Renalase inhibition for treatment of unresectable melanoma

治疗转移性黑色素瘤的最新进展，特别是免疫检查点抑制剂，已经改善了总生存期。尽管如此，由于发病率增加以及对免疫疗法的原发性和获得性抗性，黑素瘤的死亡率继续上升。在以前的研究中，我们发现分泌的黄素蛋白肾酶（RNLS）在黑色素瘤细胞中上调，并且在人类肿瘤中的高表达与肿瘤进展和存活率降低相关。在鼠模型中通过RNLS抗体和抑制肽敲低RNLS抑制肿瘤生长，减少肿瘤细胞中通过MAPK和PI 3 K途径的信号传导，并引起CD 163+肿瘤相关巨噬细胞（TAM）的显著减少。此外，我们表明，通过激活STAT 3，CD 163 + TAM增加RNLS的产生促进黑色素瘤生长，这表明RNLS的抑制可能是通过直接肿瘤细胞靶向和通过抑制CD 163 + TAM治疗黑色素瘤的有用方法。后一种机制特别令人感兴趣，因为已知CD 163 + TAM促进对效应T细胞杀死肿瘤细胞的抗性。 我们建议验证黑色素瘤中RNLS表达和信号传导失调调节肿瘤生长和扩散的假设，并将进一步研究其机制。此外，我们推测肿瘤细胞和TAM中的RNLS调节先天性和适应性免疫应答以促进癌症生长和扩散。因此，与单独的任一种方法相比，CD 8 + T细胞中RNLS和PD-1或CTLA 4的共抑制可导致增强的抗肿瘤活性。我们将利用我们团队可用的独特资源，包括新型动物模型，新型RNLS抑制剂的可用性，以及使用免疫检查点抑制剂治疗患者的黑色素瘤标本，这些研究将由基础，临床和翻译研究人员组成的跨学科团队进行。我们提出了三个相互关联但不相互依赖的目标。在目标1中，我们将确定RNLS抑制影响黑色素瘤细胞内信号传导途径的机制，以及RNLS调节体外和体内对黑色素瘤的先天性和适应性免疫应答的机制。在目标2中，我们将充分表征转移性黑色素瘤肿瘤的大队列中以BRAF和NRAS突变、TAM（CD 163+或CD 86+）、CD 4和CD 8+细胞为特征的肿瘤细胞中RNLS的表达，包括来自接受免疫检查点抑制剂治疗的患者的肿瘤。在目的3中，我们将检验以下假设：在具有与人肿瘤相关的遗传畸变（BRaf或NRAS突变）的免疫活性鼠黑素瘤模型中，抗RNLS抗体和抗CTLA 4或抗PD-1的联合治疗具有协同作用。这些研究的潜在影响很大-我们正在开发一种用于临床的RNLS抑制剂。如果成功，这些研究将导致未来的抗RNLS与免疫检查点抑制剂组合的临床试验。这种方法也可以在其他疾病中进行研究，因此可能对癌症治疗产生深远的影响。

Recent advances in treating metastatic melanoma, particularly with immune checkpoint inhibitors, have resulted in improved overall survival. Despite this, the mortality from melanoma continues to rise due to the increased incidence and to primary and acquired resistance to immunotherapy. In previous studies we showed that the secreted flavoprotein Renalase (RNLS) is upregulated in melanoma cells and high expression in human tumors is associated with tumor progression and decreased survival. Knock-down of RNLS by RNLS antibodies and an inhibitory peptide in murine models inhibited tumor growth, decreased signaling through the MAPK and PI3K pathways in tumor cells, and caused a marked reduction in CD163+ Tumor Associated Macrophages (TAMs). Furthermore, we showed that increased RNLS production by CD163+ TAMs facilitates melanoma growth by activating STAT3, suggesting that inhibition of RNLS might be a useful approach for treating melanoma by direct tumor cell targeting and by inhibition of CD163+ TAMs. The latter mechanism is particularly interesting as CD163+ TAMs are known to facilitate resistance to tumor cell kill by effector T cells. We propose to test the hypothesis that dysregulated RNLS expression and signaling in melanomas modulates tumor growth and spread and will further study the mechanism thereof. Furthermore, we surmise that RNLS in tumor cells and TAMs modulates both the innate and adaptive immune response to promote cancer growth and spread. Therefore, co-inhibition of RNLS and PD-1 or CTLA4 in CD8+ T cells could result in enhanced anti-tumor activity compared to either approach alone. We will capitalize on unique resources available to our team that include novel animal models, availability of a novel RNLS inhibitor, and access to melanoma specimens from patients treated with immune checkpoint inhibitors, and the studies will be conducted by an interdisciplinary team of basic, clinical and translational researchers. We propose three inter-related but not inter-dependent aims. In Aim 1 we will determine the mechanism by which RNLS inhibition affects signaling pathways within melanoma cells and the mechanism by which RNLS modulates the innate and adaptive immune response to melanoma in vitro and in vivo. In Aim 2 we will fully characterize expression of RNLS in tumor cells characterized for BRAF and NRAS mutations, TAMs (CD163+ or CD86+), CD4 and CD8+ cells in large cohorts of metastatic melanoma tumors, including tumors from patients treated with immune checkpoint inhibitors. In Aim 3 we will test the hypothesis that combination therapy with anti-RNLS antibodies and anti-CTLA4 or anti-PD-1 are synergistic in immune competent murine melanoma models with genetic aberrations relevant to human tumors (mutations in BRaf or NRas). The potential impact of these studies is high – we are developing a RNLS inhibitor for clinical use. If successful, these studies will lead to future clinical trials of the combination anti-RNLS with an immune checkpoint inhibitor. This approach can be studied in other diseases as well and can thus have far-reaching implications for cancer therapy.