Proliferative glomerulonephritis is a severe condition often leading to kidney failure. There is a significant lack of effective treatment for these disorders. Here, following the identification of a somatic PIK3CA gain-of-function mutation in podocytes of a patient, we demonstrate using multiple genetically engineered mouse models, single-cell RNA sequencing and spatial transcriptomics the crucial role played by this pathway for proliferative glomerulonephritis development by promoting podocyte proliferation, dedifferentiation and inflammation. Additionally, we show that alpelisib, a PI3Kα inhibitor, improves glomerular lesions and kidney function in different mouse models of proliferative glomerulonephritis and lupus nephritis by targeting podocytes. Surprisingly, we determined that pharmacological inhibition of PI3Kα affects B and T lymphocyte population in lupus nephritis mouse models with decrease in the production of proinflammatory cytokines, autoantibodies and glomerular complement deposition, which are all characteristic features of PI3K delta (PI3Kδ) inhibition, the primary PI3K isoform expressed in lymphocytes. Importantly, PI3Kα inhibition does not impact lymphocyte function under normal conditions. These findings were then confirmed in human lymphocytes isolated from patients with active lupus nephritis. In conclusion, we demonstrate the major role played by PI3Kα in proliferative glomerulonephritis and show that in this condition, alpelisib acts on both podocytes and the immune system.
Junna Yamaguchi, Pierre Isnard, Noémie Robil, Pierre de la Grange, Clément Hoguin, Alain Schmitt, Aurélie Hummel, Jérôme Mégret, Nicolas Goudin, Marine Luka, Mickaël M. Ménager, Cécile Masson, Mohammed Zarhrate, Christine Bôle-Feysot, Michalina Janiszewska, Kornelia Polyak, Julien Dairou, Sara Baldassari, Stéphanie Baulac, Christine Broissand, Christophe Legendre, Fabiola Terzi, Guillaume Canaud
Aberrant activation of RAS-MAPK signaling is common in cancer, and efforts to inhibit pathway components have yielded drugs with promising clinical activities. Unfortunately, treatment-provoked adaptive resistance mechanisms inevitably develop, limiting their therapeutic potential. As a central node essential for receptor tyrosine kinase mediated RAS activation, SHP2 has emerged as an attractive cancer target. Consequently, many SHP2 allosteric inhibitors are now in clinical testing. Here we discovered a previously unrecognized off-target effect associated with SHP2 allosteric inhibitors. We found that these inhibitors accumulate in the lysosome and block autophagic flux in a SHP2-independent manner. We showed that off-target autophagy inhibition by SHP2 allosteric inhibitors contributes to their anti-tumor activity. We also demonstrated that SHP2 allosteric inhibitors harboring this off-target activity not only suppress oncogenic RAS signaling but also overcome drug resistance such as MAPK rebound and protective autophagy in response to RAS-MAPK pathway blockage. Finally, we exemplified a therapeutic framework that harnesses both the on- and off-target activities of SHP2 allosteric inhibitors for improved treatment of mutant RAS driven and drug resistant malignancies such as pancreatic and colorectal cancers. Brief Summary: SHP2 allosteric inhibitors elicit off-target autophagy blockade that can be exploited for improved treatment of RAS-driven and drug-resistant cancers.
