Citations

4157 Background: IDH1/2 mutations occur in ~25% of cholangiocarcinomas (CCAs). Preclinical studies suggest synergy between PARP inhibition and PD-L1 blockade. We previously conducted a phase II trial of olaparib plus durvalumab in advanced IDH -mutant CCA. We report matched baseline tissue and serial cell-free DNA (cfDNA)-based correlative analysis of pts samples. The primary objective was to develop a cfDNA methylome signature as a non-invasive biomarker. Methods: This single-arm, open-label phase II study (NCT03991832) enrolled patients with unresectable/metastatic IDH -mutant CCA and ≤2 prior lines of systemic therapy. Patients received olaparib 300 mg orally twice daily and durvalumab 1500 mg IV every 4 weeks. Blood samples were collected at baseline and monthly while on treatment for planned correlatives to examine R2HG, S2HG, and artemin levels using HPLC (High-Performance Liquid Chromatography). Cell-free methylated DNA immunoprecipitation and high-throughput sequencing (cfMeDIP-seq) was performed on pts cfDNA samples and case-matched IDH -wildtype pts. Formalin-fixed, paraffin-embedded blocks of baseline tumor biopsies were used for multispectral fluorescent IHC (FL-IHC) using a 6-marker panel composed of pan-cytokeratin, CD4, CD8, CD163, PD-L1, and CD68. Results: Ten patients were enrolled (median age 63.5 y; 50% female; 90% received prior platinum). Median treatment duration was 1.95 mo (range 1.8–13.5). No complete or partial responses were observed. DCR was 30% (3 patients with stable disease); one patient remained on therapy for 13.5 mo before progression. Median PFS was 1.97 mo (95% CI 1.73–3.93). HPLC from baseline plasma showed lower baseline artemin levels correlated with stable disease (p=0.038), while R2HG/S2HG ratios showed no association with outcome. FL-IHC showed enriched immune cell populations, specifically CD4 + T cells (p=0.0032) and a trend for CD68 + macrophages (p=0.062), in the tumor stroma compared with tumor core. 54 plasma samples (from 8 patients) were profiled with cfMeDIP-seq along with 8 IDH -wildtype controls. Using top 100 differentially methylated regions, the circulating tumour DNA methylome exhibited a highly specific signature, accurately discriminating IDH -mutant and IDH -wildtype CCA regardless of treatment status. The specific differentially methylated gene pathways between groups were comprehensively examined in both on-treatment and baseline plasma samples. Conclusions: Advanced IDH -mutant CCA harbors an immune-excluded phenotype, which may underlie primary resistance to immunotherapy. Plasma cfDNA methylome signature can accurately discriminate CCA with IDH mutations. Clinical trial information: NCT03991832 .

5579 Background: Adavosertib (AZD-1775) is a potent small molecule inhibitor of Wee-1, currently in clinical development. In a double-blind, placebo-controlled, phase 2 trial (NCT02151292), adavosertib and gemcitabine significantly prolonged progression-free survival (PFS) and overall survival (OS) in patients with recurrent platinum-resistant or platinum-refractory high grade serous ovarian cancer (HGSOV) compared to gemcitabine alone. We investigated whether plasma and intra-tumoral adavosertib concentrations correlated with survival in these patients. Methods: Adavosertib was administered orally on Days 1, 2, 8, 9, 15 and 16 at 175 mg per day, and gemcitabine was administered on Days 1, 8 and 15 at 1000 mg/m2 every 28 days. Serial blood samples were collected on Day 1 of cycle 1 after adavosertib administration. Tumor biopsies were taken 1-2 weeks after initiation of study treatments. Plasma and tumor adavosertib concentrations were determined using validated HPLC-MS/MS. Patients were divided into groups with plasma or tumor adavosertib concentrations above or below the biologically active concentration (BAC) of 125 ng/ml (Leijen et al, J Clin Onco 2016). Survival was described using the Kaplan-Meier method. Results: Among 61 HGSOV patients who received adavosertib, plasma samples were available in 47, and tumor samples were available in 31 patients. Among 25 non-HGSOV patients (exploratory cohort), plasma and tumor samples were available in 21 and 17 patients respectively. The mean maximum adavosertib concentration (Cmax) was 355.3 ± 120.9 ng/ml and 358.6 ± 117.9 ng/ml respectively. Cmax was above BAC in all patients. The mean tumor adavosertib concentration was 609.2 ± 1129.2 ng/ml (range: 0.47 – 5501 ng/ml) for HGSOV patients, and 964.2 ± 1611.2 ng/ml (range: 0.22 – 6116 ng/ml) for non-HGSOV patients. There was no correlation between Cmax and tumor adavosertib concentrations. In HGSOV, the median PFS was 5.8 months for patients with tumor concentrations above BAC, and 3.5 months for those with tumor concentrations below BAC (Hazard ratio (HR): 0.46, 95% confidence interval: 0.19 – 1.14, p = 0.06). No difference in PFS was seen in non-HGSOV patients according to tumor adavosertib concentration. Tumor adavosertib concentration did not correlate with OS. Conclusions: Although Cmax was above BAC in all patients, there was a high variability in tumor adavosertib concentrations. In HGSOV, higher tumor adavosertib concentration was associated with a trend towards improved PFS, but not OS. Our results indicate that the current adavosertib dosing regimen may not produce the desired concentrations in tumors for some patients, and further optimization may be needed.

Background Combining immunotherapy and antiangiogenic agents is a promising treatment strategy in endometrial cancer. To date, no biomarkers for response have been identified and data on post-immunotherapy progression are lacking. We explored the combination of a checkpoint inhibitor (nivolumab) and an antiangiogenic agent (cabozantinib) in immunotherapy-naïve endometrial cancer and in patients whose disease progressed on previous immunotherapy with baseline biopsy for immune profiling. Patients and methods In this phase II trial (ClinicalTrials.gov NCT03367741, registered December 11, 2017), women with recurrent endometrial cancer were randomized 2:1 to nivolumab with cabozantinib (Arm A) or nivolumab alone (Arm B). The primary endpoint was Response Evaluation Criteria in Solid Tumors-defined progression-free survival (PFS). Patients with carcinosarcoma or prior immune checkpoint inhibitor received combination treatment (Arm C). Baseline biopsy and serial peripheral blood mononuclear cell (PBMC) samples were analyzed and associations between patient outcome and immune data from cytometry by time of flight (CyTOF) and PBMCs were explored. Results Median PFS was 5.3 (90% CI 3.5 to 9.2) months in Arm A (n=36) and 1.9 (90% CI 1.6 to 3.4) months in Arm B (n=18) (HR=0.59, 90% CI 0.35 to 0.98; log-rank p=0.09, meeting the prespecified statistical significance criteria). The most common treatment-related adverse events in Arm A were diarrhea (50%) and elevated liver enzymes (aspartate aminotransferase 47%, alanine aminotransferase 42%). In-depth baseline CyTOF analysis across treatment arms (n=40) identified 35 immune-cell subsets. Among immunotherapy-pretreated patients in Arm C, non-progressors had significantly higher proportions of activated tissue-resident (CD103+CD69+) ɣδ T cells than progressors (adjusted p=0.009). Conclusions Adding cabozantinib to nivolumab significantly improved outcomes in heavily pretreated endometrial cancer. A subgroup of immunotherapy-pretreated patients identified by baseline immune profile and potentially benefiting from combination with antiangiogenics requires further investigation.

Abstract Leukemia stem cells (LSCs) are linked to relapse in acute myeloid leukemia (AML). The LSC17 gene expression score robustly captures LSC stemness properties in AML and can be used to predict survival outcomes and response to therapy, enabling risk-adapted, upfront treatment approaches. The LSC17 score was developed and validated in a research setting. To enable widespread use of the LSC17 score in clinical decision making, we established a laboratory-developed test (LDT) for the LSC17 score that can be deployed broadly in clinical molecular diagnostic laboratories. We extensively validated the LSC17 LDT in a College of American Pathologists/Clinical Laboratory Improvements Act (CAP/CLIA)-certified laboratory, determining specimen requirements, a synthetic control, and performance parameters for the assay. Importantly, we correlated values from the LSC17 LDT to clinical outcome in a reference cohort of patients with AML, establishing a median assay value that can be used for clinical risk stratification of individual patients with newly diagnosed AML. The assay was established in a second independent CAP/CLIA-certified laboratory, and its technical performance was validated using an independent cohort of patient samples, demonstrating that the LSC17 LDT can be readily implemented in other settings. This study enables the clinical use of the LSC17 score for upfront risk-adapted management of patients with AML.

