Ana Kojic

All Publications

• Generation of two iPSC lines from ALS patients harboring C9orf72 hexanucleotide repeat expansions. Stem cell research Wu, D., Kojic, A., Ross, J. P., Li, D., Walther, P., Goyal, N., Sampson, J., Wu, J. C. 2026; 95: 104085

  Abstract

  The GGGGCC hexanucleotide repeat expansion (HRE) within the C9orf72 gene constitutes the leading genetic driver of amyotrophic lateral sclerosis (ALS). This fatal neurodegenerative disorder is characterized by the systematic loss of both the upper and lower motor neurons across both the central and peripheral nervous systems. This work describes the successful reprogramming of two human induced pluripotent stem cell (iPSC) lines originating from two independent ALS patients, both of whom carry a C9orf72 HRE mutation. Validation of the two established iPSC lines confirmed the expression of pluripotency markers, normal karyotypes, and successful trilineage differentiation. Consequently, these lines provide a robust in vitro platform to model ALS and study C9orf72-mediated disease mechanisms.

  View details for DOI 10.1016/j.scr.2026.104085

  View details for PubMedID 42648119

• Beta-Adrenergic Stimulation and MYH7 G256E Mutant Gene Dosage Drive Hypertrophic Cardiomyopathy Phenotype Penetrance. bioRxiv : the preprint server for biology Heinrich, P., Jung, R. M., Achter, J. S., Nguyen, V. X., Lee, C. A., Sailer, C., Domian, H., Vander Roest, A., Suchy, F. P., Jahng, J. W., Kojic, A., Lee, D., Paasche, A., Roberts, B., Nakauchi, H., Zhu, H., Wu, J. C., Bernstein, D., Moretti, A., Lundby, A., Lee, S., Wu, S. M. 2026

  Abstract

  Hypertrophic cardiomyopathy (HCM) is the most prevalent genetic heart disorder, characterized by significant phenotypic variability even among individuals with identical MYH7 mutations. This study aims to elucidate factors contributing to this variability and identify drivers of phenotype penetrance. We compared the baseline phenotypes of a highly penetrant MYH7 H251N mutation and the variably penetrant MYH7 G256E mutation and investigated the impact of adding beta-adrenergic stimulation and homozygosity on disease phenotype penetrance using cardiomyocytes from an isogenic line of human induced pluripotent stem cells (hiPSC-CMs).Isogenic hiPSCs with MYH7 H251N and MYH7 G256E mutations were generated using CRISPR/Cas9 technology and differentiated into cardiomyocytes (CMs). Single-cell RNA sequencing (scRNAseq) and functional analysis of contractile function revealed consistent HCM phenotype presentation in H251N CMs, whereas G256E CMs exhibited a subtle and more variable phenotype. Beta-adrenergic stimulation induced a distinct metabolic stress response in G256E CMs, characterized by impaired mitochondrial ATP upregulation. Increasing mutant gene dosage from hetero- to homozygosity led to consistent increase in hypertrophic and structural gene expression changes in G256E CMs at RNA and protein levels. These changes were distinct from the changes observed with stress response. Importantly, homozygous G256E CMs exhibited a hypercontractile functional and disorganized structural phenotype. Across multiple experimental conditions, we identified consistent increase in cardiomyocyte specific transcriptomic markers such as NPPB, APOE, PDLIM3 and ANKRD1.Our study highlights the use of a variably penetrant MYH7 mutation to investigate factors that influence HCM phenotype penetrance. Specifically, we found that mutant gene dosage and beta-adrenergic stimulation induce distinct HCM disease phenotypes, providing novel insights into mechanisms that may contribute to variable disease expression in HCM.

