Heart disease affects millions of individuals and prime editing (PE) may enable curative therapies that address the underlying drivers of heart disease. Here we describe the establishment and optimization of an in vivo cardiac PE platform which mediates efficient editing in the heart with no detectable editing in the liver. We performed a proof-of-concept test on RNA binding motif protein 20 (RBM20) , which if mutated, can cause dilated cardiomyopathy (DCM) in humans. Our dual-AAV based PE therapeutic rescued cardiomyopathy phenotypes in the heterozygous Rbm20R636Q mouse model. To further develop PE targeting human RBM20 , we introduced a novel humanized mouse model carrying human RBM20 wildtype (WT) or R634Q mutant sequences and displaying RBM20 cardiomyopathy phenotypes. Our human RBM20 PE therapeutic efficiently corrected the pathogenic mutation and rescued phenotypes in the humanized RBM20 mouse model. Our findings demonstrate the potential of in vivo cardiac PE in treating heart disease, offer a valuable humanized DCM mouse model for developing various therapies, and present an optimized in vivo PE platform that can be adopted for targeting other organs and tissues. ### Competing Interest Statement W.L., L.M.R., E.E., H.Z., C.D.H., A.G.S., T.H., L.M.L., K.N.I., and Z.C. performed this research as part of their employment at Tenaya Therapeutics. W.L., L.M.R., E.E., H.Z., and Z.C. are inventors of patent applications filed by Tenaya Therapeutics relating to the work described in this article.

Hypertrophic cardiomyopathy (HCM) affects approximately 600,000 people in the United States. Loss-of-function mutations in Myosin Binding Protein C3, MYBPC3, are the most common genetic cause of HCM, with the majority of mutations resulting in haploinsufficiency. To restore cardiac MYBPC3, we use an adeno-associated virus (AAV9) vector and engineer an optimized expression cassette with a minimal promoter and cis-regulatory elements (TN-201) to enhance packaging efficiency and cardiomyocyte expression. Rather than simply preventing cardiac dysfunction preclinically, we demonstrate in a symptomatic MYBPC3-deficient murine model the ability of AAV gene therapy to reverse cardiac hypertrophy and systolic dysfunction, improve diastolic dysfunction, and prolong survival. Dose-ranging efficacy studies exhibit restoration of wild-type MYBPC3 protein levels and saturation of cardiac improvement at the clinically relevant dose of 3E13 vg/kg, outperforming a previously published construct. These findings suggest that TN-201 may offer therapeutic benefits in MYBPC3-associated cardiomyopathy, pending further validation in clinical settings.

Background SYNPO2L is a component of the sarcomere Z-disk with two distinct isoforms (SYNPO2L\_A and SYNPO2L\_B) described to regulate actin bundle size. Common and rare variations in SYNPO2L have been implicated in risk for heart failure and atrial fibrillation (AF); however, little is known about the function or pathophysiological mechanism upon disease risk. Methods The role of SYNPO2L was explored with human induced pluripotent stem cell cardiomyocyte (hiPSC-CM) models of a rare splicing mutation rs766868752 differentiated into both atrial and ventricular lineages. Electrophysiology was assessed via multi-electrode arrays (MEAs) and contractility was assessed using engineered heart tissues (EHTs). Genetic survival analyses were performed to explore the potential therapeutic role of the SYNPO2L_A isoform in AF risk after myocardial infarction. Results hiPSC-CMs carrying the splice mutation rs766868752 resulted in preferential expression of the SYNPO2L\_B isoform and absence of SYNPO2L\_A isoform, and atrial hiPSC-CMs displayed significant differences in action potential durations when unpaced, and spontaneous extra-systolic beats when paced. Contractility of atrial hiPSC-CMs carrying the splice mutation was severely compromised. Electrophysiological differences were normalized and contractility was partially restored by overexpression with AAV:SYNPO2L\_A. Absence of SYNPO2L\_A in the mutant led to an observed decreased phosphorylation of YAP along with significant downstream transcriptional effects. A direct interaction between SYNPO2L\_A and LATS2, a known regulator of YAP phosphorylation was found to occur in hiPSC-CMs. Additionally, a common variant time-to-event analysis may suggest a beneficial effect of the SYNPO2L\_A isoform to lower the risk of atrial fibrillation after myocardial infarction. Conclusions A rare splice mutation conferring isoform predominance of SYNPO2L\_B appears to disturb the electrophysiology and contractility of atrial hiPSC-CMs via LATS2 phospho-regulatory effects upon YAP signaling. Supplementation of the SYNPO2L\_A isoform can restore many functional deficits and supports a specific gene regulatory role for SYNPO2L in modulating the risk of AF and heart failure. ### Competing Interest Statement All authors were employees or contractors of Tenaya Therapeutics, Inc. at the time the work in this study was performed. All authors except for L.Z. and A.E.T. also hold equity in Tenaya Therapeutics, Inc.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease associated with ventricular arrhythmias and an increased risk of sudden cardiac death. Alterations in the desmosome gene plakophilin-2 (PKP2) lead to compromised contractility and electrical instability of cardiomyocytes. In this study, we utilized ARVC mouse and human induced pluripotent stem cell-derived cardiomyocyte models to confirm impaired energy metabolism that concorded with the human data. Our results supported an intrinsic cellular link between PKP2 and energy metabolism. TN-401-mediated PKP2 expression improved mitochondrial and glycolytic energetics and rescued cardiomyocyte functions that are dictated by mechanical and structural integrity of desmosome. Therefore, maintaining energy metabolism of cardiomyocytes is an integral part of PKP2 and desmosome functions, adding a new layer of understanding to ARVC disease mechanism.

