Role of Brain Macrophages in the Pathogenesis and Treatment of Globoid Cell Leukodystrophy

球形细胞白质营养不良(GLD)，或Krabbe，是一种致命的儿童神经退行性疾病，由 GALC基因突变。之所以这样命名，是因为球状细胞巨噬细胞的出现。面临的主要障碍是 治疗GLD是对中枢神经系统(CNS)病理的治疗。造血干细胞移植(HSCT) 是唯一的治疗方法，但不能治愈，必须在婴儿早期进行症状前治疗。HSCT 被认为是通过治疗性植入供体巨噬细胞和替换球状细胞来发挥作用，但在 大脑，这样做效率很低。尽管是GLD的病原体，但对大脑中的球状细胞知之甚少 --它们的功能、起源和形成。目前尚不清楚球形细胞是否起源于胚胎组织。 驻留的小胶质细胞或HSC来源的浸润性巨噬细胞，它们的致病程度，以及它们的 替代治疗是治疗GLD的关键。这是一个严重的知识鸿沟，限制了更多 有效的GLD疗法，是这项提案的重点。我们的中心假设是球状细胞是 独一无二反应性小胶质细胞和由“真正的”小胶质细胞强有力的替代足以治疗GLD CNS 神经病理学。我们是通过在小鼠体内直接移植中枢神经系统来研究脑巨噬细胞的专家。这个 抽动(Twitcher，GALCKO)小鼠是广为接受的GLD模型。我们创造了新的方法来区分 小胶质细胞、浸润性巨噬细胞和移植的供者巨噬细胞相互取代2)宿主 脑巨噬细胞直接高效注射细胞而不需要造血干细胞移植，包括通过工程第一次 脑巨噬细胞的生存受体CSF1R的小分子抗药性变体。在这份提案中， 我们将在GALCKO模型中应用这些新方法来确定脑巨噬细胞在GLD中的作用 发病机制及治疗。在目标1中，我们将定义所有反应性脑的起源和转录识别 巨噬细胞，包括球状细胞，在GALCKO，包括HSCT后。这一知识有望揭示新的 GLD的治疗靶点。在目标2中，我们将检验GALCKO小胶质细胞直接替换的假设 与健康的替代品，消除球状细胞，并推动HSCT的神经治疗效果。如果为真，则这 方法具有巨大的翻译潜力，最大限度地提高植入效率，扩大治疗范围 细胞治疗的窗口。最后，在目标3中，我们将确定HSC来源的细胞是否为有效的小胶质细胞 代理人，鉴于非小胶质巨噬细胞在中枢神经系统中的不同功能。这将指导今后的工作 提高脑部疾病细胞疗法的疗效。这些目标的实现将填补长期存在的空白 关于小胶质细胞和其他巨噬细胞在急性髓细胞白血病发病和治疗中的作用的知识空白 儿童神经退行性疾病。

Globoid cell leukodystrophy (GLD), or Krabbe, is a fatal pediatric neurodegenerative disease caused by mutations in GALC. It is so-named due to the appearance of globoid cell macrophages. The major hurdle to curing GLD is treatment of central nervous system (CNS) pathology. Hematopoietic stem cell transplant (HSCT) is the only treatment, but is not curative, and must be administered presymptomatically in early infancy. HSCT is thought to work by therapeutic engraftment of donor macrophages and replacement of globoid cells, but in the brain, does so inefficiently. Despite being pathognomonic for GLD, little is known about globoid cells in the brain - their function, origin, and formation. It is unknown if globoid cells arise from embryonically-derived tissue resident microglia or HSC-derived infiltrating macrophages, the degree to which they are pathogenic, and if their replacement is key to GLD treatment. This is a critical knowledge gap that has limited the advancement of more effective GLD therapies and is the focus of this proposal. Our central hypothesis is that globoid cells are unique reactive microglia and that robust replacement by “true” microglia is sufficient to treat GLD CNS neuropathology. We are experts in the study of brain macrophages by direct CNS transplantation in mice. The twitcher (GALCKO) mouse is a widely accepted model of GLD. We created new methods to 1) distinguish microglia, infiltrating macrophages, and transplanted donor macrophages from each other and 2) replace host brain macrophages with directly injected cells at high efficiency without HSCT, including by engineering the first small molecule inhibitor-resistant variant of CSF1R, a survival receptor for brain macrophages. In this proposal, we will apply these new methods in the GALCKO model to determine the role of brain macrophages in GLD pathogenesis and treatment. In Aim 1, we will define the origin and transcriptomic identity of all reactive brain macrophages, including globoid cells, in GALCKO, including after HSCT. This knowledge promises to reveal new therapeutic targets for GLD. In Aim 2, we will test the hypothesis that direct replacement of GALCKO microglia with healthy surrogates eliminates globoid cells and drives the neurotherapeutic effects of HSCT. If true, this approach has great translational potential to maximize engraftment efficiency and broaden the therapeutic window for cell therapy. Finally in Aim 3, we will determine if HSC-derived cells are effective microglial surrogates, given the distinct functions of non-microglial macrophages in the CNS. This will guide future work to enhance the efficacy of cell therapies for brain diseases. Completion of these aims will fill longstanding knowledge gaps about the role of microglia and other macrophages in the pathogenesis and treatment of pediatric neurodegenerative diseases.