  View details for DOI 10.64898/2026.06.02.729411

  View details for PubMedID 42282692

  View details for PubMedCentralID PMC13252407

• Unbiased Proteome of Human Myocardium and Serum Sampled at Sudden Death for Discovery of Biomarkers for Near-term Risk of Lethal Arrhythmia Caudal, A., Belbachir, N., Kojic, A., Kinkead, B., Tsan, J., Hu, E., Yee, M., Moffatt, E., Connolly, A., Padmanabhan, A., Wu, J., Tseng, Z. LIPPINCOTT WILLIAMS & WILKINS. 2025

  View details for DOI 10.1161/circ.152.suppl_3.4354588

  View details for Web of Science ID 001620173200002

• Cardiometabolic disease and cardio-oncology: Insights from iPSC models and tissue engineering. Cell reports. Medicine Kojic, A., Moslehi, J., Ky, B., Wu, J. C. 2025: 102261

  Abstract

  Heart disease and cancer share common risk factors, genetic predispositions, and metabolic and inflammatory components. Metabolic reprogramming can drive disease progression in both, with cardiometabolic syndrome-marked by obesity, insulin resistance, dyslipidemia, and hypertension-contributing to cancer development. Studies link around 20% of cancer cases to obesity, while elevated glucose and triglyceride levels increase the risk of liver, thyroid, and respiratory cancers. Beyond treatment-related cardiotoxicity, cancer patients often have pre-existing cardiovascular disease (CVD) at diagnosis, highlighting their bidirectional relationship. Patient-specific induced pluripotent stem cells (iPSCs) offer a powerful platform to study these links at a personalized level. iPSC models help explore shared molecular mechanisms, metabolic dysregulation, inflammation, and cardiotoxicity. This review examines emerging themes in cardio-oncology and cardio-metabolism, emphasizing how iPSC-based approaches can reveal disease connections and inform new therapies.

  View details for DOI 10.1016/j.xcrm.2025.102261

  View details for PubMedID 40752492

• Selective inhibition of stromal mechanosensing suppresses cardiac fibrosis. Nature Cho, S., Rhee, S., Madl, C. M., Caudal, A., Thomas, D., Kim, H., Kojic, A., Shin, H. S., Mahajan, A., Jahng, J. W., Wang, X., Thai, P. N., Paik, D. T., Wang, M., Mullen, M., Baker, N. M., Leitz, J., Mukherjee, S., Winn, V. D., Woo, Y. J., Blau, H. M., Wu, J. C. 2025

  Abstract

  Matrix-derived biophysical cues are known to regulate the activation of fibroblasts and their subsequent transdifferentiation into myofibroblasts1-6, but whether modulation of these signals can suppress fibrosis in intact tissues remains unclear, particularly in the cardiovascular system7-10. Here we demonstrate across multiple scales that inhibition of matrix mechanosensing in persistently activated cardiac fibroblasts potentiates-in concert with soluble regulators of the TGFβ pathway-a robust transcriptomic, morphological and metabolic shift towards quiescence. By conducting a meta-analysis of public human and mouse single-cell sequencing datasets, we identify the focal-adhesion-associated tyrosine kinase SRC as a fibroblast-enriched mechanosensor that can be targeted selectively in stromal cells to mimic the effects of matrix softening in vivo. Pharmacological inhibition of SRC by saracatinib, coupled with TGFβ suppression, induces synergistic repression of key profibrotic gene programs in fibroblasts, characterized by a marked inhibition of the MRTF-SRF pathway, which is not seen after treatment with either drug alone. Importantly, the dual treatment alleviates contractile dysfunction in fibrotic engineered heart tissues and in a mouse model of heart failure. Our findings point to joint inhibition of SRC-mediated stromal mechanosensing and TGFβ signalling as a potential mechanotherapeutic strategy for treating cardiovascular fibrosis.

