Research
Abstract
Sarcomas are a heterogeneous group of cancers with few shared therapeutic targets. We show that PI3K signaling is frequently activated in sarcomas due to PTEN loss (in 30-60%), representing a common therapeutic target. The PI3K pathway has lacked a downstream oncogenic transcription factor. We show TAZ and YAP are transcriptional co-activators regulated by PI3K and drive a transcriptome necessary for tumor growth in a PI3K-driven sarcoma mouse model. This PI3K-TAZ/YAP axis exists in parallel to the known PI3K-Akt-mTORC1 axis providing a rationale for combination therapy targeting the TAZ/YAP-TEAD interaction and mTORC1. Combination therapy using IK-930 (TEAD inhibitor) and everolimus (mTORC1 inhibitor) synergistically diminished proliferation and anchorage dependent growth of PI3K-activated sarcoma cell lines at low, physiologically achievable doses. Furthermore, this combination therapy showed a synergistic effect in vivo, suggesting that an integrated view of PI3K and Hippo signaling can be leveraged therapeutically in PI3K activated sarcomas.
Authors
Keith C. Garcia, Ali A. Khan, Krishnendu Ghosh, Souradip Sinha, Nicholas Scalora, Gillian DeWane, Colleen Fullenkamp, Nicole Merritt, Yuliia Drebot, Samuel Y. Yu, Mariah Leidinger, Michael D. Henry, Patrick J. Breheny, Michael S. Chimenti, Munir R. Tanas
Abstract
Hirschsprung disease (HSCR) is a congenital intestinal disorder characterized by the absence of ganglia in the distal intestine. Despite surgical resection of the aganglionic intestine and pull-through surgery, HSCR patients still experience bowel dysfunction, indicating that latent abnormalities may also exist in the proximal ganglionic intestine. To elucidate possible causes of postoperative bowel dysfunction in HSCR, we investigated differences in the proximal ganglionic intestine using an animal model of HSCR (Ednrb-null mice) and validated our findings in tissue from human HSCR patients. We found that the proximal ganglionic colon of HSCR mice exhibited greater stiffness and fibrosis than their wild-type littermates. Similarly, submucosal fibrosis was significantly greater in the proximal ganglionic intestine of HSCR patients than in intestinal tissue from age and site-matched controls. Furthermore, we observed dysregulated expression of extracellular matrix (ECM)-related genes in the proximal ganglionic intestine of HSCR mice compared to controls. We conclude that increased fibrosis, stiffness, and alterations in ECM composition may contribute to persistent dysfunction of the ganglionic intestine in HSCR. These findings add to the growing body of literature that describe abnormalities in the proximal ganglionic intestine of HSCR and suggest that HSCR is not limited to the aganglionic intestine alone.
Authors
Prisca C. Obidike, Britney A. Hsu, Chioma Moneme, Oluyinka O. Olutoye II, Walker D. Short, Mary Hui Li, Swathi Balaji, Yuwen Zhang, Sundeep G. Keswani, Lily S. Cheng
Abstract
Dysfunctional white adipose tissue contributes to the development of obesity-related morbidities, including insulin resistance, dyslipidemia, and other metabolic disorders. Adipose tissue macrophages (ATMs) accumulate in obesity and play both beneficial and harmful roles in the maintenance of adipose tissue homeostasis and function. Despite their importance, the molecules and mechanisms that regulate these diverse functions are not well understood. Lipid-associated macrophages (LAMs), the dominant subset of obesity-associated ATMs, accumulate in crown-like structures and are characterized by a metabolically activated and proinflammatory phenotype. We previously identified CD9 as a surface marker of LAMs. However, the contribution of CD9 to the activation and function of LAMs during obesity is unknown. Using a myeloid-specific CD9 knockout model, we show that CD9 supports ATM-adipocyte adhesion and crown-like structure formation. Furthermore, CD9 promotes the expression of pro-fibrotic and extracellular matrix remodeling genes. Loss of myeloid CD9 reduces adipose tissue fibrosis, increases visceral adipose tissue accumulation, and improves global metabolic outcomes during diet-induced obesity. These results identify CD9 as a causal regulator of pathogenic LAM functions, highlighting CD9 as a potential therapeutic target for treating obesity-associated metabolic disease.
