Substitution of the m,p-chloro groups of bis-benzylidinepiperidone
RA190 for p-nitro, generating RA183, enhanced covalent drug binding to Cys88 of RPN13. Treatment of
cancer cell lines with RA183 inhibited
ubiquitin-mediated protein degradation, resulting in rapid accumulation of high-molecular-weight polyubiquitinated
proteins, blockade of NFκB signaling, endoplasmic reticulum stress, an unfolded protein response, production of
reactive oxygen species, and apoptotic cell death. High-grade
ovarian cancer,
triple-negative breast cancer, and
multiple myeloma cell lines were particularly vulnerable to RA183. RA183 stabilized a tetraubiquitin-linked
firefly luciferase reporter
protein in
cancer cell lines and mice, demonstrating in vitro and in vivo proteasomal inhibition, respectively. However, RA183 was rapidly cleared from plasma, likely reflecting its rapid degradation to the active compound RA9, as seen in human liver microsomes. Intraperitoneal administration of RA183 inhibited
proteasome function and orthotopic
tumor growth in mice bearing human
ovarian cancer model ES2-luc
ascites or syngeneic ID8-luc
tumor.