Ananthakrishnan Ganesan

All Publications

• Multicohort Analysis of Bronchial Epithelial Cell Expression in Healthy Subjects and Patients with Asthma Reveals Four Clinically Distinct Clusters. American journal of respiratory cell and molecular biology Lee, I., Ganesan, A., Kalesinskas, L., Zheng, H., Ahn, H. C., Christenson, S., Erzurum, S. C., Zein, J., Bleecker, E. R., Meyers, D. A., Castro, M., Fahy, J. V., Israel, E., Jarjour, N. N., Moore, W., Wenzel, S. E., Mauger, D. T., Levy, B. D., Woodruff, P. G., Ortega, V. E., Khatri, P. 2024

  Abstract

  Asthma is a heterogeneous disease with variable presentation and characteristics. There is a critical need to identify underlying molecular endotypes of asthma. We performed the largest transcriptomic analysis of 808 bronchial epithelial cell (BEC) samples across 11 independent cohorts, including 3 cohorts from the Severe Asthma Research Program (SARP). Using 7 datasets (218 asthma patients, 148 healthy controls) as discovery cohorts, we identified 505 differentially expressed genes (DEGs), which we validated in the remaining four datasets. Unsupervised clustering using the 505 DEGs identified four reproducible clusters of patients with asthma across all datasets corresponding to healthy controls, mild/moderate asthma, and severe asthma with significant differences in several clinical markers of severity, including pulmonary function, T2 inflammation, FeNO, and max bronchodilator reversibility. Importantly, we found the same clusters in pediatric patients using nasal lavage fluid cells, demonstrating the gene signature and clusters are not confounded by age and conserved in both lower and upper airways. The four asthma clusters may represent a unifying framework for understanding the molecular heterogeneity of asthma. Further study could potentially enable a precision medicine approach of matching therapies with asthma patients most likely to benefit.

  View details for DOI 10.1165/rcmb.2024-0125OC

  View details for PubMedID 39700523

• Multi-modal analysis reveals tumor and immune features distinguishing EBV-positive and EBV-negative post-transplant lymphoproliferative disorders. Cell reports. Medicine Toh, J., Reitsma, A. J., Tajima, T., Younes, S. F., Ezeiruaku, C., Jenkins, K. C., Peña, J. K., Zhao, S., Wang, X., Lee, E. Y., Glass, M. C., Kalesinskas, L., Ganesan, A., Liang, I., Pai, J. A., Harden, J. T., Vallania, F., Vizcarra, E. A., Bhagat, G., Craig, F. E., Swerdlow, S. H., Morscio, J., Dierickx, D., Tousseyn, T., Satpathy, A. T., Krams, S. M., Natkunam, Y., Khatri, P., Martinez, O. M. 2024: 101851

  Abstract

  The oncogenic Epstein-Barr virus (EBV) can drive tumorigenesis with disrupted host immunity, causing malignancies including post-transplant lymphoproliferative disorders (PTLDs). PTLD can also arise in the absence of EBV, but the biological differences underlying EBV(+) and EBV(-) B cell PTLD and the associated host-EBV-tumor interactions remain poorly understood. Here, we reveal the core differences between EBV(+) and EBV(-) PTLD, characterized by increased expression of genes related to immune processes or DNA interactions, respectively, and the augmented ability of EBV(+) PTLD B cells to modulate the tumor microenvironment through elaboration of monocyte-attracting cytokines/chemokines. We create a reference resource of proteins distinguishing EBV(+) B lymphoma cells from EBV(-) B lymphoma including the immunomodulatory molecules CD300a and CD24, respectively. Moreover, we show that CD300a is essential for maximal survival of EBV(+) PTLD B lymphoma cells. Our comprehensive multi-modal analyses uncover the biological underpinnings of PTLD and offer opportunities for precision therapies.

  View details for DOI 10.1016/j.xcrm.2024.101851

  View details for PubMedID 39657667

• International multi-cohort analysis identifies novel framework for quantifying immune dysregulation in critical illness: results of the SUBSPACE consortium. bioRxiv : the preprint server for biology Moore, A. R., Zheng, H., Ganesan, A., Hasin-Brumshtein, Y., Maddali, M. V., Levitt, J. E., van der Poll, T., Scicluna, B. P., Giamarellos-Bourboulis, E. J., Kotsaki, A., Martin-Loeches, I., Garduno, A., Rothman, R. E., Sevransky, J., Wright, D. W., Atreya, M. R., Moldawer, L. L., Efron, P. A., Marcela, K., Karvunidis, T., Giannini, H. M., Meyer, N. J., Sweeney, T. E., Rogers, A. J., Khatri, P. 2024

