Human organoid model of PCH2a recapitulates brain region-specific pathology

Abstract: Pontocerebellar hypoplasia type 2 a (PCH2a) is a rare, autosomal recessive neurogenetic disorder. Affected individuals present with early and severe neurological impairment. The anatomical hallmark of PCH2a is the hypoplasia of the cerebellum and pons accompanied by progressive microcephaly over the first years of life (OMIM #277470). Treatment options are limited and symptomatic. PCH2a results from a homozygous founder variant in the TSEN54 gene (OMIM *608755), which encodes a tRNA splicing endonuclease complex subunit. A recent study revealed altered tRNA pools in fibroblasts from affected individuals with PCH2a. However, the pathological mechanism underlying the hypoplasia of the cerebellum and the progressive microcephaly is unknown due to a lack of a model system. Leveraging recent progress in organoid generation, we developed human models of PCH2a using brain region-specific organoids. We, therefore, obtained skin biopsies from three affected males with genetically confirmed PCH2a and derived induced pluripotent stem cells (iPSCs). Cerebellar and neocortical organoids were differentiated from control and affected iPSCs and showed expression of TSEN54. In line with neuroimaging findings in affected individuals, PCH2a cerebellar organoids are reduced in size compared to controls starting early in differentiation. While neocortical PCH2a organoids also show growth deficits, they are less pronounced than those in cerebellar organoids, reminiscent of the progressive microcephaly in PCH2a. In addition, we do not find evidence of increased apoptosis in PCH2a organoids compared to controls in contrast to what has been suggested in loss-of-function animal models. We have generated a human model of PCH2a, which will allow to decipher disease mechanisms in depth in order to explain how a variant in a ubiquitously expressed gene involved in tRNA metabolism causes pathology in specific brain regions.