Christopher H. Chen, Ph.D. - US grants

ABSTRACT Heart failure (HF), a leading cause for cardiac transplantation and premature death, is usually preceded by ventricular dilatation and diminished systolic performance or dilated cardiomyopathy (DCM). DCM has many etiologies, including damaging variants in genes with diverse functions in cardiac biology. During the prior funding period we showed that the most common genetic cause of DCM was truncating variants in titin (TTNtv). These account for 25% of familial and 12% of sporadic DCM and for ~10% of DCM that occurs with pregnancy, alcohol abuse, and after cancer therapies. In addition, ~0.2% of the general population carries a TTNtv; these individuals have substantially higher lifelong risks for developing DCM and heart failure. We also identified mechanisms by which TTNtv and other recently recognized DCM genes (FLNC and ALPK3) cause disease. With this competitive renewal we propose to focus on the discovery of genes and mechanisms that account for unexplained DCM, which remains an unmet need. We propose that some unexplained DCM is mechanistically related to established genetic causes and results from sequence variants that are not routinely interrogated, or that have unclear functional consequences. We will study the roles of somatic variants, non-coding regulatory variants, mitochondrial variants, and variants of unknown significance (VUS) in established and newly identified DCM genes. We will also define cell populations and transcriptional profiles of all cells in human hearts with unexplained DCM and DCM with established genetic etiologies, so as to identify shared or distinct pathways that may inform therapeutic opportunities. Our analyses will employ state-of-the art technologies. We will exploit whole genome sequencing (WGS) from blood- and cardiac tissue-derived DNAs obtained from unexplained DCM subjects. We will use single nuclear RNA sequencing (NucSeq) to define how cell populations and transcription change in DCM hearts in comparison to normal hearts, using our recently completed normal human heart NucSeq data. We will perturb new identified variants and mechanisms in iPSC-CMs and mouse models. These studies will improve knowledge of the molecules and pathways that enable normal heart function, the molecular causes and mechanisms of DCM, information that will improve diagnosis and inform precision therapies to prevent heart failure. Our analyses will also contribute functional insights into noncoding sequences. To accomplish these goals, we will: 1) Identify coding and non-coding, germline and somatic variants that contribute to unexplained DCM; 2) Define perturbed cell populations and associated transcriptional profiles in hearts from variant-positive and unexplained DCM; 3) Define DCM mechanisms using engineered iPSC-CMs and mouse models.

The cerebellum is no longer just a motor structure. Human imaging studies have pointed to links between the cerebellum and cognition, language, and affect. Preclinical work has also indicated that the cerebellum has many nonmotor functions ranging from aggression to sleep. Expression of autism related proteins within cerebellar principle neurons (Purkinje cells) is sufficient to recapitulate many hallmarks of the condition in mice. These Purkinje cells send their projections to the deep cerebellar nuclei (DCN) and vestibular nuclei (VN), often considered the only cerebellar output nuclei. However, the known output pathways that connect the cerebellum with the forebrain seem insufficient to explain the diversity of behaviors now associated with it. We found that there is an additional, underappreciated output pathway through the parabrachial nucleus that receives direct Purkinje cell input and projects to the forebrain. Unlike the conventional cerebellar outputs, this pathway has significant projections to the amygdala, basal forebrain, prefrontal cortex and others. The aim of this project is to characterize cerebellar inputs to the parabrachial and identify these novel cerebellar output targets. We will focus on the projection to the amygdala. There is a rich behavioral literature describing the cerebellum as a core component in fear extinction, though there is no known neural substrate underlying this. In humans, trauma to the cerebellum is a strong predictor of post- traumatic stress disorder. We will test whether this unconventional cerebellar output can potentially mediate this nonmotor behavior.