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease associated with ventricular arrhythmias and an increased risk of sudden cardiac death. Mutations in the desmosome gene Plakophilin-2, PKP2, lead to reduction in PKP2 protein and collapse of desmosomes that is known to compromise contractility and electrical stability of cardiomyocytes. Our previous studies demonstrated the efficacy of adeno-associated virus 9 (AAV9)-mediated restoration of PKP2 expression in a cardiac specific knock-out mouse model of Pkp2 and revealed profound changes in mRNA signatures of metabolic enzymes that were reversed by the gene replacement approach. In this study, we used PKP2-deficient mouse hearts and human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to identify changes in steady-state metabolite levels associated with impaired lipid homeostasis, glycolysis, and glucose oxidation. These metabolic phenotypes align with human ARVC metabolic data and reflect an intrinsic impairment of cellular energy metabolism. Here we showed for the first time that these intracellular metabolic defects were associated specifically with poor contractility of cardiomyocytes. AAV9:PKP2 restored contractility, improved electrophysiological properties and Ca2+ transients. In contrast, we observed that treating PKP2-deficient cardiomyocytes pharmacologically with small molecule metabolic enhancers improved contractility but not electrophysiological properties and Ca2+ transients, suggesting differential sensitivity of structure-mediated functions in response to metabolic perturbance. Our study modeled and revealed a direct intracellular connection between compromised PKP2 function and metabolic impairment. We proposed that an increased risk of decoupling energy-responsive contractility from less energy-responsive electrical activities can be a new arrhythmogenic mechanism, potentially responsible for exercise-triggered cardiac adversity in ARVC disease development and progression. ### Competing Interest Statement The authors have declared no competing interest.

The I-bar protein MTSS1 has been implicated in heart failure and contractility by multiple genome-wide association studies. Human genetic analyses suggested that a variant lowering cardiac MTSS1 expression was associated with significantly improved survival in individuals with TTN dilated cardiomyopathy (DCM). Experimental knockdown of MTSS1 via small interfering RNA (siRNA) in induced pluripotent stem cell–derived cardiomyocytes deficient in TTN, CSRP3, or RBM20 led to improved increased sarcomere number and enhanced contractility. Engineered heart tissue models confirmed increased twitch force following MTSS1 siRNA knockdown across these genetic forms of DCM. Unbiased mass-spectrometry suggests that MTSS1 was found to interact with MYO18A, a protein critical for sarcomere assembly, and siRNA knockdown of MTSS1 up-regulated MYH7 and other sarcomere-related genes. These findings may suggest that MTSS1 impacts contractility as a negative regulator of sarcomere formation or turnover, and that reduction of MTSS1 may be a therapeutic target in select forms of genetic DCM.

