Molecular Mechanisms and Treatment Of Autoimmunity In Man And Animal Models

在这个项目中，我们正在开发免疫调节失调疾病的新诊断和治疗方法。首先，我们追求新的治疗方法，重点是多发性硬化症(MS)以及我们已经确定其遗传基础的新的免疫学疾病。我们的主要方法是利用抗原本身来编程特定的同源T细胞，使其通过一种称为再刺激诱导细胞死亡(RICD)的过程通过凋亡而死亡。这种死亡是克隆性的，代表了一种通过使用抗原消除致病T细胞的方法。此外，根据疾病的分子和遗传机制，我们还将研究使用其他小分子或蛋白质为基础的治疗方法。关键是准确地根据疾病的病理生理机制量身定做治疗方法，以使其高度有效。这是“精准”医学的首要目标。 目前，我们选择专注于利用免疫学和遗传学方法开发新的、高效的治疗多发性硬化症的方法。有证据表明，髓鞘蛋白抗原是自身免疫攻击的目标，但这些特异性如何决定进展性和复发缓解型多发性硬化症的疾病结局尚不清楚。通过编程让识别这种抗原的T细胞死亡，可以证明消除这些细胞对疾病的影响。尽管双胞胎研究表明多发性硬化症的易感性有显著的遗传成分，但对涉及的基因或途径几乎没有具体的了解。全基因组关联研究已经发现了多种单核苷酸多态(SNPs)，这些SNPs在统计学上与疾病相关。国际多发性硬化症遗传学联合会已经进行了大量的遗传多样性分析，但发现的与多发性硬化症密切相关的SNPs的数量远远低于预期。然而，这些研究表明，相关的SNPs主要位于免疫基因内或周围，而不是中枢神经系统基因。然而，在多发性硬化症的不同亚型中具体途径的含义还没有达到。因此，我们得出结论，通过搜索孟德尔或从头开始的遗传变异来解决这个问题可能会提供关于发病机制的更明确的信息。使用下一代测序(NGS)技术，如全外显子组测序(WES)和全基因组测序(WGS)，我们现在能够识别罕见和严重疾病的原因基因，这些疾病没有足够的适合度在人群中广泛遗传，并被GWAS检测到。此外，这些高渗透性和有害的变种可能极大地促进新生和孟德尔形式的自身免疫性疾病以及对常见自身免疫性疾病的易感性。罕见遗传变异的鉴定通常提供有关疾病机制、生物学途径和临床治疗新途径的关键信息。我们已经与加拿大不列颠哥伦比亚大学的研究人员建立了合作关系，他们已经收集了近14,000个样本的DNA生物库，其中包括超过450个多发性多发性硬化症家庭：1.有3个或更多受影响的成员患有多发性硬化症的家庭(400个家庭)；2.具有首发年龄和12岁的索引病例的家庭(极其罕见，在本研究中可用)。大约35%的样本来自受影响的患者，大多数被诊断为21岁至40岁之间的多发性硬化症，平均发病年龄为31.4岁。这些患者、家属和亲属将通过与Regeneron测序中心的合作接受WES。我们期望NGS在这些患者中发现的基因变异，特别是在前两类中，将揭示MS的主要致病途径。 我们也一直在追求治疗的方向，特别是针对一种我们已经发现的独特的疾病，我们发现了一种名为“p110增量激活突变导致衰老T细胞、淋巴结病和免疫缺陷”(Pasli)疾病。我们研究了来自7个不相关家系的9名患者以及PI3K调节亚基严重突变的9个患者的PI3K p110Delta催化亚基的种系、杂合、显性、功能获得突变。我们研究了PASLI患者中特定的p110Delta酶抑制剂，并与诺华公司签订了合作研发协议，在NIH临床中心进行临床试验，测试他们的p110Delta抑制剂，一种名为leniolisib(CDZ173)的化合物。初步证据表明，Leniolisib是一种有效和选择性的p110 Delta抑制剂，可以抑制野生型和过度活性的酶，从而有效地抑制疾病，副作用很少或没有副作用。我们完成了一项更大规模的三盲、随机、安慰剂对照研究的第三阶段。我们发现，与安慰剂相比，接受Leniolisib治疗的患者在共同原发终点方面取得了显著的好处，减少了淋巴结病，增加了原始B细胞的百分比，并且对帕斯利综合征患者的耐受性很好。这导致FDA批准Leniolisib用于治疗帕斯利病。

In this project, we are developing new diagnostic and therapeutic approaches to immune dysregulatory diseases. First, we have pursued novel therapeutic approaches focusing on multiple sclerosis (MS) as well as new immunological diseases for which we have determined the genetic basis. Our principle approach is to use antigen itself to program the specific cognate T cells to die through apoptosis via a process termed restimulation-induced cell death (RICD). The death is clonally specific and represents a way to eliminate disease-causing T cells by using antigen. Additionally, depending on the molecular and genetic mechanism of disease, we will also investigate using other small molecules or protein-based therapeutics. The key is to precisely tailor the therapy to the pathophysiological mechanism of the disease so that it will be highly effective. This is the overarching goal of "precision" medicine. At the present time, we have chosen to focus on developing new, highly effective therapeutics against MS using immunological and genetic approaches. Evidence suggests that myelin proteins antigens are targets of the autoimmune attack, but how the specificities determine disease outcome in progressive and relapsing-remitting MS is unclear. By programming the T cells that recognize such antigens to die, the effect of eliminating these cells on the disease can be demonstrated. Although twin studies show that there is a significant genetic component to MS susceptibility, there is little concrete knowledge about the genes or pathways involved. Genome-wide association studies (GWAS) have unearthed a variety of single nucleotide polymorphisms (SNPs) that are statistically associated with disease. The International MS Genetics Consortium