High grade adverse event reporting and enrolment in gynecologic oncology clinical trials.

OBJECTIVE: The primary objective is to assess factors associated with treatment related high grade (CTCAE grade ≥ 3) adverse event (AE) reporting among participants in gynecologic oncology clinical trials. METHODS: All AEs recorded in the Princess Margaret Clinical Trial adverse event database between 01/2016 and 12/2018 were evaluated. Gynecologic oncology clinical trials assessing systemic therapy were included. Inferential statistics on risk factors of related grade ≥ 3 adverse event reporting and GEE logistic models with Odds Ratios (OR) were performed. Multivariable analysis adjusting for age, clinical trial phase, sponsor, and therapy type. RESULTS: The gynecology cancer clinical trials accrued 317 unique patients (359 nested on trials) in 42 systemic therapy trials. In the period, 17,175 related AEs were reported in the gynecological cancer trials, 7.4% were grade ≥ 3. On multivariable analysis, no odds differences of grade ≥ 3 related AEs were detected according to study phase. Patients in immunotherapy clinical trials had lower odds of related grade ≥ 3 AEs than patients on targeted or other therapy (adjusted OR [aOR] 0.43; 95% CI 0.24-0.75). There was greater odds of related grade ≥ 3 AEs in clinical trials assessing combination vs single therapeutics (aOR 2.26, 95% CI 1.34-3.80). Patients aged ≥65 (aOR 1.77; 95% CI 1.08-2.89) had greater odds of related grade ≥ 3 AEs than patients aged 50 to 65 years. When compared to other disease sites, the odds of having a grade  ≥ 3 related AE reported in gynecology clinical trials was no different. CONCLUSIONS: In this cohort, factors influencing the odds of related grade ≥ 3 AE reporting in gynecologic trials included type of therapy and age. The study phase did not correlate with odds of high-grade AE reporting.

Electronic malignant bowel obstruction symptom monitoring smartphone application for patients with gynecologic cancers.

OBJECTIVES: Implementation of an interprofessional program at Princess Margaret Cancer Centre, including nurse-led proactive calls to support patients with gynecologic cancers with malignant bowel obstruction, demonstrated improved outcomes compared with historical controls. The aim of the study was to convert the proactive calls into an electronic monitoring program to assess it's feasibility and scalability in patients with gynecologic cancers with or at risk of malignant bowel obstruction. METHODS: 'My Bowels on Track' smartphone application included weekly/biweekly electronic patient-reported outcomes (PROs), educational materials, and a secure messaging system. Based on PRO answers, an alerting system flagged patients with symptoms or uncompleted PROs. Nurses tracked and called patients on receiving clinical or compliance alerts. The primary objective was to assess adherence (≥70% PRO completion per patient considered an adherent patient) in the first 2 months on the program. A secondary objective was to assess the positive predictive value (PPV) of the alerts to trigger recommendations. RESULTS: Forty patients were enrolled between August 2021 and September 2022. Median age was 64.5 years (range 29-79 years). Primary diagnosis was ovarian (75%), endometrial (17.5%), or cervical (7.5%) cancer, and 92.5% of patients were receiving systemic therapy. Median duration on the program was 55 days (range 8-121 days). The 2-month adherence was 65% (95% CI 50% to 80%) and the overall adherence was 60% (95% CI 43% to 75%). Sixty-five symptom-related alerts (75% severe, 25% moderate) were reported in 60% (24/40) of patients. There were 59 recommendations triggered by the alerts. The PPV of the alerts to trigger actions was 72% (95% CI 58% to 82%). CONCLUSIONS: This pilot electronic malignant bowel obstruction monitoring program with real-time PRO assessment was feasible, and 65% of participants were adherent during the first 2 months on the program. The PRO response-based alerting system flagged concerning symptoms in 60% of participants, with a PPV of 72% to trigger nurse-led actions and/or management recommendations. TRIAL REGISTRATION NUMBER: NCT03260647.

Adavosertib plus gemcitabine for platinum-resistant or platinum-refractory recurrent ovarian cancer: a double-blind, randomised, placebo-controlled, phase 2 trial.

