Combination of IGF-1R Targeted Alpha Therapy with Olaparib Results in Synergistic Efficacy Against Colorectal and Lung Cancer Xenografts

Abstract Abstract Number: LB130 Combination of IGF-1R Targeted Alpha Therapy with Olaparib Results in Synergistic Efficacy Against Colorectal and Lung Cancer Xenografts. M Hu1, J Forbes1, R Simms1, Y Storozhuk1, J Valliant1, E Burak1 1Fusion Pharmaceuticals, Hamilton, Ontario, Canada Objectives: FPI-1434 is a targeted alpha-particle therapeutic consisting of an IGF- 1R targeting antibody radiolabeled with actinium-225. The primary mechanism of action for FPI-1434 is induction of double-stranded DNA breaks (DSB) in targeted tumors resulting in cell death. Poly (ADP-ribose) polymerase (PARP) is part of the cellular mechanism that repairs DSB. In cancer patients with genetic defects in DSB repair (e.g. BRCA1/2), the PARP pathway becomes a primary repair system and its inhibition results in cell death. This mechanism has been leveraged as a therapy against DNA-repair deficient tumors leading to FDA-approval of PARP inhibitors (PARPi) including olaparib. Treatment with PARPi to block repair of DNA damage driven by FPI-1434 may act synergistically to increase the lethal DNA damage load. To that end, Fusion has performed studies to combine FPI-1434 and olaparib against tumor models with no pre-disposing defects in DNA repair. Methods: Colorectal (Colo-205) or lung (A549) cancer xenografts were established in Balb/c nude mice. For efficacy studies, a dose combination matrix for FPI-1434 and olaparib was tested. FPI-1434 was dosed (i.v.) once at 20-200 nCi followed with olaparib (i.p.) at doses of 0-50 mg/kg. Olaparib was dosed 24h after FPI-1434 using a 5 day on/2 day off schedule (5 mice/group). DSB formation (γH2AX) and apoptosis (cleaved caspase 3) were evaluated in treated tumors by IHC staining. Results: DSB formation and induction of apoptosis were detected in FPI-1434 treated tumors in a time and dose dependent manner. DSB formation was observed in all areas of tumor containing intact or apoptotic cancer cells confirming FPI-1434 mechanism. In tumor efficacy studies, olaparib had no single-agent efficacy in the Colo-205 or A549 models. Single doses of FPI-1434 at 20 nCi had no effect on Colo-205 tumors, suppressed growth at 50 nCi and caused regression at 100 nCi. Combination efficacy was seen in the 20 and 50 nCi dose groups, including olaparib. Doses of 20 nCi FPI-1434 and 25 mg/kg olaparib had the strongest combined effect in this model. In the A549 model, single doses of FPI-1434 had no effect at 20 or 50 nCi, caused growth suppression at 100 nCi and regression at 200 nCi. Combination efficacy was seen in the 20, 50 and 100 nCi dose groups, including olaparib. Doses of 50 nCi FPI-1434 and 25 mg/kg olaparib had the strongest combined effect in this model. In general, the strongest efficacy was observed by combining ineffective single-agent doses of the two compounds. Strong efficacy of high dose FPI-1434 masked observable combination effects suggesting that FPI-1434 was the therapeutic driver. Conclusions: Olaparib co-dosing enhanced FPI-1434 efficacy which supports consideration of this combination for clinical use. Ineffective single-agent doses, when combined, resulted in synergistic efficacy in both Colo-205 and A549 models suggesting that the mechanism can be applied to multiple cancer types where predisposing mutations in DNA repair are lacking. The strongest combination effect appeared to occur at the low single-agent doses suggesting that PARP inhibition may potentiate efficacy at lower clinical doses of FPI-1434. 5 30 39 37 10 9 11 53 0 10 20 30 40 50 60 Untreated 24H 96H 168H Normalized Nuclei Count (%) Foci γH2AX Pan-Nuclear γH2AX 14 39 50 90 8 15 27 36 0 10 20 30 40 50 60 70 80 90 100 Untreated 24H 96H 168H Normalized Nuclei Count (%) Total yH2AX CC3 Background Emw [CC BY-SA 3.0 (https://creativecommons.org/licenses/by-sa/3.0)] IGF-1R is expressed on nearly all tumor types Breast Colorectal Prostate Recurrent Thyroid Sarcomas NSCLC Pancreatic / NETs Adrenocortical carcinoma HCC H&N Type I insulin-like growth factor receptor (IGF-1R) is a transmembrane protein which is overexpressed in solid tumors including HNSCC, gynecological cancers, and Her2-negative breast cancers. FPI-1434 is a radioimmunoconjugate consisting of a humanized monoclonal antibody