Human genetics is an important tool for identifying genes as potential drug targets, and the extensive genetic study of cardiovascular disease provides an opportunity to leverage genetics to match specific patient populations to specific drug targets to improve prioritization of patient selection for clinical studies. We selected well described genetic variants in the region of PCSK9 (rs11591147 and rs562556), ADRB1 (rs7076938), ACE (rs4968782 and rs4363), GLP1R (rs10305492) and ABCC8 (rs757110) for use as proxies for the effects of drugs. Time-to-event analyses were utilized to evaluate their effects on atrial fibrillation (AF) and heart failure (HF) death and/or re-hospitalization using real-world longitudinal dataset. To mitigate the effect of confounding factors for cardiovascular (CV) outcomes, we employed propensity score matching. After matching, a genetic proxy for PCSK9 inhibition (rs11591147) improved survival from CV death/heart transplant in individuals following a diagnosis of ischemic heart disease (Hazard Ratio (HR) 0.78, P = 0.03). A genetic proxy for beta-blockade (rs7076938) improved freedom from rehospitalization or death in individuals with AF (HR 0.92, P = 0.001), and a genetic proxy of ACE inhibition (rs7076938) improved freedom from rehospitalization for HF or death (HR 0.8, P = 0.017) and AF (HR 0.85, P = 0.0014). A protective variant in GLP1R (rs10305492) showed decreased risk of developing HF or CV death after diagnosis of ischemic heart disease (HR = 0.82, P = 0.031) and a protective variant in ABCC8 (rs757110) showed decreased risk of CV mortality since ischemic disease diagnosis (HR = 0.88, P = 0.04) and decreased risk of AF in diabetic patients with ischemic heart disease (HR = 0.68, P = 0.001). Notably, despite smaller cohort sizes after matching, we often observed numerically smaller HRs and reduced P, indicating more pronounced effects and increased statistical association. However, not all genetic proxies replicated known treatment effects. Genetic proxies for well-known drugs corroborate findings from clinical trials in cardiovascular disease. Our results demonstrate a useful analytical approach that leverages genetic evidence from a large cohort with longitudinal outcomes data to effectively select patient populations where specific drug targets may be most effective.

Background Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a familial cardiac disease associated with ventricular arrhythmias and an increased risk of sudden cardiac death. Currently, there are no approved treatments that address the underlying genetic cause of this disease, representing a significant unmet need. Mutations in Plakophilin-2 (PKP2 ), encoding a desmosomal protein, account for approximately 40% of ARVC cases and result in reduced gene expression. Methods Our goal is to examine the feasibility and the efficacy of adeno-associated virus 9 (AAV9)-mediated restoration of PKP2 expression in a cardiac specific knock-out mouse model of Pkp2 . Results We show that a single dose of AAV9:PKP2 gene delivery prevents disease development before the onset of cardiomyopathy and attenuates disease progression after overt cardiomyopathy. Restoration of PKP2 expression leads to a significant extension of lifespan by restoring cellular structures of desmosomes and gap junctions, preventing or halting decline in left ventricular ejection fraction, preventing or reversing dilation of the right ventricle, ameliorating ventricular arrhythmia event frequency and severity, and preventing adverse fibrotic remodeling. RNA sequencing analyses show that restoration of PKP2 expression leads to highly coordinated and durable correction of PKP2 -associated transcriptional networks beyond desmosomes, revealing a broad spectrum of biological perturbances behind ARVC disease etiology. Conclusions We identify fundamental mechanisms of PKP2-associated ARVC beyond disruption of desmosome function. The observed PKP2 dose-function relationship indicates that cardiac-selective AAV9:PKP2 gene therapy may be a promising therapeutic approach to treat ARVC patients with PKP2 mutations.

