic therapy resulted in prolonged overall survival in vivo compared with other groups (P < 0.001). Tube formation was significantly inhibited in RF-24 endothelial cells exposed to media conditioned with metronomic nab-paclitaxel alone and media conditioned with combination metronomic nab-paclitaxel and metronomic topotecan. The combination of metronomic nab-paclitaxel and metronomic topotecan offers a novel, highly effective therapeutic approach for ovarian carcinoma that merits further clinical development. Mol Cancer Ther; 14(12); 2677–86. ©2015 AACR.

CRM1 inhibitors have demonstrated antitumor effects in ovarian and other cancers; however, rational combinations are largely unexplored. We performed a high-throughput drug library screen to identify drugs that might combine well with selinexor in ovarian cancer. Next, we tested the combination of selinexor with the top hit from the drug screen in vitro and in vivo. Finally, we assessed for mechanisms underlying the identified synergy using reverse phase protein arrays (RPPA). The drug library screen assessing 688 drugs identified olaparib (a PARP inhibitor) as the most synergistic combination with selinexor. Synergy was further demonstrated by MTT assays. In the A2780luc ip1 mouse model, the combination of selinexor and olaparib yielded significantly lower tumor weight and fewer tumor nodules compared with the control group (P < 0.04 and P < 0.03). In the OVCAR5 mouse model, the combination yielded significantly fewer nodules (P = 0.006) and markedly lower tumor weight compared with the control group (P = 0.059). RPPA analysis indicated decreased expression of DNA damage repair proteins and increased expression of tumor suppressor proteins in the combination treatment group. Collectively, our preclinical findings indicate that combination with selinexor to expand the utility and efficacy of PARP inhibitors in ovarian cancer warrants further exploration.

Supplementary Figure 1. Representative OVCAR3-inoculated mice after aflibercept, Dll4-Fc, and the combination. Supplementary Figure 2. In vivo study of Dll4 blockade, aflibercept, and docetaxel, alone and in combinations, in A2780 and HeyA8 models. Supplementary Figure 3. Immunohistochemical staining of CX3CL1 in the tumor stroma of mice inoculated with A2780 ovarian cancer cells. Supplementary Figure 4. Quantitative real-time PCR of CX3CL1 expression in A2780 cells cultured with primary dendritic cells isolated from C57/BL6 mice. Supplementary Figure 5. ELISA analysis of plasma IFN-gamma in the mice inoculated with A2780 ovarian cancer cells. Supplementary Figure 6. RT-PCR analysis of E-cadherin expression in tumors (tumor cells and stroma) obtained from A2780 mice treated with a single agent or a combination.

Supplementary Figure 1. In vitro effects of PR silencing on onapristone sensitivity in PR-high-expressing uterine cancer cell line (Ishikawa). Supplementary Figure 2. In vitro effects of onapristone and trametinib in PR-weak-expressing uterine cancer cell line (SKUT2). Supplementary Figure 3. In vivo effects of therapy with onapristone in uterine cancer model. Supplementary Figure 4. In vivo effect of onapristone, trametinib, and the combination of both drugs on total PR expression in the ISHIKAWA model.

Primate coagulation testing

Supplementary Table S1. The IC50s of EP-100 against ovarian cancer cells after 4-, 24-, and 72-h treatment.

A key resistance mechanism to platinum-based chemotherapies and PARP inhibitors in BRCA-mutant cancers is the acquisition of BRCA reversion mutations that restore protein function. To estimate the prevalence of BRCA reversion mutations in high-grade ovarian carcinoma (HGOC), we performed targeted next-generation sequencing of circulating cell-free DNA (cfDNA) extracted from pretreatment and postprogression plasma in patients with deleterious germline or somatic BRCA mutations treated with the PARP inhibitor rucaparib. BRCA reversion mutations were identified in pretreatment cfDNA from 18% (2/11) of platinum-refractory and 13% (5/38) of platinum-resistant cancers, compared with 2% (1/48) of platinum-sensitive cancers (P = 0.049). Patients without BRCA reversion mutations detected in pretreatment cfDNA had significantly longer rucaparib progression-free survival than those with reversion mutations (median, 9.0 vs. 1.8 months; HR, 0.12; P < 0.0001). To study acquired resistance, we sequenced 78 postprogression cfDNA, identifying eight additional patients with BRCA reversion mutations not found in pretreatment cfDNA.Significance:BRCA reversion mutations are detected in cfDNA from platinum-resistant or platinum-refractory HGOC and are associated with decreased clinical benefit from rucaparib treatment. Sequencing of cfDNA can detect multiple BRCA reversion mutations, highlighting the ability to capture multiclonal heterogeneity.This article is highlighted in the In This Issue feature, p. 151

