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Abstract
Dedifferentiated liposarcoma (DDLS), myxofibrosarcoma (MFS), and undifferentiated pleomorphic sarcoma (UPS) are the most common types of genetically complex sarcoma. There is an urgent need to develop effective targeted therapy for these deadly sarcoma types. Despite their genetic complexity, these sarcomas share genomic alterations causing PI3K/Akt/mTOR and MAPK pathway activation, and both pathways control translation mediated by the RNA helicase eIF4A. We therefore investigated eIF4A inhibition as a therapeutic strategy. The eIF4A inhibitor CR-1-31B effectively suppressed tumor growth and induced apoptosis in DDLS, MFS, and UPS patient-derived cell lines and mouse xenografts. Transcriptome-scale ribosome footprinting identified eIF4A-dependent mRNAs such as the Hippo pathway transcriptional coactivators YAP1 (YAP) and WWTR1 (TAZ). Combined knockdown of YAP and TAZ induced apoptosis in DDLS, MFS, and UPS cell lines, and their ectopic expression partially rescued cells from apoptosis induced by CR-1-31B. Genomic analysis of patient tumors revealed that YAP and WWTR1 were frequently amplified or gained in DDLS, MFS, and UPS and were associated with worse clinical outcomes. Together, our findings identify a new strategy for targeting the Hippo pathway in incurable forms of sarcoma based on inhibition of eIF4A-dependent translation of the key oncogenic transcription factors YAP and TAZ.
Authors
Young-Mi Kim, Prathibha Mohan, Urmila Sehrawat, Evan Seffar, Rafaela Muniz De Queiroz, Kalyani Chadalavada, Nikita Persaud, Tomoyo Okada, Anirudh Kulkarni, Jianan Lin, Nathalie Lailler, Shaleigh Smith, Bhumika Jadeja, Nicholas D. Socci, Zhengqing Ouyang, Hans-Guido Wendel, Samuel Singer
Abstract
Hypoxia critically restricts the effectiveness of immunotherapy in triple-negative breast cancer (TNBC). Comprehensive bioinformatics analyses demonstrated that highly hypoxic TNBC tumors exhibited elevated T cell exhaustion, increased immune checkpoint molecule expression, and diminished responsiveness to immune checkpoint blockade (ICB). Consequently, strategies aimed at alleviating tumor hypoxia may effectively augment ICB therapy. Although ultrasound-targeted microbubble cavitation (UTMC) has been shown to reduce tumor hypoxia, the precise molecular mechanisms remain unclear. Here, we provided evidence that UTMC activated endothelial nitric oxide synthase (eNOS) through G protein–coupled signaling, resembling pathways induced by fluid shear stress. UTMC-induced eNOS activation was largely Ca²⁺-dependent and resulted in increased nitric oxide production. Enhanced nitric oxide generation was associated with improved tumor perfusion and reduced hypoxia. Combining UTMC with anti–PD-L1 therapy markedly improved the tumor immune microenvironment, characterized by increased CD8+ T cell infiltration, reduced T cell exhaustion, diminished regulatory T cell infiltration, increased macrophage polarization from an M2 to M1 phenotype, and elevated production of pro-inflammatory cytokines. Collectively, our findings identified UTMC as a promising adjunctive therapeutic approach to mitigate hypoxia and enhance the efficacy of anti–PD-L1 immunotherapy in TNBC. These results support further translational evaluation of UTMC-based combination strategies in hypoxic TNBC.
Authors
Zhiyu Zhao, Li Ba, Siwei Li, Jianxin Wang, Yuzhou Luo, Sihan Wang, Yan Jin, Changjun Wu
Abstract
Pancreatic ductal adenocarcinoma (PDAC) shows profound resistance to immunotherapy due to its immunosuppressive tumor microenvironment. Here, we studied the relationship between T cell infiltration and innate immune signaling in PDAC, identifying Toll-like receptor 2 (TLR2) as a key regulator of T cell exclusion. TLR2 expression correlated with T cell infiltration in both human and mouse PDAC tumors. Using genetic knockout models and adoptive T cell transfer experiments, we found that TLR2 expression in both T cells and non-T cells contributes to T cell exclusion in PDAC. Notably, successful infiltration of adoptively transferred tumor-specific T cells required TLR2 deletion in both the transferred cells and the recipient host. The therapeutic implications of these findings are demonstrated through both genetic deletion and pharmacological inhibition of TLR2 using AAV-mediated and antibody-based approaches in murine models, resulting in decreased tumor growth and extended survival. Collectively, these findings identify TLR2 as a key modulator of T cell trafficking and immune suppression within the PDAC microenvironment, suggesting its potential as a therapeutic target for improving treatment outcomes.