  Abstract

  Progress in the management of critical care syndromes such as sepsis, Acute Respiratory Distress Syndrome (ARDS), and trauma has slowed over the last two decades, limited by the inherent heterogeneity within syndromic illnesses. Numerous immune endotypes have been proposed in sepsis and critical care, however the overlap of the endotypes is unclear, limiting clinical translation. The SUBSPACE consortium is an international consortium that aims to advance precision medicine through the sharing of transcriptomic data. By evaluating the overlap of existing immune endotypes in sepsis across over 6,000 samples, we developed cell-type specific signatures to quantify dysregulation in these immune compartments. Myeloid and lymphoid dysregulation were associated with disease severity and mortality across all cohorts. This dysregulation was not only observed in sepsis but also in ARDS, trauma, and burn patients, indicating a conserved mechanism across various critical illness syndromes. Moreover, analysis of randomized controlled trial data revealed that myeloid and lymphoid dysregulation is linked to differential mortality in patients treated with anakinra or corticosteroids, underscoring its prognostic and therapeutic significance. In conclusion, this novel immunology-based framework for quantifying cellular compartment dysregulation offers a valuable tool for prognosis and therapeutic decision-making in critical illness.

  View details for DOI 10.1101/2024.11.12.623298

  View details for PubMedID 39605502

  View details for PubMedCentralID PMC11601436

• Improved Prediction of Stabilizing Mutations in Proteins by Incorporation of Mutational Effects on Ligand Binding. Proteins Ganesan, S., Mittal, N., Bhat, A., Adiga, R. S., Ganesan, A., Nagarajan, D., Varadarajan, R. 2024

  Abstract

  While many computational methods accurately predict destabilizing mutations, identifying stabilizing mutations has remained a challenge, because of their relative rarity. We tested ΔΔG0 predictions from computational predictors such as Rosetta, ThermoMPNN, RaSP, and DeepDDG, using 82 mutants of the bacterial toxin CcdB as a test case. On this dataset, the best computational predictor is ThermoMPNN, which identifies stabilizing mutations with a precision of 68%. However, the average increase in Tm for these predicted mutations was only 1°C for CcdB, and predictions were poorer for a more challenging target, influenza neuraminidase. Using data from multiple previously described yeast surface display libraries and in vitro thermal stability measurements, we trained logistic regression models to identify stabilizing mutations with a precision of 90% and an average increase in Tm of 3°C for CcdB. When such libraries contain a population of mutants with significantly enhanced binding relative to the corresponding wild type, there is no benefit in using computational predictors. It is then possible to predict stabilizing mutations without any training, simply by examining the distribution of mutational binding scores. This avoids laborious steps of in vitro expression, purification, and stability characterization. When this is not the case, combining data from computational predictors with high-throughput experimental binding data enhances the prediction of stabilizing mutations. However, this requires training on stability data measured in vitro with known stabilized mutants. It is thus feasible to predict stabilizing mutations rapidly and accurately for any system of interest that can be subjected to a binding selection or screen.

  View details for DOI 10.1002/prot.26738

  View details for PubMedID 39166462

• 42-gene Severe-or-Mild (SOM) gene expression signature is conserved across viral and bacterial infections and originates from developing neutrophils Moore, A., Freedman, M., Zheng, H., Ganesan, A., Rogers, A., Khatri, P. AMER ASSOC IMMUNOLOGISTS. 2024

  View details for DOI 10.4049/jimmunol.212.supp.0635.5089

  View details for Web of Science ID 001368989500047

• Epigenetic Profiling of PTPN11 Mutant JMML Hematopoietic Stem and Progenitor Cells Reveals an Aberrant Histone Landscape. Cancers Sinha, R., Dvorak, M., Ganesan, A., Kalesinskas, L., Niemeyer, C. M., Flotho, C., Sakamoto, K. M., Lacayo, N., Patil, R. V., Perriman, R., Cepika, A. M., Liu, Y. L., Kuo, A., Utz, P. J., Khatri, P., Bertaina, A. 2023; 15 (21)

  Abstract

  Juvenile myelomonocytic leukemia (JMML) is a deadly pediatric leukemia driven by RAS pathway mutations, of which >35% are gain-of-function in PTPN11. Although DNA hypermethylation portends severe clinical phenotypes, the landscape of histone modifications and chromatin profiles in JMML patient cells have not been explored. Using global mass cytometry, Epigenetic Time of Flight (EpiTOF), we analyzed hematopoietic stem and progenitor cells (HSPCs) from five JMML patients with PTPN11 mutations. These data revealed statistically significant changes in histone methylation, phosphorylation, and acetylation marks that were unique to JMML HSPCs when compared with healthy controls. Consistent with these data, assay for transposase-accessible chromatin with sequencing (ATAC-seq) analysis revealed significant alterations in chromatin profiles at loci encoding post-translational modification enzymes, strongly suggesting their mis-regulated expression. Collectively, this study reveals histone modification pathways as an additional epigenetic abnormality in JMML patient HSPCs, thereby uncovering a new family of potential druggable targets for the treatment of JMML.

