Seribantumab Poster AACR 03.28.21
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Preclinical activity of seribantumab in gastrointestinal cancers with NRG1 fusions Igor Odintsova, MD, Renate I. Kurtha, Allan J.W. Luia, MD, Paul R. Blooma, Morana Vojnica, MD, Shawn Lelandb, PharmD, Marc Ladanyia, MD, and Romel Somwara, PhD aMemorial Sloan Kettering Cancer Center, New York, NY, USA; bElevation Oncology, New York, NY, USA Abstract Seribantumab inhibits phospho-HER3 and phospho-AKT Targeted combinations inhibit growth of NRG1-rearranged NRG1 Background. Oncogenic rearrangements of the neuregulin 1 gene (NRG1) consist of a 5’ partner fused to - activated by overexpression of NRG1 fusions in immortalized NRG1 cholangiocarcinoma PDX model harboring additional known - NRG1 NRG1 - - NRG1 - DOC4 SLC3A2 a 3’ NRG1 sequence that retains the epidermal growth factor (EGF)-like domain, and are found in ≈0.2% of EV ATP1B1 - H6C7 human pancreatic ductal epithelial cells - - - driver alterations (CH-17-0068, RBPMS-NRG1) EV ATP1B1 DOC4H6C7 SLC3A2 solid tumors including lung, breast, and gastrointestinal (GI) cancers.1 Carcinomas of GI origin, including H6C7 - H6C7 - H6C7 - - P-EGFR pancreatic and cholangiocarcinoma, represent around 20% of solid tumors harboring NRG1 fusions2 and A B C Y1068 H6C7 H6C7 H6C7 H6C7 A B H6C7-EV P-HER3 there is no approved therapy for this group of cancers. The chimeric NRG1 oncoproteins bind to human EGFR ERBB2 ERBB3 Y1289 Gene Alteration Investigational Targeted Therapy 15 epidermal growth factor receptor 3 (HER3/ERBB3) leading to trans-activation of other ERBB family P-AKT Seribantumab (anti-HER3 mAb); S473 NRG1 RBPMS-NRG1 fusion Afatinib (pan-ERBB inhibitor) members and trigger a signaling cascade that culminates in oncogenesis. Although targeting HER3 10 P-AKT T308 represents a rational therapeutic strategy for cancers harboring NRG1 fusions, this has remained relatively P-AKT S473 ERBB4 E542K Afatinib AKT unexplored for GI malignancies with NRG1 alterations. In this study, we investigated the efficacy of the anti- 5 P-PRAS40 200X IDH1 R132C AG-120 (IDH inhibitor) T246 HER3 monoclonal antibody seribantumab in preclinical models of NRG1-driven GI cancers. H6C7-ATP1B1-NRG1 Relative phosphorylation PRAS40 Methods. We developed models of isogenic pancreatic cancer cells with NRG1 fusions by lentiviral- 0 C Monotherapy Combination therapy P-ERK 800 Afatinib (5 mg/kg QD) mediated cDNA expression of ATP1B1-NRG1 and SLC3A2-NRG1 fusions in immortalized human H6C7-EV Seribantumab (2.5 mg BIW) pancreatic ductal cells (H6c7). Seribantumab efficacy was evaluated in isogenic cell lines and in patient- ERK Seribantumab (5 mg BIW) 600 Afatinib 5mg/kg QD only P-AKT S473 Seribantumab (10 mg BIW) H6C7-DOC4-NRG1 derived xenograft (PDX) models of pancreatic adenocarcinoma (CTG-0943, APP-NRG1 fusion) and H6C7-ATP1B1-NRG1 H6C7-SLC3A2-NRG1 GAPDH intrahepatic cholangiocarcinoma (CH-07-0068, RBPMS-NRG1 fusion). Western blotting analysis was used 400 Seribantumab 5 mg + AG-120 150 mg/kg BID to evaluate protein phosphorylation and expression. The presence of NRG1 fusions was confirmed by D E Seribantumab 2.5 mg + Afatinib 5 mg/kg QD 200 reverse transcription polymerase chain reaction (RT-PCR) and next-generation sequencing (NGS). Seribantumab 10 mg + Afatinib 