Multiplexed and dynamically targeted photoimmunotherapy of heterogeneous, chemoresistant micrometastases guided by online in vivo optical imaging of cell-surface biomarkers

项目总结 尽管在手术、化疗和靶向治疗方面取得了进展，但转移性卵巢上皮癌的存活率 癌症仍然令人沮丧，部分原因是肿瘤的异质性和残留的显微疾病无法被 传统的成像方式。这种恶性肿瘤与晚期腹膜癌有关；也就是说， 盆腔及其常驻器官广泛的肿瘤聚集。为了解决这些“看不见的”肿瘤，我们 介绍一系列分子靶向和细胞激活的成像和治疗剂，与 新研制的用于体内多重分子成像的微型显微镜--细胞分辨率 高光谱荧光显微内窥镜--独一无二地能够可视化 体内深处的肿瘤细胞。多路生物标志物成像功能是由关键需求推动的 定量监测突出治疗耐药和逃逸的癌细胞表型(例如，癌症干细胞 细胞由多个细胞表面生物标志物定义)。与此同时，我们还开发了近红外 针对癌细胞过度表达的细胞膜分子的光细胞毒性免疫结合物(PIC)， 包括表皮生长因子受体(EGFR)，以创建光动力和受体相结合 肿瘤靶向、可激活的光免疫疗法(TAPIT)的拮抗剂。光动力技术 而荧光成分在癌细胞结合时变得去猝灭(激活)，细胞 内化和溶酶体抗体蛋白分解。这一策略克服了偏离目标的光毒性，包括 肠光毒性，在复杂部位进行光激活治疗的主要临床障碍，如 骨盆腔。在这里，我们建立在之前工作的基础上，表明PIC在卵巢微转移瘤中被激活 在使用EGFR过表达的异种移植小鼠模型中，体内敏感性和特异性分别为93%和93% 人卵巢上皮性癌细胞，能够准确识别小至30μm的肿瘤并具有选择性 破坏播散性腹膜微转移瘤。我们建议进一步发展这一平台，以 解决潜伏在残留肿瘤沉积中的肿瘤异质性和化疗耐药表型， 代表了癌症治疗中的一个关键利基市场。目前的临床影像技术无法分辨显微镜 手术和化疗留下的肿瘤沉积，患者的治疗选择有限 患有复发的化疗耐药肿瘤。我们预计，这种显微内窥镜引导下的Tapit新范式 将补充目前对晚期疾病患者和接受治疗的患者的治疗方式 抢救疗法，为个性化医疗开辟了新的途径。新功能将应用于 引导动态靶向Tapit以适应治疗和监测的表型演变 微小残留病的治疗反应。

PROJECT SUMMARY Despite advances in surgery, chemotherapy and targeted therapies, survival of metastatic epithelial ovarian cancer remains dismal due in part to tumor heterogeneity and residual microscopic disease undetectable by traditional imaging modalities. This malignancy involves peritoneal carcinomatosis at advanced stages; that is, extensive tumor studding of the pelvic cavity and its resident organs. To address these “invisible” tumors, we introduce a series of molecular-targeted and cell-activatable imaging and therapeutic agents integrated with a newly developed miniaturized microscope for multiplexed molecular imaging in vivo—a cellular-resolution hyperspectral fluorescence microendoscope—that uniquely enable visualization of microscopic deposits of tumor cells deep inside the body. The multiplexed biomarker imaging feature is motivated by the critical need to quantitatively monitor cancer cell phenotypes salient to treatment resistance and escape (e.g., cancer stem cells are defined by multiple cell-surface biomarkers). In parallel, we have also developed near-infrared photocytotoxic immunoconjugates (PICs) that target cell membrane molecules overexpressed by cancer cells, including the epidermal growth factor receptor (EGFR), to create a combined photodynamic and receptor antagonist therapeutic agent for tumor-targeted, activatable photoimmunotherapy (taPIT). The photodynamic and fluorescence components become de-quenched (activated) upon cancer cell binding, cellular internalization and lysosomal antibody proteolysis. This strategy overcomes off-target phototoxicity, including bowel phototoxicity, the major clinical obstacle for photoactivated treatments in complex sites such as the pelvic cavity. Here, we build on our prior work that shows the PIC is activated within ovarian micrometastases with 93% sensitivity and 93% specificity in vivo, in a xenograft mouse model using EGFR overexpressing human epithelial ovarian cancer cells, enabling accurate recognition of tumors as small as 30 μm and selective destruction of disseminated peritoneal micrometastases. We propose to further develop this platform to address tumor heterogeneity and chemoresistant phenotypes lurking within residual tumor deposits, which represents a critical niche in cancer therapy. Current clinical imaging technologies cannot resolve microscopic tumor deposits left behind by surgery and chemotherapy, and there are limited treatment options for patients with recurrent, chemoresistant tumors. We anticipate that this new paradigm for microendoscopy-guided taPIT will complement current treatment modalities for patients with advanced-stage disease and those receiving salvage therapies, opening new avenues for personalized medicine. The new capabilities will be applied to guide dynamically targeted taPIT adaptive to phenotype evolution in response to therapy and for monitoring treatment response of microscopic residual disease.