Background Hypertrophic cardiomyopathy (HCM) affects an estimated 600,000 people in the U.S. and is the leading cause of sudden cardiac arrest in those under 18. Loss-of-function mutations in Myosin Binding Protein C3 , MYBPC3 , are the most common genetic cause of HCM. The majority of MYBPC3 mutations causative for HCM result in truncations. The sarcomeric pathophysiology of the majority of HCM patients with MYBPC3 mutations appears to be due to haploinsufficiency, as the total amount of MYBPC3 protein incorporated into sarcomeres falls significantly below normal.Methods A clear path for the treatment of haploinsufficiency is the restoration of the insufficient gene product; in this case wild-type MYBPC3. To achieve this, we engineered an AAV vector (TN-201) with superior properties for mediating cardiomyocyte-selective expression of MYBPC3 after systemic delivery.Results We have demonstrated for the first time with AAV gene therapy the ability of both a mouse surrogate and TN-201, which encodes human MYBPC3 to reverse cardiac hypertrophy and systolic dysfunction and to improve diastolic dysfunction and survival in a symptomatic MYBPC3-deficient murine model of disease. Dose-ranging efficacy studies exhibited restoration of wild-type MYBPC3 protein levels and saturation of cardiac improvement at the clinically relevant dose of 3E13 vg/kg, outperforming a previously published construct. Further, we have established stable cardiac benefit for greater than one year post-injection, as well as reversal of cardiac dysfunction even in late-stage models of disease.Conclusions Our data suggest that by restoring MYBPC3 to the sarcomere, TN-201 has the potential to slow and even reverse the course of the disease in patients with MYBPC3 -associated HCM.### Competing Interest StatementThe authors have declared no competing interest.

Introduction TN-301 is a highly selective, orally available histone deacetylase 6 (HDAC6) inhibitor being developed for the potential treatment of heart failure with preserved ejection fraction (HFpEF). Studies in preclinical models show reversal of HFpEF disease by selective HDAC6 inhibition. This First-in-Human (FiH) Phase 1 clinical trial evaluates the safety and biological activity of TN-301 at a broad range of single doses and multiple daily doses over 2 weeks in healthy adult participants. Hypothesis Multiple preclinical models of HFpEF suggest that selective HDAC6 inhibition has direct and systemic effects on multiple pathways linked to HFpEF pathogenesis, including mitochondrial dysfunction, fibrosis and inflammation, while avoiding undesirable effects of non-selective HDAC inhibitors. This FiH study is designed to identify a dose range for further development and to demonstrate potential clinical utility through use of relevant biomarkers. Methods The double-blinded, randomized clinical trial plans to enroll 72 participants in 6 single-ascending dose (SAD) and 3 multiple-ascending dose (MAD) cohorts, each comprised of 8 participants (6 active, 2 placebo). MAD cohorts are dosed once daily for 14 consecutive days. Endpoints include safety, tolerability, pharmacokinetic (PK) and pharmacodynamic (PD) assessments. A key PD biomarker of HDAC6 inhibition is the level of acetylated tubulin in PMBCs. Results Comprehensive synthesis of unblinded safety, tolerability, PK, and PD results, including additional planned cohorts, are anticipated by the time of presentation. As of the submission of this abstract, 48 participants have been randomized into the SAD stage and received TN-301 or placebo per protocol at oral doses from 1-700mg. An additional 16 participants have been treated in 2 MAD cohorts at 25mg and 100mg. Among these participants, there were no SAEs, DLTs or premature withdrawals due to tolerability. Interim analysis showed a slightly more than dose-proportional PK profile with a half-life consistent with once-daily dosing. Plasma concentrations were attained that corresponded with those observed in mouse models that demonstrated robust PD affects and reversal of HFpEF phenotype. Target engagement was demonstrated at doses >5mg and was sustained above baseline throughout the dosing interval in the MAD cohorts at steady state. Conclusion Initial data from this FiH clinical trial of TN-301 in healthy adult participants demonstrated clear evidence of target engagement. To date, TN-301 has been generally well tolerated with PK consistent with once-daily dosing. TN-301 represents a promising new therapeutic candidate for the potential treatment of HFpEF and other indications.

