Survival of the fittest HSPC repopulating clones by anti-HIV-1 gene-modification

描述（由申请人提供）：项目总结本项目的总体目标是了解抗HIV-1转基因对造血干祖细胞（HSPC）进行基因修饰的后果，这些基因修饰可保护携带细胞免受HIV-1感染或复制。我们现在知道，稳定的长期再增殖是一个高度复杂的过程，其中数百至数千个具有不同寿命和谱系输出潜力的HSPC在移植后做出贡献。我们将在克隆水平上研究人类再生细胞的行为，并将抗HIV-1 shRNA转基因的基因修饰用作我们的模型。我们将测试的假设，在体内的选择性压力驱动的HIV-1或化学手段可以扩大克隆群体的基因标记的细胞通常逆转赤字造成的HIV-1。最早的造血干细胞移植研究表明，造血干细胞（HSC）是一个相对同质的群体，其中任何一个都有潜力完全重建造血系统。然而，最近，这种观点被证明是简单化的。干细胞克隆可以聚集成具有广泛不同的利用和谱系定型动力学的组。人们可以认为干细胞的再增殖是一个过程，其中数百或数千个具有不同特性的移植细胞以高度受控的方式竞争以维持稳态。 通常认为，体内富集基因工程改造的HSPC或成熟T细胞是关键的，以便提供足够的受保护细胞来抵抗HIV-1破坏。研究表明，当未受保护的细胞被HIV-1杀死时，HIV-1感染对受保护的细胞施加了选择压力，使其在人源化小鼠中存活。已经在动物中研究了使用化学选择剂进一步富集基因修饰的细胞。然而，考虑到HSPC代表如此多样的克隆群体，关于HIV-1和化学试剂对不同HSPC克隆施加选择性压力的机制和程度知之甚少。除了柏林的病人，100%的受体细胞被供体细胞取代，以前的临床研究没有建立足够水平的“保护”T细胞来控制HIV-1。因此，如果我们要成功开发基于干细胞的HIV-1治疗，我们必须充分研究HIV-1和/或化学试剂在移植HSPC后富集基因修饰细胞的程度和机制。

DESCRIPTION (provided by applicant): PROJECT SUMMARY The overall goal of this project is to understand the consequences of gene modification of hematopoietic stem progenitor cell (HSPC) by anti-HIV-1 transgenes which protect the harboring cells from HIV-1 infection or replication. We now know that stable long-term repopulation is a highly complex process whereby hundreds to thousands of HSPCs with differing life-spans and lineage output potentials contribute after transplant. We will investigate the behavior of human repopulating cells at the clonal level and use as our model, gene modification with anti-HIV-1 shRNA transgenes. We will test the hypothesis that in vivo selective pressures driven by HIV-1 or chemical means can expand clonal populations of gene-marked cells normally to reverse deficits caused by HIV-1. The earliest studies of hematopoietic stem cell transplant suggested that hematopoietic stem cells (HSC) were a relatively homogeneous population, any one of which has the potential to fully repopulate the hematopoietic system. However, more recently, this view has proven to be simplistic. Stem cell clones can be clustered into groups with widely differing kinetics of utilization and lineage commitment. One can consider repopulation by the stem cells as a process whereby hundreds or thousands of engrafted cells with differing properties compete in a highly controlled fashion to maintain homeostasis. It is generally thought that in vivo enrichment for the gene engineered HSPC or mature T-cells is critical in order to provide sufficient protected cells to withstand HIV-1 destruction. Studies have shown that HIV-1 infection imposes selection pressure for protected cells to survive in humanized mice when unprotected cells are killed by HIV-1. Further enrichment of gene-modified cells using chemoselection agents have been studied in animals. However, given that HSPC represent such diverse clonal populations, little is understood regarding the mechanism and degree to which HIV-1 and chemical agents exert selective pressure upon different HSPC clones. With the exception of the Berlin patient, where 100% of recipient cells were replaced by donor cells, no previous clinical studies established sufficient levels of "protected" T-cells to control HIV-1. Therefore, if we are to be successful in developing a stem cell-based therapy for HIV-1, we must fully investigate the extent and the mechanism by which HIV-1 and/or chemical agents can enrich for gene-modified cells after transplant of HSPC.