BACKGROUND: The Wee1 (WEE1hu) inhibitor adavosertib and gemcitabine have shown preclinical synergy and promising activity in early phase clinical trials. We aimed to determine the efficacy of this combination in patients with ovarian cancer. METHODS: In this double-blind, randomised, placebo-controlled, phase 2 trial, women with measurable recurrent platinum-resistant or platinum-refractory high-grade serous ovarian cancer were recruited from 11 academic centres in the USA and Canada. Women were eligible if they were aged 18 years or older, had an Eastern Cooperative Oncology Group performance status of 0-2, a life expectancy of more than 3 months, and normal organ and marrow function. Women with ovarian cancer of non-high-grade serous histology were eligible for enrolment in a non-randomised exploratory cohort. Eligible participants with high-grade serous ovarian cancer were randomly assigned (2:1), using block randomisation (block size of three and six) and no stratification, to receive intravenous gemcitabine (1000 mg/m2 on days 1, 8, and 15) with either oral adavosertib (175 mg) or identical placebo once daily on days 1, 2, 8, 9, 15, and 16, in 28-day cycles until disease progression or unacceptable toxicity. Patients and the team caring for each patient were masked to treatment assignment. The primary endpoint was progression-free survival. The safety and efficacy analysis population comprised all patients who received at least one dose of treatment. The trial is registered with ClinicalTrials.gov, NCT02151292, and is closed to accrual. FINDINGS: Between Sept 22, 2014, and May 30, 2018, 124 women were enrolled, of whom 99 had high-grade serous ovarian cancer and were randomly assigned to adavosertib plus gemcitabine (65 [66%]) or placebo plus gemcitabine (34 [34%]). 25 women with non-high-grade serous ovarian cancer were enrolled in the exploratory cohort. After randomisation, five patients with high-grade serous ovarian cancer were found to be ineligible (four in the experimental group and one in the control group) and did not receive treatment. Median age for all treated patients (n=119) was 62 years (IQR 54-67). Progression-free survival was longer with adavosertib plus gemcitabine (median 4·6 months [95% CI 3·6-6·4] with adavosertib plus gemcitabine vs 3·0 months [1·8-3·8] with placebo plus gemcitabine; hazard ratio 0·55 [95% CI 0·35-0·90]; log-rank p=0·015). The most frequent grade 3 or worse adverse events were haematological (neutropenia in 38 [62%] of 61 participants in the adavosertib plus gemcitabine group vs ten [30%] of 33 in the placebo plus gemcitabine group; thrombocytopenia in 19 [31%] of 61 in the adavosertib plus gemcitabine group vs two [6%] of 33 in the placebo plus gemcitabine group). There were no treatment-related deaths; two patients (one in each group in the high-grade serous ovarian cancer cohort) died while on study medication (from sepsis in the experimental group and from disease progression in the control group). INTERPRETATION: The observed clinical efficacy of a Wee1 inhibitor combined with gemcitabine supports ongoing assessment of DNA damage response drugs in high-grade serous ovarian cancer, a TP53-mutated tumour type with high replication stress. This therapeutic approach might be applicable to other tumour types with high replication stress; larger confirmatory studies are required. FUNDING: US National Cancer Institute Cancer Therapy Evaluation Program, Ontario Institute for Cancer Research, US Department of Defense, Princess Margaret Cancer Foundation, and AstraZeneca.

Patient self-reporting of tolerability using PRO-CTCAE in a randomized double-blind, placebo-controlled phase II trial comparing gemcitabine in combination with adavosertib or placebo in patients with platinum resistant or refractory epithelial ovarian carcinoma.

BACKGROUND: A double-blind, randomized, placebo-controlled, phase 2 trial assessed gemcitabine in combination with the wee1 inhibitor adavosertib or placebo in platinum resistant or refractory high grade serous ovarian cancer (HGSOC), demonstrating improved progression free and overall survival favouring the adavosertib/gemcitabine arm. An exploratory objective of the study included the PRO-CTCAE assessment, to capture self-reporting of frequency, severity and/or interference of symptomatic adverse events (syAEs). METHODS: PRO-CTCAE items at baseline, days 1 and 15 of each cycle and off treatment, were completed in two centres, with the objective of characterizing syAEs in the first three months of therapy. The maximum post-baseline score proportion for each syAE was tabulated per patient. The 12-week area under the curve (AUC12w) as a measure of syAE over-time and incremental AUC12w (iAUC12w) for adjustment to baseline syAEs. RESULTS: Sixty-one patients were approached for PRO-CTCAE surveys and 55 were evaluable. Among patients with HGSOC, 28 received gemcitabine/adavosertib (arm A) and 19 gemcitabine/placebo (arm B). Survey completion rates were high. The proportion of participants with positive (≥1) PRO-CTCAE scores was higher for difficulty swallowing with gemcitabine/adavosertib (arm A 35.7% vs arm B 5.3%, p = 0.02). The high score (≥3) syAEs showed more frequent diarrhea with gemcitabine/adavosertib (arm A 25% vs arm B 0%, p = 0.03). The proportions of worsening syAEs over time were higher in patients receiving gemcitabine/adavosertib for difficulty swallowing (arm A 35.7% vs arm B 5.3%; p = 0.03) and fatigue severity (arm A 71.43% vs arm B 42.1%; p = 0.04). CONCLUSIONS: The longitudinal assessment of patient self-reported tolerability showed greater difficulty swallowing and fatigue severity in patients receiving gemcitabine/adavosertib, compared to gemcitabine/placebo. PRO-CTCAE provides complementary and objective assessment of drug tolerability from a patient's perspective.

