Therapeutics
Abstract
Mediator kinases CDK19 and CDK8, pleiotropic regulators of transcriptional reprogramming, are differentially regulated by androgen signaling but both kinases are upregulated in castration-resistant prostate cancer (CRPC). Genetic or pharmacological inhibition of CDK8 and CDK19 reverses the castration-resistant phenotype and restores the sensitivity of CRPC xenografts to androgen deprivation in vivo. Prolonged CDK8/19 inhibitor treatment combined with castration not only suppresses the growth of CRPC xenografts but also induces tumor regression and cures. Transcriptomic analysis revealed that Mediator kinase inhibition amplifies and modulates the effects of castration on gene expression, disrupting CRPC adaptation to androgen deprivation. Mediator kinase inactivation in tumor cells also affects stromal gene expression, indicating that Mediator kinase activity in CRPC molds the tumor microenvironment. The combination of castration and Mediator kinase inhibition downregulates the MYC pathway, and Mediator kinase inhibition suppresses a MYC-driven CRPC tumor model even without castration. CDK8/19 inhibitors show efficacy in patient-derived xenograft models of CRPC, and a gene signature of Mediator kinase activity correlates with tumor progression and overall survival in clinical samples of metastatic CRPC. These results indicate that Mediator kinases mediate androgen-independent in vivo growth of CRPC, supporting the development of CDK8/19 inhibitors for the treatment of this presently incurable disease.
Authors
Jing Li, Thomas A. Hilimire, Liu Yueying, Lili Wang, Jiaxin Liang, Balázs Győrffy, Vitali Sikirzhytski, Hao Ji, Li Zhang, Chen Cheng, Xiaokai Ding, Kendall R. Kerr, Charles E. Dowling, Alexander A. Chumanevich, Zachary T. Mack, Gary P. Schools, Chang-uk Lim, Leigh Ellis, Xiaolin Zi, Donald C. Porter, Eugenia V. Broude, Campbell McInnes, George Wilding, Michael B. Lilly, Igor B. Roninson, Mengqian Chen
Abstract
Despite widespread utilization of immunotherapy, challenge to treat immune-cold tumors needs to be resolved. Multiomic analyses and experimental validation identified the OTUD4-CD73 proteolytic axis as a promising target in treating immune-suppressive triple negative breast cancer (TNBC). Mechanistically, deubiquitylation of CD73 by OTUD4 counteracted its ubiquitylation by TRIM21, resulting in CD73 stabilization that inhibits tumor immune responses. We further demonstrated the importance of TGF-β signaling for orchestrating the OTUD4-CD73 proteolytic axis within tumor cells. Spatial transcriptomics profiling discovered spatially resolved features of interacting malignant and immune cells pertaining to expression levels of OTUD4 and CD73. In addition, ST80, a newly developed inhibitor, specifically disrupted proteolytic interaction between CD73 and OTUD4, leading to reinvigoration of cytotoxic CD8+ T cell activities. In preclinical models of TNBC, ST80 treatment sensitized refractory tumors to anti-PD-L1 therapy. Collectively, our findings uncover a novel strategy for targeting immunosuppressive OTUD4-CD73 proteolytic axis in treating immune-suppressive breast cancers with the inhibitor ST80.
Authors
Yueming Zhu, Anupam Banerjee, Ping Xie, Andrey A. Ivanov, Amad Uddin, Qiao Jiao, Junlong J. Chi, Lidan Zeng, Ji Young Lee, Yifan Xue, Xinghua Lu, Massimo Cristofanilli, William J. Gradishar, Curtis J. Henry, Theresa W. Gillespie, Manali Ajay Bhave, Kevin Kalinsky, Haian Fu, Ivet Bahar, Bin Zhang, Yong Wan
Abstract
Chimeric antigen receptor (CAR) designs that incorporate pharmacologic control are desirable, however designs suitable for clinical translation are needed. We designed a fully human, rapamycin-regulated, drug product for targeting CD33+ tumors called dimerization agent regulated immunoreceptor complex (DARIC33). T cell products demonstrated target specific and rapamycin-dependent cytokine release, transcriptional responses, cytotoxicity, and in vivo antileukemic activity in the presence of as little as 1nM rapamycin. Rapamycin withdrawal paused DARIC33-stimulated T cell effector functions, which were restored following re-exposure to rapamycin, demonstrating reversible effector function control. While rapamycin-regulated DARIC33 T cells were highly sensitive to target antigen, CD34+ stem cell colony forming capacity was not impacted. We benchmarked DARIC33 potency relative to CD19 CAR T cells to estimate a T cell dose for clinical testing. In addition, we integrated in vitro and preclinical in vivo drug concentration thresholds for OFF-ON state transitions, as well as murine and human rapamycin pharmacokinetics, to estimate a clinically applicable rapamycin dosing schedule. A phase 1 DARIC33 trial has been initiated (PLAT-08, NCT05105152), with initial evidence of rapamycin-regulated T cell activation and anti-tumor impact. Our findings provide evidence that the DARIC platform exhibits sensitive regulation and potency needed for clinical application to other important immunotherapy targets.
