Impact of Endogenous DOPA Signaling on Melanocyte Homeostasis and Melanoma Susceptibility

摘要 黑色素瘤的风险是相当高的人与浅色皮肤相比， 深色皮肤的人。虽然这种差异传统上归因于紫外线 黑色素的辐射屏蔽作用，有越来越多的证据表明， 可能无法解释这些差异的全部。我们的初步数据显示， 黑素细胞的基线色素沉着与抑制增殖和限制 致瘤潜力这种效应似乎不是由黑色素本身引起的，而是由黑色素本身引起的。 黑色素合成中间体二羟基苯丙氨酸（DOPA）。在浅色黑素细胞中， 在黑色素瘤细胞中，加入外源性多巴诱导了如所定义的更分化的细胞状态， 通过增加明确的黑素细胞分化抗原的表达， 色素生成，增殖能力降低，c-Myc表达降低， FOXM 1是一种重要的增殖相关转录因子，在细胞中过表达， 许多癌症。我们的研究结果表明，多巴可能抑制胆碱能受体毒蕈碱 1（CHRM 1），这是一种Gq偶联G蛋白偶联受体，似乎促进黑色素瘤 增殖CHRM 1基因切除阻断了多巴的抗增殖作用， 在具有低基线CHRM 1的黑素瘤细胞系中CHRM 1的过表达促进了CHRM 1的表达， 使细胞对多巴重新敏感。 在目标1中，我们将验证试验数据，表明CHRM 1介导原发性肝癌中的多巴效应。 黑色素细胞和黑色素瘤，并使用新的生物发光共振能量转移（BRET） 生物传感器平台，以定义CHRM 1和DOPA影响的信号传导机制 黑素细胞功能和黑色素瘤增殖。在目标2中，我们将使用医学相关的 临床前黑色素瘤模型，以测试药理学CHRM 1拮抗作用抑制 作为单一疗法，以及与标准护理免疫和 黑色素瘤的靶向治疗为此，我们将使用干递送的多巴/卡比多巴， 目前FDA批准的帕金森病药物组合，在试点研究中， 耐受并有效抑制黑色素瘤生长的小鼠模型。这项工作将共同定义 皮肤色素沉着，黑色素瘤风险，DOPA和CHRM 1之间的机械联系， 提供关键的临床前体内数据，以支持新的人体试验， 将帕金森病药物重新用于治疗黑色素瘤

Abstract The risk of melanoma is substantially higher in people with lightly pigmented skin, compared to those with darkly pigmented skin. While this discrepancy is traditionally attributed to the ultraviolet radiation shielding effect of melanin pigment, there is accumulating evidence suggesting that this may not account for the totality of these differences. Our preliminary data show that high levels of baseline pigmentation in melanocytes correlates with inhibited proliferation and limited tumorigenic potential. This effect appears to result not from melanin itself, but rather, from the melanin synthesis intermediate dihydroxyphenylalanine (DOPA). In light melanocytes and melanoma cells, addition of exogenous DOPA induced a more differentiated cell state as defined by increased expression of well-established melanocyte differentiation antigens, increased pigment production, decreased proliferative capacity, and decreased expression of c-Myc and FOXM1, which are critical proliferation-associated transcription factors that are overexpressed in many cancers. Our results indicate that DOPA likely inhibits the cholinergic receptor muscarinic 1 (CHRM1), which is a Gq coupled G Protein-Coupled Receptor that seems to promote melanoma proliferation. Genetic ablation of CHRM1 blocked DOPA’s anti-proliferative effect, while overexpression of CHRM1 in melanoma cell lines with low baseline CHRM1 promoted and also rendered cells newly sensitive to DOPA. In Aim 1 we will validate pilot data suggesting that CHRM1 mediates DOPA effects in primary melanocytes and melanoma, and use a new bioluminescence resonance energy transfer (BRET) biosensor platform to define the signaling mechanisms by which CHRM1 and DOPA impact melanocyte function and melanoma proliferation. In Aim 2 will we will use medically relevant preclinical melanoma models to test the idea that pharmacologic CHRM1 antagonism inhibits melanoma in vivo, both as monotherapy, and in combination with standard of care immune and targeted melanoma therapeutics. For this we will use stemically delivered DOPA/carbidopa, a currently FDA-approved drug combination for Parkinson’s disease, which in pilot studies was well tolerated and effectively inhibited melanoma growth mouse models. Together this work will define a mechanistic link between skin pigmentation, melanoma risk, DOPA and CHRM1, and is likely to provide critical preclinical in vivo data that would support new human trials to test the efficacy of repurposing a Parkinson’s disease drug as a melanoma therapeutic.