Yiming Miao, Yunpeng Bai, Jinmin Miao, Allison A. Murray, Jianping Lin, Jiajun Dong, Zihan Qu, Ruo-Yu Zhang, Quyen D. Nguyen, Shaomeng Wang, Jingmei Yu, Frederick Nguele Meke, Zhong-Yin Zhang
BACKGROUND. Clinical trials have suggested antitumor activity from PARP inhibition beyond homologous recombination deficiency (HRD). RNASEH2B loss is unrelated to HRD and preclinically sensitizes to PARP inhibition. The current study reports on RNASEH2B protein loss in advanced prostate cancer and its association with RB1 protein loss, clinical outcome and clonal dynamics during treatment with PARP inhibition in a prospective clinical trial. METHODS. Whole tumor biopsies from multiple cohorts of patients with advanced prostate cancer were interrogated using whole-exome sequencing (WES), RNA sequencing (bulk and single nucleus) and immunohistochemistry (IHC) for RNASEH2B and RB1. Biopsies from patients treated with olaparib in the TOPARP-A and TOPARP-B clinical trials were used to evaluate RNASEH2B clonal selection during olaparib treatment. RESULTS. Shallow co-deletion of RNASEH2B and adjacent RB1, co-located at chromosome 13q14, was common, deep co-deletion infrequent, and gene loss associated with lower mRNA expression. In castration-resistant PC (CRPC) biopsies, RNASEH2B and RB1 mRNA expression correlated, but single nucleus RNA sequencing indicated discordant loss of expression. IHC studies showed that loss of the two proteins often occurred independently, arguably due to stochastic second allele loss. Pre- and post-treatment metastatic CRPC (mCRPC) biopsy studies from BRCA1/2 wildtype tumors, treated on the TOPARP phase II trial, indicated that olaparib eradicates RNASEH2B-loss tumor subclones. CONCLUSION. PARP inhibition may benefit men suffering from mCRPC by eradicating tumor subclones with RNASEH2B loss. TRIAL REGISTRATION. Clinicaltrials.gov NCT01682772 FUNDING. AstraZeneca; Cancer Research UK; Medical Research Council; Cancer Research UK; Prostate Cancer UK; Movember Foundation; Prostate Cancer Foundation.
Juliet Carmichael, Ines Figueiredo, Bora Gurel, Nick Beije, Wei Yuan, Jan Rekowski, George Seed, Suzanne Carreira, Claudia Bertan, Maria de Los Dolores Fenor de la Maza, Khobe Chandran, Antje Neeb, Jon Welti, Lewis Gallagher, Denisa Bogdan, Mateus Crespo, Ruth Riisnaes, Ana Ferreira, Susana Miranda, Jinqiu Lu, Michael M. Shen, Emma Hall, Nuria Porta, Daniel Westaby, Christina Guo, Rafael Grochot, Christopher J. Lord, Joaquin Mateo, Adam Sharp, Johann de Bono
Cutaneous leishmaniasis caused by Leishmania parasites exhibits a wide range of clinical manifestations. Although parasites influence disease severity, cytolytic CD8 T cell responses mediate disease. While these responses originate in the lymph node, we found that expression of the cytolytic effector molecule granzyme B was restricted to lesional CD8 T cells in Leishmania-infected mice, suggesting that local cues within inflamed skin induced cytolytic function. Expression of Blimp-1 (Prdm1), a transcription factor necessary for cytolytic CD8 T cell differentiation, was driven by hypoxia within the inflamed skin. Hypoxia was further enhanced by the recruitment of neutrophils that consumed oxygen to produce reactive oxygen species and ultimately increased the hypoxic state and granzyme B expression in CD8 T cells. Importantly, lesions from cutaneous leishmaniasis patients exhibited hypoxia transcription signatures that correlated with the presence of neutrophils. Thus, targeting hypoxia-driven signals that support local differentiation of cytolytic CD8 T cells may improve the prognosis for patients with cutaneous leishmaniasis, as well as other inflammatory skin diseases where cytolytic CD8 T cells contribute to pathogenesis.