that binds to the external domain of IGF-1R, a proprietary bifunctional chelate, and the alpha-emitting radionuclide actinium-225 (Ac-225). Internalization of the radioimmunoconjugate and decay of Ac-225 causes tumor cell death primarily through induction of double-stranded DNA breaks. FPI-1434 Mechanism of Action FPI-1434 (Single dose, 400 nCi) Induced Double-Stranded DNA Breaks in Colo-205 Xenografts as Determined by γH2AX FPI-1434 (Single dose, 400 nCi) Induced Apoptosis in Colo- 205 Xenografts as Determined by Cleaved Caspase 3 (CC3) 20 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 40 50 0 2 4 6 8 10 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (20 nCi) 20 nCi + 25 mg/kg 20 nCi + 50 mg/kg 50 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 40 50 0 2 4 6 8 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (50 nCi) 50 nCi + 25 mg/kg 50 nCi + 50 mg/kg 100 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 40 50 0 2 4 6 8 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (100 nCi) 100 nCi + 25 mg/kg 100 nCi + 50 mg/kg FPI-1434 Dose Level 20 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 0 2 4 6 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (20 nCi) 20 nCi + 25 mg/kg 20 nCi + 50 mg/kg 50 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 0 2 4 6 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (50 nCi) 50 nCi + 25 mg/kg 50 nCi + 50 mg/kg 100 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 0 2 4 6 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (100 nCi) 100 nCi + 25 mg/kg 100 nCi + 50 mg/kg 200 nCi FPI-1434 + Olaparib Days after treatment initiation Relative tumor volume (RTV) 0 10 20 30 0 2 4 6 Vehicle Olaparib (25 mg/kg) Olaparib (50 mg/kg) FPI-1434 (200 nCi) 200 nCi + 25 mg/kg 200 nCi + 50 mg/kg FPI-1434 Dose Level FPI-1434 + Olaparib Therapy: Combined Efficacy at Low Single-Agent Doses (Colo-205, Colorectal Cancer Xenograft Model) FPI-1434 + Olaparib Therapy: Combined Efficacy at Low Single-Agent Doses (A549, NSCLC Xenograft Model) Summary and Conclusions IGF-1R has been implicated in:  Increased cellular proliferation  Metastatic potential  Cell survival  Chemotherapy and radiotherapy resistance 0 4 8 12 16 20 24 28Day FPI-1434: 20, 50 or 100 nCi (i.v.) Olaparib: 0, 25 or 50 mg/kg (i.p.) Dose Schedule 0 4 8 12 16 20 24 28Day FPI-1434: 20, 50, 100 or 200 nCi (i.v.) Olaparib: 0, 25 or 50 mg/kg (i.p.) Dose Schedule Delivery of alpha-particle radiation by FPI-1434 resulted in dose-dependent regression and induced double-stranded DNA breaks and apoptosis in tumor xenografts Combination of FPI-1434 and olaparib resulted in efficacy in the Colo-205 model and in the radioresistant A549 model ■ Olaparib treatment alone was not efficacious; models tested lack mutations in DNA repair pathways required for sensitivity to PARPi monotherapy ■ Strongest combination effects occurred at doses where FPI-1434 was not effective as a single agent Colo-205: 20 nCi FPI-1434 + 25 mg/kg olaparib A549: 50 nCi FPI-1434 + 25 mg/kg olaparib FPI-1434 effect dominated at the highest single agent doses suggesting it was the therapeutic driver of the combination Olaparib co-dosing enhanced FPI-1434 therapeutic efficacy which supports consideration of this combination for clinical use ■ PARP inhibition while using FPI-1434 as the therapeutic driver of DSB formation can be applied to multiple cancer types where predisposing mutations in DNA repair are lacking ■ PARP inhibition may potentiate efficacy at lower clinical doses of FPI-1434 Delivery of alpha-particle radiation by FPI-1434 induced DSB in treated tumor xenografts (γH2Ax detected by IHC on fixed tumor sections) ■ DSB formation was time-dependent, dose dependent, and was consistent with kinetics of FPI-1434 tumor accumulation (data not shown) Delivery of alpha-particle radiation by FPI-1434 induced apoptosis in treated tumor xenografts (CC3 detected by IHC on fixed tumor sections) ■ Apoptosis was time dependent, following kinetics of tumor accumulation by FPI-1434 (data not shown), and was concomitant with induction of DSB ■ Apoptosis increased in conjunction with FPI-1434 dosage (data not shown) Untreated Control PARP inhibition via olaparib co-therapy is a rational approach to potentiate tumor DSB formation driven by FPI-1434 treatment Untreated Control

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