5 mg/kg QD + AG-120 150 mg/kg BID 0 Results. Expression of NRG1 fusions in H6c7 cells resulted in enhanced phosphorylation of HER3 and (%) Volume Change in Tumor 0 10 20 30 40 50 60 AKT when compared with empty vector control cells (H6c7-EV). Treatment of H6c7-ATP1B1-NRG1 and Treatment Time (Days) H6c7-SLC3A2-NRG1 pancreatic cells with seribantumab resulted in a dose-dependent inhibition of HER3 D E Afatinib 5 mg/kg and AKT phosphorylation. Tumor growth inhibition was observed after administration of 5 mg or 10 mg Vehicle Afatinib (5 mg/kg QD) Seribantumab 2.5 mg + Afatinib 5 mg/kg 800 Seribantumab (2.5 mg BIW) 800 Seribantumab-5 mg+AG120-150 mg/kg Seribantumab (5 mg BIW) Seribantumab-10 mg BIW+Afatinib 5 mg/kg + twice weekly [BIW] seribantumab to a PDX mouse model of pancreatic adenocarcinoma with an APP-NRG1 600 600 Seribantumab (10 mg BIW) AG120-150mg/kg rearrangement (CTG-0943). The two doses of seribantumab were more effective than afatinib (5 mg/kg 400 400 200 200 QD), a pan-ERBB inhibitor, in this model, causing tumor shrinkage of up to 55% (23-77% range). There was 0 0 0 0 no regression of afatinib-treated pancreatic PDX tumors. After treatment, residual CTG-0943 tumors were -20 -20 -40 -40 extracted for Western blotting analysis (day 24 for vehicle, and day 31 or 32 for the seribantumab and -60 -60 -80 -80 Change in Tumor Volume (%) Volume Change in Tumor Change in Tumor Volume (%) Volume Change in Tumor -100 afatinib-groups, respectively). Loss of phosphorylated and total EGFR, HER2 and HER3, cyclin D1, etc. in -100 seribantumab-treated tumors at the end of the study suggests loss of the majority of human tumor cells in Figure 2. A. H6C7-EV (empty vector) and H6C7-ATP1B1-NRG1 cells were profiled for activated intracellular kinases using phospho-proteomic Figure 4. A. Representative H&E-stained CH-17-0068 PDX tumor. B. Genomic alterations identified by RNAseq and the xenograft tumors. This was confirmed using a human-specific GAPH antibody. arrays. B. H6C7-EV and H6C7 cells with the indicated NRG1 fusions were profiled for activated receptor tyrosine kinases (RTK) using phospho-RTK corresponding investigational targeted therapies. C. Mice bearing CH-17-0068 PDX tumors (5-6 mice per group) were arrays. Quantitation of phosphorylated EGFR, HER2, and HER3 is shown. C. Western blot analysis of cell extracts from H6C7-EV and H6C7 cells Seribantumab was further evaluated in an intrahepatic cholangiocarcinoma PDX model with an RPBMS- treated with seribantumab or afatinib monotherapy for 30 days. Afatinib (5 mg/kg QD) or AG-120 (isocitrate dehydrogenase expressing NRG1 fusions. D and E. Cells were treated with the indicated concentrations of seribantumab and then whole-cell lysates profiled for NRG1 fusion as well as mutations in ERBB4 and IDH1 (CH-17-0068). While monotherapy seribantumab (5 [IDH] inhibitor, 150 mg/kg twice daily [BID]) were then added to the indicated groups. Results represent the mean ± SEM. the indicated phospho- and total proteins by Western blotting. All cells were serum-started for 24 hours prior to experimentation. mg and 10 mg per dose, BIW) was equally effective as afatinib (5 mg/kg once daily [QD]) in this model, Change in the volume of individual tumors on day 30 (D) or at the end of the study (E). enhanced tumor regression