Erin A. Fowler, Camila Farias Amorim, Klauss Mostacada, Allison Yan, Laís Amorim Sacramento, Rae A. Stanco, Emily D.S. Hales, Aditi Varkey, Wenjing Zong, Gary D. Wu, Camila I. de Oliveira, Patrick L. Collins, Fernanda O. Novais
BACKGROUND. Predicting Immune-effector Cell Associated Neurotoxicity Syndrome (ICANS) in patients infused with Chimeric Antigen Receptor T cells (CAR-T) is still a conundrum. This complication, thought to be consequent to CAR-T cell activation, arises a few days after infusion, when circulating CAR-T cells are scarce and specific CAR-T cell-derived biomarkers are lacking. METHODS. Human CD19.CAR-T cells were generated to gain insight into CAR+ extracellular vesicle (CAR+EV) release upon target engagement. A prospective cohort of 100 B-cell lymphoma patients infused with approved CD19.CAR-T cell products (axi-cel, brexu-cel and tisa-cel) was assessed for plasma CAR+EVs as potential biomarkers of in vivo CD19.CAR-T cell activation and predictors of ICANS. Human induced pluripotent stem cells (iPSCs)-derived neural cells were used as a model for CAR+EV-induced neurotoxicity. RESULTS. In vitro, exosome-like CAR+EVs were released by CD19.CAR-T cells upon target engagement. In vivo, CAR+EVs were detectable as early as 1 hour in the plasma of patients. A concentration > 132.8 CAR+EVs/μl at hour +1 or > 224.5 CAR+EVs/μl at day +1 predicted ICANS in advance of 4 days, with a sensitivity up to 96.55% and a specificity up to 80.36%, outperforming other potential ICANS predictors. Enolase 2 (ENO2+) nanoparticles were released by iPSCs-derived neural cells upon CAR+EVs exposure and were increased in the plasma of ICANS patients. CONCLUSIONS. These results convey that plasma CAR+EVs are an immediate signal of CD19.CAR-T cell activation, are suitable predictors of neurotoxicity, and may be involved in ICANS pathogenesis. TRIAL REGISTRATION. NCT04892433, NCT05807789.
Gianluca Storci, Francesco De Felice, Francesca Ricci, Spartaco Santi, Daria Messelodi, Salvatore Nicola Bertuccio, Noemi Laprovitera, Michele Dicataldo, Lucrezia Rossini, Serena De Matteis, Beatrice Casadei, Francesca Vaglio, Margherita Ursi, Francesco Barbato, Marcello Roberto, Maria Guarino, Gian Maria Asioli, Mario Arpinati, Pietro Cortelli, Enrico Maffini, Enrica Tomassini, Marta Tassoni, Carola Cavallo, Francesco Iannotta, Maria Naddeo, Pier Luigi Tazzari, Elisa Dan, Cinzia Pellegrini, Serafina Guadagnuolo, Matteo Carella, Barbara Sinigaglia, Chiara Pirazzini, Caterina Severi, Paolo Garagnani, Katarzyna Malgorzata Kwiatkowska, Manuela Ferracin, Pier Luigi Zinzani, Massimiliano Bonafè, Francesca Bonifazi
Despite effective antiretroviral therapy (ART), persons living with HIV (PWH) harbor reservoirs of persistently infected CD4+ cells, which constitute a barrier to cure. Initiation of ART during acute infection reduces the size of the HIV reservoir, and we hypothesized that in addition, it would favor integration of proviruses in HIV-specific CD4+ T cells, while initiation of ART during chronic HIV infection would favor relatively more proviruses in herpesvirus-specific cells. We further hypothesized that proviruses in acute-ART-initiators would be integrated into antiviral genes, whereas integration sites in chronic-ART-initiators would favor genes associated with cell proliferation and exhaustion. We found the HIV DNA distribution across HIV-specific vs. herpesvirus-specific CD4+ T cells was as hypothesized. HIV integration sites (IS) in acute-ART-initiators were significantly enriched in gene sets controlling lipid metabolism and HIF-1α-mediated hypoxia, both metabolic pathways active in early HIV infection. Persistence of these infected cells during prolonged ART suggests a survival advantage. IS in chronic-ART-initiators were enriched in a gene set controlling EZH2 histone methylation; and methylation has been associated with diminished LTR transcription. These differences we found in antigen specificities and IS distributions within HIV-infected cells might be leveraged in designing cure strategies tailored to the timing of ART initiation.