Research biopsies in patients with gynecologic cancers: patient-reported outcomes, perceptions, and preferences.

BACKGROUND: Despite the growing integration of mandatory biopsies for correlative endpoints within oncology clinical trials, there are sparse data on patient-reported outcomes, perceptions, and preferences. OBJECTIVE: This study aimed to prospectively assess the impact of research biopsies on the quality of life in patients with gynecologic cancer, evaluate patient-reported outcomes, and determine factors associated with patients' willingness to undergo sequential biopsies. STUDY DESIGN: We conducted a prospective study in patients with gynecologic malignancies undergoing research biopsies between 2015 and 2019 at Princess Margaret Cancer Centre (ClinicalTrials.gov Identifier: NCT02334761). Here, we report the results of the paper-based surveys performed before and 1 week after biopsy. Although the questionnaires each assessed the impact of anxiety using a modified version of the Hospital Anxiety and Depression Scale, the postbiopsy questionnaire specifically assessed the likelihood of future biopsies, postbiopsy symptoms, complications, and perceptions. RESULTS: A total of 129 patients were enrolled, of which 91 (70.5%) completed at least 1 questionnaire. These patients had either ovarian (89%; 81 of 91) or endometrial cancer (11%; 10 of 91). Of all biopsies taken, 75% were from the abdomen or pelvis (67 of 89). There was 1 clinician-reported complication, a perihepatic hematoma (1%). Pain during the biopsy and physical discomfort were experienced by 60.3% (41 of 68) and 61.8% (42 of 68), respectively. Embarrassment and loss of dignity were experienced by 13.2% (9 of 68) and 11.8% (8 of 68), respectively. Although the mean Hospital Anxiety and Depression Scale score was in the normal range before and after biopsy, there was a significant decline in the total score after the biopsy (prebiopsy, 5.3 [standard deviation, 4.7] vs postbiopsy, 3.7 [standard deviation, 4.5]; P=.005); 84% of subjects (58 of 69) stated that they would definitely or likely consent to another biopsy. There was no impact on patients' willingness for future biopsies based on Eastern Cooperative Oncology Group status, biopsy site, age, number of cores, and pain during the biopsy; however, subjects who reported feeling physically uncomfortable (odds ratio, 0.14; P=.005), embarrassed (odds ratio, 0.03; P=.004) or experienced loss of dignity (odds ratio, 0.05; P=.01) during the biopsy and those who experienced flu-like symptoms (odds ratio, 0.2; P=.018) or felt feverish (odds ratio, 0.2; P=.035) 1 week after biopsy, were less likely to undergo a sequential biopsy. Similarly, those with higher Hospital Anxiety and Depression Scale scores before biopsy (odds ratio, 0.83; P=.008) and after biopsy (odds ratio, 0.8; P=.003) were less likely to consent for another biopsy. CONCLUSION: Research biopsies were generally well accepted. Most patients (83%) were willing to undergo serial biopsies if necessary. Addressing the potentially modifiable psychosocial aspects of the procedure may improve the experience with research biopsies for patients with gynecologic cancers.

Biomarkers of outcome to weekly paclitaxel in epithelial ovarian cancer.