Authors
Jacob Appelbaum, April E. Price, Kaori Oda, Joy Zhang, Wai-Hang Leung, Giacomo Tampella, Dong Xia, Pauline P.L. So, Sarah K. Hilton, Claudya Evandy, Semanti Sarkar, Unja Martin, Anne-Rachel Krostag, Marissa Leonardi, Daniel E. Zak, Rachael Logan, Paula Lewis, Secil Franke-Welch, Njabulo Ngwenyama, Michael Fitzgerald, Niklas Tulberg, Stephanie Rawlings-Rhea, Rebecca A. Gardner, Kyle Jones, Angelica Sanabria, William Crago, John Timmer, Andrew Hollands, Brendan Eckelman, Sanela Bilic, Jim Woodworth, Adam Lamble, Philip D. Gregory, Jordan Jarjour, Mark Pogson, Joshua A. Gustafson, Alexander Astrakhan, Michael C. Jensen
Abstract
Neurofibromatosis Type 1 (NF1) is caused by mutations in the NF1 gene that encodes neurofibromin, a RAS GTPase-Activating Protein. Inactivating NF1 mutations cause hyperactivation of RAS-mediated signaling, resulting in development of multiple neoplasms, including Malignant Peripheral Nerve Sheath Tumors (MPNSTs). MPNSTs are an aggressive tumor and the main cause of mortality in NF1 patients. MPNSTs are difficult to resect and refractory to chemo- and radiotherapy, and no molecular therapies currently exist. Immune Checkpoint Blockade (ICB) is an approach to treat inoperable, undruggable cancers like MPNST, but successful outcomes require an immune cell-rich tumor microenvironment (TME). While MPNSTs are non-inflamed “cold” tumors, here, we turned MPNSTs into T cell-inflamed “hot” tumors by activating “stimulator of interferon genes” (STING) signaling. Mouse genetic and human xenograft MPNST models treated with STING agonist plus ICB exhibited growth delay via increased apoptotic cell death. This strategy offers a potential treatment regimen for MPNST.
Authors
Bandarigoda N. Somatilaka, Laasya Madana, Ali Sadek, Zhiguo Chen, Sanjay Chandrasekaran, Renee M. McKay, Lu Q. Le
Abstract
Macrophage immune checkpoint inhibitors, such as anti-CD47 antibodies, show promise in clinical trials for solid and hematologic malignancies. However, the best strategies to use these therapies remain unknown, and ongoing studies suggest they may be most effective when used in combination with other anticancer agents. Here, we developed a novel screening platform to identify drugs that render lung cancer cells more vulnerable to macrophage attack, and we identified therapeutic synergy exists between genotype-directed therapies and anti-CD47 antibodies. In validation studies, we found the combination of genotype-directed therapies and CD47 blockade elicited robust phagocytosis and eliminated persister cells in vitro and maximized anti-tumor responses in vivo. Importantly, these findings broadly applied to lung cancers with various RTK/MAPK pathway alterations—including EGFR mutations, ALK fusions, or KRASG12C mutations. We observed downregulation of β2-microglobulin and CD73 as molecular mechanisms contributing to enhanced sensitivity to macrophage attack. Our findings demonstrate that dual inhibition of the RTK/MAPK pathway and the CD47/SIRPa axis is a promising immunotherapeutic strategy. Our study provides strong rationale for testing this therapeutic combination in patients with lung cancers bearing driver mutations.
Authors
Kyle Vaccaro, Juliet Allen, Troy W. Whitfield, Asaf Maoz, Sarah Reeves, José Velarde, Dian Yang, Anna Meglan, Juliano Ribeiro, Jasmine Blandin, Nicole Phan, George W. Bell, Aaron Hata, Kipp Weiskopf
Abstract
Craniofacial anomalies, especially midline facial defects, are among the most common birth defects in patients associated with increased mortality or require lifelong treatment. During mammalian embryogenesis, specific instructions arising at genetic, signaling, and metabolic levels are important for stem cell behaviors and fate determination, but how these functionally relevant mechanisms are coordinated to regulate craniofacial morphogenesis remain unknown. Here, we report that BMP signaling in cranial neural crest cells (CNCCs) is critical for glycolytic lactate production and subsequent epigenetic histone lactylation, thereby dictating craniofacial morphogenesis. Elevated BMP signaling in CNCCs through constitutively activated ACVR1 (ca-ACVR1) suppressed glycolytic activity and blocked lactate production via a p53-dependent process that resulted in severe midline facial defects. By modulating epigenetic remodeling, BMP signaling-dependent lactate generation drived histone lactylation levels to alter essential genes of Pdgfra thus regulating CNCC behavior in vitro as well as in vivo. These findings define an axis wherein the BMP signaling controls a metabolic-epigenetic cascade to direct craniofacial morphogenesis, thus providing a conceptual framework for understanding the interaction between genetic and metabolic cues operative during embryonic development. These findings indicate potential preventive strategies of congenital craniofacial birth defects via modulating metabolic-driven histone lactylation.