Authors
Jacqueline Plesset, Meredith L. Stone, John C. McVey, Heather Coho, Kelly Markowitz, Kayjana Coho, Jesse Lee, Anna S. Thickens, Devora Delman, Gregory L. Beatty
Abstract
HIV infection rapidly impairs the gastrointestinal (GI) barrier, contributing to persistent mucosal immune dysfunction, microbial translocation, and systemic inflammation despite antiretroviral therapy (ART). Using SIV-infected rhesus macaques on long-term ART, we investigated mechanisms underlying impairment in gut barrier-protective IL-17/IL-22 responses and the potential modulation of this pathway by dietary indoles. Longitudinal profiling of colonic epithelial and lamina propria cells revealed a selective loss of IL-17/IL-22–producing γδT cells and type 3 innate lymphoid cells (ILC3s). This loss correlated with reduced expression of the transcription factors AHR and RORγt and was associated with elevated plasma markers of intestinal epithelial barrier disruption (IEBD), including intestinal fatty acid–binding protein (iFABP), zonulin, and LPS-binding protein (LBP). Targeting this transcriptional deficiency, dietary indole supplementation for one month restored colonic AHR⁺IL-22-producing γδ T cells, RORγt⁺ ILC3s, and Vδ1 T cells, and was associated with reduced iFABP and zonulin levels. Immunohistochemical analyses further demonstrated enrichment of AHR/RORγt-co-expressing cells in the colon of indole-supplemented animals during chronic SIV infection on ART. Collectively, these findings indicate that disruption of the AHR-RORγt axis is a key pathogenic mechanism underlying persistent IEBD in chronic SIV/HIV infection. Modulation of AHR and RORγt signaling pathways in the gut may therefore represent a promising therapeutic strategy to reinforce mucosal barrier function and mitigate chronic inflammation in people living with HIV.
Authors
Siva Thirugnanam, Alison R. Van Zandt, Alexandra B. McNally, Victoria A. Hart, Isabelle Berthelot, Cecily C. Midkiff, Lara A. Doyle-Meyers, David A. Welsh, Robert V. Blair, Andrew G. MacLean, Namita Rout
Abstract
Spinocerebellar Ataxia Type 14 (SCA14) is an autosomal dominant neurodegenerative disease caused by mutations in the gene encoding protein kinase C gamma (PKCγ), a Ca2+/diacylglycerol (DG)-dependent serine/threonine kinase dominantly expressed in cerebellar Purkinje cells. These mutations impair autoinhibitory constraints to increase the basal activity of the kinase, resulting in deficits in the cerebellum that are not observed upon simple deletion of the gene, and severe ataxia. To better understand the impact of aberrant PKCγ signaling in disease pathology, we developed a knock-in murine model of the SCA14 mutation ΔF48 in PKCγ. This fully-penetrant mutation is severe in humans and is mechanistically informative as it has high basal activity but is unresponsive to agonist stimulation. Genetic, behavioral, and molecular testing revealed that ΔF48 PKCγ mice have ataxia-related phenotypes and an altered cerebellar phosphoproteome driven primarily by enhanced Ca2+/calmodulin-dependent Kinase II (CaMKII) signaling, effects that were more severe in male mice. Analysis of existing human data revealed that SCA14 has a significantly earlier age of onset for males compared with females. Data from this clinically relevant mutation suggested that enhanced basal activity of PKCγ is sufficient to cause ataxia and that treatment strategies to modulate aberrant PKCγ may be particularly beneficial in males.