OBJECTIVE: We sought to evaluate the role of intrinsic chromosomal aberrations in determining favorable outcome to weekly paclitaxel (WP) in patients with epithelial ovarian cancer (EOC). METHODS: We evaluated the common genomic aberrations of two patients with EOC and exceptional WP response in the GENIUS study (NCT03740503). We then searched for potential markers of unusual outcomes to WP in a validation cohort. We performed shallow whole genome sequencing (sWGS) in the tumor tissue of women with EOC considered as short-responders (SR; progression with ≤3 cycles) and long-responders (LR; response at ≥8 cycles) to WP monotherapy. RESULTS: We identified two women with exceptional response to WP, lasting over four years, who shared chromosome 8 gain as a common genomic aberration. In order to validate our findings, we reviewed 188 patients with EOC treated with WP and selected 61 women (39 SR, 22 LR) with unusual responses. By sWGS, there was no differential alterations in the copy number changes in chromosome 8, or in genes related to angiogenesis, tubulin superfamily, cell-cycle, apoptosis and paclitaxel metabolism or transportation pathways. Amongst the LR group, we identified six exceptionally long responders (ExLR), with responses lasting over a year. In an exploratory analysis, there was increased amplification of angiogenesis (VEGFB, MMP9), tubulin superfamily (TSC2) and apoptosis related genes (BCL2L1, BAD) in ExLR compared to SR. We identified one patient with a complete response to WP for over 7 years. Molecular profiling identified unique amplifications in interleukin related genes (CXCR1, CXCR2, IL1A, IL1B), not detected in other patients. CONCLUSION: Intrinsic tumor pathways may impact outcome with weekly paclitaxel monotherapy and further investigations are required.

TP53 mutations in high grade serous ovarian cancer and impact on clinical outcomes: a comparison of next generation sequencing and bioinformatics analyses.

OBJECTIVE: Mutations in TP53 are found in the majority of high grade serous ovarian cancers, leading to gain of function or loss of function of its protein product, p53, involved in oncogenesis. There have been conflicting reports as to the impact of the type of these on prognosis. We aim to further elucidate this relationship in our cohort of patients. METHODS: 229 patients with high grade serous ovarian cancer underwent tumor profiling through an institutional molecular screening program with targeted next generation sequencing. TP53 mutations were classified using methods previously described in the literature. Immunohistochemistry on formalin-fixed paraffin embedded tissue was used to assess for TP53 mutation. Using divisive hierarchal clustering, we generated patient clusters with similar clinicopathologic characteristics to investigate differences in outcomes. RESULTS: Six different classification schemes of TP53 mutations were studied. These did not show an association with first platinum-free interval or overall survival. Next generation sequencing reliably predicted mutation in 80% of cases, similar to the proportion detected by immunohistochemistry. Divisive hierarchical clustering generated four main clusters, with cluster 3 having a significantly worse prognosis (p<0.0001; log-rank test). This cluster had a higher concentration of gain of function mutations and these patients were less likely to have undergone optimal debulking surgery. CONCLUSIONS: Different classifications of TP53 mutations did not show an impact on outcomes in this study. Immunohistochemistry was a good predictor for TP53 mutation. Cluster analysis showed that a subgroup of patients with gain of function mutations (cluster 3) had a worse prognosis.

Comparison of Practice Guidelines, BRCAPRO, and Genetic Counselor Estimates to Identify Germline BRCA1 and BRCA2 Mutations in Pancreatic Cancer.

Germline BRCA1 and BRCA2 (BRCA) mutation carriers with pancreatic ductal adenocarcinoma (PDAC) may benefit from precision therapies and their relatives should undergo tailored cancer prevention. In this study, we compared strategies to identify BRCA carriers with PDAC. Incident cases of PDAC were prospectively recruited for BRCA sequencing. Probands were evaluated using the National Comprehensive Cancer Network (NCCN) and the Ontario Ministry of Health and Long-Term Care (MOHLTC) guidelines. The probability of each proband carrying a mutation was estimated by surveying genetic counselors and using BRCAPRO. BRCA mutations were detected in 22/484 (4.5%) probands. 152/484 (31.2%) and 16/484 (3.3%) probands met the NCCN and MOHLTC guidelines, respectively. The NCCN guidelines had higher sensitivity than the MOHLTC guidelines (0.864 versus 0.227, P < 0.001) but lower specificity (0.712 versus 0.976, P < 0.001). One hundred and nineteen genetic counselors completed the survey. Discrimination was similar between genetic counselors and BRCAPRO (area-under-the-curve: 0.755 and 0.775, respectively, P = 0.702). Genetic counselors generally overestimated (P = 0.008), whereas BRCAPRO severely underestimated (P < 0.001), the probability that each proband carried a mutation. Our results indicate that the NCCN guidelines and genetic counselors accurately identify BRCA mutations in PDAC, while the MOHLTC guidelines and BRCAPRO should be updated to account for the association between BRCA and PDAC.

FOLFIRINOX for advanced pancreatic cancer: the Princess Margaret Cancer Centre experience.

