Vaccine and radiation for the therapy of human cancers

使用螯合放射性核素(Sm-153-EDTMP)调节肿瘤细胞的表型和增强T细胞介导的杀伤。将肿瘤细胞暴露于亚致死剂量的外照射可上调肿瘤抗原和辅助分子的表达，使肿瘤细胞更容易受到抗原特异性细胞毒性T淋巴细胞(CTL)的杀伤。该项目探索了一种可能性，即暴露于姑息性剂量的放射性药物可以改变肿瘤细胞的表型，使它们更容易受到T细胞介导的杀伤。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)暴露在剂量增加的放射性药物samarium-153-ethylenediaminetetramethylenephosphonate(153Sm-EDTMP)中，该药物用于癌症患者治疗骨转移引起的疼痛。对前列腺癌相关的5种表面分子和几种肿瘤相关抗原的表达进行了荧光激活细胞分选分析和实时定量聚合酶链式反应(PCR)分析。将人前列腺癌LNCaP细胞暴露于153Sm-EDTMP，并与肿瘤相关抗原特异性CTL孵育，进行CTL杀伤实验，以确定暴露于153Sm-EDTMP是否使LNCaP细胞对T细胞介导的杀伤更敏感。153Sm-EDTMP使肿瘤细胞表面分子Fas(100%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%)和细胞间黏附分子-1(40%)表达上调。实时定量聚合酶链式反应分析揭示了更多上调的肿瘤抗原。暴露于153Sm-EDTMP使LNCaP细胞更容易受到前列腺特异性抗原、癌胚抗原(CEA)和粘蛋白-1特异性CTL的杀伤。153Sm-EDTMP相当于骨内姑息性给药的剂量改变了肿瘤细胞的表型，提示153Sm-EDTMP可能与免疫治疗协同作用，增加肿瘤细胞对CTL杀伤的敏感性。图像引导射频消融术(RFA)是一种局部局部治疗方法，越来越多地用于治疗多种恶性肿瘤。然而，复发是经常发生的(肿瘤复发15%，肿瘤复发3厘米)。RFA肿瘤消融术引起的高温诱导的细胞死亡可以诱导和/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组成部分，以及(B)RFA能否与疫苗免疫疗法相结合，进一步提高抗肿瘤活性，潜在地防止肿瘤复发和/或治疗更远的疾病。我们检测了体外亚致死温度对表达肿瘤相关抗原(TAA)CEA(MC38-CEA)的小鼠结肠癌细胞表面表型和对CTL介导的杀伤敏感性的影响。我们还评估了体内RFA对MC38-CEA肿瘤表型的影响，以及对诱导肿瘤特异性免疫反应的影响。最后，我们评估了RFA和表达CEA和三种共刺激分子CEA/Tricom的重组病毒疫苗之间的潜在协同作用。这些研究表明，MC38-CEA细胞在体外暴露于亚致死温度下，细胞表面CEA、HSP-70和ICAM-1的表达增加，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)增加脾对CEA和其他内源性TAA(包括gp70)的应答，以及(C)促进CD8+T细胞向肿瘤的募集。抗肿瘤研究表明，RFA联合疫苗消除了原发的MC38-CEA sc肿瘤，并显著控制了远处疾病的生长。综上所述，这些数据表明，在患有多个肿瘤病变的患者中，将RFA与疫苗联合使用，以利用可能作用于未经治疗的远处转移的不同免疫反应的诱导和扩展，具有潜在的临床益处。

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.