Background: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a genetic cardiac disorder characterized by severe arrhythmias and heart dysfunction. Mutations in desmosome gene Plakophilin-2 ( PKP2 ) account for 40% of ARVC cases, with current palliative therapies failing to address the genetic cause. Herein, we generated an in vitro ARVC model using human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) harboring a pathogenic PKP2 variant (c.2146G>C), as a platform to test a PKP2 -gene replacement approach developed at Tenaya Therapeutics for the treatment of PKP2 -associated ARVC patients. Methods: Three isogenic human iPSC lines (wild-type, heterozygous and homozygous PKP2 mutant) were differentiated to iPSC-CM; and monolayers and engineered heart tissues (EHTs) were generated. Gene and protein expression were assessed by RNA-Seq/RT-qPCR, and immunocytochemistry, respectively. Contractility, electrophysiology and calcium handling were measured. A proprietary adeno-associated virus gene therapy (AAV9: PKP2 ) was used to transduce human iPSC-CM, and changes in gene and protein expression, and contractile function were evaluated. Results: A human iPSC-CM beating monolayer was achieved for all three lines. PKP2 expression was depleted in a genotype-dependent manner, and desmosome structure was disrupted in the mutant lines. Functional assessment of human iPSC-CM monolayers showed impaired contractile properties and abnormal electrophysiological and calcium transients in the mutant lines, such as decreased contraction amplitude and prolonged field and action potential duration. Transcriptional analysis revealed changes in desmosome, gap junctions, sarcomere, ion channels, metabolic and apoptosis gene expression. Characterization of EHTs displayed a deficit in contractility, slower action and field potential kinetics and irregular calcium homeostasis in the mutant lines. The administration of AAV9: PKP2 to the mutant lines restored ion channel and desmosome gene and protein expression, and contractile function. Conclusions: Our PKP2 human iPSC-CM disease model recapitulated the main hallmarks of ARVC phenotype. Administration of AAV9: PKP2 restored desmosome protein expression and contractility. This model lays a foundation for the understanding of the underlying molecular pathophysiological mechanisms of PKP2 -ARVC, and the potential for AAV9: PKP2 as a one-time dose to correct the genetic cause of disease in individuals with PKP2 -associated ARVC.

Background HFpEF is a form of heart failure characterized by diastolic dysfunction and associated with high morbidity, mortality and significant unmet need. Previously, we have demonstrated that selective inhibition of histone deacetylase 6 (HDAC6) has positive effects on diastolic dysfunction and left ventricular thickness of the heart, as well as overall improvements in systemic inflammation and metabolism, in a mouse model of HFpEF using a high-fat diet (HFD) and L-NAME. Empagliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor approved by the FDA for HFpEF patients, works as expected in this model, confirming the potential clinical translatability of results seen with HDAC6 inhibitors. In head-to-head studies, the beneficial effects of HDAC6 inhibition have been shown to be comparable to empagliflozin, while demonstrating a distinct mechanism of action in gene expression analysis. TN-301, a highly selective HDAC6 inhibitor has been advanced into clinical development for the potential treatment of HFpEF. Purpose In this study, we sought to investigate the additive or synergistic effects of combining HDAC6 and SGLT2 inhibition to improve cardiac function in a two-hit mouse model of HFpEF. Methods and Results Low doses of TYA-018 (a highly selective HDAC6 inhibitor) and empagliflozin were co-administered in our HFpEF model. We then assessed functional measures and cardiac gene expression in hearts from treated and control HFpEF mice. In our mouse model of HFpEF, pharmacological inhibition of TYA-018 results in extensive cardiac functional and structural improvements, including diastolic dysfunction and LV mass. Co-administration of TYA-018 and empagliflozin resulted in additive cardiac functional and structural measures vs. the single agents. Of note, multiple measures of diastolic dysfunction (e.g., E/e’) were returned to baseline values by combination treatment in HFpEF mice similar to those in WT mice. Gene expression analysis is being conducted using RNA-seq to elucidate the potential mechanisms underlying the efficacy of HDAC6 and SGLT2 inhibition by characterizing the pathway-level modulation by combination treatment compared to each therapy alone. Results of gene expression analysis comparing single-agent and combination activity are planned for inclusion at the time of presentation. Conclusion These studies demonstrate an additive benefit on diastolic dysfunction and elucidate key molecular mechanisms supporting the rationale for the potential use of HDAC6 inhibition as a single agent or in combination with SGLT2 inhibition for the treatment of HFpEF. HFpEF is a form of heart failure characterized by diastolic dysfunction and associated with high morbidity, mortality and significant unmet need. Previously, we have demonstrated that selective inhibition of histone deacetylase 6 (HDAC6) has positive effects on diastolic dysfunction and left ventricular thickness of the heart, as well as overall improvements in systemic inflammation and metabolism, in a mouse model of HFpEF using a high-fat diet (HFD) and L-NAME. Empagliflozin, a sodium-glucose cotransporter 2 (SGLT2) inhibitor approved by the FDA for HFpEF patients, works as expected in this model, confirming the potential clinical translatability of results seen with HDAC6 inhibitors. In head-to-head studies, the beneficial effects of HDAC6 inhibition have been shown to be comparable to empagliflozin, while demonstrating a distinct mechanism of action in gene expression analysis. TN-301, a highly selective HDAC6 inhibitor has been advanced into clinical development for the potential treatment of HFpEF. In this study, we sought to investigate the additive or synergistic effects of combining HDAC6 and SGLT2 inhibition to improve cardiac function in a two-hit mouse model of HFpEF. Low doses of TYA-018 (a highly selective HDAC6 inhibitor) and empagliflozin were co-administered in our HFpEF model. We then assessed functional measures and cardiac gene expression in hearts from treated and control HFpEF mice. In our mouse model of HFpEF, pharmacological inhibition of TYA-018 results in extensive cardiac functional and structural improvements, including diastolic dysfunction and LV mass. Co-administration of TYA-018 and empagliflozin resulted in additive cardiac functional and structural measures vs. the single agents. Of note, multiple measures of diastolic dysfunction (e.g., E/e’) were returned to baseline values by combination treatment in HFpEF mice similar to those in WT mice. Gene expression analysis is being conducted using RNA-seq to elucidate the potential mechanisms underlying the efficacy of HDAC6 and SGLT2 inhibition by characterizing the pathway-level modulation by combination treatment compared to each therapy alone. Results of gene expression analysis comparing single-agent and combination activity are planned for inclusion at the time of presentation. These studies demonstrate an additive benefit on diastolic dysfunction and elucidate key molecular mechanisms supporting the rationale for the potential use of HDAC6 inhibition as a single agent or in combination with SGLT2 inhibition for the treatment of HFpEF.