BACKGROUND: FOLFIRINOX has been shown to significantly increase both overall survival (OS) and progression-free survival (PFS) in metastatic pancreas cancer. There is limited data regarding the treatment of locally advanced pancreatic cancer. We present a retrospective study of patients with both locally advanced and metastatic pancreas cancer using FOLFIRINOX as first-line therapy in our centre. METHODS: This is a retrospective review of patients treated with FOLFIRINOX for pancreatic cancer at Princess Margaret Cancer Centre, between December 2011 and July 2014. The primary objective was to evaluate the efficacy and safety of FOLFIRINOX when used with dose modifications. RESULTS: One hundred two patients were identified; 66 metastatic and 36 locally advanced. Sixty-eight per cent of patients initiated treatment with a dose reduction. The median (95% CI) OS in the metastatic group was 13.1 (6.3-16.1) months with full dose and 12.9 (10.3-30.1) months with modified dose. The median (95% CI) OS in the locally advanced group was 11.1 (6.1-not reached) months with full dose and 23 (not reached-not reached) months with modified dose. The median (95% CI) PFS in the metastatic group was 6.2 (4.9-15.2) months with full dose and 8.7 (5.7-12.9) months with modified dose. The median (95% CI) PFS in the locally advanced group was 11.1 (3.1-not reached) months with full dose and 10.4 (6.8-not reached) months with modified dose. Grade 3/4 haematologic adverse events were observed in 43% of patients. Grade 3/4 non-haematologic adverse events were observed in 28% of patients. Patient well-being significantly improved from baseline to cycle 4 (P=0.002). CONCLUSIONS: Efficacy was achievable with dose-modified FOLFIRINOX in daily setting. The safety of FOLFIRINOX remains a concern with a high rate of grades 3 and 4 neutropaenia despite dose reduction.

Rerouting the GPS Directing Immunotherapy in Endometrial Cancer.

Mismatch repair (MMR) status alone is insufficient to guide the use of PD-(L)1 monotherapy in patients with endometrial cancer. Additional biomarkers, including tumor mutational burden and combined positive score, may help to identify patients with MMR-proficient tumors with a high probability of benefit from PD-(L)1 monotherapy, and those with MMR-deficient tumors who might require combination strategies. See related article by Oaknin et al., p. 4564.

A rapidly evolving landscape: immune checkpoint inhibitors in pretreated metastatic endometrial cancer.

Background and objectives: Endometrial cancer is a common malignancy and recurrences can be fatal. Although platinum-pretreated endometrial tumors are commonly treated with anthracyclines and taxanes, there is no current standard of care. Both immune checkpoint inhibitors (ICIs) and tyrosine kinase inhibitors (TKIs) have been extensively assessed in this setting, including tumors selected for DNA mismatch repair (MMR)/microsatellite instability (MSI) and programmed death-ligand 1 expression status. This review will provide evidence-based guidance on use of ICIs alone or in combination with TKIs in patients with pretreated advanced, persistent, or recurrent metastatic endometrial cancer. Data sources and methods: Randomized phase II-III trials in unselected populations pretreated, recurrent, or metastatic endometrial cancer and phase I-II trials in biomarker selected populations were identified from PubMed as well as conference proceedings using the key search terms 'immune checkpoint inhibitors', 'endometrial cancer', and 'advanced'. Results: A total of nine eligible studies were identified assessing ICI monotherapy for biomarker-selected or ICI plus TKI combinations and a dual ICI regimen for biomarker-unselected patients with pretreated recurrent or metastatic endometrial cancer. In MMR/MSI-selected tumors, five phase I/II studies evaluated ICI monotherapy indicating benefit in these patients. Only the phase III KEYNOTE-775 trial reported a statistically significant overall survival improvement for the combination of pembrolizumab plus lenvatinib compared with docetaxel or paclitaxel regardless of MMR/MSI status. Conclusions: Pembrolizumab plus lenvatinib is indicated for patients with unselected pretreated metastatic endometrial cancer and pembrolizumab monotherapy is a preferred option for patients with MMRd/MSI-H tumors.

FBXW7-loss Sensitizes Cells to ATR Inhibition Through Induced Mitotic Catastrophe.

