ABCB1-Mediated Drug Efflux Drives Resistance to VpreB1-Targeted Antibody-Drug Conjugates in B-cell Lymphoblastic Leukemia

Although B-cell acute lymphoblastic leukemia (B-ALL) is highly responsive to antigen-directed immunotherapies, treatment resistance remains a major barrier to achieving durable responses in patients. We recently developed a novel VpreB1 (CD179a)-directed antibody-drug conjugate with calicheamicin (VpreB1-ADC) that exploits the restricted expression of VpreB1 within the surrogate light chain in early B cells, including B-ALL. In the present work, we investigated mechanisms of resistance to the VpreB1-ADC. Mechanisms of resistance were evaluated using a TCF3::HLF B-ALL model, assessing target engagement parameters including VpreB1 surface expression and antibody internalization. The role of the multidrug resistance transporter ABCB1 (P-glycoprotein) was evaluated via pharmacologic inhibition, using tariquidar and zosuquidar, and enforced overexpression across multiple B-ALL cell lines. Sensitivity to alternative non-ABCB1 substrate payloads exatecan and PNU-159682 was also assessed. Resistant TCF3::HLF cells retained VpreB1 expression and efficient antibody internalization. Instead, resistance was driven by elevated ABCB1 expression and activity. ABCB1 inhibition with tariquidar or zosuquidar restored VpreB1-ADC sensitivity. Conversely, enforced ABCB1 overexpression conferred ADC resistance, which was reversed by ABCB1 inhibition. Cells with high ABCB1 activity remained fully sensitive to alternative payloads, including exatecan and PNU-159682, which are not ABCB1 substrates. ABCB1-mediated drug efflux drives intrinsic resistance to calicheamicin-conjugated ADCs in B-ALL. Combining ADCs with ABCB1 inhibitors or selecting payloads non-susceptible to ABCB1 efflux offer viable strategies to overcome resistance and optimize future ADC therapies.