Development of Targeted Damaging Agents for the Treatment of Drug-Resistant Gliomas

项目摘要 O 6-甲基鸟嘌呤甲基转移酶（MGMT）表达的缺失在癌症中很常见，并赋予敏感性 DNA烷化剂，如替莫唑胺（TMZ）。通过启动子超甲基化实现MGMT的表观遗传沉默 在约50%的胶质母细胞瘤（GBM）和大多数低级别胶质瘤中发现异柠檬酸脱氢酶-1/2 (IDH1/2）突变。MGMT在其他癌症中也是沉默的，包括高达40%的结肠癌，35%的小细胞肺癌，40%的结肠癌和40%的结肠癌。 非小细胞肺癌占25%。在缺乏MGMT表达的细胞中（称为MGMT-1）， 细胞），TMZ衍生的O 6-甲基鸟嘌呤（O 6 MeG）损伤与胸腺嘧啶错配，在DNA复制过程中，由于 改变氢碱基配对，导致错配修复途径（MMR）激活。MMR试图 通过切除新合成的链来修复这些损伤，但是胸腺嘧啶再次插入到 O6 MeG。这种重新插入再次触发MMR，导致DNA修复的反复“无效循环”，并最终 凋亡在临床上，MGMT启动子去甲基化是罕见的，而MMR突变经常发生， 在许多肿瘤类型中，TMZ耐药的主要机制。因为MGMT沉默在许多 癌症，克服MMR抗性的DNA病变（同时仍可通过MGMT解决，以便 维持治疗指数（TI））将产生广泛的影响。此外，由于这种生物标志物即使在 治疗难治性环境（即，在MMR缺陷的背景下），我们认为MGMT表达的缺失 没有被充分利用来获得治疗效果。基于上述研究结果，我们寻求开发一种新的 在Ranjit Bindra和Seth Herzon博士的实验室中发现的一类能产生O 6 MeG损伤的物质 在健康细胞中对MGMT去除敏感（“MGMT依赖性”），但可以克服MMR 耐药性（“MMR独立”）。为此，宾德拉博士和赫尔松共同创立了KL 50 Therapeutics，LLC， 他们的研究导致了KL 50的鉴定，KL 50是一种对MMR细胞更有效的新型烷基化剂 细胞系比MMR+细胞系，同时保留MGMT可分解性。这种分子表现出了极高的灵敏度 在MMR状态无关的MGMT缺陷细胞中，在MGMT-熟练细胞中具有可忽略的活性，并且具有 TI约为TMZ的30倍。在这些成就的基础上，在这个快速SBIR项目中，我们 建议进行电极导线优化，以改善中枢神经系统（CNS）渗透， 小分子在高级别胶质瘤（HGG）小鼠模型中具有体内功效，并进一步开发这些 用于1期临床试验的化合物。这些MGMT依赖性-MMR非依赖性烷化剂是 预计将拥有TMZ的积极属性，同时避免不可避免的MMR损失介导 耐药机制，从而对我们治疗GBM和其他缺乏耐药机制的肿瘤的方式产生重大影响。 获管理这些分子可以通过显着改善其生物学特性来代表肿瘤学的范式转变。 治疗指数如果成功，我们的方法将显著提高DNA烷化剂的安全性和有效性。 并将扩大其用于更广泛的复发性神经胶质瘤和许多其他癌症。

PROJECT SUMMARY Loss of O6-methylguanine methyltransferase (MGMT) expression is common in cancers and confers sensitivity to DNA alkylators, such as temozolomide (TMZ). Epigenetic silencing of MGMT via promoter hypermethylation is found in ~50% of glioblastomas (GBMs), and in most lower grade gliomas with isocitrate dehydrogenase-1/2 (IDH1/2) mutations. MGMT is also silenced in other cancers, including up to 40% of colon cancers, 35% of small cell lung cancers, and 25% of non-small cell lung cancers. In cells that lack MGMT expression (termed MGMT- cells), TMZ-derived O6-methylguanine (O6MeG) lesions mispair with thymine, during DNA replication, due to altered hydrogen base pairing, leading to activation of the mismatch repair pathway (MMR). MMR attempts to repair these lesions by resecting the newly synthesized strand, but thymine once again is inserted opposite of O6MeG. This reinsertion again triggers MMR, leading to iterative “futile cycles” of DNA repair and ultimately apoptosis. Clinically, MGMT promoter demethylation is rare, whereas MMR mutations occur frequently as a dominant mechanism of resistance to TMZ in many tumor types. Because MGMT silencing is found in many cancers, DNA lesions that overcome the MMR resistance (while still being resolvable by MGMT, so as to maintain a therapeutic index (TI)) will have a broad impact. Furthermore, as this biomarker persists even in the treatment-refractory setting (i.e., in the context of MMR defects), we argue that loss of MGMT expression has not been fully exploited for therapeutic gain. Based on the findings presented above, we seek to develop a new class of agents discovered in the laboratory of Drs. Ranjit Bindra and Seth Herzon that generate O6MeG lesions that are susceptible to MGMT removal (“MGMT dependent”) in healthy cells, but which can overcome MMR resistance (“MMR independent”). To this end, Drs. Bindra and Herzon have co-founded KL50 Therapeutics, LLC, and their studies lead to the identification of KL50, a novel alkylation agent that is more active against MMR- cell lines than MMR+ cell lines, while retaining MGMT resolvability. This molecule demonstrates exquisite sensitivity in MGMT-deficient cells independent of MMR status, with negligible activity in MGMT-proficient cells, and has a TI approximately 30 times greater than TMZ. Building on these achievements, in this fast-track SBIR project, we propose to conduct lead optimization to improve central nervous system (CNS) penetration, identify a collection of small molecules with in vivo efficacy in mouse models of high-grade gliomas (HGG), and further develop these compounds for use in a Phase 1 clinical trial. These MGMT dependent–MMR independent alkylating agents are anticipated to possess the positive attributes of TMZ, while circumventing the unavoidable MMR loss mediated resistance mechanism and, thereby, have a major impact on the way we treat GBMs and other tumors lacking MGMT. These molecules could represent a paradigm shift in oncology by dramatically improving their therapeutic index. If successful, our approach will significantly increase the safety and efficacy of DNA alkylators and will expand their use for a broader range of recurrent gliomas and many other cancers.