Search

Abstract Hypertrophic cardiomyopathy (HCM) afflicts humans, cats, pigs, and rhesus macaques. Disease sequelae include congestive heart failure, thromboembolism, and sudden cardiac death (SCD). Sarcomeric mutations explain some human and cat cases, however, the molecular basis in rhesus macaques remains unknown. RNA-Seq of the LV tissues of five HCM-affected and seven healthy control rhesus macaques was employed for differential transcriptomic analyses. DNA from 15 severely HCM-affected and 21 healthy geriatric rhesus macaques were selected for whole-genome sequencing. A genome-wide association study (GWAS) of disease status and SCD outcome was performed. 614 down- and 1,065 upregulated differentially expressed genes (DEGs) were identified between groups. The top DEG (MAFF) was overexpressed in affected animals (log2FoldChange = 4.71; P Adjusted-value = 1.14E-133). Channelopathy-associated enriched terms were identified in ~ 57% of downregulated DEGs providing transcriptomic evidence of hypertrophic and arrhythmic disease processes. For GWAS, no putative variant withstood segregation. Polygenic modeling analysis resulted in poor prediction power and burden testing could not explain HCM by an association of multiple variants in any gene. Neither single nor compound genetic variant(s), or identified polygenic profile, suggest complex genotype–phenotype interactions in rhesus macaques. Brought forth is an established dataset of robustly phenotyped rhesus macaques as an open-access resource for future cardiovascular disease genetic studies.

Background Hypertrophic cardiomyopathy (HCM) is a common heritable heart disease where the most frequently associated mutations occur in the myosin‐binding protein C (MYBPC3) sarcomere‐associated gene. HCM is also a common veterinary clinical problem in certain cat breeds such as Maine Coons and Ragdolls, also most associated with mutations in MYBPC3. Mouse models of HCM in which Mybpc3 mutations are introduced recapitulate some, but not all, features of human HCM. Methods and Results To elucidate the common and distinctive pathological pathways across species and foster a greater understanding of the concordance of mouse HCM models to clinical mybpc3‐associated HCM, we generated single nuclei RNA‐sequencing data sets from feline, human, and murine heart tissue carrying MYBPC3 variants. Numerous genes were differentially expressed between mutation positive and mutation negative cell types within each species, identified using the model‐based analysis of single‐cell transcriptomics algorithm. Gene Ontology enrichment analysis of differentially expressed genes in cardiomyocytes across species revealed alterations in genes involved in muscle development, muscle contraction, muscle hypertrophy, regulation of sarcoplasmic calcium release, ATP metabolic process, and oxidative phosphorylation. Conclusions These common biological processes across species are consistent with known phenotypic aspects of HCM such as hypertrophy, hypercontractility, diastolic dysfunction, and altered energy metabolism. Surprisingly, among conserved biological processes within cardiomyocytes across species, the individual genes driving the biological processes were distinct. This work to identify common and species‐specific disease‐promoting pathway differences will allow development of targeted therapies for both human and veterinary application and will facilitate an understanding of the idiosyncrasies of mouse models.

Abstract Hypertrophic cardiomyopathy (HCM) remains the most common cardiomyopathy in humans and cats with few preclinical pharmacologic interventional studies. Small-molecule sarcomere inhibitors are promising novel therapeutics for the management of obstructive HCM (oHCM) patients and have shown efficacy in left ventricular outflow tract obstruction (LVOTO) relief. The objective of this study was to explore the 6-, 24-, and 48-hour (h) pharmacodynamic effects of the cardiac myosin inhibitor, CK-586, in six purpose-bred cats with naturally occurring oHCM. A blinded, randomized, five-treatment group, crossover preclinical trial was conducted to assess the pharmacodynamic effects of CK-586 in this oHCM model. Dose assessments and select echocardiographic variables were assessed five times over a 48-h period. Treatment with oral CK-586 safely ameliorated LVOTO in oHCM cats. CK-586 treatment dose-dependently eliminated obstruction (reduced LVOTOmaxPG), increased measures of systolic chamber size (LVIDs Sx), and decreased select measures of heart function (LV FS% and LV EF%) in the absence of impact on heart rate. At all tested doses, a single oral CK-586 dose resulted in improved or resolved LVOTO with well-tolerated, dose-dependent, reductions in LV systolic function. The results from this study pave the way for the potential use of CK-586 in both the veterinary and human clinical setting.

Abstract Background Glanzmann's thrombasthenia (GT) is a congenital platelet disorder affecting approximately 1:1 000 000 people globally and characterized by impaired platelet aggregation and clot retraction. Autosomal recessive, loss‐of‐function, variants in ITGA2B or ITGB3 of the αIIbβ3 receptor cause the disease in humans. A cat affected by Glanzmann's and macrothrombocytopenia was presented to the UC Davis VMTH. Hypothesis/Objectives Severe thrombopathia in this cat has an underlying genetic etiology. Animals A single affected patient, 2 age‐matched clinically healthy controls, and a geriatric population (n = 20) of normal cats. Methods Physical examination and clinical pathology tests were performed on the patient. Flow cytometry and platelet aggregometry analyses for patient phenotyping were performed. Patient and validation cohort gDNA samples were extracted for Sanger sequencing of a previously identified ITGA2B (c.1986delC) variant. Reverse transcriptase PCR was performed on patient and healthy control PRP samples to verify ITGA2B variant consequence. Results A novel c.1986_1987insCC autosomal recessive variant in ITGA2B was identified. This variant was absent in a population of 194 unrelated cats spanning 44 different breeds. Complete loss of ITGA2B transcript and protein expression was verified by RT‐PCR and flow cytometry, explaining the underlying etiology of GT, and likely macrothrombocytopenia, in this cat. Conclusions and Clinical Importance This study emphasizes the role of precision medicine in cardiovascular disease of cats and identified yet another variant that may be of utility for screening in the feline population. This study provides a small‐volume, standardized, successful protocol for adequate platelet RNA isolation and subsequent molecular assessment of gene expression in cats.