  View details for DOI 10.1038/s41586-025-08945-9

  View details for PubMedID 40307543

  View details for PubMedCentralID 5749528

• Trifunctional lipid derivatives: PE's mitochondrial interactome CHEMICAL COMMUNICATIONS Thomas, A., Mueller, R., Farley, S., Kojic, A., Stein, F., Haberkant, P., Schultz, C. 2025; 61 (12): 2564-2567

  Abstract

  Phosphatidylethanolamine (PE) is a ubiquitous lipid species in higher eukaryotes. Here, we synthesized a multifunctionalized PE derivative (1) designed to identify PE-binding proteins in intact cells through photo-crosslinking and subsequent isolation and proteomic analysis of the PE-protein conjugates. We show that the tool is also useful for tracking PE translocation to mitochondria after uncaging. A trifunctional phosphatidic acid derivative was rapidly metabolized and is of more limited use.

  View details for DOI 10.1039/d4cc03599b

  View details for Web of Science ID 001396852000001

  View details for PubMedID 39817617

• Generation of two induced pluripotent stem cell lines from healthy patients of African American ancestry. Stem cell research Mullen, M., Kojic, A., Alamana, C., Canel, G., Lai, C., Knowles, J. W., Wu, J. C. 2024; 76: 103322

  Abstract

  Stem cells are a resourceful tool for investigating cardiovascular disease in the context of race and gender. Once derived from blood or skin cells, the reprogrammed induced pluripotent stem cells (iPSCs) adopt an embryonic-like pluripotent state, enabling researchers to develop drug screening or disease modeling platforms. Here, we generated two iPSC lines from peripheral blood mononuclear cells (PBMCs) of two healthy African American patients. Both lines display the usual morphology of pluripotent stem cells, demonstrate elevated expression of pluripotent markers, show normal karyotype, and differentiate into all three germ layers in vitro.

  View details for DOI 10.1016/j.scr.2024.103322

  View details for PubMedID 38359472

• Structurally distinct PARP7 inhibitors provide new insights into the function of PARP7 in regulating nucleic acid-sensing and IFN-(3 signaling CELL CHEMICAL BIOLOGY Sanderson, D. J., Rodriguez, K. M., Bejan, D. S., Olafsen, N. E., Bohn, I. D., Kojic, A., Sundalam, S., Siordia, I. R., Duell, A. K., Deng, N., Schultz, C., Grant, D. M., Matthews, J., Cohen, M. S. 2023; 30 (1): 43-+

  Abstract

  The mono-ADP-ribosyltransferase PARP7 has emerged as a key negative regulator of cytosolic NA-sensors of the innate immune system. We apply a rational design strategy for converting a pan-PARP inhibitor into a potent selective PARP7 inhibitor (KMR-206). Consistent with studies using the structurally distinct PARP7 inhibitor RBN-2397, co-treatment of mouse embryonic fibroblasts with KMR-206 and NA-sensor ligands synergistically induced the expression of the type I interferon, IFN-β. In mouse colon carcinoma (CT-26) cells, KMR-206 alone induced IFN-β. Both KMR-206 and RBN-2397 increased PARP7 protein levels in CT-26 cells, demonstrating that PARP7's catalytic activity regulates its own protein levels. Curiously, treatment with saturating doses of KMR-206 and RBN-2397 achieved different levels of PARP7 protein, which correlated with the magnitude of type I interferon gene expression. These latter results have important implications for the mechanism of action of PARP7 inhibitors and highlights the usefulness of having structurally distinct chemical probes for the same target.

  View details for DOI 10.1016/j.chembiol.2022.11.012

  View details for Web of Science ID 000994359000001

  View details for PubMedID 36529140

  View details for PubMedCentralID PMC9868104

• Generation of two induced pluripotent stem cell lines from dilated cardiomyopathy patients caused by heterozygous mutations in the HCN4 gene. Stem cell research Yildirim, Z., Kojic, A., Yan, C. D., Wu, M. A., Vagelos, R., Wu, J. C. 2022; 65: 102951

  Abstract

  Dilated cardiomyopathy (DCM) is a progressive heart muscle disease that can culminate with heart failure and death. Mutations in several genes can cause DCM, including hyperpolarization-activated cyclic nucleotide-gated channel (HCN4), which has a critical function in the autonomic control of the heart rate. Here, we generated two human induced pluripotent stem cell (iPSC) lines generated from two DCM patients carrying variants in the HCN4 gene (c.2587G > T and c.2846G > A). Both lines display normal karyotype, typical morphology of pluripotent stem cells, and differentiate into all three germ layers in vitro. These lines are valuable resources for studying the pathological mechanisms of DCM.