has performed numerous GWAS but the number of identified SNPs with strong association with MS has been much lower than anticipated. However, these studies that the correlated SNPs are in or around mainly immune genes rather than central nervous system genes. Nonetheless, the implication of specific pathways in various subtypes of MS has not been achieved. Therefore, we have concluded that approaching the problem by searching for Mendelian or de novo genetic variants may provide more definitive information about pathogenesis. Using next generation sequencing (NGS) techniques such as whole exome sequencing (WES) and whole genomic sequencing (WGS) we are now capable of identifying causal genes in rare and severe disorders that do not have enough fitness to be inherited broadly in the population and detected by GWAS. Moreover, these highly penetrant and deleterious variants likely contribute strongly to de novo and Mendelian forms of autoimmune disease as well as susceptibility to common autoimmune diseases. Identification of rare genetic variants generally provides key information about disease mechanisms, biological pathways, and novel avenues for clinical treatments. We have established a collaboration with investigators at the University of British Columbia, Canada who have assembled a DNA biobank of nearly 14,000 samples, including over 450 Multiplex MS families: 1. Families with 3 or more affected members with MS (400 families); 2. Families with index cases with age of onset < 12 years old (extremely rare and available under this study). Approximately 35% of the samples are from affected patients, and most were diagnosed with MS between the ages of 21 and 40 years old, with the average age of onset being 31.4 years old. These patients, family members, and relatives will be subjected to WES through a collaboration with the Regeneron Sequencing Center. We expect that gene variants found by NGS in these patients, particular in the first two categories, will unveil major pathogenic pathways for MS. We have also pursued the direction of therapeutics particularly for a unique disorder we have discovered called "p110delta activating mutations causing Senescent T cells, Lymphadenopathy, and Immunodeficiency" (PASLI) disease. We studied the germline, heterozygous, dominant, gain-of- function mutations in the p110delta catalytic subunit of PI3K in nine patients from seven unrelated families and also families with severe mutations in the regulatory subunit of PI3K. We investigated specific p110delta enzyme inhibitors in PASLI patients and entered into a Cooperative Research and Development Agreement with Novartis to carry out a clinical trial testing their inhibitor of p110delta, a compound called leniolisib (CDZ173), in the NIH Clinical Center. Preliminary evidence showed that leniolisib is a potent and selective inhibitor of p110delta and can inhibit the wild-type as well as overactive forms of the enzyme thereby potently suppressing disease with few or no side effects. We completed a phase III of a larger triple-blinded, randomized, placebo-controlled study. We found that patients who received leniolisib achieved a significant benefit over placebo with respect to the co-primary endpoints, reducing lymphadenopathy and increasing the percentage of naive B cells, and was well tolerated in patients suffering from PASLI. This resulted in FDA-approval of leniolisib for treatment of PASLI disease.