Hypoxia-Activated O6-Benzylguanine Prodrugs

描述（由申请人提供）：将DNA鸟嘌呤的O-6位氯乙基化的烷基化剂前药包括氯雷他嗪，该药物目前处于III期临床试验；BCNU， FDA批准临床使用的亚硝基脲；KS119W，一种选择性激活实体肿瘤缺氧细胞的水溶性磺酰肼前药。这种DNA损伤很容易被o6 -烷基鸟嘌呤-DNA烷基转移酶（ACT）修复，这是一种将鸟嘌呤O-6位置的烷基转移到ACT分子的蛋白质。这一作用代表了肿瘤和宿主组织对氯噻嗪、BCNU和KS119W耐药的主要机制。已知的最有效的ACT抑制剂之一是o6 -苄基鸟嘌呤（06-BG），它与ACT反应在蛋白的活性部位形成S-苄基半胱氨酸。结果，O6-BG消耗ACT，增加肿瘤和宿主细胞对氯噻嗪、BCNU和KS119W的敏感性。无毒性剂量的全身性O6-BG已在患者中被证明可以消耗肿瘤中的ACT含量。然而，这种作用也会使宿主组织对联合使用BCNU致敏，由于骨髓抑制，需要减少80%的剂量，导致BCNU水平无效。该应用的具体目的是：(a)设计和合成通过在实体瘤缺氧细胞中还原酶选择性激活的O6-BG前药，从而选择性地消耗肿瘤中的AGT，同时保留充氧正常组织；(b)体外评价O6-BG前药后氯噻嗪、BCNU和KS119W对缺氧和氧合肿瘤细胞的预处理效果；(c)在体内评估这些组合对多种含有高水平AGT活性的移植人类肿瘤的作用；(d)合成前药作用机制的药理学和生化研究。主要总体目标是选择一种O6-BG前药用于临床开发，与BCNU、氯噻嗪和KS119W联合使用。所选择的O6-BG前药应选择性地消耗实体肿瘤中的AGT，从而允许在接近全治疗剂量的氯乙基化剂的顺序组合中使用，而不会增加骨髓抑制或对其他正常组织的毒性。我们的预期是，由于AGT的组成性高，对氯噻嗪、KS119W和BCNU的细胞毒性作用产生耐药性的实体肿瘤，将通过O6-BG前药预处理选择性地消耗诱导耐药性的蛋白，导致氯乙基化剂耐药肿瘤转化为药物敏感肿瘤，从而增加可能受益于氯噻嗪、KS119W和BCNU的恶性肿瘤的范围。

DESCRIPTION (provided by applicant): Alkylating agent prodrugs that chloroethylate the O-6 position of DNA guanine include Cloretazine, an agent currently in Phase III clinical trial; BCNU, an FDA approved clinically used nitrosourea; and KS119W, a water-soluble sulfonyl hydrazine prodrug selectively activated in hypoxic cells of solid tumors. This DNA lesion is susceptible to repair by O6-alkylguanine-DNA alkyltransferase (ACT), a protein that transfers alkyl groups from the O-6 position of guanine to the ACT molecule. This action represents the primary mechanism of tumor and host tissue resistance to Cloretazine, BCNU, and KS119W. One of the most potent known inhibitors of ACT is O6-benzylguanine (06-BG), which reacts with ACT to form S- benzylcysteine in the active site of the protein. As a result, O6-BG depletes ACT and increases the sensitivity of tumor and host cells to Cloretazine, BCNU and KS119W. Non-toxic doses of systemic O6-BG have been shown in patients to deplete the ACT content of tumors. This action, however, also sensitizes host tissues to BCNU used in combination, necessitating an 80% decrease in the dosage of this agent because of myelosuppression, leading to an ineffective level of BCNU. The Specific Aims of this application are the (a) design and synthesis of O6-BG prodrugs activated selectively by reducing enzymes in hypoxic cells of solid tumors, thereby selectively depleting tumors of AGT while sparing oxygenated normal tissues; (b) evaluation of pretreatment of hypoxic and oxygenated tumor cells with O6-BG prodrugs in vitro followed by Cloretazine, BCNU, and KS119W; (c) evaluation in vivo of these combinations against a variety of transplanted human tumors containing high levels of AGT activity; and (d) pharmacological and biochemical studies of the mechanism of action of synthesized prodrugs. The primary overall objective is the selection of an O6-BG prodrug for clinical development to use in combination with BCNU, Cloretazine and KS119W. The O6-BG prodrug selected should deplete AGT selectively in solid tumors, thereby permitting use in sequential combination at close to full therapeutic dosage of a chloroethylating agent without increased myelosuppression or toxicities to other normal tissues. The expectation is that solid tumors, resistant to the cytotoxic actions of Cloretazine, KS119W and BCNU because of constitutively high levels of AGT, will be selectively depleted of the resistance inducing protein by pretreatment with the O6-BG prodrug, resulting in conversion of chloroethylating agent resistant neoplasms to drug sensitive ones, thereby increasing the spectrum of malignancies that may benefit from Cloretazine, KS119W, and BCNU.