Ectonucleotide phosphodiesterase 1 (ENPP1) Inhibitor: Treating anti-PD-1/-L1 resistant cancers
Assays Completed
Owned and Available for Licensing
In vivo efficacy studies –Combo study with
anti-PD-1, or anti-PD-L1
In vivo efficacy studies – Single agent
In vivo PK-PD
In vivo PK studies
In vitro ADMET studies
In vitro cell-based
Toxicology studies
Target Rationale
Therapeutic monoclonal antibodies targeting immune checkpoints (ICPs) have changed the treatment landscape of many tumors. However, response rates remain relatively low in most cases. A major factor involved in initial resistance to ICP inhibitors is the lack or paucity of tumor T cell infiltration, which characterizes the so-called "cold tumor" subset. The main mechanisms which contribute to the absence of T cell infiltration include lack of tumor-associated antigens, defects in antigen presentation, absence of T cell activation and deficits in T cell homing to the tumor bed.

ENPP1, an ecto-nucleotide pyrophosphatase, was discovered as the predominant enzyme hydrolyzing cGAMP, the activator of STING. ENPP1 inhibitors have been designed to inhibit two distinct signaling pathways in the tumor microenvironment: A. ENPP1-mediated downregulation of STING dependent anti-tumor innate immune responses by hydrolysis of cGMAP, a crucial second messenger and potent activator of the STING pathway, and B. ENPP1-mediated production of adenosine, a key immune suppressive molecule in the TME that promotes tumor progression.
ENPP1 inhibition would achieve the dual purpose of reducing extracellular levels of the immune suppressor adenosine, while simultaneously increasing extracellular levels of the immunostimulatory STING pathway activator, cGAMP.
PNAS (2023) 120 (52) e2313693120 ; Cancer Discov (2021) 11 (5): 1212–1227.

Insilico Medicine ENPP1 Inhibitor Summary – IND-Enabling
Novel structure generated by AI
Differentiated structure vs known competitor molecules
Potent in vitro activity and in vivo efficacy
  • The most potent candidate in both cGAMP/ATP hydrolysis inhibition
  • ISM candidate showed efficacy in both MC38 and CT26 syngeneic mice models
Promising drug-ability as an oral agent
  • Good in vitro ADME profiles
  • Promising PK profile across different preclinical animal species
Safety margin
  • Promising safety margin in 28-day rat and dog DRF studies
  • No significant body weight change in MC38 mouse models at 30 mpk p.o. BID
  • No effect on mortality, hematology, serum chemistry, ECG or gross pathology

ICB (Immune Checkpoint Blockade) works in only 10-30% of all cancer patients. In colorectal cancer (CRC), PD-1/-L1 antibodies are efficacious in MSI-high patients, which make up around 15% of CRC tumors. Up to 80% of CRC is microsatellite stable (MSS) and considered to be chromosomally unstable (CIN+). Tumors with high chromosome instability generate micronuclei, the rupture of which releases DNA into the cytosol, which is sensed by cGAS, leading to the production of the anti-tumor STING pathway activator, cGAMP. ENPP1 (which hydrolyses cGAMP, thus suppressing STING activation) is often highly expressed in these tumors and correlates with poor patient outcomes and resistance to anti PD-1/PD-L1 antibodies.

By driving STING pathway activity, inhibition of ENPP1 may therefore represent a useful treatment for a subset of anti-PD-1/L-1 resistant tumors that exhibit reduced tumor T cell activity and infiltration.

The combination of ENPP1 inhibitors with anti-PD-1/PD-L1 antibodies has great potential in immuno-oncology.
Combination therapy with anti-PD-1/PD-L1 antibodies:
  • ENPP1 inhibitor as a tumor microenvironment modulator resulted in increased T cell infiltration into solid tumors
  • Enhanced efficacy for anti-PD-1/PD-L1 antibodies in both MSS and MSI-h cancers

Project Status – IND-Enabling
In vitro data showed Insilico Medicine's ENPP1 inhibitor potently inhibiting the target and simultaneously causing IFNβ induction in both human and mice cell lines with excellent selectivity. In vivo efficacy studies in CT26 and MC38 syngeneic mouse models showed strong anti-tumor activity and robust tumor regression in combination with immune checkpoint inhibition.
In a MC38 syngeneic mice model, a single dose of Insilico Medicine's (ISM) compound showed 67% tumor growth inhibition.
In combination with anti-PD-L1 antibody, ISM cpd showed dose dependence from 3mpk to 30mpk PO. BID.
Tumor free animals were found in the combo group.