Search

Summary Background PD-1 and PD-L1 inhibitors are active in metastatic urothelial carcinoma, but positive randomised data supporting their use as a first-line treatment are lacking. In this study we assessed outcomes with first-line pembrolizumab alone or combined with chemotherapy versus chemotherapy for patients with previously untreated advanced urothelial carcinoma. Methods KEYNOTE-361 is a randomised, open-label, phase 3 trial of patients aged at least 18 years, with untreated, locally advanced, unresectable, or metastatic urothelial carcinoma, with an Eastern Cooperative Oncology Group performance status of up to 2. Eligible patients were enrolled from 201 medical centres in 21 countries and randomly allocated (1:1:1) via an interactive voice-web response system to intravenous pembrolizumab 200 mg every 3 weeks for a maximum of 35 cycles plus intravenous chemotherapy (gemcitabine [1000 mg/m2] on days 1 and 8 and investigator's choice of cisplatin [70 mg/m2] or carboplatin [area under the curve 5] on day 1 of every 3-week cycle) for a maximum of six cycles, pembrolizumab alone, or chemotherapy alone, stratified by choice of platinum therapy and PD-L1 combined positive score (CPS). Neither patients nor investigators were masked to the treatment assignment or CPS. At protocol-specified final analysis, sequential hypothesis testing began with superiority of pembrolizumab plus chemotherapy versus chemotherapy alone in the total population (all patients randomly allocated to a treatment) for the dual primary endpoints of progression-free survival (p value boundary 0·0019), assessed by masked, independent central review, and overall survival (p value boundary 0·0142), followed by non-inferiority and superiority of overall survival for pembrolizumab versus chemotherapy in the patient population with CPS of at least 10 and in the total population (also a primary endpoint). Safety was assessed in the as-treated population (all patients who received at least one dose of study treatment). This study is completed and is no longer enrolling patients, and is registered at ClinicalTrials.gov , number NCT02853305 . Findings Between Oct 19, 2016 and June 29, 2018, 1010 patients were enrolled and allocated to receive pembrolizumab plus chemotherapy (n=351), pembrolizumab monotherapy (n=307), or chemotherapy alone (n=352). Median follow-up was 31·7 months (IQR 27·7–36·0). Pembrolizumab plus chemotherapy versus chemotherapy did not significantly improve progression-free survival, with a median progression-free survival of 8·3 months (95% CI 7·5–8·5) in the pembrolizumab plus chemotherapy group versus 7·1 months (6·4–7·9) in the chemotherapy group (hazard ratio [HR] 0·78, 95% CI 0·65–0·93; p=0·0033), or overall survival, with a median overall survival of 17·0 months (14·5–19·5) in the pembrolizumab plus chemotherapy group versus 14·3 months (12·3–16·7) in the chemotherapy group (0·86, 0·72–1·02; p=0·0407). No further formal statistical hypothesis testing was done. In analyses of overall survival with pembrolizumab versus chemotherapy (now exploratory based on hierarchical statistical testing), overall survival was similar between these treatment groups, both in the total population (15·6 months [95% CI 12·1–17·9] with pembrolizumab vs 14·3 months [12·3–16·7] with chemotherapy; HR 0·92, 95% CI 0·77–1·11) and the population with CPS of at least 10 (16·1 months [13·6–19·9] with pembrolizumab vs 15·2 months [11·6–23·3] with chemotherapy; 1·01, 0·77–1·32). The most common grade 3 or 4 adverse event attributed to study treatment was anaemia with pembrolizumab plus chemotherapy (104 [30%] of 349 patients) or chemotherapy alone (112 [33%] of 342 patients), and diarrhoea, fatigue, and hyponatraemia (each affecting four [1%] of 302 patients) with pembrolizumab alone. Six (1%) of 1010 patients died due to an adverse event attributed to study treatment; two patients in each treatment group. One each occurred due to cardiac arrest and device-related sepsis in the pembrolizumab plus chemotherapy group, one each due to cardiac failure and malignant neoplasm progression in the pembrolizumab group, and one each due to myocardial infarction and ischaemic colitis in the chemotherapy group. Interpretation The addition of pembrolizumab to first-line platinum-based chemotherapy did not significantly improve efficacy and should not be widely adopted for treatment of advanced urothelial carcinoma. Funding Merck Sharp and Dohme, a subsidiary of Merck, Kenilworth, NJ, USA.

