Dual Blockade of IGF1R and MEK synergistically inhibits pediatric cancers

与LASP合作，我们在Ras突变横纹肌肉瘤的小鼠异种移植模型中进行了曲美替尼和ganitumab的体内研究。我们已经发现，曲美替尼和加尼妥单抗的组合与单独的任一种药物相比，以及与标准护理化疗剂长春新碱相比，提供了生存优势。我们还发现，该组合在肿瘤内提供了预期的药效学作用，即与对照相比，在用曲美替尼和加尼妥单抗组合治疗的肿瘤中ERK磷酸化和IGF1R表达降低。我们还确定了人Ras突变的横纹肌肉瘤细胞系SMS-CTR对曲美替尼和加尼妥单抗的组合具有特殊的应答。为了确定SMS-CTR对这种组合反应如此良好的原因，我们发现在Ras突变的横纹肌肉瘤细胞系中，SMS-CTR具有最高的IGF1R表达。我们的目的是确定共同靶向RAS-RAF-MEK-ERK途径和IGF1R-IRS1/2-PI3K-AKT-mTOR途径的机制，这些机制在IGF1R表达不高的细胞系中有效，包括对IGF1R配体、IGF1/2和IRS1/2抑制剂具有特异性的单克隆抗体。这项工作目前正在进行中。重要的是，我们还在RAS突变RMS的几种PDX模型中测试了曲美替尼和ganitumab的组合。我们在由NRAS和HRAS两者驱动的细胞系异种移植物和PDX模型中显示了组合的功效。具有对组合的相对内在抗性的模型是也具有PI3激酶途径的基因改变（PIK3CA突变或PTEN表达丧失）的模型。目前的工作旨在更好地了解PI3激酶下游可能介导这种耐药性的效应子。我们正在与CTEP和儿童肿瘤学小组讨论在儿科患者中启动曲美替尼和ganitumab的I/II期临床试验。我们还在与ImmunityBio讨论由制药商发起的单机构试验。为了促进这种组合的临床转化，我们正在评估曲美替尼/ganitumab在其他疾病组织学中的疗效。我们已经在RAS突变的神经母细胞瘤的两种细胞系异种移植物中显示了曲美替尼/ganitumab的疗效，并且正在继续评估由MAPK通路（不是RAS本身）的改变驱动的神经母细胞瘤模型以及由NF 1或BRAF的改变驱动的MPNST中的组合。在曲美替尼和ganitumab组合的动物研究过程中，我们创建了对曲美替尼具有抗性的SMS-CTR细胞系。我们目前正在确定该细胞系对曲美替尼耐药的机制。我们已经确定，解释曲美替尼和ganitumab之间协同作用的一种机制是ganitumab阻断了由曲美替尼单药治疗诱导的ERK磷酸化反弹和AKT磷酸化增加。然而，trametinib和ganitumab诱导与细胞信号变化相关的代谢，表观遗传和基因表达变化，我们正在研究中。

In collaboration with LASP, we conducted an in vivo study of trametinib and ganitumab in mouse xenograft models of Ras-mutated rhabdomyosarcoma. We have found that the combination of trametinib and ganitumab provided a survival advantage compared to either agent alone, and also compared to a standard of care chemotherapeutic, vincristine. We also identified that this combination provided the expected pharmacodynamic effects within the tumors, i.e. ERK phosphorylation and IGF1R expression was decreased in tumors that were treated with the combination of trametinib and ganitumab as compared to controls. We also identified that the human Ras-mutated rhabdomyosarcoma cell line, SMS-CTR, is an exceptional responder to the combination of trametinib and ganitumab. In an effort to identify the reason why SMS-CTR responds so well to this combination, we showed that of the Ras-mutated rhabdomyosarcoma cell lines, SMS-CTR has the highest expression of IGF1R. We aimed to identify mechanisms of co-targeting the RAS-RAF-MEK-ERK pathways and the IGF1R-IRS1/2-PI3K-AKT-mTOR pathways that would be effective in cell lines that do not have high IGF1R expression, including monoclonal antibodies specific for the ligand of IGF1R, IGF1/2, and an inhibitor of IRS1/2. This work is currently ongoing. Importantly, we have also tested the combination of trametinib and ganitumab in several PDX models of RAS-mutated RMS. We show efficacy of the combination in cell line xenograft and PDX models driven by both NRAS and HRAS. The models with relative intrinsic resistance to the combination are models that also have an alteration in genes of the PI3 kinase pathway, either PIK3CA mutation or loss of expression of PTEN. Current work is aimed at better understanding the effectors downstream of PI3 kinase that might mediate this resistance. We are in discussions with CTEP and the Children's oncology group regarding the initiation of a phase I/II clinical trial of trametinib and ganitumab in pediatric patients. We are also in discussions with ImmunityBio regarding an investigator-initiated single institution trial. In order to facilitate the clinical translation of this combination, we are evaluating the efficacy of trametinib/ganitumab in other disease histologies. We have shown efficacy of trametinib/ganitumab in two cell line xenografts of RAS-mutated neuroblastoma, and are continuing to evaluate the combination in neuroblastoma models driven by alterations in the MAPK pathway that are not RAS itself, as well as in MPNST, driven by alterations in NF1 or BRAF. In the course of the animal study of the trametinib and ganitumab combination, we created an SMS-CTR cell line that is resistant to trametinib. We are currently in the process of identifying the mechanism of resistance to trametinib in this cell line. We have identified that one mechanism that explains the synergy between trametinib and ganitumab is that ganitumab blocks the rebound ERK phosphorylation and increase in AKT phosphorylation induced by trametinib monotherapy. However, trametinib and ganitumab induce metabolic, epigenetic and gene expression changes related to the changes in cell signaling, which we are in the process of investigating.