97P - Bcl-xL prevents the arginine starvation induced by PEGylated arginine deiminase (ADI-PEG20) from inducing apoptosis

Background

Multiple clinical trials are using PEGylated arginine deiminase (ADI-PEG20) to induce arginine starvation in cancers that are argininosuccinate synthetase (ASS1) low. ASS1 low tumors include sarcomas, melanomas, non-small cell and small cell lung cancers, triple-negative breast, pancreatic cancer, and many others. While ADI-PEG20 clearly induces an arginine starvation response, no cell death is initiated at a meaningful level. While many studies have seen low levels of apoptosis (<10%), the primary effect of ADI-PEG20 is cytostatic, as tumors metabolically reprogram themselves by upregulating ASS1, gaining the ability to produce arginine.

Methods

Cell lines were treated with ADI-PEG20 and A-1331852 (Bcl-xL inhibitor) and monitored for cell death using the Incucyte. CRISPR models confirmed that the Bcl-xL effects of A-1331852 were on target. CDK2 activity, Bcl-xL and Mcl-1 were measured in response to ADI-PEG20 using Wes capillary electrophoresis. These cell lines were grafted into mice and treated with ADI-PEG20 and A-1331852.

Results

Arginine starvation inhibits CDK2, resulting in the loss of Mcl-1 expression. Neither Bcl-2 nor Mcl-1 inhibitors initiated cell death when combined with ADI-PEG20. Dual treatment of Bcl-xL inhibitor A-1331852 with ADI-PEG20 led to mitochondrial BAX levels increasing fivefold, corresponding to a drop in cytoplasmic levels that suggests translocation to the mitochondria. A similar but less dramatic pattern of significant increase in mitochondrial levels was observed with BAK. Inhibition of Bcl-xL with ADI-PEG20 induced CASP3/7 and PARP cleavage, resulting in apoptosis in a panel of ASS1 low cell lines. This was confirmed in vivo.

Conclusions

ADI-PEG20 does not induce cell death as a monotherapy, but the arginine starvation instead results in metabolic adaptation. This begins to explain the lack of efficacy seen in early ADI-PEG20 monotherapy trials. Bcl-xL, but not Mcl-1 or Bcl-2, prevents ADI-PEG20 from inducing apoptosis. Bcl-xL inhibition synergies with ADI-PEG20 to induce apoptosis. This study provides the preclinical rationale for combining ADI-PEG20 with next generation Bcl-xL inhibitors such as the Bcl-xL PROTAC in a phase I clinical trial.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Polaris, NIH.

Disclosure

B.A. Van Tine: Financial Interests, Personal, Invited Speaker, Educational Speaker: Targeted Oncology; Financial Interests, Personal, Advisory Board, Also, Travel paid to conference to present abstract: Adaptimmune; Financial Interests, Personal, Invited Speaker, Also attended an Ad Board meeting, Travel was paid to present abstract at conference: GSK; Financial Interests, Personal, Other, Consulting, Also attended and Ad Board meeting, Travel was paid to attend an Ad board meeting: Epizyme; Financial Interests, Personal, Other, Consulting- ADRx working on a cancer project and they are requesting my expertise: ADRx; Financial Interests, Personal, Advisory Board, tenosynovial giant cell tumors (TGCT): Ayala Pharmaceuticals; Financial Interests, Personal, Other, Consulting/Advisor: Cytokinetics; Financial Interests, Personal, Other, Consulting: Bayer; Financial Interests, Personal, Other, 60 minute interview regarding Synovial Sarcoma: Bionest Partners; Financial Interests, Personal, Other, OncLive Virtual Workshop: Intellisphere; Financial Interests, Personal, Advisory Board, Attended Advisory Board Meeting: Apexigen; Financial Interests, Personal, Advisory Board, Attended an Advisory Board Meeting: Daiichi Sankyo, Deciphera Pharmaceuticals; Financial Interests, Personal, Advisory Board, Attended an Advisory Board Meeting: Novartis; Financial Interests, Personal, Advisory Board: Lilly, PTC Therapeutics; Financial Interests, Personal, Invited Speaker, Sigma-2 Receptor Ligands and Therapeutic uses therefor (006766), Modular Platform for Targeted Therapeutic Delivery (006755), Sigma-2 Receptor Ligand Drug Conjugates as Antitumor Compounds, Methods of synthesis and Uses Thereof (014229): Accuronix Therapeutics; Financial Interests, Institutional, Research Grant: Pfizer, Merck, Tracon Pharm, GSK; Non-Financial Interests, Personal, Invited Speaker: Polaris. All other authors have declared no conflicts of interest.