Although blocking the androgen receptor (AR) pathway is a core treatment strategy in early prostate cancer, most patients progress to metastatic castration-resistant prostate cancer(mCRPC). Immunotherapy with PD-1/PD-L1 inhibition has limited activity in mCRPC. AR signaling modulates CD8+ T cells and antitumor immune responses in mCRPC. In a companion presentation (Schwermann et al., submitted AACR 2023), we demonstrate that AR blockade activates NK cell killing of prostate cancer (PC) cells. We hypothesize this activation requires IFN-γ and TRAIL pathways. We investigated the mechanisms of AR inhibitors (ARi)enzalutamide (Enza) and darolutamide (Daro) on the antitumor function of NK cells and potential enhancement with checkpoint inhibitors. Material and Methods: We performed ATP-based CellTiterGlo viability assays to investigate the effects of Enza and Daro on PC cells (LNCap, 22Rv1 [ARv7mutation], DU145, PC3 [AR-]) and NK-92 cells. We performed co-culture experiments with PC, NK, and T cells, at a 1:1 ratio, using IFN-γ blocking mAb (10 μg/ml), RIK-2 (TRAIL blocking mAb- 10 μg/ml) and analyzed immune cell-mediated tumor cell killing (ImageXpress Confocal HT). The NK cell immune checkpoint inhibitor monalizumab that targets NKG2A was combined with ARi. Results: Co-cultures of PC plus NK-92 cells with ARi significantly increased immune-mediated PCkilling within 24hrs (control[C]: 20%±1.9; Enza: 40%±4.3; Daro: 36%±3.72, p=0.0001). This immune enhancement effect was abolished by IFN-γ mAb. The combination of TRAIL inhibitory mAb with Enza decreased NK immune-mediated killing by 16±2.4% ([C]: 40%±4.3; Enza+RIK-2:25.6%±2.1, p=0.002). Treatment of PC cells plus NK cells with Enza, IFN-γ mAb, and TRAIL-blocking RIK-2 significantly reduced NK cell-mediated killing of PCs (control[C]: 20%±1.9, [T]:3.2%±2.6, p=0.003). ARi immune enhancement effect was increased by NKG2A blockade with monalizumab ([C]: 31.2%±3.1, [T]: 43.92%±4.52, p=0.0045). Conclusions: ARi promotes NK cell killing of PC cells via IFN-γ gamma and TRAIL. NK cell activation is enhanced by the combination of ARi and the checkpoint inhibitor monalizumab. Validation of these results in patient-derived organoids paired with tumor samples from patients with PC is ongoing to further understand the innate immune response and direct novel therapeutic strategies. Citation Format: Maximilian Schwermann, Lindsey Carlsen, Kelsey E. Huntington, Lanlan Zhou, Andrew George, Praveen Srinivasan, Vida Tajiknia, Arielle De La Cruz, Andre De Souza, Anthony E. Mega, Howard P. Safran, Benedito A. Carneiro, Wafik S. El-Deiry. IFN-γ and TRAIL involvement in androgen signaling-mediated NK cell killing of prostate cancer cells and further enhancement by NKG2A checkpoint inhibition with monalizumab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1822.

550 Background: 24% of patients with high-risk urothelial carcinoma have pathogenic germline mutations (Nassar, Genetics in Medicine 2020). In that study, demographics, metachronous/synchronous tumors, and family history did not differ between germline and sporadic cases of bladder cancer. We report herein the prevalence of actionable mutations and de novo metastatic disease in patients with germline mutations who developed metastatic urothelial carcinoma. Methods: We retrospectively analyzed a database of 90 patients with metastatic urothelial carcinoma (urethral, bladder, and upper tract disease) at our institution who had genetic testing performed on tumor specimens. T-student test was performed to calculate statistical significance for the distribution of age, while Chi-square test evaluated the frequency distribution of gender, actionable mutations, and de novo metastatic disease. Patients’ tumors were sequenced by a 700 gene panel for both somatic and germline mutations. Comprehensive chart review was performed to extract clinical data. Results: Out of the 90 patients reviewed, 11 (11.1%) had germline mutations. Of these patients, 5 had upper tract urothelial carcinoma, 5 had bladder cancer, and 1 had urethral cancer. Nine patients had pathogenic germline mutations: MUTYH, BRCA2 (each representing 1.8% of patients); APC, BRCA1, CDKN2A, FH, MSH2 (each representing 0.9% of patients). Two patients had germline mutations of unknown significance ( APC, CHEK2). Age (T-value 1.62053, p=1.08453), gender (Chi-square 0.0024, p=0.961037) or de novo metastatic presentation (Chi-square 0.5, p=0.4795) were not statistically significant between patients with germline and sporadic mutations. Somatic actionable mutations included ATR, BRCA2, BRAF, CDK12, ERBB2, FBXW7, FGFR3, HRAS, MTAP, and PIK3CA. Microsatellite instability high (MSI-H) status was only present in the patient with germline MSH2 mutation. PD-L1 expression was high (CPS ≥10) in 4 patients with germline mutations. Tumor mutational burden ranged from 1.1 to 28.4 mutations per Megabase. Conclusions: Our findings further define the clinical and genomic characteristics of patients with metastatic urothelial carcinoma and germline mutations in a tertiary center. Further investigation is warranted to validate these findings in national sequencing databases.

