We have defined the mechanism of action of lurbinectedin, a marine-derived drug exhibiting a potent antitumor activity across several cancer cell lines and tumor xenografts. This drug, currently undergoing clinical evaluation in ovarian, breast, and small cell lung cancer patients, inhibits the transcription process through (i) its binding to CG-rich sequences, mainly located around promoters of protein-coding genes; (ii) the irreversible stalling of elongating RNA polymerase II (Pol II) on the DNA template and its specific degradation by the ubiquitin/proteasome machinery; and (iii) the generation of DNA breaks and subsequent apoptosis. The finding that inhibition of Pol II phosphorylation prevents its degradation and the formation of DNA breaks after drug treatment underscores the connection between transcription elongation and DNA repair. Our results not only help to better understand the high specificity of this drug in cancer therapy but also improve our understanding of an important transcription regulation mechanism. Mol Cancer Ther; 15(10); 2399-412. Ó2016 AACR.

PurposeTrabectedin induces synthetic lethality in tumor cells carrying defects in homologous recombinant DNA repair. We evaluated the effect of concomitant inhibition of nucleotide-excision repair and poly (ADP-ribose) polymerase (PARP) activity with trabectedin and PARP inhibitors, respectively, and whether the synthetic lethality effect had the potential for a synergistic effect in breast cancer cell lines. Additionally, we investigated if this approach remained effective in BRCA1-positive breast tumor cells.MethodsWe have evaluated the in vitro synergistic effect of combinations of trabectedin and three different PARP inhibitors (veliparib, olaparib, and iniparib) in four breast cancer cell lines, each presenting a different BRCA1 genetic background. Antiproliferative activity, DNA damage, cell cycle perturbations and poly(ADP-ribosyl)ation were assessed by MTT assay, comet assay, flow cytometry and western blot, respectively.ResultsThe combination of trabectedin and olaparib was synergistic in all the breast cancer cell lines tested. Our data indicated that the synergy persisted regardless of the BRCA1 status of the tumor cells. Combination treatment was associated with a strong accumulation of double-stranded DNA breaks, G2/M arrest, and apoptotic cell death. Synergistic effects were not observed when trabectedin was combined with veliparib or iniparib.ConclusionCollectively, our results indicate that the combination of trabectedin and olaparib induces an artificial synthetic lethality effect that can be used to kill breast cancer cells, independent of BRCA1 status.

Enhancement of adiponectin level has been shown to have beneficial effects, including antiobesity, antidiabetic, and hepatoprotective effects. This evidence supports the therapeutic utility of adiponectin in complicated obesity. The present study characterized the in vivo effects of sustained adiponectin release by NP-1, a new class of thiazol derivative that increases adiponectin levels. Acute administration of NP-1 reduced feeding, increased plasma adiponectin, and improved insulin sensitivity without inducing malaise, as revealed by conditioned taste aversion studies. Short-term (7 days) treatment with NP-1 also reduced feeding and body weight gain and increased phosphorylation of AMPK in muscle, a main intracellular effector of adiponectin. NP-1 was also evaluated in diet-induced obesity, and adult male Wistar rats were fed two different types of diet: a standard high-carbohydrate/low-fat diet (SD) and a high-fat diet (HFD). Once obesity was established, animals were treated daily with NP-1 (5 mg/kg) for 14 consecutive days. Chronic NP-1 induced body weight loss and reduction of food intake and resulted in both a marked decrease in liver steatosis and an improvement of biochemical indexes of liver damage in HFD-fed rats. However, a marked induction of tolerance in adiponectin gene transcription and release was observed after chronic NP-1 with respect to the acute actions of this drug. The present results support the role of adiponectin signaling in diet-induced obesity and set in place a potential use of compounds able to induce adiponectin release for the treatment of obesity and nonalcoholic fatty liver, with the limits imposed by the induction of pharmacological tolerance.