Authors
Jingwen Yang, Lingxin Zhu, Haichun Pan, Hiroki Ueharu, Masako Toda, Qian Yang, Shawn A. Hallett, Lorin E. Olson, Yuji Mishina
Abstract
Development of effective strategies to manage the inevitable acquired resistance to osimertinib, an approved 3rd generation EGFR inhibitor for the treatment of EGFR mutant (EGFRm) non-small cell lung cancer (NSCLC), is urgently needed. This study reported that the DNA topoisomerase II (Topo II) inhibitors, doxorubicin and etoposide (VP-16) synergistically decreased cell survival with enhanced induction of DNA damage and apoptosis in osimertinib-resistant cells, suppressed the growth of osimertinib-resistant tumors, and delayed the emergence of osimertinib acquired resistance. Mechanistically, osimertinib decreased Topo IIα levels in EGFRm NSCLC cells by facilitating FBXW7-mediated proteasomal degradation, resulting in induction of DNA damage; these effects were lost in osimertinib-resistant cell lines possessing elevated levels of Topo IIα. Topo IIα elevation was also detected in the majority of EGFRm NSCLC tissues relapsed from EGFR-TKI treatment. Enforced expression of an ectopic TOP2A gene in sensitive EGFRm NSCLC cells conferred resistance to osimertinib, whereas knockdown of TOP2A in osimertinib-resistant cell lines restored their response to undergo osimertinib-induced DNA damage and apoptosis. Together, these results reveal an essential role of Topo IIα inhibition in mediating the therapeutic efficacy of osimertinib against EGFRm NSCLC, providing scientific rationale for targeting Topo II to manage acquired resistance to osimertinib.
Authors
Zhen Chen, Karin A. Vallega, Dongsheng Wang, Zihan Quan, Songqing Fan, Qiming Wang, Ticiana Leal, Suresh S. Ramalingam, Shi-Yong Sun
Abstract
Nicotinamide adenine dinucleotide (NAD) is essential for embryonic development. To date, biallelic loss-of-function variants in 3 genes encoding nonredundant enzymes of the NAD de novo synthesis pathway — KYNU, HAAO, and NADSYN1 — have been identified in humans with congenital malformations defined as congenital NAD deficiency disorder (CNDD). Here, we identified 13 further individuals with biallelic NADSYN1 variants predicted to be damaging, and phenotypes ranging from multiple severe malformations to the complete absence of malformation. Enzymatic assessment of variant deleteriousness in vitro revealed protein domain–specific perturbation, complemented by protein structure modeling in silico. We reproduced NADSYN1-dependent CNDD in mice and assessed various maternal NAD precursor supplementation strategies to prevent adverse pregnancy outcomes. While for Nadsyn1+/– mothers, any B3 vitamer was suitable to raise NAD, preventing embryo loss and malformation, Nadsyn1–/– mothers required supplementation with amidated NAD precursors (nicotinamide or nicotinamide mononucleotide) bypassing their metabolic block. The circulatory NAD metabolome in mice and humans before and after NAD precursor supplementation revealed a consistent metabolic signature with utility for patient identification. Our data collectively improve clinical diagnostics of NADSYN1-dependent CNDD, provide guidance for the therapeutic prevention of CNDD, and suggest an ongoing need to maintain NAD levels via amidated NAD precursor supplementation after birth.