4571 Background: p53 target and cell cycle inhibitor CDKN1A/p21(WAF1) was initially not found to be mutated in cancer. TCGA analysis identified CDKN1A mutations are present but rare with frequencies of < 1%, but enrich in bladder cancer (̃8%). Truncating WAF1 mutations are associated with sensitivity to cisplatin and are associated with truncating Rb mutations in bladder cancer (RW+). We hypothesized RW+ bladder cancers may represent a unique subgroup with sensitivity to therapeutics. Methods: A total of 1104 urothelial tumors underwent molecular profiling at Caris Life Sciences (Phoenix, AZ) utilizing NGS of DNA (592 Gene Panel, NextSeq, or WES, NovaSeq) and RNA (NovaSeq, WTS). Wilcoxon, Fisher’s exact were used for statistical significance (p value without and q value with multi comparison correction). Immune cell fraction (QuanTIseq) and pathway analysis (ssGSEA) were assessed by mRNA analysis. Immune epitope prediction was performed using the NetMHCpan v4.0 method in the Immune Epitope Database. Results: Concurrent truncating mutation (frameshift, nonsense) for RB1 and WAF1 were detected in 47 tumors (RW+, 4.25%) and tumors with wild-type status for both RB1 and WAF1 genes were classified as RW- group (54.08%). Tumors harboring only one RB1 or WAF1 mutation were excluded for further analysis. When compared to RW- group, RW+ tumors showed lower mutation rate of TP53 (54.5% vs 80.9%, q < 0.05), ARID1A (23.5% vs 38.3%, p < 0.05), and PIK3CA (18.4% vs 31.9%, p < 0.05). Interestingly, RW+ was mutually exclusive with FGFR3 mutation (18.0% vs 0%, p < 0.05). We further evaluated RNA expression of DNA repair and checkpoint arrest pathways. Notably, E2F pathway (Normalized Enrichment Scores, NES: 0.89 vs 0.86, q < 0.01) and DNA G2M checkpoint (NES: 0.89 vs 0.86, q < 0.01) were found to be the most enriched in RW+ with respect to RW- group. In addition, mRNA levels of FANCC/A, CHEK1, WEE1, CDC25A/C, PALB2 and BRCA1/2 were found to be overexpressed in RW+ group (q < 0.05). RW+ tumors also displayed a distinct immunological profile: They were associated with higher PD-L1 status (63.8% vs 37.3%, q < 0.01), higher median TMB (11 mut/Mb vs 8 mut/Mb, q < 0.01) and with less frequent loss of heterozygosity for HLA-DPA1 (51.1% vs 66.7%, p < 0.05), with more high-binding-affinity neoantigen load (4.78 vs 3.89, p < 0.05) to MHC proteins, consistent with the significantly more myeloid dendritic cells in in RW+ group (0.3 vs 0.04, q < 0.001). Conclusions: Concurrent truncating mutation in RB1 and WAF1 (RW+) bladder carcinomas have fewer p53, ARID1A, and PIK3CA mutations but are enriched for E2F targets, G2/M checkpoint genes, FANCC/A, CHEK1, WEE1, CDC25A/C, PALB2 and BRCA1/2 and have a distinct immunological profile. The findings suggest therapeutic strategies for RW+ bladder cancers including Chk1/Wee1, PARP inhibitors, -/+ immunotherapy that may impact on clinical outcomes.

