PROJECT SUMMARY
Autism-spectrum disorders impact millions of individuals worldwide, representing a heavy toll on affected
children, their families, and the health care system. Pitt–Hopkins Syndrome (PTHS) is an ASD caused by de
novo mutations in the TCF4 gene. PTHS is characterized by severe intellectual disability, pronounced
developmental and motor delays, absence of speech, repetitive behaviors, peculiar facial gestalt, and
gastrointestinal manifestations. While the genetic etiology of PTHS is well established, the cellular and neural
phenotypic alterations in human patients are still not fully understood, nor is it clear how TCF4 mutations
cause such abnormalities. Lack of understanding about PTHS's molecular and cellular mechanisms is a
problem because, until this information becomes available, specific altered pathways cannot be therapeutically
targeted. Moreover, without neuropathological knowledge, it is impossible to treat and eventually cure PTHS
by directly correcting the mutation in the genome.
Our long-term goal is to understand how specific genetic defects and altered pathways in the brain result
in the debilitating phenotypes exhibited by autistic children. The objectives of this application are to: (a) use
human models of neural development in vitro to define the cellular and neural pathological consequences of
clinically relevant TCF4 mutations in PTHS; and (b) provide proof-of-concept that correctional molecular
strategies can be used to fix TCF4 expression, an approach that could eventually be used as gene therapy for
PTHS. Our central hypothesis is that TCF4 mutations cause aberrant phenotypes in specific cell types of the
nervous system, leading to the patients' neurological symptoms. We postulated that patient-derived in vitro
models of PTHS can better recapitulate the pathophysiology than mouse models, because brain structure,
genome architecture and development vary greatly between rodents and humans, and current PTHS animal
models do not closely mimic all the disease's clinically relevant aspects. In preliminary experiments, we
obtained patient-derived brain organoids and cultured neural cell types in vitro and used them as human
models to show that PTHS neural progenitor cells exhibit senescence and decreased proliferation,
accompanied by downregulation of Wnt signaling and SOX3 expression. Moreover, we observed that PTHS
brain organoids fail to develop normal anatomically organized progenitor structures and that PTHS neurons
display severely impaired firing properties. Our anticipated results/deliverables include the identification and
manipulation of specific altered molecular pathways and neural cell types and the testing of genetic
correctional strategies for the disease, which could propel future research on pharmacological and gene
therapy for PTHS.