Direct inhibition of mutant K-Ras for the treatment of cancer

直接抑制突变型 K-​​Ras 治疗癌症

• 批准号: 8591442
• 负责人: Matt Patricelli
• 金额: $ 22.5万
• 依托单位: WELLSPRING BIOSCIENCES, LLC
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-02 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8591442
• 关键词: Active Sites; Address; Affect; Affinity; Binding; Biochemical; Biodistribution; Biological Assay; Cells; Cellular Assay; Colonic Neoplasms; Cysteine; Data; Development; Drug Formulations; Drug Targeting; Frequencies; Glycine; Growth; Guanosine Triphosphate Phosphohydrolases; Human; Imatinib; In Vitro; Inhibitory Concentration 50; K-ras Oncogene; KRAS2 gene; Laboratories; Lead; Libraries; Lung; Lung Neoplasms; Malignant Neoplasms; Malignant neoplasm of lung; Mass Spectrum Analysis; Measures; Medical; Membrane Proteins; Mutate; Mutation; Nucleotides; Oncogenes; Patients; Pharmaceutical Preparations; Phase; Positioning Attribute; Proteins; Proto-Oncogenes; RAS genes; Refractory; Relative (related person); Roentgen Rays; Safety; Structure; Sulfhydryl Compounds; Surface; Testing; Therapeutic; Work; base; cancer therapy; cancer type; counterscreen; drug development; inhibitor/antagonist; mutant; neoplastic cell; novel; pancreatic neoplasm; phase 2 study; professor; public health relevance; ras Proteins; screening; small molecule; success; tumor

DESCRIPTION (provided by applicant): The development of drugs that inhibit the K-Ras oncogene represents one of the greatest unmet needs in the treatment of human cancer. The Ras gene is the most frequently mutated oncogene in cancer, with a greater than 30% cumulative mutation frequency across all cancer types. Cancers with Ras mutations are aggressive and respond poorly to standard therapies. Previous attempts to target K-Ras have failed due to the difficulty of competing with the picomolar nucleotide affinity for the active sit and due to the high similarity of most GTPases. Our scientific co-founder, Professor Kevan Shokat, has discovered a novel, small molecule approach to target the most chemically tractable K-Ras mutant that contains a glycine-12 to cysteine mutation. The G12C mutation is the most common K-Ras mutation in lung cancer. Indeed, 43% of all lung cancers with K-Ras mutations contain the G12C mutation. This mutation positions a chemically reactive sulfhydryl group on the surface of K-Ras. We have carried out a preliminary 500 compound library screen based on mass spectrometry for molecules which bind covalently to K-rasG12C, H-rasG12C and counterscreened against WT K-Ras. 17 hits were identified from the screening library, and the first round optimization of the initial hits led to the discovery of a potent inhibitor JO-01-18. We have now solved the crystal structure of JO-01-148 bound to K-Ras G12C and identified a previously undescribed allosteric pocket on the surface of the protein adjacent to the cysteine moiety. This pocket makes it possible to identify irreversible inhibitors that bind in the pocket and selectively target the cysteine at position 12. Importantly, these small molecules inhibit only mutant K-Ras and not the normal protein. We have now solved more than 10 X-ray crystal structures of irreversible inhibitors bound to this allosteric pocket and synthesized more than 120 compounds. A clear SAR has been established. We are now proposing to further validate our lead G12C compounds in biochemical and cellular assays. The Phase I specific aims are: (1) Develop assays to evaluate K-Ras effector binding; (2) Demonstrate that the G12C irreversible binders can disrupt K-Ras effector binding; and (3) Demonstrate that the G12C irreversible binders can differentially affect tumor cells with G12C mutation compared to cells with wild type K-Ras and other K-Ras mutations. The Phase I milestone is the identification of K-ras G12C inhibitors that suppress proliferation of tumor cells with G12C K-ras mutation five-fold more potently (as measured by IC50 values) relative to tumor cells with other K-Ras mutations or wild type Ras. Collectively, we expect the Phase I results to demonstrate that we can generate a small molecule inhibitor that will specifically inhibit the growth of tumor cells wih K-Ras G12C mutation. If our approach is successful, our Phase II studies will more fully examine the safety, efficacy, and PK/biodistribution of a lead formulation for advancement to an IND application.

描述（由申请人提供）：抑制K-Ras癌基因的药物开发代表了人类癌症治疗中最大的未满足需求之一。Ras基因是癌症中最常突变的癌基因，在所有癌症类型中具有大于30%的累积突变频率。Ras突变的癌症具有侵袭性，对标准疗法的反应很差。先前靶向K-Ras的尝试由于难以与皮摩尔核苷酸亲和力竞争活性位点以及由于大多数GTP酶的高度相似性而失败。我们的科学联合创始人Kevan Shokat教授发现了一种新的小分子方法，可以靶向化学上最易处理的K-Ras突变体，该突变体含有甘氨酸-12到半胱氨酸的突变。G12 C突变是肺癌中最常见的K-Ras突变。事实上，所有K-Ras突变的肺癌中有43%含有G12 C突变。该突变在K-Ras表面上定位化学反应性巯基。我们已经基于质谱法对共价结合K-rasG 12 C、H-rasG 12 C的分子进行了初步的500个化合物文库筛选，并针对WT K-Ras进行了反筛选。从筛选文库中鉴定了17个命中，并且初始命中的第一轮优化导致发现了有效的抑制剂JO-01-18。我们现在已经解决了与K-Ras G12 C结合的JO-01-148的晶体结构，并在蛋白质表面上与半胱氨酸部分相邻的位置鉴定了先前未描述的变构口袋。该口袋使得有可能鉴定结合在口袋中并选择性靶向位置12处的半胱氨酸的不可逆抑制剂。重要的是，这些小分子仅抑制 突变的K-Ras而不是正常的蛋白质。我们现在已经解决了10多个X射线晶体结构的不可逆抑制剂绑定到这个变构口袋，并合成了120多个化合物。建立了明确的SAR。我们现在正计划在生化和细胞检测中进一步验证我们的G12 C先导化合物。I期的具体目标是：（1）开发评价K-Ras效应物结合的测定法;（2）证明G12 C不可逆结合剂可破坏K-Ras效应物结合;（3）证明G12 C不可逆结合剂与具有野生型K-Ras和其他K-Ras突变的细胞相比可差异性地影响具有G12 C突变的肿瘤细胞。I期研究的里程碑是确定K-ras G12 C抑制剂，其抑制具有G12 C K-ras突变的肿瘤细胞增殖的效力是具有其他K-Ras突变或野生型Ras的肿瘤细胞的5倍（通过IC 50值测量）。总的来说，我们期望I期结果证明我们可以产生特异性抑制具有K-Ras G12 C突变的肿瘤细胞生长的小分子抑制剂。如果我们的方法是成功的，我们的II期研究将更全面地检查安全性，有效性和PK/生物分布的先导制剂推进到IND申请。