  View details for DOI 10.1016/j.scr.2022.102951

  View details for PubMedID 36332467

• Generation of two induced pluripotent stem cell lines from dilated cardiomyopathy patients carrying heterozygous FLNC mutations. Stem cell research Kojic, A., Kim, H., Guevara, J. V., Ravada, S., Sallam, K., Wu, J. C. 2022; 64: 102928

  Abstract

  Dilated cardiomyopathy (DCM) is a heterogeneous cardiac disorder characterized by left ventricular dilatation and dysfunction. Mutations in dozens of cardiac genes have been connected to the development of DCM including the filamin C gene (FLNC). We generated two induced pluripotent stem cell (iPSCs) lines from DCM patients carrying single missense heterozygote FLNC mutations (c.6689G > A and c.3745G > A). Both lines expressed high levels of pluripotency markers, differentiated into derivatives of the three germ layers and possessed normal karyotypes. The derived iPSC lines can serve as powerful tools to model DCM in vitro and as a platform for therapeutic development.

  View details for DOI 10.1016/j.scr.2022.102928

  View details for PubMedID 36194907

• Synthesis and Cellular Labeling of Multifunctional Phosphatidylinositol Bis- and Trisphosphate Derivatives ANGEWANDTE CHEMIE-INTERNATIONAL EDITION Mueller, R., Kojic, A., Citir, M., Schultz, C. 2021; 60 (36): 19759-19765

  Abstract

  We synthesized the first multifunctionalized phosphoinositide polyphosphate derivatives featuring a photo-removable protecting group ("cage"), a photo-crosslinkable diazirine group, and a terminal alkyne group useful for click chemistry. We demonstrate that the lipid derivatives readily enter cells. After photo-crosslinking, cell fixation and fluorescent tagging via click chemistry, we determined the intracellular location of the lipid derivatives before and after uncaging of the lipids. We find that there is rapid trafficking of PI(3,4)P2 and PI(3,4,5)P3 derivatives to the plasma membrane, opening the intriguing possibility that there is active transport of these lipids involved. We employed the photo-crosslinking and click chemistry functions to analyze the proteome of PI(3,4,5)P3 -binding proteins. From the latter, we validated by RNAi that the putative lipid binding proteins ATP11A and MPP6 are involved in the transport of PI(3,4,5)P3 to the plasma membrane.

  View details for DOI 10.1002/anie.202103599

  View details for Web of Science ID 000678863300001

  View details for PubMedID 34075669

  View details for PubMedCentralID PMC8390440

• amTCO, a new <i>trans</i>-cyclooctene derivative to study drug-target interactions in cells CHEMICAL COMMUNICATIONS Echalier, C., Rutkowska, A., Kojic, A., Thomson, D. W., Edwards, L. J., McKay, B. S. J., Muelbaier, M., Schultz, C., Bergamini, G. 2021; 57 (14): 1814-1817

  Abstract

  Click chemistry probes have improved the study of drug interactions in live cells and relevant disease models. Proper design of the probes, including the choice of the click moiety coupled to the drug, is crucial to ensure good performance and broad application. A new trans-cyclooctene derivative, amTCO, was synthesised via a novel route using a phthalimide protecting group as a built-in photosensitiser for the cyclooctene isomerization. amTCO improved the physical chemical properties of click chemistry probes compared to standard TCO moieties. An amTCO probe targeting indoleamine 2,3-dioxygenase (IDO1) was a superior tool for visualizing IDO1 and measuring the binding affinities of small molecule inhibitors to IDO1 in cells.