Abstract We sought to establish a large animal model of inherited hypertrophic cardiomyopathy (HCM) with sufficient disease severity and early penetrance for identification of novel therapeutic strategies. HCM is the most common inherited cardiac disorder affecting 1 in 250–500 people, yet few therapies for its treatment or prevention are available. A research colony of purpose-bred cats carrying the A31P mutation in MYBPC3 was founded using sperm from a single heterozygous male cat. Cardiac function in four generations was assessed by periodic echocardiography and measurement of blood biomarkers. Results showed that HCM penetrance was age-dependent, and that penetrance occurred earlier and was more severe in successive generations, especially in homozygotes. Homozygosity was also associated with progression from preclinical to clinical disease. A31P homozygous cats represent a heritable model of HCM with early disease penetrance and a severe phenotype necessary for interventional studies aimed at altering disease progression. The occurrence of a more severe phenotype in later generations of cats, and the occasional occurrence of HCM in wildtype cats suggests the presence of at least one gene modifier or a second causal variant in this research colony that exacerbates the HCM phenotype when inherited in combination with the A31P mutation.

Abstract Hypertrophic cardiomyopathy (HCM) is the most prevalent inherited cardiac disease in humans and cats and lacks efficacious pharmacologic interventions in the preclinical phase of disease. LV outflow tract obstruction (LVOTO) is commonly observed in HCM-affected patients and is a primary driver of heart failure symptoms and reduced quality of life. Novel small-molecule cardiac myosin inhibitors target actin-myosin interactions to alleviate overactive protein interactions. A prospective, randomized, controlled cross-over study was performed to evaluate pharmacodynamic effects of two doses (0.3 and 1 mg/kg) of a next-in-class cardiac myosin inhibitor, aficamten (CK-3773274, CK-274), on cardiac function in cats with the A31P MYBPC3 mutation and oHCM. Dose-dependent reductions in LV systolic function, LVOT pressure gradient, and isovolumetric relaxation times compared to baseline were observed. Promising beneficial effects of reduced systolic function warrant further studies of this next-in-class therapeutic to evaluate the benefit of long-term administration in this patient population.

Hypertrophic cardiomyopathy (HCM) remains the single most common cardiomyopathy in cats, with a staggering prevalence as high as 15%. To date, little to no direct therapeutical intervention for HCM exists for veterinary patients. A previous study aimed to evaluate the effects of delayed-release (DR) rapamycin dosing in a client-owned population of subclinical, non-obstructive, HCM-affected cats and reported that the drug was well tolerated and resulted in beneficial LV remodeling. However, the precise effects of rapamycin in the hypertrophied myocardium remain unknown. Using a feline research colony with naturally occurring hereditary HCM (n = 9), we embarked on the first-ever pilot study to examine the tissue-, urine-, and plasma-level proteomic and tissue-level transcriptomic effects of an intermittent low dose (0.15 mg/kg) and high dose (0.30 mg/kg) of DR oral rapamycin once weekly. Rapamycin remained safe and well tolerated in cats receiving both doses for eight weeks. Following repeated weekly dosing, transcriptomic differences between the low- and high-dose groups support dose-responsive suppressive effects on myocardial hypertrophy and stimulatory effects on autophagy. Differences in the myocardial proteome between treated and control cats suggest potential anti-coagulant/-thrombotic, cellular remodeling, and metabolic effects of the drug. The results of this study closely recapitulate what is observed in the human literature, and the use of rapamycin in the clinical setting as the first therapeutic agent with disease-modifying effects on HCM remains promising. The results of this study establish the need for future validation efforts that investigate the fine-scale relationship between rapamycin treatment and the most compelling gene expression and protein abundance differences reported here.

Idiopathic hypocalcemia in Thoroughbred (TB) foals causes tetany and seizures and is invariably fatal. Based upon the similarity of this disease with human familial hypoparathyroidism and occurrence only in the TB breed, we conducted a genetic investigation on two affected TB foals. Familial hypoparathyroidism was identified, and pedigree analysis suggested an autosomal recessive (AR) mode of inheritance. We performed whole-genome sequencing of the two foals, their unaffected dams and four unaffected, unrelated TB horses. Both homozygosity mapping and an association analysis were used to prioritize potential genetic variants. Of the 2,808 variants that significantly associated with the phenotype using an AR mode of inheritance (PA,p.Ser875*) was significantly associated with the hypoparathyroid phenotype (Pallelic = 0.008). Affected foals were homozygous for the variant, with two additional affected foals subsequently confirmed in 2019. Necropsies of all affected foals failed to identify any histologically normal parathyroid glands. Because the nonsense mutation in RAPGEF5 was near the C-terminal end of the protein, the impact on protein function was unclear. Therefore, we tested the variant in our Xenopus overexpression model and demonstrated RAPGEF5 loss-of-function. This RAPGEF5 variant represents the first genetic variant for hypoparathyroidism identified in any domestic animal species.