Authors
Julia Chini, Nicole DeMarco, Dana V. Mitchell, Sam J. McCright, Kaitlyn M. Shen, Divyansi Pandey, Rachel L. Clement, Jessica Miller, Rajan Jain, Deanne M. Taylor, Mitchell A. Lazar, David A. Hill
Abstract
Latently infected cells persist in people living with HIV (PWH) despite suppressive antiretroviral therapy (ART) and evade immune clearance. Shock and Kill cure strategies are hampered by insufficient enhancement of targeted immune responses following latency reversal. We previously demonstrated autologous Vδ2 T cells from PWH retain anti-HIV activity and can reduce CD4+ T cell reservoirs, although their use in cure approaches is limited due to their dual role as a viral reservoir. However, promising clinical data in oncology shows their unique MHC- unrestricted antigen recognition affords potent on-target cytotoxicity in the absence of graft-versus-host disease when used as an allogeneic adoptive cell therapy modality. Here, we found expanded allogeneic Vδ2 T cells specifically eliminated HIV-infected CD4+ T cells and monocyte-derived macrophages (MDM), overcoming inherent resistance to killing by other cell types such as NK and CD8+ T cells. Notably, we demonstrated allogeneic Vδ2 T cells recognized and eliminated the HIV-latent CD4+ T cell reservoir following latency reversal. Our study provides evidence for developing an allogeneic γδ T cell therapy for HIV cure and warrants pre-clinical investigation in combination approaches.
Authors
Brendan T. Mann, Marta Sanz, Alisha Chitrakar, Kayley Langlands, Marc Siegel, Natalia Soriano-Sarabia
Abstract
Infectious diseases remain a global health challenge, driven by increasing antimicrobial-resistance and the threat of emerging epidemics. Mycobacterium tuberculosis and Staphylococcus aureus are leading causes of mortality worldwide. Trained immunity—a form of innate immune memory—offers a promising approach to enhance pathogen clearance. Here, we demonstrate that IFN-γ induces trained immunity in human monocytes through a mechanism involving mTORC1 activation, glutaminolysis, and epigenetic remodeling. Macrophages derived from IFN-γ–trained monocytes exhibited increased glycolytic activity with enhanced cytokine and chemokine responses upon stimulation or infection. Crucially, trained macrophages had increased production of reactive oxygen species which mediated enhanced bactericidal activity against methicillin-resistant S. aureus. Furthermore, ATAC-sequencing analysis of IFN-γ trained macrophages revealed increased chromatin accessibility in regions associated with host defence. Lastly, IFN-γ training restored impaired innate responses in macrophages from individuals homozygous for the TIRAP 180L polymorphism, a genetic variant associated with increased susceptibility to infection. These findings establish IFN-γ as a potent inducer of trained immunity in human monocytes and support its potential as a host-directed strategy to strengthen antimicrobial defenses, particularly in genetically susceptible individuals and high-risk clinical contexts.
Authors
Dearbhla M. Murphy, Isabella Batten, Aoife O'Farrell, Simon R. Carlile, Sinead A. O'Rourke, Chloe Court, Brenda Morris, Gina Leisching, Gráinne Jameson, Sarah A. Connolly, Adam H. Dyer, John P. McGrath, Emma McNally, Olivia Sandby-Thomas, Anjali Yennemadi, Conor M. Finlay, Clíona Ni Cheallaigh, Jean Dunne, Cilian Ó Maoldomhnaigh, Laura E. Gleeson, Aisling Dunne, Nollaig Bourke, Reinout van Crevel, Donal J. Cox, Niall Conlon, Arjun Raj, Rachel M. McLoughlin, Joseph Keane, Sharee A. Basdeo
Abstract
Nicotinamide adenine dinucleotide (NAD⁺) is essential for cellular metabolism, DNA repair, and stress responses. NAD+ is synthesized from nicotinamide, nicotinic acid (collectively termed niacin), and tryptophan. In humans, deficiencies in these nutrients result in pellagra, marked by dermatitis, diarrhea, and dementia. The dermatitis associated with pellagra typically manifests as photodermatosis in sun-exposed areas. This study examined the effects of NAD+ deficiency on skin homeostasis using epidermis-specific Nampt conditional knockout (cKO) mice. These mice displayed substantial NAD⁺ depletion, reduced poly(ADP-ribose) polymerase (PARP) activity, and increased DNA damage. Consequently, Nampt cKO mice developed spontaneous skin inflammation and epidermal hyperplasia. RNA sequencing and immunohistochemical analyses demonstrated increased interleukin-36 (IL-36) cytokine expression, suggesting that DNA repair-related genomic stress triggers keratinocyte-driven IL-36 production, which promotes inflammation. Furthermore, reduced collagen17A1 expression and elevated thymic stromal lymphopoietin (TSLP) levels were observed. NAD+ repletion by transdermal supplementation of nicotinamide mononucleotide (NMN) suppressed the rise of IL-36 levels and skin inflammation. These findings underscore the importance of Nampt-mediated NAD⁺ metabolism for epidermal stability and indicate that NAD⁺ depletion may contribute to IL-36-mediated skin inflammation, offering insights for therapeutic strategies in inflammatory skin disorders.