  View details for DOI 10.1039/d0cc06709a

  View details for Web of Science ID 000619625700020

  View details for PubMedID 33480895

• Cloning and expression profiling of muscle regulator ANKRD2 in domestic chicken<i>Gallus gallus</i> HISTOCHEMISTRY AND CELL BIOLOGY Stamenkovic, N., Jasnic, J., Novkovic, M., Milosevic, E., Boskovic, S., Kojic, A., Popic, K., Stankovic, M., Wang, Y., Milenkovic, S., Radojkovic, D., Ma, G., Kojic, S. 2020; 154 (4): 383-396

  Abstract

  Striated muscle signaling protein and transcriptional regulator ANKRD2 participates in myogenesis, myogenic differentiation, muscle adaptation and stress response. It is preferentially expressed in slow, oxidative fibers of mammalian skeletal muscle. In this study, we report on characterization of chicken ANKRD2. The chicken ANKRD2 coding region contains 1002 bp and encodes a 334-amino acid protein which shares approximately 58% identity with human and mouse orthologs, mostly in the conserved region of ankyrin repeats. Comprehensive analysis of the ANKRD2 gene and protein expression in adult chicken demonstrated its predominant expression in red muscles of thigh and drumstick, compared to white muscle. It was not detected in heart and white pectoral muscle. Uneven expression of ANKRD2 in chicken skeletal muscles, observed by immunohistochemistry, was attributed to its selective expression in slow, oxidative, type I and fast, oxidative-glycolytic, type IIA myofibers. Association of chicken ANKRD2 with phenotypic differences between red and white muscles points to its potential role in the process of myofiber-type specification. In addition to expression in slow oxidative myofibers, as demonstrated for mammalian protein, chicken ANKRD2 was also detected in fast fibers with mixed oxidative and glycolytic metabolism. This finding suggests that ANKRD2 is responsive to metabolic differences between types of avian myofibers and orientates future studies towards investigation of its role in molecular mechanisms of myofiber-type-specific gene expression.

  View details for DOI 10.1007/s00418-020-01899-1

  View details for Web of Science ID 000547349500002

  View details for PubMedID 32653935

  View details for PubMedCentralID 1197752

• A Potent and Selective PARP11 Inhibitor Suggests Coupling between Cellular Localization and Catalytic Activity CELL CHEMICAL BIOLOGY Kirby, I. T., Kojic, A., Arnold, M. R., Thorsell, A., Karlberg, T., Vermehren-Schmaedick, A., Sreenivasan, R., Schultz, C., Schuler, H., Cohen, M. S. 2018; 25 (12): 1547-+

  Abstract

  Poly-ADP-ribose polymerases (PARPs1-16) play pivotal roles in diverse cellular processes. PARPs that catalyze poly-ADP-ribosylation (PARylation) are the best characterized PARP family members because of the availability of potent and selective inhibitors for these PARPs. There has been comparatively little success in developing selective small-molecule inhibitors of PARPs that catalyze mono-ADP-ribosylation (MARylation), limiting our understanding of the cellular role of MARylation. Here we describe the structure-guided design of inhibitors of PARPs that catalyze MARylation. The most selective analog, ITK7, potently inhibits the MARylation activity of PARP11, a nuclear envelope-localized PARP. ITK7 is greater than 200-fold selective over other PARP family members. Using live-cell imaging, we show that ITK7 causes PARP11 to dissociate from the nuclear envelope. These results suggest that the cellular localization of PARP11 is regulated by its catalytic activity.

  View details for DOI 10.1016/j.chembiol.2018.09.011

  View details for Web of Science ID 000454180900012

  View details for PubMedID 30344052

• Characterization of zebrafish (<i>Danio rerio</i>) muscle ankyrin repeat proteins reveals their conserved response to endurance exercise PLOS ONE Boskovic, S., Marin-Juez, R., Jasnic, J., Reischauer, S., El Sammak, H., Kojic, A., Faulkner, G., Radojkovic, D., Stainier, D. Y. R., Kojic, S. 2018; 13 (9): e0204312