FBXW7 is a commonly mutated tumor suppressor gene that functions to regulate numerous oncogenes involved in cell-cycle regulation. Genome-wide CRISPR fitness screens identified a signature of DNA repair and DNA damage response genes as required for the growth of FBXW7-knockout cells. Guided by these findings, we show that FBXW7-mutant cells have high levels of replication stress, which results in a genotype-specific vulnerability to inhibition of the ATR signaling pathway, as these mutant cells become heavily reliant on a robust S-G2 checkpoint. ATR inhibition induces an accelerated S-phase, leading to mitotic catastrophe and cell death caused by the high replication stress present in FBXW7-/- cells. In addition, we provide evidence in cell and organoid studies, and mining of publicly available high-throughput drug screening efforts, that this genotype-specific vulnerability extends to multiple types of cancer, providing a rational means of identifying responsive patients for targeted therapy. SIGNIFICANCE: We have elucidated the synthetic lethal interactions between FBXW7 mutation and DNA damage response genes, and highlighted the potential of ATR inhibitors as targeted therapies for cancers harboring FBXW7 alterations.

Clinical outcome and biomarker assessments of a multi-centre phase II trial assessing niraparib with or without dostarlimab in recurrent endometrial carcinoma.

This multi-centre, non-randomized, open-label, phase II trial (NCT03016338), assessed niraparib monotherapy (cohort 1, C1), or niraparib and dostarlimab (cohort 2, C2) in patients with recurrent serous or endometrioid endometrial carcinoma. The primary endpoint was clinical benefit rate (CBR), with ≥5/22 overall considered of interest. Secondary outcomes were safety, objective response rate (ORR), duration of response, progression free survival and overall survival. Translational research was an exploratory outcome. Potential biomarkers were evaluated in archival tissue by immunohistochemistry and next generation sequencing panel. In C1, 25 patients were enrolled, and CBR was 20% (95% CI: 9-39) with median clinical benefit duration of 5.3 months. The ORR was 4% (95% CI: 0-20). In C2, 22 patients were enrolled, and the CBR was 31.8% (95% CI: 16-53) with median clinical benefit duration of 6.8 months. The ORR was 14% (95% CI: 3-35). No new safety signals were detected. No significant association was detected between clinical benefit and IHC markers (PTEN, p53, MMR, PD-L1), or molecular profiling (PTEN, TP53, homologous recombination repair genes). In conclusion, niraparib monotherapy did not meet the efficacy threshold. Niraparib in combination with dostarlimab showed modest activity.

Identifying Mechanisms of Resistance by Circulating Tumor DNA in EVOLVE, a Phase II Trial of Cediranib Plus Olaparib for Ovarian Cancer at Time of PARP Inhibitor Progression.

PURPOSE: To evaluate the use of blood cell-free DNA (cfDNA) to identify emerging mechanisms of resistance to PARP inhibitors (PARPi) in high-grade serous ovarian cancer (HGSOC). EXPERIMENTAL DESIGN: We used targeted sequencing (TS) to analyze 78 longitudinal cfDNA samples collected from 30 patients with HGSOC enrolled in a phase II clinical trial evaluating cediranib (VEGF inhibitor) plus olaparib (PARPi) after progression on PARPi alone. cfDNA was collected at baseline, before treatment cycle 2, and at end of treatment. These were compared with whole-exome sequencing (WES) of baseline tumor tissues. RESULTS: At baseline (time of initial PARPi progression), cfDNA tumor fractions were 0.2% to 67% (median, 3.25%), and patients with high ctDNA levels (>15%) had a higher tumor burden (sum of target lesions; P = 0.043). Across all timepoints, cfDNA detected 74.4% of mutations known from prior tumor WES, including three of five expected BRCA1/2 reversion mutations. In addition, cfDNA identified 10 novel mutations not detected by WES, including seven TP53 mutations annotated as pathogenic by ClinVar. cfDNA fragmentation analysis attributed five of these novel TP53 mutations to clonal hematopoiesis of indeterminate potential (CHIP). At baseline, samples with significant differences in mutant fragment size distribution had shorter time to progression (P = 0.001). CONCLUSIONS: Longitudinal testing of cfDNA by TS provides a noninvasive tool for detection of tumor-derived mutations and mechanisms of PARPi resistance that may aid in directing patients to appropriate therapeutic strategies. With cfDNA fragmentation analyses, CHIP was identified in several patients and warrants further investigation.

Phase II study of cytarabine in men with docetaxel-refractory, castration-resistant prostate cancer with evaluation of TMPRSS2-ERG and SPINK1 as serum biomarkers.

