Exploiting Cancer Metabolism and Drug Efflux with Bystander-Assisted Immunotherapy

摘要。癌症耐药性的两个标志是不规则代谢和药物外排。在多种药物中- 耐药的癌症，这两个过程解除化疗药物的疗效，最终导致去- 化疗效果增加和死亡率增加。发展中的几项战略尝试， 通过调节特定的代谢途径或破坏药物外排来控制耐药性的影响。具体- 具体而言，这些策略包括抑制剂、干扰RNA和纳米医学方法。然而，基金会- 对这些策略的精神挑战是由破坏代谢或药物外排引起的脱靶毒性 由P-糖蛋白（P-gp）介导，因为这些机制对许多健康过程也至关重要 在整个身体。为了解决这个问题，我们的长期目标是开发一种利用 这两种耐药机制串联产生治疗性抗癌免疫应答。 我们的中心假设是，合理设计的前药可以协同癌细胞代谢和药物外排， 通过我们称为旁观者辅助免疫的作用机制引起抗癌免疫应答， noTherapy（BAIT）.在BAIT中，酶导向的前药首先代谢为免疫代谢物 由多重耐药癌细胞的不规则代谢引起。接下来，免疫球蛋白通过 P-gp介导的药物外排至细胞外。这导致了免疫系统中旁观者免疫细胞的激活。 局部接近，这引发了抗癌免疫反应。因为诱饵需要串联代谢， 药物外排，我们预计一个独特的增强特异性的多药耐药表型，表现出这两个 这些过程。为了开发合理设计的BAIT前药，我们首先鉴定了小分子免疫疗法， 易受药物外排影响的药物。在同时的研究中，我们还开发了合成酶指导的 调节免疫治疗剂的活性并被在细胞中表达的酶特异性去除的基团。 多重耐药癌细胞的不规则代谢。结合这两个研究领域， 酶导向的BAIT前药，其赋予多药耐药癌症免疫原性。在体外，这是康- 在表达这些代谢酶和P-gp的免疫细胞和癌细胞系的共培养物中得到证实。在- 体内，我们使用前列腺癌的小鼠模型系统（TRAMP-C2同种异体移植物）来证明BAIT pro- 相对于化疗药物，化疗药物导致毒性降低、肿瘤体积减小和无进展生存期增加。 免疫活性小鼠中的常规免疫治疗剂。综上所述，我们设想这项研究将 建立BAIT作为一种治疗策略，通过耐药性增强而不是解除。是我们 长期的愿景是，这种策略可以广泛应用于逃避行动的多药耐药癌症 通过改变代谢和药物外排来改善传统疗法。

ABSTRACT. Two hallmarks of drug resistance in cancers are irregular metabolism and drug efflux. In multidrug- resistant cancers, both of these processes disarm the efficacy of chemotherapeutics, ultimately resulting in de- creased chemotherapeutic efficacy and increased mortality. Several strategies in development attempt to miti- gate the effects of drug resistance by modulating specific metabolic pathways or disrupting drug efflux. Specifi- cally, these strategies include inhibitors, interference RNAs, and nanomedicine approaches. However, a funda- mental challenge to these strategies is the off-target toxicity that arises from disrupting metabolism or drug efflux mediated by P-glycoprotein (P-gp), as these mechanisms are also critical to a number of healthy processes throughout the body. To address this, our long-term objective is to develop a therapeutic strategy that exploits both of these mechanisms of drug resistance in tandem to generate a therapeutic anti-cancer immune repsonse. Our central hypothesis is that rationally designed prodrugs can co-opt cancer cell metabolism and drug efflux to cause an anti-cancer immune response via a mechanism of action we have termed Bystander Assisted Immu- noTherapy (BAIT). In BAIT, an enzyme-directed prodrug is first metabolized to an immunotherapeutic metabolite by the irregular metabolism of multidrug-resistant cancer cells. Next, the immunotherapeutic is transported, via P-gp-mediated drug efflux, to the extracellular space. This results in the activation of bystander immune cells in local proximity, which initiate an anti-cancer immune response. Because BAIT requires tandem metabolism and drug efflux, we anticipate a uniquely enhanced specificity for multidrug-resistant phenotypes that exhibit both of these processes. To develop rationally designed BAIT prodrugs, we first identify small-molecule immunothera- peutics that are susceptible to drug efflux. In concurrent studies, we also develop synthetic enzyme-directing groups that modulate the activity of immunotherapeutics and are specifically removed by enzymes expressed in the irregular metabolism of multidrug-resistant cancer cells. Combining these two research areas, we generate enzyme-directed BAIT prodrugs that confer immunogenicity to multidrug-resistant cancers. In-vitro, this is con- firmed in co-cultures of immune cells and cancer cell lines that express these metabolic enzymes and P-gp. In- vivo, we use a murine model system for prostate cancer (TRAMP-C2 allograft) to demonstrate that BAIT pro- drugs result in lowered toxicity, decreased tumor volume, and increased progression-free survival, relative to conventional immunotherapeutics in immunocompetent mice. Taken together, we envision that this research will establish BAIT as a therapeutic strategy that is enhanced, rather than disarmed, by drug resistance. It is our long-term vision that this strategy could be widely applicable to multidrug-resistant cancers that evade the action of conventional therapies through altered metabolisms and drug efflux.