We have discovered and developed a series of molecules (thiazole benzenesulfonamides). HA15, the lead compound of this series, displayed anti-cancerous activity on all melanoma cells tested, including cells isolated from patients and cells that developed resistance to BRAF inhibitors. Our molecule displayed activity against other liquid and solid tumors. HA15 also exhibited strong efficacy in xenograft mouse models with melanoma cells either sensitive or resistant to BRAF inhibitors. Transcriptomic, proteomic, and biochemical studies identified the chaperone BiP/GRP78/HSPA5 as the specific target of HA15 and demonstrated that the interaction increases ER stress, leading to melanoma cell death by concomitant induction of autophagic and apoptotic mechanisms.

Metformin was reported to inhibit the proliferation of many cancer cells, including melanoma cells. In this report, we investigated the effect of metformin on melanoma invasion and metastasis development. Using different in vitro approaches, we found that metformin inhibits cell invasion without affecting cell migration and independently of antiproliferation action. This inhibition is correlated with modulation of expression of proteins involved in epithelial-mesenchymal transition such as Slug, Snail, SPARC, fibronectin, and Ncadherin and with inhibition of MMP-2 and MMP-9 activation. Furthermore, our data indicate that this process is dependent on activation of AMPK and tumor suppressor protein p53. Finally, we showed that metformin inhibits melanoma metastasis development in mice using extravasation and metastasis models. The presented data reinforce the fact that metformin might be a good candidate for clinical trial in melanoma treatment.

Fetal testis is a major target of endocrine disruptors (EDs). During the last 20 years, we have developed an organotypic culture system that maintains the function of the different fetal testis cell types and have used this approach as a toxicological test to evaluate the effects of various compounds on gametogenesis and steroidogenesis in rat, mouse and human testes. We named this test rat, mouse and human fetal testis assay. With this approach, we compared the effects of six potential EDs ((mono-(2-ethylhexyl) phthalate (MEHP), cadmium, depleted uranium, diethylstilboestrol (DES), bisphenol A (BPA) and metformin) and one signalling molecule (retinoic acid (RA)) on the function of rat, mouse and human fetal testis at a comparable developmental stage. We found that the response is similar in humans and rodents for only one third of our analyses. For instance, RA and MEHP have similar negative effects on gametogenesis in the three species. For another third of our analyses, the threshold efficient concentrations that disturb gametogenesis and/or steroidogenesis differ as a function of the species. For instance, BPA and metformin have similar negative effects on steroidogenesis in human and rodents, but at different threshold doses. For the last third of our analyses, the qualitative response is species specific. For instance, MEHP and DES affect steroidogenesis in rodents, but not in human fetal testis. These species differences raise concerns about the extrapolation of data obtained in rodents to human health risk assessment and highlight the need of rigorous comparisons of the effects in human and rodent models, when assessing ED risk.

Data from experiments conducted almost exclusively in the rat have established that some phthalates have deleterious effects on the fetal testis probably due to their antiandrogenic and/or estrogenic effects, but their mechanisms of action remain unknown. A recent study reported that phthalates also have deleterious effects on human fetal testis with germ cell number, but not steroidogenesis altered. Therefore, we used organ culture of fetal testes at different stages of development to analyze the direct effects of phthalates on both steroidogenesis and gonocyte development and to determine if the effects of MEHP on these functions reported in the rat can be extended to other mammalian species. We defined specific periods of sensitivity of the fetal mouse testis to MEHP for these two functions and showed that the effects of phthalates on steroidogenesis vary with the developmental stage. Conversely, the strong deleterious effects of phthalates on germ cells were constantly present during the active phases of gonocyte development and thus share no relationship with the steroidogenic status. Moreover, all the effects of phthalates were unchanged in testes from mice deficient for estrogen (ERαKO or ERβKO) or androgen (Tfm) receptors. In conclusion, our results demonstrate that phthalates impair mouse fetal germ cell number similarly to other mammalian species, but are neither estrogenic nor antiandrogenic molecules because their effects do not involve, directly or indirectly, ER or AR.

Specific BRAFV600E inhibitors (BRAFi) are highly effective in the treatment of melanoma. However, acquired drug resistances invariably develop after the initial response. Therefore, the identification of new mechanisms of acquired resistance gives important clues towards the development of therapies that could elicit long lasting responses. Here we report that CD271 confers resistance to BRAFi in melanoma cells. The expression of CD271 is increased by BRAFi through a stimulation of tumor necrosis factor-alpha (TNFα) secretion that leads to NF-κB signaling pathway activation. CD271 is upregulated in a subset of BRAFi-resistant melanoma cells. The inhibition of TNFα/NF-κB pathway and CD271 silencing restore the BRAFi sensitivity of resistant melanoma cells. Finally, increase of CD271 expression is validated in BRAFi-resistant xenografts tumors and also in tumors from the patients who relapsed under BRAFi. In summary, these results reveal a novel TNFα/NF-κB/CD271 axis whose activation contributes to the acquisition of resistance to BRAFi and therefore may represent a novel therapeutic target to improve the efficacy of therapy in melanoma.

Several reports have demonstrated the inhibitory effect of metformin, a widely used drug in the treatment of type 2 diabetes, on the proliferation of many cancers including melanoma. Recently, it has been shown that metformin is able to modulate the cAMP level in the liver. As cAMP has a crucial role in melanin synthesis and skin pigmentation, we investigated the effect of metformin on melanogenesis both in vitro and in vivo. We showed that metformin led to reduced melanin content in melanoma cells and in normal human melanocytes by decreasing cAMP accumulation and cAMP-responsive element-binding protein phosphorylation. This inhibitory effect is correlated with decreased expression of master genes of melanogenesis, microphthalmia-associated transcription factor, tyrosinase, dopachrome tautomerase, and tyrosinase-related protein 1. Furthermore, we demonstrated that the antimelanogenic effect of metformin is independent of the AMPK pathway. Interestingly, topical application of metformin induced tail whitening in mice. Finally, we confirmed the antimelanogenic effect of metformin on reconstituted human epidermis and on human skin biopsies. These data emphasize the depigmenting effect of metformin and suggest a clinical strategy for using metformin in the topical treatment of hyperpigmentation disorders.

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The present study was conducted to determine whether exposure to the mono-(2-ethylhexyl) phthalate (MEHP) represents a genuine threat to male human reproductive function. To this aim, we investigated the effects on human male fetal germ cells of a 10⁻⁵ M exposure. This dose is slightly above the mean concentrations found in human fetal cord blood samples by biomonitoring studies. The in vitro experimental approach was further validated for phthalate toxicity assessment by comparing the effects of in vitro and in vivo exposure in mouse testes. Human fetal testes were recovered during the first trimester (7-12 weeks) of gestation and cultured in the presence or not of 10⁻⁵ M MEHP for three days. Apoptosis was quantified by measuring the percentage of Caspase-3 positive germ cells. The concentration of phthalate reaching the fetal gonads was determined by radioactivity measurements, after incubations with ¹⁴C-MEHP. A 10⁻⁵ M exposure significantly increased the rate of apoptosis in human male fetal germ cells. The intratesticular MEHP concentration measured corresponded to the concentration added in vitro to the culture medium. Furthermore, a comparable effect on germ cell apoptosis in mouse fetal testes was induced both in vitro and in vivo. This study suggests that this 10⁻⁵ M exposure is sufficient to induce changes to the in vivo development of the human fetal male germ cells.