6010 Background: The efficacy of treatment for recurrent endometrial cancer (EC) remains limited. Vascular endothelial growth factor and inflammatory chemokines are proangiogenic factors and immune modulators involved in immune suppression. Reprogramming the tumor microenvironment by combining antiangiogenic and immunotherapy (IO) could enhance antitumor responses. Methods: A 2:1 randomized phase 2 trial compared the combination of cabozantinib and nivolumab (Arm A) versus nivolumab (Arm B) in recurrent EC. Primary endpoint was progression free survival (PFS) assessed by RECIST 1.1 (NCT03367741). Women with recurrent measurable EC were eligible. There were no limits on prior therapy, but at least one prior platinum-based chemotherapy was required. Patients (pts) were stratified according to MSI status and assessed by CT every 8 weeks. Cabozantinib was given at 40 mg daily (Arm A) and nivolumab at 240 mg, on D1 and D15 of a 28-day cycle for 4 cycles, followed by 480 mg every 4 weeks (Arms A & B). Pts with carcinosarcoma or prior IO were enrolled in an exploratory cohort and received combination treatment (Arm C). A baseline biopsy was required for all pts. CyTOF analysis was performed on fresh biopsies. Results: 76 evaluable pts were enrolled (Arm A: 36, Arm B: 18, Arm C: 9 carcinosarcoma, and 20 post IO including 7 pts crossed over from Arm B). 55% of pts had received ≥3 prior lines of therapy. Two pts were MSI high in Arm A and none in Arm B. The Kaplan-Meier estimated median PFS was 5.3 (95% CI: 3.5-9.5) months in Arm A and 1.9 (95% CI: 1.6-3.8) months in Arm B, with a log-rank p = 0.07, which met the significance level of 0.1 used for sample size calculation. Objective response rate (ORR) was 25% for Arm A and 16.7% for Arm B; stable disease (SD) was seen in 44.4% vs 11.1%, respectively. Clinical benefit (ORR+SD) was significantly higher in arm A vs B (p < 0.001). In Arm C-carcinosarcoma, one patient had a partial response (11.9 months duration) and four SD. In Arm C-prior IO, six pts responded and eight had SD. The most common related AEs in Arm A were diarrhea (47.2%), elevated liver enzymes (44.4%), fatigue (38.9%), anorexia, hypertension, and nausea (30.6%), mainly grade 1/2. Preliminary CyTOF analysis across treatment arms identified multiple immune subsets for further interrogation including activated CD8+ and CD4+ T cells. Conclusions: Cabozantinib plus nivolumab demonstrates improved PFS compared to nivolumab in heavily pre-treated women with recurrent EC. In-depth CyTOF analysis of the tumor microenvironment to identify predictive immune biomarkers of response is ongoing. Clinical trial information: NCT03367741.