Authors
Sarah A. Wolfe, Yuliang Ma, Tomer M. Yaron-Barir, Carly Chang, Caila A. Pilo, Majid Ghassemian, Amanda J. Roberts, Sang Ryeul Lee, Benjamin A. Henson, Kristen Jepsen, Jared L. Johnson, Lewis C. Cantley, Susan S. Taylor, George Gorrie, Alexandra C. Newton
Abstract
Authors
William J. Crisler, Noor Sohail, Samuel J. Steuart, Maria Vazquez-Machado, Arjun Mahajan, Maureen Whittelsey, Alex Pickering, Michael J. Martinez, Theresa Hutchins, Jessica E. Teague, Qian Zhan, Shannan Ho Sui, Ruth Ann Vleugels, Kathryn S. Torok, Heidi Jacobe, Rachael A. Clark, Avery LaChance
Abstract
Huntington’s disease (HD) is a fatal neurodegenerative disease caused by an expanded polyglutamine (CAG) repeat in the N-terminal of the Huntingtin protein (HTT). Microglial activation and elevated pro-inflammatory cytokines are observed in HD brains, but the mechanisms regulating neuroinflammation and microglial activation are poorly understood. Metformin-mediated neuroprotection has been demonstrated in experimental models of neurodegeneration, including HD. We found that metformin inhibits mitochondrial DNA (mtDNA) release and subsequent neuroinflammation in the cortex and striatum of a mouse model of HD. Moreover, elevated pro-inflammatory cytokines and microglial activation are inhibited by metformin in HD transgenic mice brain. Metformin reduced pathological microglial clusters and shifted towards a quiescent, homeostatic phenotype. Metformin improved aberrant immunometabolism in HD mouse brain and primary microglia. Mechanistically found that metformin regulates mitochondrial fission, reprograms deregulated metabolism in HD microglia, and controls microglial activation and inflammation in HD transgenic mice.
Authors
Abhishek Jauhari, Adam C. Monek, Olena S. Abakumova, Tanisha Singh, Sukhman Singh, Xiaomin Wang, Carley S. Clise, Diane L. Carlisle, Robert M. Friedlander
Abstract
The RhoBTB1-Cullin3 (CUL3) pathway in smooth muscle cells (SMCs) controls the ubiquitination and proteasomal degradation of target proteins that regulate vasodilation, vasoconstriction, and the actin cytoskeleton, and through this blood pressure (BP) and arterial stiffness. Using proximity labelling coupled with mass spectrometry in A7R5 SMCs, we identified proteins which bound to the C-terminal half of RhoBTB1 which functions as an adapter to deliver substrates to CUL3. We examined the physiological relevance of one of these substrates, RbFox2. Co-immunoprecipitation validated the interaction of RbFox2 with RhoBTB1. RbFox2 expression was elevated in response to inhibition of the ubiquitination-proteasomal pathway, CUL3-deficiency, and RhoBTB1 inhibition by either siRNA or angiotensin II (ANG). RbFox2 was ubiquitinated in a RhoBTB1- and CUL3-dependent manner suggesting its regulation through the RhoBTB1-CUL3-dependent ubiquitin-proteasome pathway. Inhibition of RbFox2 impaired the actin cytoskeleton in A7R5 cells and in primary SMC from RbFox2Flox/Flox (RbFox2F/F) mice and decreased the levels of globular and filamentous actin. ANG increased BP and arterial stiffness of RbFox2F/F mice, but the progression of arterial stiffness was halted after SMC-specific RbFox2 deletion despite a continued rise in BP. We conclude that RhoBTB1 and RbFox2 are important regulators of arterial stiffness through a mechanism that influences cytoskeletal integrity.