Authors
Taiki Seki, Jun-Dal Kim, Yasuhito Yahara, Hitoshi Uchida, Keisuke Yaku, Mariam Karim, Teruhiko Makino, Tadamichi Shimizu, Takashi Nakagawa
Abstract
Adaptive remodeling of retrodiscal tissue following anterior disc displacement (ADD) of the temporomandibular joint (TMJ) has been recognized for decades, yet the underlying cellular dynamics and molecular mechanisms remain unclear. Using a porcine ADD model, this study investigated the cellular and molecular basis driving retrodiscal tissue adaptation. Histological staining revealed adaptive remodeling of retrodiscal tissue after ADD induction, with dense connective tissue and cartilaginous masses replacing loose connective tissue. Furthermore, single-cell RNA sequencing (scRNA-seq) captured pronounced fibroblast expansion during tissue remodeling, notably the FB2 subcluster with high developmental potential, and the emergence of a mural cell subcluster MC4 associated with extracellular matrix (ECM) remodeling. CellChat analysis highlighted MC4-FB2 crosstalk via FGF2 and BMP5 signaling. The combination of pathway-aware multi-layered hierarchical network (P-NET) and Seurat with drug database screening identified five promising compounds. Among them, Zaprinast demonstrated the most robust effects by enhancing the remodeling capability of fibroblasts in vitro, and also alleviated TMJ deformation in vivo. Collectively, fibroblast activation is pivotal for early retrodiscal tissue adaptation following ADD, which is driven by MC4-derived FGF2/BMP5 signaling. Zaprinast treatment potentiates this remodeling process. These findings provide new insights into cellular basis of TMJ adaptation and identify potential therapeutic targets for ADD management.
Authors
Wenlin Yuan, Yilin Chen, Ruojin Yan, Wei Liu, Chenyu Wang, Ying Wang, Qiaoli Dai, Wen Li, Mengqi Zhu, Xiao Chen, Jiejun Shi
Abstract
The chronic inflammation of Crohn’s disease frequently leads to fibrosis and muscular hypertrophy of the intestinal wall. This often culminates in strictures, a serious condition lacking directed therapy. Severe pathological changes occur in the submucosa and muscularis propria intestinal wall layers of strictures, yet stricture-associated proteome changes in these layers is unexplored. We perform unbiased proteomics on submucosa and muscularis propria microdissected from transmural sections of strictured and non-strictured ileum. Proteome changes in stricture submucosa reflect a transition from homeostasis to tissue remodeling, inflammation and smooth muscle alterations. Top submucosa features include reduced vascular components and lipid metabolism proteins accompanied by increased proteins with immune-, matrix- or stress functions including CTHRC1, TNC, IL16, MZB1 and TXNDC5. In parallel, predominant changes in stricture muscularis propria include increased matrix (POSTN) and immune (mast cell CPA3) proteins alongside decreased proteins with lipid metabolic, mitochondrial or key muscle functions. Finally, trends of differentially expressed proteins along non-stricture submucosa suggest progressive profibrotic tissue remodeling and muscle expansion as proximity to stricture increases. The comprehensive proteome map presented here offers unique layer-resolved insight into the stricture microenvironment and potential drivers of fibrotic disease, providing a valuable resource to fuel biomarker and therapeutic target research. Keywords: Crohn’s disease strictures, proteomics, intestinal fibrosis, fibrostenosis
Authors
Johannes Alfredsson, Carina Sihlbom Wallem, Maja Östling, Hanna de la Croix, Elinor Bexe-Lindskog, Mary Jo Wick
Abstract
