Albumin-conjugated drugs attain KRAS mutant
cancer targeting through KRAS-enhanced macropinocytosis and intensified lysosomal degradation due to reduced
neonatal Fc receptor (FcRn) expression. The cytosolic delivery of active payloads relies on endocytosis and subsequent intracellular processing of
albumin delivery vehicles, wherein complex regulatory mechanisms and molecular machineries are closely involved. Despite the obvious merit of KRAS targeting, could such an endocytic process involving extra molecular regulators also bring about extra vulnerabilities to
albumin-conjugated drugs, particularly, unexpected drug resistance? To assess such risks, here we performed an unbiased drug resistance mechanism comparison in
pancreatic cancer, between free
triptolide (TP, a potent
cytotoxin) and
albumin-conjugated TP, using genome-wide CRISPR-Cas9 loss-of-function screens. GTF2H5, a subunit of GTF2H
transcription factor complex, was the only hit identified regardless of forms of TP treatment. With
drug efficacy tests on GTF2H5 knockout clones, we further concluded that GTF2H5 deficiency conferred drug resistance primarily due to the pharmacological mechanism of action (MoA) of TP. In addition, molecules previously considered to be able to affect endocytosis and intracellular processing were not enriched during the screening with
albumin-conjugated TP. With the aid of genome-wide CRISPR-Cas9 loss-of-function screens, we conclude that the pharmacological resistance of the active payload, rather than any potential loss-of-function mutations in endocytic molecular machineries, is the solely crucial drug resistance mechanism of
albumin-conjugated drugs.