  Abstract

  Muscle proteins with ankyrin repeats (MARPs) ANKRD1 and ANKRD2 are titin-associated proteins with a putative role as transcriptional co-regulators in striated muscle, involved in the cellular response to mechanical, oxidative and metabolic stress. Since many aspects of the biology of MARPs, particularly exact mechanisms of their action, in striated muscle are still elusive, research in this field will benefit from novel animal model system. Here we investigated the MARPs found in zebrafish for protein structure, evolutionary conservation, spatiotemporal expression profiles and response to increased muscle activity. Ankrd1 and Ankrd2 show overall moderate conservation at the protein level, more pronounced in the region of ankyrin repeats, motifs indispensable for their function. The two zebrafish genes, ankrd1a and ankrd1b, counterparts of mammalian ANKRD1/Ankrd1, have different expression profiles during first seven days of development. Mild increase of ankrd1a transcript levels was detected at 72 hpf (1.74±0.24 fold increase relative to 24 hpf time point), while ankrd1b expression was markedly upregulated from 24 hpf onward and peaked at 72 hpf (92.18±36.95 fold increase relative to 24 hpf time point). Spatially, they exhibited non-overlapping expression patterns during skeletal muscle development in trunk (ankrd1a) and tail (ankrd1b) somites. Expression of ankrd2 was barely detectable. Zebrafish MARPs, expressed at a relatively low level in adult striated muscle, were found to be responsive to endurance exercise training consisting of two bouts of 3 hours of forced swimming daily, for five consecutive days. Three hours after the last exercise bout, ankrd1a expression increased in cardiac muscle (6.19±5.05 fold change), while ankrd1b and ankrd2 were upregulated in skeletal muscle (1.97±1.05 and 1.84±0.58 fold change, respectively). This study provides the foundation to establish zebrafish as a novel in vivo model for further investigation of MARPs function in striated muscle.

  View details for DOI 10.1371/journal.pone.0204312

  View details for Web of Science ID 000445639700041

  View details for PubMedID 30252882

  View details for PubMedCentralID PMC6155536

• Differential expression and localization of Ankrd2 isoforms in human skeletal and cardiac muscles HISTOCHEMISTRY AND CELL BIOLOGY Jasnic-Savovic, J., Krause, S., Savic, S., Kojic, A., Kovcic, V., Boskovic, S., Nestorovic, A., Rakicevic, L., Schreiber-Katz, O., Vogel, J. G., Schoser, B. G., Walter, M. C., Valle, G., Radojkovic, D., Faulkner, G., Kojic, S. 2016; 146 (5): 569-584

  Abstract

  Four human Ankrd2 transcripts, reported in the Ensembl database, code for distinct protein isoforms (360, 333, 327 and 300 aa), and so far, their existence, specific expression and localization patterns have not been studied in detail. Ankrd2 is preferentially expressed in the slow fibers of skeletal muscle. It is found in both the nuclei and the cytoplasm of skeletal muscle cells, and its localization is prone to change during differentiation and upon stress. Ankrd2 has also been detected in the heart, in ventricular cardiomyocytes and in the intercalated disks (ICDs). The main objective of this study was to distinguish between the Ankrd2 isoforms and to determine the contribution of each one to the general profile of Ankrd2 expression in striated muscles. We demonstrated that the known expression and localization pattern of Ankrd2 in striated muscle can be attributed to the isoform of 333 aa which is dominant in both tissues, while the designated cardiac and canonical isoform of 360 aa was less expressed in both tissues. The 360 aa isoform has a distinct nuclear localization in human skeletal muscle, as well as in primary myoblasts and myotubes. In contrast to the isoform of 333 aa, it was not preferentially expressed in slow fibers and not localized to the ICDs of human cardiomyocytes. Regulation of the expression of both isoforms is achieved at the transcriptional level. Our results set the stage for investigation of the specific functions and interactions of the Ankrd2 isoforms in healthy and diseased human striated muscles.

  View details for DOI 10.1007/s00418-016-1465-0

  View details for Web of Science ID 000386695800006

  View details for PubMedID 27393496