Supplementary Figures S1-S6 from SLC46A3 Is Required to Transport Catabolites of Noncleavable Antibody Maytansine Conjugates from the Lysosome to the Cytoplasm

Hamblett, Kevin J.; Jacob, Allison P.; Gurgel, Jesse L.; Tometsko, Mark E.; Rock, Brooke M.; Patel, Sonal K.; Milburn, Robert R.; Siu, Sophia; Ragan, Seamus P.; Rock, Dan A.; Borths, Christopher J.; O'Neill, Jason W.; Chang, Wesley S.; Weidner, Margaret F.; Bio, Matthew M.; Quon, Kim C.; Fanslow, William C.

doi:10.1158/0008-5472.22406858.v1

Supplementary Figures S1-S6 from SLC46A3 Is Required to Transport Catabolites of Noncleavable Antibody Maytansine Conjugates from the Lysosome to the Cytoplasm

journal contribution

posted on 2023-03-30, 23:22 authored by Kevin J. Hamblett, Allison P. Jacob, Jesse L. Gurgel, Mark E. Tometsko, Brooke M. Rock, Sonal K. Patel, Robert R. Milburn, Sophia Siu, Seamus P. Ragan, Dan A. Rock, Christopher J. Borths, Jason W. O'Neill, Wesley S. Chang, Margaret F. Weidner, Matthew M. Bio, Kim C. Quon, William C. Fanslow

Predicted topology for SLC46A3 (S1); Effect of siRNA on mRNA expression of SLC46A3 and CD70 in 786-0 cells (S2); Effect of siRNA on CD70 protein expression in 786-0 cells (S3); Anti-CD70 Ab-DSG-Maytansine and anti-CD70 Ab-Cys-Maytansine are potent against CD70 expressing 786-0 Cells (S4); Lysine-MCC-DM1 concentration time course in lysosome and total cell lysate (S5); Confluency of 786-0 cells transfected with siRNA (S6).

History

ARTICLE ABSTRACT

Antibody–drug conjugates (ADC) target cytotoxic drugs to antigen-positive cells for treating cancer. After internalization, ADCs with noncleavable linkers are catabolized to amino acid-linker-warheads within the lysosome, which then enter the cytoplasm by an unknown mechanism. We hypothesized that a lysosomal transporter was responsible for delivering noncleavable ADC catabolites into the cytoplasm. To identify candidate transporters, we performed a phenotypic shRNA screen with an anti-CD70 maytansine-based ADC. This screen revealed the lysosomal membrane protein SLC46A3, the genetic attenuation of which inhibited the potency of multiple noncleavable antibody–maytansine ADCs, including ado-trastuzumab emtansine. In contrast, the potencies of noncleavable ADCs carrying the structurally distinct monomethyl auristatin F were unaffected by SLC46A3 attenuation. Structure–activity experiments suggested that maytansine is a substrate for SLC46A3. Notably, SLC46A3 silencing led to relative increases in catabolite concentrations in the lysosome. Taken together, our results establish SLC46A3 as a direct transporter of maytansine-based catabolites from the lysosome to the cytoplasm, prompting further investigation of SLC46A3 as a predictive response marker in breast cancer specimens. Cancer Res; 75(24); 5329–40. ©2015 AACR.