UNLABELLED: What's known on the subject? and What does the study add? To date, there has been limited impetus to examine the use of cytarabine in prostate cancer. We presented preliminary laboratory data to suggest its utility in the castration refractory prostate cancer (CRPC) population which, combined with a previous case report, suggested it may have hitherto unrecognized utility in this setting. Embedded in this study was peripheral blood sampling for TMPRSS2-ERG and SPINK1, two genes that are believed to define prostate cancer genotypes, to assess their utility as biomarkers This study suggests that at the delivered doses, cytarabine has limited efficacy and significant myelotoxicity suggesting, it does not have a role in the treatment of docetaxel-refractory CRPC. The presence of serum TMPRSS2-ERG and SPINK1 mRNA biomarkers recovered from blood suggest that their analysis is worthy of further study. OBJECTIVES: To run a phase II clinical trial of cytarabine in men with docetaxel-refractory, castration-resistant prostate cancer (CRPC), based on evidence that cytarabine might be effective in men with abnormalities of ERG oncogenes. To measure mRNA levels of prostate cancer-related genes in blood as biomarkers. PATIENTS AND METHODS: Ten of a planned maximum of 30 men received i.v. cytarabine at doses of 0.25-1g/m(2) at 21-day intervals. The primary endpoint was prostate-specific antigen (PSA) response. Archival tumour samples were assessed by fluorescence in-situ hybridization for TMPRSS2:ERG translocation, and by immunohistochemistry for serine peptidase inhibitor Kazal type 1 (SPINK1). Blood was processed for mRNA quantification of TMPRSS2:ERG (exon1:exon4), SPINK1 and PSA. RESULTS: A PSA response was not observed in any patient. The trial was stopped at the end of stage 1 of a modified Flemming design. The median number of cycles administered was 3. Grade 3-4 haematological toxicity was common. Five patients were subsequently excluded from the study for toxicity, and five for disease progression. Analysis of whole blood mRNA for T1:E4 translocation in TMPRSS2:ERG was consistent with that in the tumour in 8/9 evaluable cases (one was concordantly positive, seven were concordantly negative), SPINK1 results were concordant in 9/10 cases (two were concordantly positive, seven were concordantly negative [P = 0.047 for the predictive value]). There was no correlation between PSA or SPINK protein and their respective mRNA copy levels in blood. CONCLUSIONS: Cytarabine at the doses used is ineffective for men with CRPC. Blood mRNA levels of prostate cancer genes may represent a novel aspect of monitoring prostate cancer and have implications for the understanding of tumour-derived mRNA.

Digital quantitative tissue image analysis of hypoxia in resected pancreatic ductal adenocarcinomas.

Background: Tumor hypoxia is theorized to contribute to the aggressive biology of pancreatic ductal adenocarcinoma (PDAC). We previously reported that hypoxia correlated with rapid tumor growth and metastasis in patient-derived xenografts. Anticipating a prognostic relevance of hypoxia in patient tumors, we developed protocols for automated semi-quantitative image analysis to provide an objective, observer-independent measure of hypoxia. We further validated this method which can reproducibly estimate pimonidazole-detectable hypoxia in a high-through put manner. Methods: We studied the performance of three automated image analysis platforms in scoring pimonidazole-detectable hypoxia in resected PDAC (n = 10) in a cohort of patients enrolled in PIMO-PANC. Multiple stained tumor sections were analyzed on three independent image-analysis platforms, Aperio Genie (AG), Definiens Tissue Studio (TS), and Definiens Developer (DD), which comprised of a customized rule set. Results: The output from Aperio Genie (AG) had good concordance with manual scoring, but the workflow was resource-intensive and not suited for high-throughput analysis. TS analysis had high levels of variability related to misclassification of cells class, while the customized rule set of DD had a high level of reliability with an intraclass coefficient of more than 85%. Discussion: This work demonstrates the feasibility of developing a robust, high-performance pipeline for an automated, quantitative scoring of pimonidazole-detectable hypoxia in patient tumors.

Repurposing Itraconazole and Hydroxychloroquine to Target Lysosomal Homeostasis in Epithelial Ovarian Cancer.

