应用异基因干细胞移植进行细胞免疫治疗 Richard Champlin, M.D.

造血干细胞移植 D D Preparative Regimen D D HSCT D R R D R D RL D RL D 异基因造血干细胞移植有效，但对血液系统恶性肿瘤的毒性治疗与高发的并发症和死亡率相关 (10->50%), 限制其用于无合并症的年轻患者

移植物抗肿瘤 异基因 SCT AlloSCT的很多好处是由于GVL 的免疫效应; 因此，大剂量根治是不必要的. 低剂量非清髓预处理方案足以防止排斥. 我们假设减低剂量，非清髓异基因足以可以减低毒性并使老年患者和伴有重要并发症的患者获得成功.

非清髓移植 Dsc DT DT Preparative Regimen D DT DT DT HSCT +DLI DT DB Dsc R R DNK Dsc R DT DNK RL D R RL RL D Recipient Donor Mixed Chimera Complete Chimera

异基因 HSCT 治疗 AML 的结果 OS EFS in 96 pts depending on disease state at time of HSCT. Upper line pts traqnsplanted in CR1-CR#, middle curve pts with active disease confined to the marrow but w/o circulating blasts. Lower curve shows pts with active AML in both marrow and peripheral blood. Upper curve 43 pts – plateau in EFS seemingly at 74% at 2 years.

HSCT的主要问题 移植后自然产生的抑制物抗肿瘤作用相对较弱 GVM 与 GVHD相关 复发仍是失败的主要原因

抑制物抗肿瘤的靶点 Allo-Specific Malignancy Specific Broadly expressed minor histocompatibility antigen (GVHD) Idiotype, Fusion peptide of translocation (bcr-abl) Lineage restricted minor histocompatibility antigen (G-vs-hematopoietic), or Redirected CAR T-cells vs CD19 Overexpressed normal cellular constituent (Proteinase 3, WT1, telomerase)

供者淋巴细胞数注 有效治疗复发的 CML, 对复发的AML 和 ALL 有效率低 有计划的DLI 研究显示提高GVM 效应 经常伴有 GVHD 与相比数量和移植后时间相关

Suicide Switch Genetically modify T-cells to introduce gene to induce apoptosis upon treatment with an activating drug Herpes virus tyrosine kinase – activated with ganciclovir Modified Caspace 9

Q6. retroviral vector issues. Adenoviral vector used for transient transfection situations Retroviral vector for stably transfected situations with low oncogenic risk (e.g. T cells) Lentivirus for stem cells (in development)

自毁转换 Quickest to go straight here. Q2. ARIAD was inducing Fas ligand signaling, which is far upstream. Principal challenge was basal activity, i.e. cells die in absence of AP1903, and select for knock-out

AML抗原特异的免疫治疗 NE P3 No AML AML Molldrem et al PR1-CTL Leukemia PR1-CTL are naturally enriched (0.1-0.4%) in fetal cord blood Proteasome P3 NE TCR Leukemia PR1 peptide HLA-A2 PR1 PR1-CTL PR1/HLA-A2 No AML Clinical trials with cord blood-derived PR1-CTL are ongoing for transplant recipients (AML, CML) AML Shared Resources Flow Cytometry and Cellular Imaging Facility, Genetically Engineered Mouse Facility, Monoclonal Antibody Facility; Clinical Trials Support Resource Molldrem et al

通过嵌合抗原受体(CARs)介入的变更的 T-细胞特异性 TCR-complex Antibody a b g e e d vL z z Fab vL vH vH CL CL CH1 CH1 vH vL Chimeric antigen receptor

嵌合抗原受体 Cooper et al

嵌合抗原受体T-细胞 T-细胞特异性变更 能够靶向非免疫靶点，组织/治疗特异抗原如 B 细胞淋巴瘤的 CD19 第一、二和三代的组成，包括共刺激分子 CD28, CD137 可以提高细胞的体内生存和增殖 很好设计的 CAR 尚无 少数患者接受自体或异体CAR+细胞, 部分CR, 清除了 CD19+ 细胞

CAR+ T细胞杀死表达CD19的肿瘤细胞 Tumor Time lapse bioluminescent imaging Measuring light.. Time lapse bioluminescent imaging

CAR 存在的问题 自体 vs. 异体 体内生存、归巢 毒性,应用含有 CD137的 CARs可致“细胞因子风暴” 时间/ 费用用于制造患者特异的产物

用于过继免疫“现成的” CD19-特异的 CAR+T细胞 Cooper et al

NK 细胞 独特的一类淋巴细胞, 固有没有系统的组成 CD3- TCR-, CD16+, CD56+ 激活和抑制受体 (KIR) 介导抗肿瘤, 抗病毒l, BM 排斥 丢失配体假说调控的细胞毒作用 Cw alleles that bind to KIR2DL1 have amino acid K at position 80. Cw alleles that bind to KIR2DL2 or to KIR2DL3 have amino acid N at position 80 Bw4 or Bw6, KIR 3DL1 amino acids at positions 82-83 NK 再激活报道称可降低复发 (抗AML), 改善植入 (抗宿主 T-细胞), 降低 GVHD (抗树突细胞)

通过激活和抑制受体调控 NK 细胞反应 Farag S S et al. Blood 2002;100:1935-1947 ©2002 by American Society of Hematology

Lysis Lysis leukemia Lysis Kill recipient APCs = protection from GvHD Donor alloreactive NK cells DC DC DC NK Lysis NK NK Lysis leukemia In summary, our mouse models show the pre-transplant infusion of alloreactive NK cells eradicates leukaemia, ablates host T cells and therefore conditions the host to transplantation and ablates host antigen presenting cells thereby protecting the host from GvHD. Lysis Kill leukemia = GvL effect T T T T T T Kill recipient T cells = improved engraftment

制造 mbIL21-扩增的 NK cells Cryopreserve in aliquots

Haploidentical Allo reactive Allo match NK Cells PBPC Busulfan Fludarabine ATG 24