Vaccine and radiation for the therapy of human cancers

使用螯合放射性核素（钐-153- edtmp）来调节肿瘤细胞的表型并增强t细胞介导的杀伤。将人类肿瘤细胞暴露于亚致死剂量的外部束辐射会上调肿瘤抗原和辅助分子的表达，使肿瘤细胞更容易被抗原特异性细胞毒性T淋巴细胞（ctl）杀死。该项目探讨了暴露于姑息剂量的放射性药物制剂可能改变肿瘤细胞的表型，使其更容易受到t细胞介导的杀伤的可能性。在这里，10个人类肿瘤细胞系（4个前列腺，2个乳腺和4个肺）暴露于增加剂量的放射性药物钐-153-乙二胺四亚甲基膦酸盐（153Sm-EDTMP），用于治疗癌症患者因骨转移引起的疼痛。荧光活化细胞分选分析和实时定量聚合酶链反应（PCR）分析了5种表面分子和几种肿瘤相关抗原在前列腺癌中的表达。将LNCaP人类前列腺癌细胞暴露于153Sm-EDTMP中，并与肿瘤相关抗原特异性CTL一起培养，进行CTL杀伤实验，以确定暴露于153Sm-EDTMP是否使LNCaP细胞更容易受到t细胞介导的杀伤。肿瘤细胞对153Sm-EDTMP的反应上调了表面分子Fas（100%的细胞系上调Fas）、癌胚抗原（90%）、粘蛋白-1（60%）、主要组织相容性复合体（MHC） I类（50%）和细胞间粘附分子-1（40%）。实时荧光定量PCR分析发现了其他上调的肿瘤抗原。暴露于153Sm-EDTMP使LNCaP细胞更容易被前列腺特异性抗原、癌胚抗原（CEA）和粘蛋白-1特异性ctl杀死。153Sm-EDTMP的剂量相当于给骨的缓解剂量，可改变肿瘤细胞的表型，这表明153Sm-EDTMP可能与免疫疗法协同作用，增加肿瘤细胞对CTL杀伤的易感性。图像引导射频消融（RFA）是一种局部区域治疗越来越多地用于治疗广泛的恶性肿瘤。然而，复发是常有的（肿瘤小于3cm的占15%）。已知由RFA肿瘤消融引起的高温诱导细胞死亡可诱导和/或增强抗肿瘤免疫反应。我们试图在临床前模型中确定(a) RFA是否可以积极调节免疫系统的成分，以及(b) RFA是否可以与疫苗免疫疗法联合使用，以进一步提高抗肿瘤活性，并可能预防肿瘤复发和/或治疗更远的疾病。我们检测了体外亚致死高温对细胞表面表型的影响，以及对ctl介导的表达肿瘤相关抗原（TAA） CEA （MC38-CEA）的小鼠结肠癌细胞的杀伤敏感性。我们还评估了体内RFA对MC38-CEA肿瘤表型的影响，以及对肿瘤特异性免疫反应的诱导。最后，我们评估了RFA与表达CEA和三组共刺激分子CEA/TRICOM的重组病毒疫苗之间的潜在协同作用。这些研究表明，将MC38-CEA细胞体外暴露于亚致死高温下，会增加细胞表面CEA、HSP-70和ICAM-1的表达，并使肿瘤细胞更容易受到ctl介导的裂解。这些研究还表明，MC38-CEA肿瘤的RFA (a)诱导肿瘤细胞表面HSP-70和ICAM-1的上调，(b)增加脾脏CD4+和CD8+ T细胞对CEA和其他内源性TAAs（包括gp70）的反应，以及(c)改善CD8+ T细胞向肿瘤的募集。抗肿瘤研究表明，RFA联合疫苗可消除原发性MC38-CEA sc肿瘤，并显著控制远处病变的生长。综上所述，这些数据表明，在多发性肿瘤病变患者中，将RFA与疫苗联合使用，利用诱导和扩大多种免疫反应的潜在临床益处，这些免疫反应可能对未经治疗的远处转移起作用。

The use of chelated radionuclide (Samarium-153-EDTMP) to modulate phenotype of tumor cells and enhance T-cell-mediated killing. Exposing human tumor cells to sublethal doses of external beam radiation upregulates expression of tumor antigen and accessory molecules, rendering tumor cells more susceptible to killing by antigen-specific cytotoxic T lymphocytes (CTLs). This project explored the possibility that exposure to palliative doses of a radiopharmaceutical agent could alter the phenotype of tumor cells to render them more susceptible to T-ell-mediated killing. Here, 10 human tumor cell lines (4 prostate, 2 breast, and 4 lung) were exposed to increasing doses of the radiopharmaceutical samarium-153-ethylenediaminetetramethylenephosphonate (153Sm-EDTMP) used in cancer patients to treat pain due to bone metastasis. Fluorescence-activated cell sorting analysis and quantitative real-time polymerase chain reaction (PCR) analysis for expression of five surface molecules and several tumor-associated antigens involved in prostate cancer were done. LNCaP human prostate cancer cells were exposed to153Sm-EDTMPand incubated with tumor-associated antigen-specific CTL in a CTL killing assay to determine whether exposure to 153Sm-EDTMP rendered LNCaP cells more susceptible to T-cell-mediated killing. Tumor cells up-regulated the surface