Drug repurposing is an attractive option for oncology drug development. Itraconazole is an antifungal ergosterol synthesis inhibitor that has pleiotropic actions including cholesterol antagonism, inhibition of Hedgehog and mTOR pathways. We tested a panel of 28 epithelial ovarian cancer (EOC) cell lines with itraconazole to define its spectrum of activity. To identify synthetic lethality in combination with itraconazole, a whole-genome drop-out genome-scale clustered regularly interspaced short palindromic repeats sensitivity screen in two cell lines (TOV1946 and OVCAR5) was performed. On this basis, we conducted a phase I dose-escalation study assessing the combination of itraconazole and hydroxychloroquine in patients with platinum refractory EOC (NCT03081702). We identified a wide spectrum of sensitivity to itraconazole across the EOC cell lines. Pathway analysis showed significant involvement of lysosomal compartments, the trans-golgi network and late endosomes/lysosomes; similar pathways are phenocopied by the autophagy inhibitor, chloroquine. We then demonstrated that the combination of itraconazole and chloroquine displayed Bliss defined synergy in EOC cancer cell lines. Furthermore, there was an association of cytotoxic synergy with the ability to induce functional lysosome dysfunction, by chloroquine. Within the clinical trial, 11 patients received at least one cycle of itraconazole and hydroxychloroquine. Treatment was safe and feasible with the recommended phase II dose of 300 and 600 mg twice daily, respectively. No objective responses were detected. Pharmacodynamic measurements on serial biopsies demonstrated limited pharmacodynamic impact. In vitro, itraconazole and chloroquine have synergistic activity and exert a potent antitumor effect by affecting lysosomal function. The drug combination had no clinical antitumor activity in dose escalation. Significance: The combination of the antifungal drug itraconazole with antimalarial drug hydroxychloroquine leads to a cytotoxic lysosomal dysfunction, supporting the rational for further research on lysosomal targeting in ovarian cancer.

Amplifying the Voice of the Whispering Cancer.

Nontargeted circulating tumor DNA (ctDNA) whole-genome sequencing is a novel strategy for genomic characterization of high-grade serous ovarian cancer. Changes in ctDNA levels are a sensitive indicator of disease burden with an average lead time of 6 months to clinical progression. This presents a unique opportunity to identify pathways driving progression as molecular vulnerabilities for clinical drug development.See related article by Paracchini et al., p. 2549.

Safety and Antitumor Activity of α-PD-L1 Antibody as Monotherapy or in Combination with α-TIM-3 Antibody in Patients with Microsatellite Instability-High/Mismatch Repair-Deficient Tumors.

PURPOSE: Immune checkpoint inhibitors show high response rates and durable clinical benefit in microsatellite instability-high/mismatch repair-deficient (MSI-H/dMMR) tumors. However, 50%-60% do not respond to single-agent anti-programmed death-1/programmed death ligand 1 (PD-1/PD-L1) antibodies, and approximately 50% of responders relapse within 6-12 months. This phase Ib trial evaluated safety and antitumor activity of anti-PD-L1 antibody LY3300054 monotherapy or in combination with anti-TIM-3 antibody LY3321367 in patients with MSI-H/dMMR advanced solid tumors. PATIENTS AND METHODS: Eligible patients ≥18 years without prior anti-PD-1/PD-L1 therapy received LY3300054 monotherapy (N = 40) or combination (N = 20); patients with PD-1/PD-L1 inhibitor-resistant/refractory tumors received the combination (N = 22). LY3300054 (700 mg) and anti-TIM-3 antibody (cycles 1-2: 1,200 mg, cycle 3 onward: 600 mg) were administered intravenously every 2 weeks. Primary endpoints were safety and tolerability. RESULTS: Eighty-two patients were enrolled. Most had colorectal (n = 39, 47.6%) or endometrial (n = 14, 17.1%) tumors. More than 70% of patients in the PD-1/PD-L1 inhibitor-resistant/refractory combination cohort had received ≥3 treatment lines. Treatment-related adverse events (TRAE) occurred in 22 patients (55.0%) receiving monotherapy, 13 (65.0%) in the PD-1/PD-L1 inhibitor-naïve combination cohort, and 6 (27.3%) in the PD-1/PD-L1 inhibitor-resistant/refractory combination cohort. A total of 2 patients (5.0%) receiving monotherapy and 3 (7.1%) receiving the combination experienced grade ≥3 TRAEs. Objective responses occurred in 13 patients (32.5%) with monotherapy, 9 (45.0%) in the PD-1/PD-L1 inhibitor-naïve combination cohort, and 1 patient (4.5%) in the PD-1/PD-L1 inhibitor-resistant/refractory combination cohort. CONCLUSIONS: LY3300054 monotherapy and combined LY3300054/anti-TIM-3 had manageable safety profiles. Both regimens showed promising clinical activity against PD-1/PD-L1 inhibitor-naïve MSI-H/dMMR tumors. The combination had limited clinical benefit in patients with PD-1/PD-L1 inhibitor-resistant/refractory MSI-H/dMMR tumors.