Lentiviral-based mutagenesis to identify mutations that confer resistance to anti-cancer drugs [umi]

Identifying resistance mutations in a drug target provides crucial information. Lentiviral transduction creates multiple types of mutations due to the error-prone nature of the HIV-1 reverse transcriptase (RT) and we show this property can be leveraged to identify mutations that confer resistance to targeted anti-cancer drugs, a technique we term “LentiMutate”. First, we improved LentiMutate by making the lentiviral RT more error-prone. Next, we applied this technique to two anti-cancer drugs, imatinib and AMG 510. We find novel deletions in BCR-ABL that confer resistance to BCR-ABL inhibitors and point mutations in the AMG 510 binding pocket or oncogenic non-G12C mutations, in KRAS-G12C or wild-type KRAS, respectively, that confer resistance to AMG 510. LentiMutate may prove highly valuable to clinical and preclinical cancer drug development Overall design: HCC827 cells were infected with lentiviruses harboring the EGFRdel746-750 cDNA packaged using either the M- or WT-reverse transcriptase (RT). Lentiviral integrants in these cells were PCR amplified using primers with unique molecular identifiers to determine the mutation rate in the EGFR cargo when packaged using the M-RT or WT-RT.