  View details for DOI 10.3390/cancers15215204

  View details for PubMedID 37958378

• Ancestry-based differences in the immune phenotype are associated with lupus activity. JCI insight Slight-Webb, S., Thomas, K., Smith, M., Wagner, C. A., Macwana, S., Bylinska, A., Donato, M., Dvorak, M., Chang, S. E., Kuo, A., Cheung, P., Kalesinskas, L., Ganesan, A., Dermadi, D., Guthridge, C. J., DeJager, W., Wright, C., Foecke, M. H., Merrill, J. T., Chakravarty, E., Arriens, C., Maecker, H. T., Khatri, P., Utz, P. J., James, J. A., Guthridge, J. M. 2023; 8 (16)

  Abstract

  Systemic lupus erythematosus (SLE) affects 1 in 537 Black women, which is >2-fold more than White women. Black patients develop the disease at a younger age, have more severe symptoms, and have a greater chance of early mortality. We used a multiomics approach to uncover ancestry-associated immune alterations in patients with SLE and healthy controls that may contribute biologically to disease disparities. Cell composition, signaling, epigenetics, and proteomics were evaluated by mass cytometry; droplet-based single-cell transcriptomics and proteomics; and bead-based multiplex soluble mediator levels in plasma. We observed altered whole blood frequencies and enhanced activity in CD8+ T cells, B cells, monocytes, and DCs in Black patients with more active disease. Epigenetic modifications in CD8+ T cells (H3K27ac) could distinguish disease activity level in Black patients and differentiate Black from White patient samples. TLR3/4/7/8/9-related gene expression was elevated in immune cells from Black patients with SLE, and TLR7/8/9 and IFN-α phospho-signaling and cytokine responses were heightened even in immune cells from healthy Black control patients compared with White individuals. TLR stimulation of healthy immune cells recapitulated the ancestry-associated SLE immunophenotypes. This multiomic resource defines ancestry-associated immune phenotypes that differ between Black and White patients with SLE, which may influence the course and severity of SLE and other diseases.

  View details for DOI 10.1172/jci.insight.169584

  View details for PubMedID 37606045

• Inferring direction of associations between histone modifications using a neural processes-based framework. iScience Ganesan, A., Dermadi, D., Kalesinskas, L., Donato, M., Sowers, R., Utz, P. J., Khatri, P. 2023; 26 (1): 105756

  Abstract

  Current technologies do not allow predicting interactions between histone post-translational modifications (HPTMs) at a system-level. We describe a computational framework, imputation-followed-by-inference, to predict directed association between two HPTMs using EpiTOF, a mass cytometry-based platform that allows profiling multiple HPTMs at a single-cell resolution. Using EpiTOF profiles of >55,000,000 peripheral mononuclear blood cells from 158 healthy human subjects, we show that neural processes (NP) have significantly higher accuracy than linear regression and k-nearest neighbors models to impute the abundance of an HPTM. Next, we infer the direction of association to show we recapitulate known HPTM associations and identify several previously unidentified ones in healthy individuals. Using this framework in an influenza vaccine cohort, we identify changes in associations between 6 pairs of HPTMs 30 days following vaccination, of which several have been shown to be involved in innate memory. These results demonstrate the utility of our framework in identifying directed interactions between HPTMs.

  View details for DOI 10.1016/j.isci.2022.105756

  View details for PubMedID 36619977

  View details for PubMedCentralID PMC9813700

• Mass-cytometry-based quantitation of global histone post-translational modifications at single-cell resolution across peripheral immune cells in IBD. Journal of Crohn's & colitis Bai, L., Dermadi, D., Kalesinskas, L., Dvorak, M., Chang, S. E., Ganesan, A., Rubin, S. J., Kuo, A., Cheung, P., Donato, M., Utz, P. J., Habtezion, A., Khatri, P. 2022

  Abstract

  Current understanding of histone post-translational modifications (histone modifications) across immune cell types in patients with inflammatory bowel disease (IBD) during remission and flare is limited. The study aimed to quantify histone modifications at a single-cell resolution in IBD patients during remission and flare and how they differ compared to healthy controls.We performed a case-control study of 94 subjects (83 IBD patients and 11 healthy controls). IBD patients had either UC (n=38) or CD (n=45) in clinical remission or flare. We used epigenetic profiling by time-of-flight (EpiTOF) to investigate changes in histone modifications within peripheral blood mononuclear cells from IBD patients.We discovered substantial heterogeneity in histone modifications across multiple immune cell types in IBD patients. They had a higher proportion of less differentiated CD34 + hematopoietic progenitors, and a subset of CD56 bright NK cells and γδ T cells characterized by distinct histone modifications associated with the gene transcription. The subset of CD56 bright NK cells had increased several histone acetylations. An epigenetically defined subset of NK was associated with higher levels of CRP in peripheral blood. CD14+ monocytes from IBD patients had significantly decreased cleaved H3T22, suggesting they were epigenetically primed for macrophage differentiation.We describe the first systems-level quantification of histone modifications across immune cells from IBD patients at a single-cell resolution revealing the increased epigenetic heterogeneity that is not possible with traditional ChIP-seq profiling. Our data open new directions in investigating the association between histone modifications and IBD pathology using other epigenomic tools.

  View details for DOI 10.1093/ecco-jcc/jjac194

  View details for PubMedID 36571819