Metastatic breast cancer is an incurable disease and identification of novel therapeutic opportunities is vital. Triple negative breast cancer (TNBC) frequently metastasizes and high levels of activated RSK, a downstream MEK-ERK1/2 effector, are found in TNBC. We demonstrate using direct pharmacological and genetic inhibition of RSK1/2 that these kinases contribute to the TNBC metastatic process in vivo. Kinase profiling demonstrated that RSK1 and RSK2 are the predominant kinases targeted by the new inhibitor, which is based on the natural product, SL0101. Further evidence for selectivity was provided by the observations that silencing RSK1 and RSK2 eliminated the ability of the analogue to further inhibit survival or proliferation of a TNBC cell line. In vivo, the new derivative was as effective as the FDA-approved MEK inhibitor, trametinib, in reducing the establishment of metastatic foci. Importantly, inhibition of RSK1/2 did not result in activation of AKT, which is known to limit the efficacy of MEK inhibitors in the clinic. Our results demonstrate that RSK is a major contributor to the TNBC metastatic program and provide preclinical proof-of-concept for the efficacy of the novel SL0101 analogue in vivo.

SUMMARY In response to estrogens, estrogen receptor alpha (ERα), a critical regulator of homeostasis, is degraded through the 26S proteasome. However, despite the continued presence of estrogen before menopause, ERα protein levels are maintained. We discovered that ERK1/2-RSK2 activity oscillates during the estrous cycle. In response to high estrogen levels, ERK1/2 is activated and phosphorylates ERα to drive ERα degradation and estrogen-responsive gene expression. Reduction of estrogen levels results in ERK1/2 deactivation. RSK2 maintains redox homeostasis, which prevents sustained ERK1/2 activation. In juveniles, ERK1/2-RSK2 activity is not required. Mammary gland regeneration demonstrates that ERK1/2-RSK2 regulation of ERα is intrinsic to the epithelium. Reduced RSK2 and enrichment in an estrogen-regulated gene signature occur in individuals taking oral contraceptives. RSK2 loss enhances DNA damage, which may account for the elevated breast cancer risk with the use of exogenous estrogens. These findings implicate RSK2 as a critical component for the preservation of estrogen homeostasis.

A convergent synthesis of 5a-carbasugar analogues of the n-Pr-variant of SL0101 is described. The analogues were synthesized in an effort to find compounds with potent in vivo efficacy in the inhibition of p90 ribosomal s6 kinase (RSK1/2). The synthesis derived the desired C-4 L-rhamnose stereochemistry from quinic acid and used a highly selective cuprate addition, NaBH reduction, Mitsunobu inversion, and alkene dihydroxylation to install the remaining stereochemistry. A Pd-catalyzed cyclitolization stereoselectively installed the aglycon at the anomeric position. The analogues were evaluated as RSK1/2 inhibitors and found to have 3- to 6-fold improved activity.

An asymmetric synthesis of two analogues of SL0101 (1) has been achieved. The effort is aimed at the discovery of inhibitors of the p90 ribosomal S6 kinase (RSK) with improved bioavailability. The route relies upon the use of the Taylor catalyst to regioselectively install C-3″ acetyl or carbamate functionality. This study led to the identification of a third-generation analogue of SL0101 with a C-4″ n-Pr-carbamate and a C-3″ acetate with improved RSK inhibitory activity.

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