Authors
Gaurav Kumar, Nisita Chaihongsa, Daniel T. Brozoski, Daria Golosova, Ibrahim Vazirabad, Ko-Ting Lu, Kelsey K. Wackman, Ravi K. Singh, Curt D. Sigmund
Abstract
Authors
Anna J. Son, Emmanuel Rapp, Alex Wiezorek, Max G. Leung, Ronadip R. Banerjee, Thomas H. Leung
Abstract
VIC-1911 (formerly TAS-119) is a next-generation, ATP-competitive Aurora kinase A (AURKA) inhibitor with a favorable biosafety profile. However, it has not been evaluated in prostate cancer (PC), wherein AURKA is highly expressed in advanced stages and represents a critical therapeutic target. Here, we demonstrate that VIC-1911 potently inhibits AURKA activity with high selectivity over AURKB/C across diverse PC cell lines. Treatment with VIC-1911, even at nanomolar concentrations, substantially inhibits the growth of both androgen receptor (AR)-positive and AR-negative PC cells. VIC-1911 triggers mitotic failure, induces DNA double-strand breaks (DSBs), and activates the p53 pathway, halting cell division and inducing cell death. Notably, VIC-1911 showed synergistic effects in inhibiting PC cell growth in vitro and xenograft tumor growth in vivo with poly (ADP-ribose) polymerase inhibitors (PARPi), which have proven effective in PC with a deficiency in Homologous Recombination (HR) repair. Mechanistically, VIC-1911 disabled HR-mediated repair of DSBs in otherwise HR-proficient PC cells, leading to a “BRCAness” phenotype and pronounced accumulation of DNA damage and mitotic catastrophe. In summary, our study uncovers what we believe a novel mechanism to functional “BRCAness” by inducing mitotic arrest and highlights VIC-1911 as a promising therapeutic agent for advanced PC, either as a single agent or in combination, sensitizing HR-proficient tumors to PARP inhibitors.
Authors
Galina Gritsina, Sandip Kumar Rath, Hongshun Shi, Qi Chu, Wanqing Xie, Que Thanh Thanh Nguyen, Sambhavi Senthil, Thomas J. Myers, Mehmet A. Bilen, Sarah E. Fenton, Maha Hussain, David S. Yu, Jonathan C. Zhao, Jindan Yu
Abstract
β-arrestins are ubiquitously expressed cytosolic adaptor proteins that regulate G protein-coupled receptor-dependent and -independent pathways essential for numerous physiological functions. This study investigated the role of β-arrestin1 and -2 in embryonic lymphatic vessel development and survival by generating and characterizing mice with lymphatic, tamoxifen-inducible loss of the genes encoding β-arrestin-1 and -2 (Arrb1/2ΔiLEC). At embryonic day15.5 (E15.5), Arrb1/2ΔiLEC embryos exhibit profound hydrops fetalis and increased embryonic mortality compared to control Arrb1/2fl/fl embryos. Edematous Arrb1/2ΔiLEC embryos, which were more often represented by the female sex, showed growth restriction and decreased lymphatic endothelial cell (LEC) proliferation in the jugular lymphatic sac compared to controls. In vitro knockdown of β-arrestin1 in LECs increased proliferation and increased activation of AKT, while knockdown of β-arrestin2 decreased proliferation and decreased activation of both ERK and CREB. Arrb1/2ΔiLEC embryos also exhibited dilated dermal lymphatics with decreased continuous VE-Cadherin adherens junctions compared to controls. These results were recapitulated in vitro in β-arrestin1 and/or -2 knockdown human LECs, which showed a decrease in membrane VE-Cadherin and β-catenin levels, and prevention of adrenomedullin-induced linearization of VE-cadherin at endothelial cell–cell junctions. Collectively, these results demonstrate that loss of β-arrestin1/2 in lymphatics causes hydrops fetalis, mid-gestational growth arrest and embryonic demise associated with reduced LEC proliferation and disrupted VE-Cadherin adherens junctions.
Authors
Yanna Tian, D. Stephen Serafin, Monserrat Avila-Zozaya, Alyssa M. Tauro, Natalie M. Torres-Valle, Bryan M. Kistner, Danielle M. Dy, Elizabeth S. Douglas, Kathleen M. Caron
Abstract
Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder caused by mutations in the survival motor neuron 1 (SMN1) gene leading to decreased SMN protein levels and motor neuron dysfunction. SMN-restoring therapies offer clinical benefit, but the downstream molecular consequences of SMN reduction remain incompletely understood. SMN deficiency resulted in downregulation of kinesin heavy chain isoform 5A (KIF5A) in human neurons and in a mouse model of SMA. SMN associated with KIF5A mRNA and contributed to its stability. Reduced SMN levels impaired axon regeneration, which was rescued by KIF5A overexpression. Because KIF5A has also been connected to ALS, these findings provide evidence of a molecular link between SMA and ALS pathophysiology, highlighting KIF5A as an SMN regulated factor. Our findings suggest SMN-independent interventions targeting KIF5A could represent a complementary therapeutic approach for SMA and other motor neuron diseases.
