Allogeneic Immunotherapy for cancer and nonmalignant hem

在过去的十(20)年里，同种异体淋巴细胞靶向和根除白血病细胞的能力已经被确立为一种真正的生物实体。这种效应是否能对非血液系统的“实体”恶性肿瘤产生，目前尚不清楚。我们启动了一项非清髓性异基因干细胞移植治疗难治性转移性肾细胞癌(RCC)的临床试验。在一些患者中观察到的大转移完全消退，证实了同种异体移植物抗肾细胞癌效应的确凿证据。最近，我们在一位结肠癌和前列腺癌患者身上发现了类似的效果。随后，我们开始研究那些表现出抗肾细胞癌效应的患者的免疫重建，试图确定介导这些退化的效应细胞群体及其靶抗原。本实验室已证实肾癌细胞表面表达次要组织相容性抗原。我们证明了对微小抗原具有特异性的细胞毒性T细胞克隆能够杀死移植后出现GVT效应的患者的肾癌细胞。我们还能够在两个有反应的患者中扩增T细胞克隆，这些患者识别患者造血细胞和肾细胞癌细胞上存在的肿瘤细胞特异性或广泛表达的抗原。这些观察结果首次揭示了非清髓性异基因移植后转移性癌症消退的免疫机制。通过c-DNA表达克隆，我们最近发现了一种新的肿瘤抗原，该抗原来源于一种在肾癌细胞上过度表达的人内源性逆转录病毒，称为CT-RCC。这种抗原在正常组织中不表达，因此可能成为未来肾癌疫苗的靶点。我们目前正试图确定调节这种新发现的肿瘤抗原表达的因素。 最近，我们建立了一种在转移性肾细胞癌宿主体内进行同种异体干细胞移植的小鼠模型，与自体移植的受者相比，可重复性的GVT效应延长了动物的存活时间。移植后的肿瘤疫苗研究正在这个模型中进行，以及在移植的最初几个月使用PDGF、VEGF和EGF-R酪氨酸激酶抑制剂抑制血管生成的影响的调查。我们的研究小组最近发现，KIR血型不合的NK细胞在体外对实体瘤细胞具有细胞毒作用。利用上述动物模型，我们发现单次输注同种异体反应性NK细胞可以显著降低肾细胞癌小鼠的移植物抗宿主病(GVHD)，延长其存活期。基于这些发现，我们计划评估是否可以通过过继输注剂量递增的供者NK细胞来增强异基因HCT后GVT对肾癌的影响。 我们还在探索通过靶向基因诱导改变肿瘤细胞表型，使实体瘤对NK细胞攻击敏感的方法。最近我们发现，硼替佐米和去脂肽通过增强NK细胞介导的TRAIL杀伤作用而使肿瘤对NK细胞的细胞毒作用增敏。这种敏化似乎克服了通过KIR-KIR配体相互作用而介导的NK抑制。我们还开发了一种方法，通过扩增来自健康捐赠者的&gt；4log NK细胞，用于在未来基于NK细胞的过继免疫治疗试验中过继输注。基于这些发现，今年我们获得了为期两年的床边长凳奖，以扩大我们的NK细胞扩增，以测试Bortezomib是否可以用于增强患者肿瘤对过继自体NK细胞输注的敏感性。 我们小组继续探索同种异体干细胞移植在非恶性疾病患者中的应用，如PNH或ATGF-难治性重型再生障碍性贫血。我们最近还表明，PNH可以通过非清髓性干细胞移植治愈。我们实验室进行的体外研究表明，PNH细胞对同种异体免疫攻击的敏感性与正常GPI阳性免疫细胞相同。目前已有29例SAA/PNH患者进行了移植，移植后100TRM为0%。

The ability of allogeneic lymphocytes to target and eradicate leukemia cells has been established as a true biological entity over the past ten (20) years. Whether such effects can be generated against non-hematological "solid" malignancies remains unexplored. We initiated a clinical trail using a non-myeloablative approach of allogeneic stem cell transplantation in patients with treatment refractory metastatic renal cell carcinoma (RCC). Definitive evidence for an allogeneic graft-versus-RCC effect has been demonstrated with complete regression of large metastasis observed in some patients. Recently we demonstrated a similar effect to have occurred in a patient with colon carcinoma and prostate cancer. We have subsequently initiated studies investigating immune reconstitution in those demonstrating an anti-RCC effect in attempts to identify both the effector cell populations mediating these regressions as well as their target antigens. Our laboratory has confirmed the expression of minor histocompatibility antigens on the surface of kidney cancer cells. We demonstrated that cytotoxic T-cell clones with specificity for minor antigens are capable of killing RCC cells in patients having a GVT effect post transplant. We also were able to expand T-cell clones in two responding patients that recognized either tumor cells specifically or broadly expressed antigens present on both patient hematopoietic cells and RCC cells. These observations provide the first insight into the immune mechanisms mediating the regression of metastatic cancer following non-myeloablative allogeneic transplantation. Using c-DNA expression cloning, we have recently identified a novel tumor antigen derived from a human endogenous retrovirus over expressed on RCC cells called CT-RCC. This antigen is not expressed on normal tissues and therefore could potentially serve as a target for a future kidney cancer vaccine. We are currently attempting to identify factors regulating expression of this newly identified tumor antigen. Recently we have developed a murine model of allogeneic SCT in hosts bearing metastatic RCC, in which reproducible GVT effects occur extending animal survival compared to recipients of autologous transplants. Post transplant tumor vaccination studies are being conducted in this model, as well as investigations into the impact of inhibiting angiogenesis in the first few months of transplantation using PDGF, VEGF, and EGF-R tyrosine kinase inhibitors. Our group has recently shown that KIR incompatible NK cells are cytotoxic to solid tumor cells in vitro. Using the above mentioned animal model, we have shown that a single infusion of alloreactive NK cells can significantly reduce GVHD and prolong survival in mice with RCC undergoing allogeneic HCT. Based on these findings, we plan to evaluate whether GVT effects aginanst RCC can be enhanced after allogeneic HCT by adoptively infusing escalating doses of donor NK cells. We are also exploring methods to sensitize solid tumors to NK cell attack by altering the phenotype of tumor cells through targeted gene induction. Recently we showed that bortezomib and depsipeptide sensitize tumors to NK cell cytotoxity by enhancing NK-cell mediated TRAIL killing. This sensitization appears to overcome NK inhibition that is mediated through KIR-KIR ligand interactions. We have also developed a method to expand by > 4 logs NK cells from healthy donors for adoptive infusion in future NK-Cell based adoptive immunotherapy trials. Based on these findings, this year we were awarded a 2 year bench to bedside award to scale up our NK cell expansions to test whether bortezomib could be used to sensitize patient's tumors to adoptive autologous NK cell infusions. Our group continues to explore the use of allogeneic SCT in patients with nonmalignant diseases such as PNH or ATGF-Refractory severe aplastic anemia. We have also recently shown that PNH can be cured following nonmyeloablative stem cell transplantation. In vitro studies conducted in our laboratory have shown PNH cells are equally sensitive to allogeneic immune attack as normal GPI-positive immune cells. At present, 29 patients with SAA/PNH have been transplanted with a day 100 TRM of 0%.