CLM296 is a potent and selective inhibitor of ALDH1A3, with an IC50 of 2 nM. It does not exhibit off-target effects on the highly homologous ALDH1A1 isoform. CLM296 is capable of inhibiting ALDH1A3-driven breast cancer metastasis and can be utilized in the study of triple-negative breast cancer (TNBC).

ALDH1A3:2 nM

CLM296, within a concentration range of 0.01 nM to 10 μM, significantly reduces Aldefluor TM fluorescence in ALDH1A3 overexpressing MDA-MB-231 cells, as well as in MDA-MB-468 and HCC1806 shRNA control cells, with IC50 values of 2.1 nM, 2.5 nM, and 2.3 nM respectively. CLM296 at 1 μM for 72 hours does not affect cell death or proliferation in MDA-MB-231, MDA-MB-468, or HCC1806 cells. Additionally, CLM296 at 100 nM for 24 hours selectively decreases the enhanced invasiveness due to ALDH1A3 overexpression in MDA-MB-231 cells. Furthermore, CLM296 at concentrations of 1-100 nM for 24 hours specifically blocks ALDH1A3-mediated gene expression changes in MDA-MB-231 cells.

CLM296 administered intraperitoneally (IP) at doses ranging from 0 to 4 mg/kg, once daily for 42 consecutive days, significantly reduced tumor volume and metastasis in mice overexpressing ALDH1A3 when started 12 or 15 days post-cell injection.