molecules Fas (100% of cell lines upregulated Fas), carcinoembryonic antigen (90%), mucin-1 (60%), major histocompatibility complex (MHC) class I (50%), and intercellular adhesion molecule-1 (40%) in response to 153Sm-EDTMP. Quantitative real-time PCR analysis revealed additional upregulated tumor antigens. Exposure to 153Sm-EDTMP rendered LNCaP cells more susceptible to killing by CTLs specific for prostate-specific antigen, carcinoembryonic antigen (CEA), and mucin-1. Doses of 153Sm-EDTMP equivalent to palliative doses delivered to bone alter the phenotype of tumor cells, suggesting that 153Sm-EDTMP may work synergistically with immunotherapy to increase the susceptibility of tumor cells to CTL killing. Image-guided radiofrequency ablation (RFA) is a local regional therapy increasingly used to treat a broad spectrum of malignancies. However, relapse often occurs (>15% for tumors < 3 cm). Hyperthermia-induced cell death resulting from RFA tumor ablation is known to induce and /or augment anti-tumor immune responses. We sought to determine in a preclinical model if (a) RFA could positively modulate components of the immune system, and (b) could RFA be combined with vaccine immunotherapy to further increase antitumor activity and potentially prevent tumor relapse and / or treat more distant disease. We examined the effect of sub-lethal hyperthermia in-vitro on cell-surface phenotype and sensitivity to CTL-mediated killing of murine colon carcinoma cells expressing the tumor-associated antigen (TAA) CEA (MC38-CEA). We also evaluated the effect of RFA in-vivo on the phenotype of MC38-CEA tumors, and on the induction of tumor-specific immune responses. Finally, we evaluated the potential synergy between RFA and a recombinant viral vaccine expressing CEA and a TRIad of COstimulatory Molecules, CEA/TRICOM. These studies demonstrated that exposure of MC38-CEA cells in-vitro to sub-lethal hyperthermia increases cell-surface expression of CEA, HSP-70, and ICAM-1, and renders tumor cells more susceptible to CTL-mediated lysis. These studies also demonstrated that RFA of MC38-CEA tumors (a) induces upregulation of HSP-70 and ICAM-1 on the cell-surface of tumor cells, (b) increases splenic CD4+ and CD8+ T cell responses to CEA and to additional endogenous TAAs, including gp70, and (c) improves recruitment of CD8+ T cells to the tumor. Antitumor studies showed that RFA combined with vaccine eliminated primary MC38-CEA sc tumors and significantly controlled growth of distant disease. Taken together, these data indicate a potential clinical benefit in combining RFA with vaccine in patients with multiple tumor lesions to capitalize on the induction and expansion of diverse immune responses that may act on untreated distant metastases.