Authors
Tetsuya Akiyama, Yi Zeng, Caiwei Guo, Olivia Gautier, Lauren Koepke, Heankel Lyons, Elana Molotsky, Juliane S. Bombosch, Odilia Sianto, Jay P. Ross, Phuong Hoang, Luke Zhao, Cole Spencer, Charlotte J. Sumner, Michelle Monje, John W. Day, Aaron D. Gitler
Abstract
Thoracic Aortic Aneurysm and Dissections (TAAD) is a progressive dilation of the aortic wall associated with degradation of the extracellular matrix (ECM), cystic medial degeneration, smooth muscle cell (SMC) dysfunction, and rarefaction. TAAD etiology and pathogenesis suggest that alteration of mechanical force propagation may contribute to SMC dysfunction. This study aims to determine the role of SMC focal adhesion proteins, which are key components of force transmission, in TAAD pathogenesis. scRNAseq analysis of human TAA aortas showed reduced expression of intracellular focal adhesion components, including PTK2 (FAK), VCL, ILK, and TES transcripts, in SMCs. Additionally, protein levels of FAK, ILK, and VCL were decreased in the aorta of patients with TAA. SMC-specific Ptk2, Vcl, and Ilk knockout mice treated with β-Aminopropionitrile (BAPN) exhibited increased mortality, aortic dilation, ECM breakdown, and SMC loss. Mechanistically, knocking down FAK, ILK, and VCL exacerbated gliotoxin-induced SMC anoikis, whereas overexpressing full-length wild-type (WT) and dead-kinase FAK conferred resistance to apoptosis and cell detachment, indicating that FAK's protective effects depend on its expression rather than its enzymatic activity. Inhibition of FAK kinase activity did not affect SMC apoptosis in vitro or aortic dilation in vivo. Our findings demonstrated that the expression of focal adhesion proteins protects against TAAD progression and SMC anoikis independently of FAK kinase activity.
Authors
Zhenyuan Zhu, Mingjun Liu, Jianxin Wei, Deepa Suryanarayan, Parya Behzadi, Robert Edgar, Julie A. Phillippi, Cynthia St. Hilaire, Cristina Espinosa-Diez, Delphine Gomez
Abstract
Glioblastoma (GBM) is the most malignant primary brain tumor. The presence of glioma stem/initiating cells (GICs) is known to cause strong treatment resistance; therefore, GICs are a major target for GBM therapy, although there are no therapies targeting GICs clinically. To identify novel treatments for GBMs, we performed drug repositioning screening using GICs and identified T-type calcium channel blocker lomerizine—a migraine prophylactic drug. Lomerizine inhibited proliferation, migration, invasion, and cell cycle progression and induced apoptosis in GICs and differentiated glioma cells. Lomerizine had antitumor effects by inactivating STAT3 in all cell lines. Furthermore, lomerizine also dephosphorylated AKT and ERK only in GICs and strong tumor suppressive ability. Lomerizine also reduced tumor volume and prolonged overall survival in vivo. Based on our data from in vitro and in vivo experiments, lomerizine has potential as a novel GBM therapeutic agent targeting against both GICs and differentiated glioma cells and could benefit for GBM patients.