was observed with combination therapy. The triple combination of seribantumab 10 mg BIW with afatinib and AG-120, an IDH inhibitor, led to regressions in the majority of Seribantumab inhibits growth of NRG1-rearranged pancreatic Summary and Conclusion tumors. Allometric scaling (based on FDA guidelines) indicates that 5 mg/kg afatinib in mice is equivalent to adenocarcinoma PDXVehicle model Vehicle(CTG-0943, APP-NRG1) a human dose of approximately 50 mg daily. • NRG1 fusions are rare but recurrent oncogenic drivers in GI cancers.1,2 Afatinib (5 mg/kg QD)Afatinib (5 mg/kg QD) Conclusions. Our results here suggest that seribantumab is effective at reducing tumor growth in A B VehicleSeribantumab (5 mg SeribantumabBIW) (5 mg BIW) D • Overexpression of NRG1 fusions in immortalized human pancreatic ductal epithelial H6C7 2500 AfatinibSeribantumab (5 mg/kg2500 QD) (10 mgSeribantumab BIW) (10 mg BIW) preclinical models of GI cancers with NRG1 fusions. These data support the use of monotherapy Seribantumab) (5 mg BIW) 3 cells activated HER3 and AKT. ) 2000 3 25002000 Seribantumab (10 mg BIW) seribantumab to treat GI and other cancers uniquely driven by an NRG1 fusion in the ongoing phase 2 • Seribantumab inhibits HER3 and AKT phosphorylation in H6C7 cells with NRG1 fusions. ) 3 1500 CRESTONE study (NCT#04383210). 20001500 1000 • Treatment of NRG1 fusion-positive pancreatic PDX model with seribantumab inhibits 1500 1000 500 tumor growth at clinically achievable doses. Residual tumor xenografts show depleted Prevalence of NRG1 fusions in GI tumors 40X 1000 Tumor Volume (mm 500 0 Tumor Volume (mm human tumor cell content when assessed by Western blotting. 500 Tumor Volume (mm 0 5 10 15 20 25 30 35 0 Treatment Time (Days) 0 • Investigation of a cholangiocarcinoma PDX model with three genomic alterations (NRG1 0 0 5 5 10 1015 1520 2520 3025 3530 35 TreatmentTreatment Time (Days)Time (Days) fusion, and ERBB4 and IDH1 mutations) suggests that treatment of NRG1 fusion-driven C Vehicle Seribantumab (5 mg BIW) tumors harboring additional oncogenic drivers may require combination therapy to Afatinib (5 mg/kg QD) Seribantumab (10 mg BIW) 100X 1000 address the contribution of each genomic alteration in disease progression. 800 • These data support the use of monotherapy seribantumab to treat GI and other cancers 600 400 uniquely driven by an NRG1 fusion in the ongoing phase 2 CRESTONE study 200 0 (NCT#04383210). 0 -50 200X -100 Change in Tumor Volume (%) Volume Change in Tumor References Acknowledgements Figure 3. Mice bearing CTG-0943 PDX tumors (5-8 mice per group) were treated with indicated agents. A. Representative H&E-stained slides of a 1. Jonna S et al. Clin Cancer Res. 2019; This study was partially funded by grants vehicle-treated tumor. B. Tumor volume, results represent the mean ± SEM. Animals in the seribantumab 5 mg and 10 mg groups were administered seribantumab 5 mg/kg and 10 mg/kg, respectively, for the first two doses. C. Change in the volume of individual tumors (vehicle: day 25:4865‒4867. from Elevation Oncology and Cycle for Figure 1. Pancreatic cancer, biliary cancer (cholangiocarcinoma, gall bladder), and colorectal 20; treated groups: day 31) compared with day 0 (%). D. Western blot analysis of vehicle, and afatinib and seribantumab-treated tumors.