Metastatic Melanoma Patient-derived Xenografts Respond to MDM2 Inhibition as a Single Agent or in Combination with BRAF/MEK Inhibition.
转移性黑色素瘤患者来源的异种移植物对 MDM2 抑制作为单药或与 BRAF/MEK 抑制联合反应。
- 作者列表："Shattuck-Brandt RL","Chen SC","Murray E","Johnson CA","Crandall H","O'Neal JF","Al-Rohil RN","Nebhan CA","Bharti V","Dahlman KB","Ayers GD","Kelley MC","Kauffman RM","Hooks M","Grau A","Johnson DB","Vilgelm AE","Richmond A
PURPOSE:Over 60% of melanoma patients respond to immune checkpoint inhibitor (ICI) therapy, but many subsequently progress on these therapies. Second-line targeted therapy is based on BRAF mutation status, but no available agents are available for NRAS, CDKN2A, PTEN, and TP53 mutations. Over 70% of melanoma tumors have activation of the MAPK pathway due to BRAF or NRAS mutations, while loss or mutation of cdkn2a occurs in ~40% of melanomas, resulting in unregulated MDM2-mediated ubiquitination and degradation of P53. Here we investigated the therapeutic efficacy of over-riding MDM2-mediated degradation of P53 in melanoma with an MDM2 inhibitor that interrupts MDM2 ubiquitination of P53, treating tumor-bearing mice with the MDM2 inhibitor alone or combined with MAPK-targeted therapy. EXPERIMENTAL DESIGN:To characterize the ability of the MDM2 antagonist, KRT-232, to inhibit tumor growth, we established patient-derived xenografts (PDX) from 15 melanoma patients. Mice were treated with KRT-232 or a combination with BRAF and/or MEK inhibitors. Tumor growth, gene mutation status, as well as protein and protein-phosphoprotein changes, were analyzed. RESULTS:100% of the 15 PDX tumors exhibited significant growth inhibition either in response to KRT-232 alone or in combination with BRAF and/or MEK inhibitors. Only BRAFV600wt tumors responded to KRT-232 treatment alone while BRAFV600E/M PDXs exhibited a synergistic response to the combination of KRT-232 and BRAF/MEK inhibitors. CONCLUSIONS:KRT-232 is an effective therapy for the treatment of either BRAFwt or PANwt(BRAFwt, NRASwt) TP53WT melanomas. In combination with BRAF and/or MEK inhibitors, KRT-232 may an effective treatment strategy for BRAFV600 mutant tumors.
目的: 超过 60% 的黑色素瘤患者对免疫检查点抑制剂 (ICI) 治疗有反应，但随后这些治疗有许多进展。二线靶向治疗基于 BRAF 突变状态，但 NRAS 、 CDKN2A 、 PTEN 和 TP53 突变尚无可用的药物。超过 70% 的黑色素瘤由于 BRAF 或 NRAS 突变而激活 MAPK 通路，而约 40% 的黑色素瘤发生 cdkn2a 缺失或突变，导致 MDM2-mediated 泛素化和 p53 降解。在这里，我们研究了阻止 P53 的 MDM2 泛素化的 MDM2 抑制剂对黑色素瘤中 P53 的过度 MDM2-mediated 降解的治疗效果,单独使用 MDM2 抑制剂或联合 MAPK 靶向治疗治疗荷瘤小鼠。 实验设计: 为了表征 MDM2 拮抗剂 KRT-232 抑制肿瘤生长的能力，我们从 15 例黑色素瘤患者建立了患者来源的异种移植 (PDX)。用 KRT-232 或 BRAF 和/或 MEK 抑制剂联合治疗小鼠。分析了肿瘤生长、基因突变状态以及蛋白质和蛋白质-磷蛋白的变化。 结果: 100% 的 15 个 PDX 肿瘤表现出显著的生长抑制，无论是对 KRT-232 单独或与 BRAF 和/或 MEK 抑制剂联合反应。只有 BRAFV600wt 肿瘤对 KRT-232 单独治疗有反应，而 BRAFV600E/M PDXs 对 KRT-232 和 BRAF/MEK 抑制剂的联合治疗表现出协同反应。 结论: KRT-232 是治疗 BRAFwt 或 PANwt(BRAFwt，NRASwt) TP53WT 黑色素瘤的有效方法。联合 BRAF 和/或 MEK 抑制剂，KRT-232 可能是 BRAFV600 突变肿瘤的有效治疗策略。
METHODS::Macrophages (MΦ) play a critical role in tumor growth, immunosuppression and inhibition of adaptive immune responses in cancer. Hence, targeting signaling pathways in MΦs that promote tumor immunosuppression will provide therapeutic benefit. PI3Kγ has been recently established by our group and others as a novel immuno-oncology target. Herein, we report that a macrophage Syk-PI3K axis drives polarization of immunosuppressive MΦs which establish an immunosuppressive tumor microenvironment in in vivo syngeneic tumor models. Genetic or pharmacological inhibition of Syk and/or PI3Kγ in MΦs promotes a pro-inflammatory MΦphenotype, restores CD8+ T cell activity, destabilizes HIF under