Revised Supplementary Data Demonstrating All Supplementary Figures 1-6 and Supplementary Table 1

Primate electrolytes and liver function tests

Hematologic effects of daily metronomic (MET) nab-paclitaxel. White blood count (WBC), hemoglobin, and platelet number were assessed from tumor bearing mice after one, two, and three weeks of treatment.

Here, we examined the role of EP-100 [luteinizing hormone-releasing hormone (LHRH) ligand joined to a lytic peptide], improving the efficacy of immune checkpoint blockade. LHRH-R–positive murine ovarian cancer cells (ID8, IG10, IF5, and 2C12) were sensitive to EP-100 and were specifically killed at low micromolar levels through LHRH-R. EP-100 increased PD-L1 levels on murine ovarian cancer cells. In vivo syngeneic mouse models (ID8 and IG10) demonstrated that single-agent EP-100 reduced tumor volume, tumor weight, and ascites volume. The greatest reductions in tumor and ascites volume were observed with the combination of EP-100 with an anti–PD-L1 antibody. Immune profiling analysis showed that the population of CD8+ T cells, natural killer cells, dendritic cells, and macrophages were significantly increased in tumor and ascitic fluid samples treated with anti–PD-L1, EP-100, and the combination. However, monocytic myeloid suppressor cells, B cells, and regulatory T cells were decreased in tumors treated with anti–PD-L1, EP-100, or the combination. In vitro cytokine arrays revealed that EP-100 induced IL1α, IL33, CCL20, VEGF, and Low-density lipoprotein receptor (LDLR) secretion. Of these, we validated increasing IL33 levels following EP-100 treatment in vitro and in vivo; we determined the specific biological role of CD8+ T-cell activation with IL33 gene silencing using siRNA and Cas9-CRISPR approaches. In addition, we found that CD8+ T cells expressed very low level of LHRH-R and were not affected by EP-100. Taken together, EP-100 treatment had a substantial antitumor efficacy, particularly in combination with an anti–PD-L1 antibody. These results warrant further clinical development of this combination.

Molecular Dynamics Modeling of WT RAD51D monofilament interaction with dsDNA.

To address the need for efficient and biocompatible delivery systems for systemic siRNA delivery, we developed 1,2-Dioleoyl-sn-Glycero-3-Phosphatidylcholine (DOPC) nanoliposomal EphA2-targeted therapeutic (EPHARNA). Here, we performed safety studies of EPHARNA in murine and primate models. Single dosing of EPHARNA was tested at 5 concentrations in mice (N = 15 per group) and groups were sacrificed on days 1, 14, and 28 for evaluation of clinical pathology and organ toxicity. Multiple dosing of EPHARNA was tested in mice and Rhesus macaques twice weekly at two dose levels in each model. Possible effects on hematologic parameters, serum chemistry, coagulation, and organ toxicity were assessed. Following single-dose EPHARNA administration to mice, no gross pathologic or dose-related microscopic findings were observed in either the acute (24 hours) or recovery (14 and 28 days) phases. The no-observed-adverse-effect level (NOAEL) for EPHARNA is considered >225 μg/kg when administered as a single injection intravenously in CD-1 mice. With twice weekly injection, EPHARNA appeared to stimulate a mild to moderate inflammatory response in a dose-related fashion. There appeared to be a mild hemolytic reaction in the female mice. In Rhesus macaques, minimal to moderate infiltration of mononuclear cells was found in some organs including the gastrointestinal tract, heart, and kidney. No differences attributed to EPHARNA were observed. These results demonstrate that EPHARNA is well tolerated at all doses tested. These data, combined with previously published in vivo validation studies, have led to an ongoing first-in-human phase I clinical trial (NCT01591356). Mol Cancer Ther; 16(6); 1114–23. ©2017 AACR.

Copy number estimation by SNP array in the archival tumor tissue of the patient identified to have a germline RAD51C mutation (c.577C