Authors
Justin O. Szot, Hartmut Cuny, Ella M.M.A. Martin, Delicia Z. Sheng, Kavitha Iyer, Stephanie Portelli, Vivien Nguyen, Jessica M. Gereis, Dimuthu Alankarage, David Chitayat, Karen Chong, Ingrid M. Wentzensen, Catherine Vincent-Delormé, Alban Lermine, Emma Burkitt-Wright, Weizhen Ji, Lauren Jeffries, Lynn S. Pais, Tiong Y. Tan, James Pitt, Cheryl A. Wise, Helen Wright, Israel D. Andrews, Brianna Pruniski, Theresa A. Grebe, Nicole Corsten-Janssen, Katelijne Bouman, Cathryn Poulton, Supraja Prakash, Boris Keren, Natasha J. Brown, Matthew F. Hunter, Oliver Heath, Saquib A. Lakhani, John H. McDermott, David B. Ascher, Gavin Chapman, Kayleigh Bozon, Sally L. Dunwoodie
Abstract
Diffuse midline glioma (DMG), including tumors diagnosed in the brainstem (diffuse intrinsic pontine glioma – DIPG), are uniformly fatal brain tumors that lack effective treatment. Analysis of CRISPR-Cas9 loss-of-function gene deletion screens identified PIK3CA and MTOR as targetable molecular dependencies across DIPG patient models, highlighting the therapeutic potential of the blood-brain barrier penetrant PI3K/Akt/mTOR inhibitor, paxalisib. At the human equivalent maximum tolerated dose, mice treated with paxalisib experienced systemic glucose feedback and increased insulin levels commensurate with patients using PI3K inhibitors. To exploit genetic dependence and overcome resistance whilst maintaining compliance and therapeutic benefit, we combined paxalisib with the anti-hyperglycemic drug, metformin. Metformin restored glucose homeostasis and decreased phosphorylation of the insulin receptor in vivo, a common mechanism of PI3K-inhibitor resistance, extending survival of orthotopic models. DIPG models treated with paxalisib increased calcium-activated PKC signaling. The brain penetrant PKC inhibitor enzastaurin in combination with paxalisib, synergistically extended the survival of multiple orthotopic patient-derived and immunocompetent syngeneic allograft models; benefits potentiated in combination with metformin and standard-of-care radiotherapy. Therapeutic adaptation was assessed using spatial transcriptomics and ATAC-sequencing, identifying changes in myelination and tumor immune microenvironment crosstalk. Together, we have identified a clinically relevant DIPG therapeutic combinatorial approach.
Authors
Ryan J. Duchatel, Evangeline R. Jackson, Sarah G. Parackal, Dylan Kiltschewskij, Izac J. Findlay, Abdul Mannan, Dilana E. Staudt, Bryce C. Thomas, Zacary P. Germon, Sandra Laternser, Padraic S. Kearney, M. Fairuz B. Jamaluddin, Alicia M. Douglas, Tyrone S. Beitaki, Holly P. McEwen, Mika L. Persson, Emily A. Hocke, Vaibhav Jain, Michael Aksu, Elizabeth E. Manning, Heather C. Murray, Nicole M. Verrills, Claire Xin Sun, Paul Daniel, Ricardo E. Vilain, David A. Skerrett-Byrne, Brett Nixon, Susan Hua, Charles E. de Bock, Yolanda Colino-Sanguino, Fatima Valdes-Mora, Maria Tsoli, David S. Ziegler, Murray J. Cairns, Eric H. Raabe, Nicholas A. Vitanza, Esther Hulleman, Timothy N. Phoenix, Carl Koschmann, Frank Alvaro, Christopher V. Dayas, Christopher L. Tinkle, Helen Wheeler, James R. Whittle, David D. Eisenstat, Ron Firestein, Sabine Mueller, Santosh Valvi, Jordan R. Hansford, David M. Ashley, Simon G. Gregory, Lindsay B. Kilburn, Javad Nazarian, Jason E. Cain, Matthew D. Dun
Abstract
Converging studies demonstrate the dysfunction of the dopaminergic neurons following chronic opioid administration. However, the therapeutic strategies targeting opioid-responsive dopaminergic ensembles that contribute to the development of opioid withdrawal remain to be elucidated. Here, we used the neuronal activity-dependent Tet-Off system to label dopaminergic ensembles in response to initial morphine exposure (Mor-Ens) in the ventral tegmental area (VTA). Fiber optic photometry recording and transcriptome analysis revealed downregulated spontaneous activity, dysregulated mitochondrial respiratory, ultrastructure, and oxidoreductase signal pathways after chronic morphine administration in these dopaminergic ensembles. Mitochondrial fragmentation and the decreased mitochondrial fusion gene mitofusin 1 (Mfn1) were found in these ensembles after prolonged opioid withdrawal. Restoration of Mfn1 in the dopaminergic Mor-Ens attenuated excessive oxidative stress and the development of opioid withdrawal. Administration of Mdivi-1, a mitochondrial fission inhibitor, ameliorated the mitochondrial fragmentation and maladaptation of the neuronal plasticity in these Mor-Ens, accompanied by attenuated development of opioid withdrawal after chronic morphine administration, without affecting the analgesic effect of morphine. These findings highlighted the plastic architecture of mitochondria as a potential therapeutic target for opioid analgesic-induced substance use disorders.
Authors
Changyou Jiang, Han Huang, Xiao Yang, Qiumin Le, Xing Liu, Lan Ma, Feifei Wang
No posts were found with this tag.
Copyright © 2025 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)