Gerhard Dürnberger

NOTCH1 activation in breast cancer confers sensitivity to inhibition of SUMOylation.

Breast cancer is genetically heterogeneous, and recent studies have underlined a prominent contribution of epigenetics to the development of this disease. To uncover new synthetic lethalities with known breast cancer oncogenes, we screened an epigenome-focused short hairpin RNA library on a panel of engineered breast epithelial cell lines. Here we report a selective interaction between the NOTCH1 signaling pathway and the SUMOylation cascade. Knockdown of the E2-conjugating enzyme UBC9 (UBE2I) as well as inhibition of the E1-activating complex SAE1/UBA2 using ginkgolic acid impairs the growth of NOTCH1-activated breast epithelial cells. We show that upon inhibition of SUMOylation NOTCH1-activated cells proceed slower through the cell cycle and ultimately enter apoptosis. Mechanistically, activation of NOTCH1 signaling depletes the pool of unconjugated small ubiquitin-like modifier 1 (SUMO1) and SUMO2/3 leading to increased sensitivity to perturbation of the SUMOylation cascade. Depletion of unconjugated SUMO correlates with sensitivity to inhibition of SUMOylation also in patient-derived breast cancer cell lines with constitutive NOTCH pathway activation. Our investigation suggests that SUMOylation cascade inhibitors should be further explored as targeted treatment for NOTCH-driven breast cancer.

A combinatorial screen of the CLOUD uncovers a synergy targeting the androgen receptor

Approved drugs are invaluable tools to study biochemical pathways, and further characterization of these compounds may lead to repurposing of single drugs or combinations. Here we describe a collection of 308 small molecules representing the diversity of structures and molecular targets of all FDA-approved chemical entities. The CeMM Library of Unique Drugs (CLOUD) covers prodrugs and active forms at pharmacologically relevant concentrations and is ideally suited for combinatorial studies. We screened pairwise combinations of CLOUD drugs for impairment of cancer cell viability and discovered a synergistic interaction between flutamide and phenprocoumon (PPC). The combination of these drugs modulates the stability of the androgen receptor (AR) and resensitizes AR-mutant prostate cancer cells to flutamide. Mechanistically, we show that the AR is a substrate for γ-carboxylation, a post-translational modification inhibited by PPC. Collectively, our data suggest that PPC could be repurposed to tackle resistance to antiandrogens in prostate cancer patients.

Abstract LB-099: A high-throughput screen of approved drugs uncovers a synergistic interaction targeting prostate cancer

Libraries of approved drugs contain some of the best studied small molecules and represent invaluable resources for drug repurposing and chemical biology studies. Access to comprehensive libraries of clinical compounds is limited and these collections are otherwise difficult to arrange. Using cheminformatics approaches we have assembled a non-redundant set of representative FDA-approved small molecules that we call the CeMM Library of Unique Drugs (CLOUD). The CLOUD covers the chemical and biological space of approved drugs, contains active forms of prodrugs, and can be screened at clinically relevant concentrations. In a combinatorial viability screen of the CLOUD we uncovered a synergistic interaction between two approved drugs. We show that the concomitant administration of these two small molecules induces apoptosis in the androgen receptor (AR)-dependent prostate cancer cell line LNCaP. Mechanistically, the combination affects AR signaling and the stability of the receptor itself. We are currently dissecting further aspects of the mechanism of action and performing in vivo experiments. Altogether, our data suggest that the combination of these two drugs can be repurposed for the treatment of prostate cancer. Citation Format: Marco P. Licciardello, Patrick Markt, Freya Klepsch, Charles-Hugues Lardeau, Gerhard Durnberger, Vladimir Ivanov, Jacques Colinge, Stefan Kubicek. A high-throughput screen of approved drugs uncovers a synergistic interaction targeting prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-099. doi:10.1158/1538-7445.AM2015-LB-099

CLOUD – CeMM library of unique drugs

Approved drugs serve as functional probes for analyzing disease-relevant biochemical pathways or are screened against proteins to identify novel medical applications resulting in a repurposing of known drugs. Despite these benefits of approved drugs, it is almost impossible to physically obtain a complete collection. Commercial vendors cover about 50-65% of drugs. Academic efforts such as the Johns Hopkins Clinical Compound Library (JHCCL) [1] and the NCGC Pharmaceutical collection [2] are only accessible via collaboration agreements. Thus, an affordable reference drug library obtainable for all screening centers is still missing. Here we report the generation of the CeMM Library of Unique Drugs (CLOUD), a focused collection of 314 systemically bioavailable small molecule drugs. The library was derived from FDA-approved drugs applying data mining and structural clustering techniques. Reduction of approved drugs to a set of systemically available small molecules and clustering based on their activity classes (e.g. dopamine receptor agonist) resulted in 244 compounds. In addition to this representative set of structural and biological space of drugs, CLOUD contains 35 drugs where the biological target is still unknown. Another 35 molecules representing the active forms of CLOUD prodrugs ensure the usability of the library for both biochemical and cell-based assays. Finally, all CLOUD drugs are delivered in a single 384-well plate in concentrations related to their therapeutic plasma levels to make high-throughput screening as comfortable as possible.