hypoxia, and stimulates an antitumor immune response. Assay for Transposase-accessible Chromatin using Sequencing (ATAC-seq) analyses on the bone marrow derived macrophages (BMDMs) show that inhibition of Syk kinase promotes activation and binding of NF-κB motif in SykMC-KO BMDMs, thus stimulating immunostimulatory transcriptional programming in MΦs to suppress tumor growth. Finally, we have developed in silico the "first in class" dual Syk/PI3K inhibitor, SRX3207, for the combinatorial inhibition of Syk and PI3K in one small molecule. This chemotype demonstrates efficacy in multiple tumor models and represents a novel combinatorial approach to activate antitumor immunity.
METHODS::Dysfunction of invariant natural killer T (iNKT) cells in tumor microenvironment hinders their anti-tumor efficacy, and the underlying mechanisms remain unclear. Here we report that iNKT cells increase lipid biosynthesis after activation, and that is promoted by PPARγ and PLZF synergically through enhancing transcription of Srebf1. Among those lipids, cholesterol is required for the optimal IFN-γ production from iNKT cells. Lactic acid in tumor microenvironment reduces expression of PPARγ in intratumoral iNKT cells and consequently diminishes their cholesterol synthesis and IFN-γ production. Importantly, PPARγ agonist pioglitazone, a thiazolidinedione drug for type 2 diabetes, successfully restores IFN-γ production in tumor-infiltrating iNKT cells from both human patients and mouse models. Combination of pioglitazone and alpha-galactosylceramide treatments significantly enhances iNKT cell-mediated anti-tumor immune responses and prolongs survival of tumor-bearing mice. Our studies provide a strategy to augment the anti-tumor efficacy of iNKT cell-based immunotherapies via promoting their lipid biosynthesis.
METHODS::Near-infrared photoimmunotherapy (NIR-PIT) is a newly developed and selective cancer treatment that induces necrotic and immunogenic cell death and utilizes a monoclonal antibody conjugated to a photo-absorber dye, IR700DX, activated by NIR light. Although CD44 is surface cancer marker associated with drug resistance, anti-CD44-IR700 NIR-PIT results in inhibited cell growth and prolonged survival in multiple tumor types. Meanwhile, anti-CD25-IR700-targeted NIR-PIT has been reported to achieve selective and local depletion of FOXP3+CD25+CD4+ regulatory T cells (Tregs), which are primary immunosuppressive cells in the tumor microenvironment (TME), resulting in activation of local antitumor immunity. Combined NIR-PIT with CD44- and CD25-targeted agents has the potential to directly eliminate tumor cells and also amplify the immune response by removing FOXP3+CD25+CD4+ Tregs from the TME. We investigated the difference in therapeutic effects of CD44-targeted NIR-PIT alone, CD25-targeted NIR-PIT alone, and the combination of CD44- and CD25-targeted NIR-PIT in several syngeneic tumor models, including MC38-luc, LL/2, and MOC1. The combined NIR-PIT showed significant tumor growth inhibition and prolonged survival compared with CD44-targeted NIR-PIT alone in all tumor models and showed prolonged survival compared with CD25-targeted NIR-PIT alone in MC38-luc and LL/2 tumors. Combined CD44/CD25 NIR-PIT also resulted in some complete remissions, whereas this was not achieved with either type of NIR-PIT alone. Therefore, combined NIR-PIT simultaneously targeting cancer antigens and immunosuppressive cells in the TME may be more effective than either type of NIR-PIT alone and may have potential to induce prolonged immune responses in treated tumors.