P-glycoprotein (P-gp)-induced drug resistance is a major road block for successful cancer chemotherapy. Through phenotypic screening, the compound 2-(2-chlorophenylimino)-5-(4-dimethylaminobenzylidene) thiazolidin-4-one (CDBT) was discovered to have potent anti-tumor activity in P-gp over-expressing drug-resistant non-small-cell lung cancer (NSCLC) H460_TaxR cells. Here, we report mechanistic investigations of the P-gp–evading anti-tumor activity of CDBT. CDBT is evidently not a P-gp substrate and escapes the P-gp efflux pump. As a novel microtubule and heat shock protein 90 (HSP90) dual targeting inhibitor, CDBT causes the destabilization of microtubules and degradation of HSP90 client proteins CRAF-1 and ERBB2, resulting in cell cycle arrest at the G2/M phase and apoptosis. Furthermore, CDBT effectively inhibits tumor growth by 60.4% relative to the vehicle control after intraperitoneal administration at 30 mg/kg for 11 days and shows no toxicity in normal tissues in the NSCLC H460_TaxR xenograft mouse model. Our data suggest a novel drug discovery strategy to combat P-gp over-expressing drug-resistant NSCLC cancer cells with a single therapeutic agent.