128 Background: mCRPC with alterations in genes associated with homologous recombination (HR) DNA repair (e.g., BRCA1/2) is sensitive to poly ADP-ribose polymerase inhibitors (PARPi). Preclinical studies showed that PI3K inhibitors (PI3Ki) impairs HR and sensitize cancer cells to PARPi even in the absence of HR gene mutations. These results support our hypothesis that dual PI3K and PARP inhibition may improve clinical outcomes in progressive mCRPC. We describe preliminary results of a phase Ib/II study investigating safety of the combination of copanlisib (pan-class I PI3Ki) and rucaparib (PARP-1, -2 and -3 inhibitor). Methods: Enrollment criteria included progressive mCRPC, prior androgen inhibitors (abiraterone, enzalutamide, and/or apalutamide); prior taxane chemotherapy was allowed. HR-deficiency was not required for the phase Ib. The phase I followed a standard 3+3 escalation design. Dose schema: rucaparib (continuous oral administration twice daily) 400mg (dose level [DL] -1, 1), 500mg (DL 2) or 600mg (DL 3,4) and intravenous copanlisib (45mg D1, D15 (DL -1, -2); 45mg, D1, D8, D15 (DL 1, 2, 3); 60mg, D1, D8, D15 (DL 4); 28-day cycle). Adverse events (AE) were graded by CTCAE v5.0. The primary aim of the phase I was to establish the MTD and the recommended phase II dose (RP2D) of copanlisib in combination with rucaparib. Results: Eleven pts were enrolled with a median age of 63 (55-78) and median PSA of 12 ng/mL (0.018–2,101). Seven pts (63%) received prior chemotherapy (docetaxel [7], cabazitaxel [3]). Pathogenic HR mutations included BRCA1 (1), BRCA2 (3), CDK12 (1), and FANCA (1). Treatment-related AE included grade 2 (G2) leukopenia (30%), G2 anemia (20%), G2 rash (20%). Two dose-limiting toxicities (DLTs) were observed in DL 1: G3 rash and G3 AST/ALT elevation attributed to both drugs. Six pts were treated at DL -1 without DLTs. The RP2D was rucaparib 400mg BID with copanlisib 45mg (D1, D15; 28-day cycle). There were 2 confirmed PSA50 responses among 7 evaluable pts (28%). One pt had BRCA2 loss and 1 had PALB2 VUS (ongoing PSA response for 14 mo). Three stable disease and 1 partial response were observed among 6 pts evaluable by RECIST 1.1. Conclusions: The combination of rucaparib and copanlisib is well tolerated. The RP2D was rucaparib 400mg BID with copanlisib 45mg (D1, D15; 28-day cycle) with signal of efficacy. Enrollment in a phase 2 expansion cohort in HR-mutated mCRPC is ongoing. Clinical trial information: NCT04253262.

5018 Background: The abundant expression of prostate specific membrane antigen (PSMA) on prostate cancer cells provides a rationale for antibody therapy. PSMA ADC is a fully human antibody to PSMA linked to the microtubule disrupting agent monomethyl auristatin E (MMAE). It binds PSMA and is internalized within the cancer cell where cleavage by lysosomal enzymes release free MMAE, causing cell cycle arrest and apoptosis. A phase 1 dose escalation study of PSMA ADC in taxane-refractory mCRPC has been completed. Methods: Patients with progressive mCRPC following taxane-containing chemotherapy and ECOG status of 0 or 1 were eligible. PSMA ADC was administered by IV infusion Q3W for up to 4 cycles. Safety, pharmacokinetics, PSA, circulating tumor cells (CTC), immunogenicity and clinical progression were assessed. Serum PSMA ADC and total anti-PSMA ADC antibodies were measured by ELISA, and free MMAE was measured by LC/MS/MS.The dosing cohorts ranged from 0.4 mg/kg to 2.8 mg/kg. Subjects who benefitted from PSMA ADC were eligible for treatment in an extension study. Results: 52 subjects were dosed in 9 dose levels. All subjects received prior docetaxel, 6 also received cabazitaxel and 3 subjects also received paclitaxel. PSMA ADC was generally well tolerated with the most commonly seen adverse events being anorexia and fatigue. 16 patients reported peripheral neuropathies, including 3 with grade 3. Dose limiting toxicities (DLT) seen at 2.8 mg/kg were neutropenia (one death) and reversible elevations in liver function tests (LFTs). Antitumor activity was manifested as reductions either in PSA or in CTCs in approximately 50% of patients at ≥ 1.8 mg/kg PSMA ADC. Exposure to PSMA ADC increased with dose and was ~1,000-fold greater than MMAE exposure. There was no accumulation. Conclusion: PSMA ADC in this study was generally well tolerated in subjects with progressive mCRPC, previously treated with taxane. Antitumor activity was seen at doses ≥ 1.8 mg/kg. DLTs were neutropenia and reversible LFT abnormalities. The maximum tolerated dose was determined to be 2.5 mg/kg. A phase 2 trial of PSMA ADC in taxane refractory mCRPC has been initiated at 2.5 mg/kg. Clinical trial information: NCT01414283.