Cardiovascular disease (CVD), both atherosclerosis-related and heart failure with preserved ejection fraction (HFpEF) linked to cardiac fibrosis, contributes to morbidity and mortality in people with HIV (PWH) receiving antiretroviral therapy (ART). In the REPRIEVE trial, pitavastatin reduces atherosclerotic CVD risk to a magnitude inconsistent with pitavastatin’s impact solely on LDL-cholesterol and inflammation. We hypothesized that HFpEF in PWH is related to HIV-induced fibrosis mediated by platelet TGFβ1, is accelerated by certain contemporary ART, and may also be inhibited by statins. ART drugs used in REPRIEVE, including a nucleoside/nucleotide, integrase inhibitor-based regimen (tenofovir (TDF), emtricitabine (FTC), and dolutegravir (DTG)), and the protease inhibitors ritonavir (RTV) and darunavir (DRV), and the impact of atorvastatin, were examined in two HIV mouse models: transgenic HIV-Tg26 mice and HIV-PDX mice engrafted with T cells isolated from PWH. HIV-Tg26 and HIV-PDX mice had higher cardiac fibrosis than littermate controls without HIV (p<0.05). Administration of TDF-FTC-DTG or RTV, but not DRV, resulted in a further ~2-fold increase in fibrosis (p<0.01). Cardiac fibrosis and intracardiac fat accumulation correlated with reduced diastolic function. Mice depleted of platelet TGFβ1 (TGFβ1Platelet-ΔTg26), or treated with atorvastatin, were partially protected from HIV- and ART-induced cardiac fibrosis, steatosis, and diastolic dysfunction. Atorvastatin effects were independent of changes in inflammatory cytokines. These effects correlated with reduced platelet activation and TGFβ signaling in cardiac endothelial cells, fibroblasts, and macrophages undergoing mesenchymal transition. Our results indicate that certain ART regimens accelerate HIV-associated CVD characterized by HFpEF via platelet TGFβ1-dependent processes, which were mitigated by atorvastatin. We postulate that our findings provide a potential mechanism for the pleiotropic effects of statins in HIV/ART-linked CVD which could be targeted by antiplatelet agents or inhibition of TGFβ signaling.
Authors
Kumar Subramani, Denys Babii, Brienne Cole, Tayyab A. Afzal, Thamizhiniyan Venkatesan, Trevor Word, Sandra Gostynska, Sixia Chen, Kar-Ming Fung, Ali Danesh, Itzayana G. Miller, Paul Klotman, Brad R. Jones, Jeffrey Laurence, Jasimuddin Ahamed
Abstract
Mutation of KRAS in endothelial cells (KRAS-EC) leads to intracerebral hemorrhage (ICH) in brain arteriovenous malformations (bAVM), resulting in severe disabilities or even death. However, it is unclear what causes this hemorrhagic conversion of bAVM. Here, using a locally established, clinically-relevant sporadic bAVM mouse model, created by overexpressing mutant KRAS (KRASG12V) in the brain EC, we demonstrate that KRAS-EC act as trigger for microglia (MG) activation and infiltration of macrophages (Mϕ). Using three-dimensional immunostaining approach with cleared human and mouse bAVM tissues, we demonstrate an abundance of MG/Mϕ around the bAVM nidus. The presence of MG/Mϕ are correlated to the blood-brain barrier leakage in bAVM area. Time-lapsed intravital imaging in Cx3cr1-gfp;Ccr2-rfp reporter mice demonstrate the dynamic activation of MG and infiltration of Mϕ toward mutant KRAS-modified dysplastic vessels. Importantly, a time course analysis showed that these activated/infiltrated MG/Mϕ are present around the bAVMs prior to hemorrhagic conversion, and controlled depletion of MG/Mϕ reduced ICH incidence in bAVM. Inhibition of MG/Mϕ with long-term minocycline treatment attenuated the incidence of ICHs around bAVMs. Our study indicates that MG/Mϕ are involved in destabilization of KRAS-induced bAVM, leading to hemorrhagic conversion/ICH. Thus, modulation of MG/Mϕ may reduce ICH risk in bAVM patients.
Authors
Hyejin Park, Jung-Eun Park, Bridger H. Freeman, Bosco Seong Kyu Yang, Shun-Ming Ting, Alexander K. Suh, Jude P.J. Savarraj, Shuning Huang, Jakob Körbelin, Huimahn Alex Choi, Sean P. Marrelli, Jaroslaw Aronowski, Peng Roc Chen, Eunhee Kim, Eun S. Park
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