Melanocytes are melanin-producing cells found in skin, hair follicles, eyes, inner ear, bones, heart and brain of humans. They arise from pluripotent neural crest cells and differentiate in response to a complex network of interacting regulatory pathways. Melanins are pigment molecules that are endogenously synthesized by melanocytes. The light absorption of melanin in skin and hair leads to photoreceptor shielding, thermoregulation, photoprotection, camouflage and display coloring. Melanins are also powerful cation chelators and may act as free radical sinks. Melanin formation is a product of complex biochemical events that starts from amino acid tyrosine and its metabolite, dopa. The types and amounts of melanin produced by melanocytes are determined genetically and are influenced by a variety of extrinsic and intrinsic factors such as hormonal changes, inflammation, age and exposure to UV light. These stimuli affect the different pathways in melanogenesis. In this review we will discuss the regulatory mechanisms involved in melanogenesis and explain how intrinsic and extrinsic factors regulate melanin production. We will also explain the regulatory roles of different proteins involved in melanogenesis.

Background:The thieno [2,3-b]pyridines were discovered by virtual high throughput screening as potential inhibitors of phospholipase C (PLC) isoforms and showed potent growth inhibitory effects in National Cancer Institute's human tumour cell line panel (NCI60). The mechanism of the anti-proliferative activity of thieno [2,3-b]pyridines is explored here. Objectives:We aimed to investigate the basis for the anti-proliferative activity of these thieno [2,3-b]pyridines and to determine whether the cellular inhibition was related to their inhibition of PLC. Methods:Four breast cancer cell lines were used to assess the anti-proliferative effects (IC 50 values) of six representative thieno [2,3-b]pyridines. The most potent compound (derivative 3; NSC768313), was further studied in MDA-MB-231 cells. DNA damage was examined by γH2AX expression level, and cell cycle arrest by flow cytometry. Cell morphology was examined by tubulin antibody staining. The growth inhibitory effect of combination treatment with derivative 3 and paclitaxel (tubulin inhibitor), doxorubicin (topoisomerase II inhibitor) or camptothecin (topoisomerase I inhibitor) was evaluated. A preliminary mouse toxicity assay was used to evaluate the pharmacological properties.Results: Addition of the thieno[2,3-b]pyridine derivative 3 to the MDA-MB-231 cells induced G2/M growth inhibition, cell cycle arrest in G2-phase, membrane blebbing and the formation of multinucleated cells. It did not induce DNA damage, mitotic arrest or changes in calcium ion flux. Combination of derivative 3 with paclitaxel showed a high degree of synergy, while combinations with doxorubicin and camptothecin showed only additive effects. A mouse pharmacokinetic study of derivative 3 showed that after intraperitoneal injection of a single does (10 mg/Kg), the C max was 0.087 μmol/L and the half-life was 4.11 h. Conclusions:The results are consistent with a mechanism in which thieno [2,3-b]pyridine derivatives interact with PLC isoforms (possibly PLC-δ), which in turn affect the cellular dynamics of tubulin-β, inducing cell cycle arrest in G2-phase. We conclude that these compounds have novelty because of their PLC target and may have utility in combination with mitotic poisons for cancer treatment.

TPS43 Background: Activation of the receptor tyrosine kinase AXL has a profound suppressive effect on the innate immune system. AXL is overexpressed on multiple cell types in the tumour immune microenvironment including dendritic cells, NK cells and tumour-associated macrophages. AXL signalling in immune cells supports tumour immune escape by downregulating dendritic cell activity, modulating efferocytosis as well as favouring an immunosuppressive chemokine profile and M-MDSC expansion. AXL is prevalent in tumours resistant to anti-PD-1 therapy (Hugo, 2016). Axl expression in tumour cells confers resistance to effector T cell cytotoxicity. Bemcentinib (BGB324) is a first-in-class, highly selective and orally bioavailable small molecule AXL inhibitor in phase II clinical development. In pre-clinical models of pancreas, breast and lung cancer, inhibition of AXL signalling with bemcentinib reversed multiple tumour immune suppressive mechanisms as evidenced by increased infiltration of cytotoxic T lymphocytes, NK and NKT cells and decreased infiltration of M-MDSCs (Wnuk-Lipinska, 2017). Bemcentinib enhanced the effect of immune checkpoint blockade via PD-1 or CTLA-4 in lung and mammary adenocarcinoma mouse models and achieved sustained tumour immunity. Methods: BGBC007 (NCT03184558) and BGBC008 (NCT03184571) are open-label, phase II studies designed to assess the anti-tumour activity of bemcentinib in combination with pembrolizumab in patients with previously treated TNBC and adenocarcinoma of the lung respectively. All patients will be treated with bemcentinib in combination with pembrolizumab continuously for up to two years. The primary endpoint is objective response rate, secondary endpoints include duration of response, progression free survival according to RECIST 1.1, pharmacokinetics, safety and tolerability. Pretreatment tumour specimens are scheduled to assess AXL expression/signalling and PD-L1 expression; the levels of circulating immune-related cytokines and soluble AXL receptor will also be measured in longitudinal patient plasma samples. Both studies are open to recruitment. Clinical trial information: NCT03184558.

Previous whole-exome sequencing has demonstrated that melanoma tumors harbor mutations in the GRIN2A gene. GRIN2A encodes the regulatory GluN2A subunit of the glutamate-gated N-methyl-d-aspartate receptor (NMDAR), involvement of which in melanoma remains undefined. Here, we sequenced coding exons of GRIN2A in 19 low-passage melanoma cell lines derived from patients with metastatic melanoma. Potential mutation impact was evaluated in silico, including within the GluN2A crystal structure, and clinical correlations were sought. We found that of 19 metastatic melanoma tumors, four (21%) carried five missense mutations in the evolutionarily conserved domains of GRIN2A; two were previously reported. Melanoma cells that carried these mutations were treatment-naïve. Sorting intolerant from tolerant analysis predicted that S349F, G762E, and P1132L would disrupt protein function. When modeled into the crystal structure of GluN2A, G762E was seen to potentially alter GluN1–GluN2A interactions and ligand binding, implying disruption to NMDAR functionality. Patients whose tumors carried non-synonymous GRIN2A mutations had faster disease progression and shorter overall survival (P < 0.05). This was in contrast to the BRAF V600E mutation, found in 58% of tumors but showing no correlation with clinical outcome (P = 0.963). Although numbers of patients in this study are small, and firm conclusions about the association between GRIN2A mutations and poor clinical outcome cannot be drawn, our results highlight the high prevalence of GRIN2A mutations in metastatic melanoma and suggest for the first time that mutated NMDARs impact melanoma progression.