Human-Enzyme Mediated, Systemic Depletion of Cystine for Cancer Treatment.

描述（申请人提供）：广泛的研究表明，与正常组织相比，前列腺癌（PCa）和其他高死亡率癌症，包括胶质母细胞瘤，而小细胞肺癌的增殖和存活严重依赖于非必需氨基酸L-半胱氨酸（L-Cys）及其氧化形式L-胱氨酸（CSSC）的摄取（由于对抗氧化剂的需求增加，并且在一些肿瘤中，因为它们显示内源性L-Cys合成的缺陷）。我们假设酶介导的血清L-Cys/CSSC池的全身性消耗（使用工程化的人酶，因此可能耐受良好）构成了用于PCa的强大且完全新颖的癌症治疗方法，作为单一疗法和作为与现有化疗方式机械协同的药剂。这一假设得到了强有力的初步数据的支持，这些数据表明，（i）导致血清L-Cys/CSSC库的几乎完全消耗;（ii）介导肿瘤生长的持续和完全停止，癌细胞经历细胞周期停滞，伴随着增加的氧化应激和自噬，和（iii）重要的是，一个多月的治疗耐受性非常好，没有体重减轻或明显毒性。在这些初步观察的基础上，我们提出了一个详细的研究计划，旨在提供临床相关的酶治疗候选药物，并通过体外和同种异体移植/异种移植PCa模型进行验证，最终将有助于支持后期临床前开发和IND申请。虽然L-Cys/CSSC消耗可适用于许多肿瘤的治疗，但我们选择关注PCa，首先是因为已知许多人PCA肿瘤经历L-Cys/CSSC稳态缺陷，其次是因为需要有效的无毒治疗，因为PCa是美国男性癌症死亡的第二大原因。实现这些目标将需要完成3个具体目标下的工作：Sp.Aim 1将集中于用于L-Cys/CSSC耗尽的候选人酶的工程化，其显示最佳的酶动力学、底物选择性、稳定性， 失活和聚集以及通过生物处理容易生产。将详细评价优化酶的PK和PD以及重复给药后的任何毒性作用。在Sp.Aim 2中，我们将寻求理解L-Cys/CSSC消耗对PCa细胞的总体机制/代谢后果，什么生物标志物可以预测易感性/抗性，并确定与标准护理化疗药物的哪些组合可以提供协同/累加效应。最后，在Sp. Aim 3中，我们将在代表不同表型（即p53状态、雄激素依赖性/非依赖性）的临床相关原位PCa动物模型和小鼠原位PCa同种异体移植物中评价优化的酶，以便在密切模拟实际疾病的模型中研究药物的疗效。

DESCRIPTION (provided by applicant): Extensive studies have revealed that in contrast to normal tissue, prostate carcinomas (PCa) and other high mortality cancers including glioblastomas, and small cell lung carcinomas are critically dependent on the uptake of the non-essential amino acid L-cysteine (L-Cys) and its oxidized form L-cystine (CSSC) for proliferation and survival (due to increased requirements for anti-oxidants and in some tumors because they display defects in endogenous L-Cys synthesis). We hypothesize that enzyme-mediated systemic depletion of the serum L-Cys/CSSC pool (using an engineered human enzyme, and thus likely well tolerated) constitutes a powerful and completely novel cancer therapeutic approach for PCa as a monotherapy and as an agent that is mechanistically synergistic with existing chemotherapeutic modalities. This hypothesis is supported by strong preliminary data demonstrating that the administration (every 4 days) of a prototypical engineered human L- Cys/CSSC depleting enzyme in an allograft model of PCa: (i) results in near complete depletion of the serum L-Cys/CSSC pool; (ii) mediates sustained and complete cessation of tumor growth with cancer cells experiencing cell cycle arrest accompanied by increased oxidative stress and autophagy and (iii) importantly treatment for over a month was very well tolerated with no weight loss or gross toxicities. To build on these preliminary observations we present a detailed research plan that is aimed at delivering a clinically relevant enzyme therapeutic candidate and providing the validation by in vitro and in allograft/xenograft PCa models that will eventually hel support late stage preclinical development and an IND application. While L-Cys/CSSC depletion may be applicable to the treatment of many tumors we elected to focus on PCa first because many human PCA tumors are known to experience defects in L-Cys/CSSC homeostasis and second because effective non-toxic treatments are needed as PCa is the second leading cause of cancer death in American men. Reaching these goals will require the completion of work under 3 Specific Aims: Sp. Aim 1 will focus on the engineering of a candidate human enzyme for L-Cys/CSSC depletion that displays optimal enzyme kinetics, substrate selectivity, stability to deactivation and aggregation and facile production by bioprocessing. The PK and PD of the optimized enzymes as well as any toxic effects upon repeated administration will be evaluated in detail. In Sp. Aim 2 we will seek to understand the global mechanistic/metabolic consequences of L-Cys/CSSC depletion on PCa cells, what biomarkers may predict susceptibility/resistance and to determine which combinations with standard of care chemotherapeutics may provide synergistic/additive effects. Finally, in Sp. Aim 3 we will evaluate the optimized enzyme in clinically relevant orthotopic PCa animal models that represent diverse phenotypes (i.e. p53 status, androgen dependence/independence) and mouse orthotopic PCa allografts such that the efficacy of the drug can be studied in models closely mimicking the actual disease.