Authors
Toshiya Ichinose, Sho Tamai, Nozomi Hirai, Takashi Maejima, Kosuke Nambu, Hemragul Sabit, Shingo Tanaka, Masashi Kinoshita, Masahiko Kobayashi, Michihiro Mieda, Atsushi Hirao, Mitsutoshi Nakada
Abstract
A large inter-individual variability in weight loss outcomes following bariatric surgery is reported. To ensure optimal patient management, it is crucial to accurately identify those most likely to benefit from the intervention. Since genetic variants largely contribute to surgery response, polygenic scores (PGS) derived from genome-wide association studies (GWAS) could constitute valuable tools for clinical decision making. We developed and evaluated PGS to predict the weight loss response in 540 patients with body mass index (BMI) ≥35kg/m2 who underwent biliopancreatic diversion with duodenal switch. Summary statistics derived from BMI-derived GWAS, together with summary statistics from previously published GWAS of BMI and adiposity features, were used to construct, evaluate, and benchmark weight-loss PGS. The full-adjusted BMI PGS model built in the entire cohort explained 39.6% of the mean-over-time excessive body weight loss (%EBWL), while the BMI-PGS built in the training dataset explained 38.9%. All benchmarked PGS based on BMI showed a significant relationship with mean-over-time %EBWL. These findings highlight the potential of BMI PGS in predicting weight loss after bariatric surgery and support their use as promising tools to improve the effectiveness of future anti-obesity treatments. Funding: Canadian Institutes of Health Research (PJT-168876).
Authors
Bastien Vallée Marcotte, Juan de Toro-Martín, André Tchernof, Louis Pérusse, Simon Marceau, Marie-Claude Vohl
Abstract
Authors
Robert Lakin, Xueyan Liu, Dana Sherrard, Mihir Parikh, Ryan Debi, Nazari Polidovitch, Markus J. Duncan, Jian Wu, Peter H. Backx
Abstract
N6-methyladenosine (m6A) modification is the most prevalent post-transcriptional epigenetic modification in mammalian mRNAs, and it has been implicated in the regulation of nervous system development by modulating mRNA metabolism. VIRMA is the largest core subunit of the m6A methyltransferase complex and essential for the assembly and stability of the m6A methyltransferase complex. In the retina, m6A methylation modification is widely distributed in various cellular layers and is essential for retinal homeostasis. Here, we demonstrate that VIRMA-mediated m6A modification is essential for retinal homeostasis. Loss of Virma in retinal rod cells resulted in abnormal reduction in m6A methylation levels, along with impaired photoreceptor function and degeneration. Mechanically, Virma depletion in photoreceptors dampened the m6A modification level of visual perception-associated genes, resulting compromised visual function and photoreceptors degeneration. Moreover, Virma interacts with splicing factor to regulate the alternative splicing events of retina function-related genes such as Polg2, which contributes to photoreceptor damage. Reintroduction of normal Virma expression colonially rescued photoreceptor degeneration. Collectively, our data elucidate the important role of Virma-mediated m6A modification in photoreceptor function and suggest that epigenetic modulation could serve as potential targets to treat these blinding diseases.
Authors
Wenjing Liu, Xiaojing Wu, Rong Zou, Fan Zhang, Yudi Fan, Kuanxiang Sun, Liping Yang, Jiang Hu, Lin Zhang, Xianjun Zhu
Abstract
We previously reported that excessive angiotensin-II (AT)->AT receptor-1 (ATR1) signaling results in sickle cell anemia (SCA)-associated nephropathy. Herein, we showed hyperangiotensinemia in SCA results from high erythroid cell-generated reactive oxygen species (ROS), which oxidized angiotensinogen (ATGN) and favored its rapid conversion to AT. Increased AT->ATR1 signaling in SCA erythroid cells generated ROS and created a positive feedback loop of ROS->oxidized ATGN->AT->ATR1-> ROS, perpetuating the hyperangiotensinemia. ATR1-blocker, losartan, reduced erythrocyte ROS, oxidized-AGTN, and AT levels. The ROS->AT->ATR1->ROS loop was driven by sickle erythropoiesis as it was reproduced when WT mice were transplanted with SCA hematopoiesis. Using SCA and WT mice with germline- and erythroid-specific ATR1-deficiency, we found that stress-erythropoiesis, but not steady-state-erythropoiesis, was critically dependent on erythroid AT->ATR1 signaling, which acted in harmony with increased erythropoietin signaling. Further, instead of the canonical AT->ATR1-> NADPH-oxidase->ROS signaling in steady-state erythropoiesis, AT->ATR1 signaling in stress-erythroid cells increased mitochondrial mass and dysfunctional mitochondria, which thereby increased ROS. SCA mice with erythroid-specific ATR1 deficiency had decreased RBC accumulation of dysfunctional mitochondria and decreased ROS, which reduced SCA-associated nephropathy. Overall, we demonstrated that AT->ATR1 signaling was essential for stress-erythropoiesis but led to increased dysfunctional mitochondria retention in mature RBCs, which generated ROS and perpetuated hyperangiotensinemia, resulting in end-organ damage.
