Genome-wide CRISPR/Cas9 Screens Reveal Shared and Bespoke Mechanisms of Resistance to SHP2 inhibition

SHP2 (PTPN11) acts upstream of SOS1/2 to enable RAS activation. Allosteric inhibitors (SHP2is) stabilize SHP2 auto-inhibition, preventing activation by upstream stimuli. SHP2is block proliferation of RTK- or cycling RAS mutant-driven cancers and overcome adaptive resistance to other RAS-ERK pathway drugs. Several SHP2is are in clinical trials. To identify potential SHP2i resistance mechanisms, we performed genome-wide CRISPR/Cas9 knockout screens on two SHP2i-sensitive AML cell lines and recovered genes expected to cause resistance, including tumor suppressor (NF1, PTEN, CDKN1B) and "RASopathy" (LZTR1, RASA2) genes, and several novel targets (INPPL1, MAP4K5, epigenetic modifiers). We then screened 14 cancer lines with a focused CRISPR library targeting common "hits" from the genome-wide screens. LZTR1 deletion conferred resistance in 12/14 lines, followed by MAP4K5 (8/14), SPRED2 (6/14), STK40 (6/14), and INPPL1 (5/14). INPPL1, MAP4K5, or LZTR1 deletion reactivated ERK signaling. INPPL1-mediated sensitization to SHP2i required its NPXY motif but not its lipid phosphatase domain. MAP4K5 acted upstream of MEK via a kinase-dependent target(s), whereas LZTR1 showed cell-dependent effects on RIT and RAS stability. INPPLI, MAP4K5, or LZTR1 deletion also conferred SHP2i resistance in mice. Our results reveal multiple SHP2i resistance genes, emphasizing the need for detailed understanding of the resistance landscape to arrive at effective combinations.