Abstract Abstract 718 Introduction: In a previously reported phase 2 study of single agent lenalidomide in 25 untreated CLL patients (pts), we reported an overall response rate (ORR) of 56% (14 pts), 40% SD (10 pts) and no CR at a median follow-up of 20.7 months (Chen et al. JCO 2010;29:1175). Although an amended protocol with conservative lenalidomide dosing was used to mitigate tumor lysis and severe myelosuppression observed in the first 2 accrued pts, we continued to observe frequent toxicities of grade 3–4 neutropenia (72%) and tumor flare (TF 88%). We now report long-term efficacy and toxicity from this study at a median follow-up of 47 months (mos). Methods: Patients were eligible if previously untreated and symptomatic (cytopenias, symptomatic adenopathy/organomegaly, constitutional symptoms, lymphocyte doubling count <12 mos). The amended starting dose for lenalidomide was 2.5mg daily on days 1–21 of a 28 day cycle, with slow monthly dose escalations (2.5mg cycle 1, 5mg cycle 2, 10mg cycle 3 and if required for response, further 5mg increments to a maximum of 25mg daily were allowed). Results: Longterm toxicities: Hematologic toxicities were common: grade 3–4 neutropenia (76%), thrombocytopenia (28%), anemia (20%). With longer term use, neutropenia tended to recur (12% of all cycles) and 10 pts required GCSF support (5 routinely during each cycle). Most common non-hematologic toxicities (all grades) were TF (88%), fatigue (76%), rash (60%), muscle cramping (40%), diarrhea (40%). All non-hematologic toxicities were mild (grade 1–2), except for 1 pt each with grade 3 rash and diarrhea. Although TF was most common during cycle 1, repeat flare symptoms upon resuming lenalidomide after the 7 day rest period of each cycle were noted in 16% of all 898 cycles administered, and as late as at cycle 28. Infections were mild (most respiratory, skin) with only 2 grade 3 events (disseminated zoster, S.pneumoniae bacteremia). Other malignancies: 2 pts developed transformed large cell lymphoma 7 and 18 mos after study discontinuation, 1 pt developed squamous cell carcinoma of skin at cycle 51, and 1 pt developed recurrence of remote non-small cell lung cancer at cycle 34. Dose modifications/discontinuation: The median highest dose achieved for all 25 pts was 15 mg (range 2.5–25 mgs); 8 pts were able to escalate to the maximal 25mg dose. Ten pts (40%) required dose reductions for grade 3 cytopenias [neutropenia (2), thrombocytopenia (2), both (2)], febrile neutropenia (2), and diarrhea (2). Of all 25 pts, the median duration on therapy was 31.1 mos (range 28 days – 60.6 mos). Twelve pts (52%) currently remain on study, receiving a median of 59 cycles of therapy (range 48–66). Causes of discontinuation for 13 pts included: treatment-related toxicity (8), lack of response/progressive disease (4), and recurrence of remote lung cancer (1). Toxicities leading to discontinuation included: prolonged cytopenias (3), recurrent infections (1), atrial fibrillation (1), disseminated herpes zoster (1), persistent grade 2 diarrhea (1), and grade 3 skin rash (1). Efficacy: With extended median follow-up from 20.7 to 47 mos, the ORR improved from 56% (14 pts) to 72% (18 pts), with 3 pts in PR upgrading to CR, and 1 SD to PR. Although the median time to response was 7.7 mos, responses occurred as quickly as 1.8 mos to as late as 27.0 mos of therapy. For the 3 CR pts, prolonged therapy with an additional 14.9, 28.3 and 40.6 mos beyond the time of first response was required to achieve CR. To date, 7 pts have progressed with 3-year PFS 68.8% (95% CI:52–91%) and OS 85.3% (95% CI:71.1–100%). Correlatives: Cereblon (CRBN), recently identified as a direct protein target of lenalidomide, was evaluated by gene expression profiling and Western blot and found to be uniformly expressed in all 19 evaluable day 1 pt samples regardless of lenalidomide response. Thus, baseline CRBN expression does not appear to be a useful predictive biomarker of response in this population. The mechanism by which CRBN is linked to response is reported by Trudel et al, ASH 2012. Conclusions: Long-term followup of this study demonstrates that when using low doses of single agent lenalidomide in CLL, prolonged therapy is feasible and may be required for the achievement of durable, high quality responses. Maximal daily doses of 25mg can be reached and may also be needed for optimal response, though recurrent myelosuppression remains limiting. Disclosures: Chen: Celgene: Honoraria, Research Funding; GlaxoSmithKline: Research Funding; Lundbeck: Consultancy; Janssen: Consultancy, Research Funding. Off Label Use: Lenalidomide is not approved for use in chronic lymphocytic leukemia. Kukreti:Roche: Honoraria; Celgene: Honoraria; Janssen Ortho: Honoraria. Trudel:Celgene: Honoraria, Research Funding, Speakers Bureau.