Authors
Parul Rai, Swarnava Roy, Paritha Arumugam, Diamantis G. Konstantinidis, Sithara Raju Ponny, Marthe-Sandrine Eiymo Mwa Mpollo, Archana Shrestha, Theodosia A. Kalfa, Punam Malik
Abstract
Cutaneous radiation injury is an unintended consequence of radiotherapy for many common cancers and can progress to debilitating radiation-induced skin fibrosis (RISF). Existing radiation injury models do not fully capture the skin toxicities observed in patients, contributing to the lack of efficacious therapies to mitigate RISF. To address this, we developed an ex vivo human skin model that recapitulates the temporal radiation injury and RISF response. Human skin explants (N=12) subjected to ionizing radiation demonstrated DNA double-strand breaks and robust p53-driven transcriptional programming of cell cycle arrest, apoptosis, and senescence compared to non-irradiated controls. Irradiated skin also exhibited induction of pro-inflammatory cytokines, epithelial-mesenchymal transition, pro-fibrotic TGF-beta1 (TGFB1)-mediated signaling, and thickened collagen over time. P53 regulators murine double minute 2 (MDM2) and microRNA (miR)-34a were induced post-irradiation and may be leveraged to modulate injury response. Notably, RNA-sequencing of breast skin from mastectomy patients post-radiotherapy showed similar p53, inflammatory, and TGFB1 signatures as the ex vivo model, supporting its translational relevance. Together, this model provides a platform for identifying biomarkers and testing therapies to prevent or mitigate cutaneous radiation toxicities. Targeting the dynamic p53-driven pro-fibrotic radiation response represents a new therapeutic avenue to improve post-radiotherapy quality of life for cancer survivors.
Authors
Caroline Dodson, Sophie M. Bilik, Gabrielle DiBartolomeo, Hannah Pachalis, Lindsey G. Siegfried, Jordan A. K. Johnson, Seth R. Thaller, Irena Pastar, Marjana Tomic-Canic, Anthony J. Griswold, Rivka C. Stone
Abstract
Myotonic Dystrophy Type 1 (DM1) is caused by an expanded CTG repeat in the DMPK gene, resulting in mutant transcripts that form expanded CUG (CUGexp) RNA foci and sequester muscleblind-like (MBNL) RNA-binding proteins. DM1 is multisystemic with progressive worsening of disease manifestations in affected tissues. Disease progression is attributed to somatic expansion of the CTG repeats with age, resulting in production of CUGexp RNA with enhanced intrinsic toxicity due to increased MBNL sequestration. To determine the degree to which cardiac disease progression can occur independently of repeat expansion, we used a transgenic DM1 mouse model with inducible heart-specific expression of a stable, interrupted 960-CUG repeat RNA. Sustained CUGexp RNA expression caused progressive cardiac enlargement, contractile dysfunction, conduction delay, myocardial fibrosis, and reduced survival, while MBNL-dependent splicing defects remained static, consistent with the stable repeat length. We also determined the degree of reversibility after different periods of CUGexp RNA expression by shutting off the repeat-containing transgene. Suppression of CUGexp RNA expression rescued cardiac abnormalities, but reversibility declined with longer exposure to the toxic RNA. These findings demonstrate that prolonged expression of stable CUGexp RNA drives progressive cardiac pathology, revealing a mechanism of disease progression in DM1 in addition to somatic expansion.
Authors
Rong-Chi Hu, Mohammadreza Tabary, Xander H.T. Wehrens, Thomas A. Cooper
