Chirality and asymmetry increase the potency of candidate ADRM1/RPN13 inhibitors

Abstract

Bortezomib and the other licensed 20S proteasome inhibitors show robust activity against liquid tumors like multiple myeloma, but have disappointed against solid tumors including ovarian cancer. Consequently, interest is mounting in alternative non-peptide based drugs targeting the proteasome’s 19S regulatory particle subunit, including its ubiquitin receptor RPN13. RA183 and RA375 are more potent analogs of the prototypic inhibitor of RPN13 (iRPN13) called RA190, and they show promise for the treatment of ovarian cancer. Here we demonstrate that rendering these candidate RPN13 inhibitors chiral and asymmetric through the addition of a single methyl to the core piperidone moiety increases their potency against cancer cell lines, with the S-isomer being more active than the R-isomer. The enhanced cancer cell cytotoxicities of these compounds are associated with improved binding to RPN13 in cell lysates, ATP depletion by inhibition of glycolysis and mitochondrial electron chain transport, mitochondrial depolarization and perinuclear clustering, oxidative stress and glutathione depletion, and rapid accumulation of high molecular weight polyubiquitinated proteins with a consequent unresolved ubiquitin proteasome system (UPS) stress response. Cytotoxicity was associated with an early biomarker of apoptosis, increased surface annexin V binding. As for cisplatin, BRCA2 and ATM deficiency conferred increased sensitivity to these iRPN13s. Ubiquitination plays an important role in coordinating DNA damage repair and the iRPN13s may compromise this process by depletion of monomeric ubiquitin following its sequestration in high molecular weight polyubiquitinated protein aggregates. Indeed, a synergistic cytotoxic response was evident upon treatment of several ovarian cancer cell lines with either cisplatin or doxorubicin and our new candidate iRPN13s, suggesting that such a combination approach warrants further exploration for the treatment of ovarian cancer.

Development and anticancer properties of Up284, a spirocyclic candidate ADRM1/RPN13 inhibitor

“…A trend for reduction in ECAR, a surrogate measure of glycolysis, was observed after 24 h Up284 treatment (Fig 4H). These observations suggest that Up284 affects ES2 cancer cells by inhibition of glycolysis and OXPHOS, thereby ultimately reducing mitochondrial ATP production as previously shown with RA413S, a bis-benylidine piperidone iRPN13 candidate [46].…”

Ubiquitin receptors play redundant roles in the proteasomal degradation of the p53 repressor MDM2

“…However, in CRISPR screens, ADRM1 was classified as essential in around 20% of cell lines. Small molecule inhibitors of ADRM1 are being developed for the treatment of cancer [ 134 , 135 , 136 , 137 ]. S5A is also an attractive potential target for cancer therapy.…”

Analogues of Anticancer Natural Products: Chiral Aspects

“…Non-peptide inhibitors targeting different components of the proteasome system appear to be a promising alternative for the treatment of solid tumours. Anchoori et al [ 117 ] presented the development of novel derivatives targeting the 19S regulatory particle unit which contains the ubiquitin receptor RPN13, RA 183, and RA375.…”

“…The mechanism of antitumour activity for the active ( S )-isomer RA413S and the racemate was further evaluated. The cancer cell toxicity was associated with improved binding to RPN13 lysates, ATP depletion, mitochondrial damage, oxidative stress, and glutathione and NF-κB inhibition [ 117 ].…”