Targeted therapy offers an opportunity for personalized medicine that's specific for a patient's tumor, but the hyper-focused treatment creates possibilities for cells to mutate and become resistant to the therapy.

"If we can understand the mechanisms of intrinsic resistance, that will help us with better patient selection. For each patient and for each specific alteration, we may be able to better select the optimal agent. We may be able to understand if combinations can help prevent resistance," Funda Meric-Bernstam, chair of the department of investigational cancer therapeutics at MD Anderson Cancer Center, told the audience at the American Association for Cancer Research Virtual Annual Meeting II.

Mirati Therapeutics Inc.'s MRTX-849 is still a ways away from regulatory approval, but the company is already looking at potential drivers of resistance to the KRAS-G12C inhibitor.

"Personally, I don't think the industry goes far enough or early enough in preclinical development to address resistance as many tumors are able to escape therapy, including targeted therapies in biomarker-selected patient populations," Lars Engstrom, principal scientist at San Diego-based Mirati, told the audience.

Engstrom used a CRISPR library to knock out 938 targeted genes in cell lines with the KRAS-G12C mutation. The cells were then selected in cell culture exposed to MRTX-849 as well as tumors grown on mice exposed to MRTX-849.

Looking at the resulting clones, Engstrom found genes that were underrepresented compared to the original library, including EGFR family, SHP2, SOS1, mTOR and CDK4/6, suggesting potential combination targets. Indeed, in cell lines, MRTX-849 had a synergistic effect with the HER inhibitor afatinib, the SHP2 inhibitor RMC-4550, the mTOR inhibitor vistusertib (Astrazeneca plc) and the CDK4/6 inhibitor Ibrance (palbociclib, Pfizer Inc.).

The screen also allowed Engstrom to find clones that were enriched, including KEAP1, NF1, Rb1, TSC1/2 and PTEN, suggesting a growth benefit and that they could confer resistance to MRTX-849 if mutated in patients.

KEAP1 is particularly worrisome because it's often seen in non-small-cell lung cancer (NSCLC), one of the tumor types Mirati is targeting since KRAS-G12C is found in 14% of NSCLC. In an animal model, KEAP1 knockout tumors regressed, but eventually demonstrated adaptative resistance after two to three weeks of treatment.

"Importantly, one of our first patients with a confirmed response on the trial harbored a KEAP1 mutation, providing clinical evidence that this mutation does not broadly confer intrinsic resistance to 849," Engstrom noted.

Resistance to one class confers sensitivity to another

To better understand the resistance to second-generation tropomyosin receptor kinase inhibitors (TRKi) that are designed to treat NTRK gene fusions, Emiliano Cocco, a postdoctoral fellow in Maurizio Scaltriti's lab at Memorial Sloan Kettering Cancer Center, looked at patients who were treated with first-generation TRKi who became resistant after generating a G595R mutation and then became resistant to the second-generation TRKi selitrectinib (Bayer AG).

Cocco found three different patients with sarcoma, breast cancer and colorectal cancer who had acquired mutations at G667 in the XDFG codon of TRKA, the TRK that's encoded by the NTRK gene. Cocco also found mutations at G667 in TRKA in the pretreated sample of two other patients who had rapid progression on selitrectinib. Modeling of the mutations found they caused resistance by generating steric hindrance that partially blocks binding of the drug.

Interestingly, the G667C change resulted in the mutant protein being more sensitive to type II kinase inhibitors Cabometyx (cabozantinib, Exelixis Inc.), foretinib and Iclusig (ponatinib, Takeda Pharmaceutical Co. Ltd.). The drugs had higher activity in kinase assays for the G667C mutant protein than the protein with the first-generation G595R mutation. Cabometyx and foretinib also bound substantially tighter to the G667C mutant protein compared to selitrectinib.

Cocco showed cell lines with the double G595R/G667C mutation were highly sensitive to all three type II kinase inhibitors. And xenografts with G667C mutations grown on animals were highly sensitive to Cabometyx.

PARP1 inhibitor resistance

Mei-Kuang Chen, a recent PhD graduate at the University of Texas MD Anderson Cancer Center, made a BRCA1-mutated triple-negative breast cancer (TNBC) cell line resistant to poly (ADP-ribose) polymerase (PARP) inhibitors by gradually increasing the dose of Talzenna (talazoparib, Pfizer).

The resulting clones had comparable resistance to other TNBC cell lines that are known to be resistant to Talzenna and were also resistant to other PARP inhibitors, Lynparza (olaparib, Astrazeneca/Merck & Co. Inc.), veliparib (Abbvie Inc.) and Rubraca (rucaparib, Clovis Oncology Inc.).

Looking at kinase activity, Chen discovered that EGFR and MET activity was highly increased in one of the resistant clones. In that clone, PARP was phosphorylated compared to the parental cell line. The combination of Talzenna and the tyrosine kinase inhibitor Xalkori (crizotinib) had a synergistic effect on survival in multiple clones that came out of the screen.

PARP phosphorylation was also seen in other cell lines that are resistant to PARP inhibitors, and treatment with Xalkori inhibited the PARP phosphorylation in breast, lung and ovarian cell lines, further supporting the mechanism of resistance.

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