Small-Cell Lung Cancer (SCLC) is an aggressive neuroendocrine malignancy with a poor prognosis. Here, we focus on the neuroendocrine SCLC subtypes, SCLC-A and SCLC-N, whose transcription addiction was driven by ASCL1 and NEUROD1 transcription factors which target E-box motifs to activate up to 40% of total genes, the promoters of which are maintained in a steadily open chromatin environment according to ATAC and H3K27Ac signatures. This leverage is used by the marine agent lurbinectedin, which preferentially targets the CpG islands located downstream of the transcription start site, thus arresting elongating RNAPII and promoting its degradation. This abrogates the expression of ASCL1 and NEUROD1 and of their dependent genes, such as BCL2, INSM1, MYC, and AURKA, which are responsible for relevant SCLC tumorigenic properties such as inhibition of apoptosis and cell survival, as well as for a part of its neuroendocrine features. In summary, we show how the transcription addiction of these cells becomes their Achilles's heel, and how this is effectively exploited by lurbinectedin as a novel SCLC therapeutic endeavor.

Lurbinectedin (PM1183) is a new synthetic compound from the tetrahydroisoquinoline family, which has demonstrated a strong antiproliferative activity against a panel of human tumor models in preclinical assays and is currently being evaluated in phase III clinical trials in platinum-resistant ovarian cancer and small cell lung cancer. Lurbinectedin binds to DNA, inhibits trans-activated transcription, induces the degradation of elongating RNA Pol II and fools nucleotide excision repair to produce dsDNA breaks that need to be repaired mainly by homologous recombination (HR)1,2. Nearly 70% of patients diagnosed with ovarian cancer are in advanced stage, and the vast majority of them will eventually relapse after a primary cytoreductive surgery and several cycles of standard adjuvant chemotherapy including a platinum drug and a taxane. After a period of treatment with platinum drugs, patients will finally develop resistance, usually mediated by mechanisms such as drug detoxification or efflux and enhanced DNA repair. IRF-1 transcription factor expression has been shown to be up-regulated by cisplatin (CDDP) in ovarian cancer cells and might be limiting the response to the drug, likely by inhibiting cell proliferation3. Here we took advantage of the A2780/A2780cis human ovarian cancer cell lines, the second being a cisplatin resistant derivative, to investigate the role of IRF1 in the response of human ovarian cancer cells to cisplatin and lurbinectedin. A2780cis cells are, indeed, more resistant to cisplatin that their parental cell line but they do not differ in their resistance to lurbinectedin. Basal IRF-1 protein levels were actually higher in A2780cis cells than in their parental cell line, contributing to their resistance to cisplatin. Furthermore, cisplatin treatment induced the overexpression and nuclear localization of IRF-1 both, in A2780 and A2780cis cell lines. Contrarily, lurbinectedin did not induce the overexpression of IRF-1 neither in A2780 nor in A2780cis, explaining why this latter cell line is not resistant to the compound. Furthermore, lurbinectedin co-treatment with cisplatin reduced the expression of IRF-1 in A2780 and, more importantly, in A2780cis cells, explaining the synergism the combination has on these tumor cell lines. Thus, lurbinectedin not only did not activate the same mechanisms of resistance as cisplatin in ovarian cancer cells, but even reversed the resistance of these resistant cells to platinum drugs. 1 Santamaría Nuñez et al, 2016. Mol Cancer Ther 15(10):2399-2412 2 Romano et al, 2013. Int J Cancer. 2013 Nov;133(9):2024-33 3 Pavan et al, 2013. Eur J Cancer 49(4):964-973 Citation Format: Gema Santamaria Nuñez, Maria Jose Guillén, Juan F. Martínez-Leal, Pablo Avilés, Carlos M. Galmarini. Lurbinectedin reverses platinum dependent IRF1 overexpression and nuclear localization, partially responsible for resistance to platinum drugs in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1211. doi:10.1158/1538-7445.AM2017-1211