5518 Background: Platinum resistant ovarian cancer (OC) remains a therapeutic challenge. High grade serous OC (HGSOC) harbors TP53 mutations leading to increased dependency on S- and G2-phase checkpoints. Wee1 inhibition with Adavosertib (AZD 1775) (A) induces G2 checkpoint escape. Gemcitabine (G) is an antimetabolite therapy and blocks the progression of cells through the G1/S phase. We hypothesized that combining G+A would be synergistic and overcome resistance. Methods: We conducted a multicentre double-blind 2:1 randomized phase 2 trial to assess the progression free survival (PFS) in women with recurrent platinum-resistant/refractory HGSOC receiving G+A or G+placebo (P) (NCT02151292). Eligibility required measurable disease and feasibility of paired tumor biopsies; no limitation in prior lines of therapy. Non HGSOC histologic subtypes were enrolled in a separate non-randomized exploratory cohort. A/P was given orally at 175mg QD on D1-2, D8-9 and D15-16 with G 1000mg/m² IV D1, D8 and D15 in a 28-day cycle until progression or unacceptable AE. Tumor staging was scheduled every 8 weeks. TP53 mutations were analyzed on archival tissue with Sanger sequencing, TAm-Seq and IHC. TP53 mutation will be also assessed in circulating tumor DNA (ctDNA). Whole exome and RNA sequencing were performed on paired tumor tissues. Results: 124 patients (pts) with median of 3 prior lines of therapy (range 1-10) from 12 centres across Canada and US were enrolled between Sep 2014 to May 2018, with 99 pts randomized (65 in Arm G+A and 34 in G+P). 5 pts were ineligible; 64 pts have died. The median follow-up was 14.3 months. Main related AE was hematologic toxicity (Anemia G≥3: 31% in G+A vs 18% in G+P; Thrombocytopenia G≥3: 31% vs 6%; Neutropenia G≥3: 62% vs 30%). PFS was significantly improved from 3.0 to 4.6 months (HR 0.56 (95%CI: 0.35-0.90, p=0.015 Log rank). There was a significant improvement in overall survival (OS) from 7.2 to 11.5 months (HR 0.56 (95%CI: 0.34-0.92, P=0.022). Partial response by RECIST 1.1 was observed in 13 (21%) and 1 (3%) pts for Arms G+A and G+P, respectively (p=0.02). From the 25 pts in the exploratory cohort, 3 (12%) partial responses were observed. Final results will be reported at the meeting. Conclusion: Addition of adavosertib to gemcitabine in women with platinum resistant/refractory OC improved response rate, PFS and OS with manageable toxicity. Clinical trial information: NCT02151292.

Background Noonan syndrome with multiple lentigines (NSML), formerly known as LEOPARD syndrome, is an autosomal-dominant disorder characterised by lentigines, EKG abnormalities, ocular hypertelorism, pulmonic stenosis, abnormal genitalia, growth retardation and deafness. There is significant clinical overlap between NSML and other disorders that result from dysregulated rat sarcoma/mitogen-activated protein kinase pathway (RASopathies). Except for neurofibromatosis type 1, other RASopathies are not known to be typically associated with neurogenic tumours. Methods and results We evaluated patients from three families with pigmentary skin lesions, progressive neuropathy, enlarged nerves, massive burden of paraspinal tumours (neurofibroma was confirmed in one patient) and a clinical diagnosis of NSML. All patients had a mutation in the protein tyrosine phosphatase catalytic domain of the PTPN11 gene; two unrelated patients had the p.Thr468Met mutation, while the family consisting of two affected individuals harboured the p.Thr279Cys mutation. Molecular analysis performed on hypertrophic nerve tissue did not disclose a second somatic hit in NF1, PTPN11, NF2 or SMARCB1 genes. Conclusions Neurogenic tumours and hypertrophic neuropathy are unusual complications of NSML and may be an under-recognised manifestation that would warrant surveillance. Our observation may also have implications for other disorders caused by RAS-pathway dysregulation.