Heart failure with preserved ejection fraction (HFpEF) poses therapeutic challenges due to the limited treatment options. Building upon our previous research that demonstrates the efficacy of histone deacetylase 6 (HDAC6) inhibition in a genetic cardiomyopathy model, we investigate HDAC6’s role in HFpEF due to their shared mechanisms of inflammation and metabolism. Here, we show that inhibiting HDAC6 with TYA-018 effectively reverses established heart failure and its associated symptoms in male HFpEF mouse models. Additionally, in male mice lacking Hdac6 gene, HFpEF progression is delayed and they are resistant to TYA-018’s effects. The efficacy of TYA-018 is comparable to a sodium-glucose cotransporter 2 (SGLT2) inhibitor, and the combination shows enhanced effects. Mechanistically, TYA-018 restores gene expression related to hypertrophy, fibrosis, and mitochondrial energy production in HFpEF heart tissues. Furthermore, TYA-018 also inhibits activation of human cardiac fibroblasts and enhances mitochondrial respiratory capacity in cardiomyocytes. In this work, our findings show that HDAC6 impacts on heart pathophysiology and is a promising target for HFpEF treatment.

Cardiovascular disease is a leading cause of death and continues to require novel forms of therapies. Mutations in sarcomeres, the basic contractile units of cardiac muscle, causing disarray and loss of function are causal in many genetic cardiomyopathies. Here, we model sarcomeric disarray in vitro by culturing human induced pluripotent stem cell-derived cardiomyocytes (human iPSC-CMs) on tissue culture plates with biomimetic patterned grooves designed to simulate extracellular matrix. This culture system models the natural structure and phenotypic development of human physiology. Next, we created an Adeno-associated virus open reading frame (AAV-ORF) library of ~100 heart failure associated genes of interest (GOI) identified using publicly available human genetic data. We developed a high throughput screening (HTS) process to test the effects of overexpression of these GOIs on sarcomere morphology. Images of human iPSC-CMs treated with overexpression libraries were run through a custom MATLAB HTS image processing script named ‘Tamarack’, developed for quantifying human iPSC-CM sarcomere morphology. Tamarack enables quantification of sarcomere count, length, and orientation. Analysis of sarcomeres from the Tamarack code in the biomimetic plates allows for enhanced quantification of sarcomere structures, in particular, alignment, a metric missing from standard 2D culture plates. We found that treating cells with siRBM20 induced a sarcomere misalignment phenotype which we looked to rescue with overexpression of our GOI. Genes that increased alignment in an RBM20 background were considered top hits and moved to further analysis.

Supplementary Figure 4 from Anti-DLL4 Inhibits Growth and Reduces Tumor-Initiating Cell Frequency in Colorectal Tumors with Oncogenic KRAS Mutations