Jaimy Joy, Ana L. Gervassi, Lennie Chen, Brent Kirshenbaum, Sheila Styrchak, Daisy Ko, Sherry McLaughlin, Danica Shao, Ewelina Kosmider, Paul T. Edlefsen, Janine Maenza, Ann C. Collier, James I. Mullins, Helen Horton, Lisa M. Frenkel
Primary lymphedema (PL), characterized by tissue swelling, fat accumulation and fibrosis, results from defective lymphatic vessels or valves caused by mutations in genes involved in development, maturation and function of the lymphatic vascular system. Pathogenic variants in various genes have been identified in about 30% of PL cases. By screening of a cohort of 755 individuals with PL, we identified two TIE1 (tyrosine kinase with immunoglobulin- and epidermal growth factor-like domains 1) missense variants and one truncating variant, all predicted to be pathogenic by bioinformatic algorithms. The TIE1 receptor, in complex with TIE2, binds angiopoietins to regulate the formation and remodelling of blood and lymphatic vessels. The premature stop codon mutant encoded an inactive truncated extracellular TIE1 fragment with decreased mRNA stability and the amino acid substitutions led to decreased TIE1 signaling activity. By reproducing the two missense variants in mouse Tie1 via CRISPR-Cas9, we showed that both cause edema and are lethal in homozygous mice. Thus, our results indicate that TIE1 loss-of-function variants can cause lymphatic dysfunction in patients. Together with our earlier demonstration that ANGPT2 loss-of-function mutations can also cause PL, our results emphasize the important role of the ANGPT2-TIE1 pathway in lymphatic function.
Pascal Brouillard, Aino Murtomäki, Veli-Matti Leppänen, Marko Hyytiäinen, Sandrine Mestre, Lucas Potier, Laurence M. Boon, Nicole Revencu, Arin K. Greene, Andrey Anisimov, Miia H. Salo, Reetta Hinttala, Lauri Eklund, Isabelle Quéré, Kari Alitalo, Miikka Vikkula
The surface receptor CD8α is present on 20-80% of human (but not mouse) NK cells, yet its function on NK cells remains poorly understood. CD8α expression on donor NK cells was associated with a lack of therapeutic responses for leukemia patients in prior studies, thus we hypothesized that CD8α may impact critical NK cell functions. Here, we discovered that CD8α- NK cells had improved control of leukemia in xenograft models, compared to CD8α+ NK cells, likely due to an enhanced capacity for proliferation. Unexpectedly, CD8α expression was induced on approximately 30% of previously CD8α- NK cells following IL-15 stimulation. These ‘induced’ CD8α+ (‘iCD8α+’) NK cells had the greatest proliferation, responses to IL-15 signaling, and metabolic activity, compared to those that sustained existing CD8α expression (‘sustained CD8α+) or those that remained CD8α- (‘persistent CD8α-‘). These iCD8α+ cells originated from an IL-15Rβ high NK cell population, with CD8α expression dependent on the transcription factor RUNX3. Moreover, CD8A CRISPR/Cas9 deletion resulted in enhanced responses through the activating receptor NKp30, possibly by modulating KIR inhibitory function. Thus, CD8α status identifies human NK cell capacity for IL-15-induced proliferation and metabolism in a time-dependent fashion and exhibits a suppressive effect on NK cell activating receptors.
Celia C. Cubitt, Pamela Wong, Hannah K. Dorando, Jennifer A. Foltz, Jennifer Tran, Lynne Marsala, Nancy D. Marin, Mark Foster, Timothy Schappe, Hijab Fatima, Michelle Becker-Hapak, Alice Y. Zhou, Kimberly Hwang, Miriam T. Jacobs, David A. Russler-Germain, Emily M. Mace, Melissa M. Berrien-Elliott, Jacqueline E. Payton, Todd A. Fehniger
Pathogenic variants in VCP cause multisystem proteinopathy (MSP), a disease characterized by multiple clinical phenotypes including inclusion body myopathy, Paget’s disease of the bone, and frontotemporal dementia (FTD). How such diverse phenotypes are driven by pathogenic VCP variants is not known. We found that these diseases exhibit a common pathologic feature, ubiquitinated intranuclear inclusions affecting myocytes, osteoclasts and neurons. Moreover, knock-in cell lines harboring MSP variants show a reduction in nuclear VCP. Given that MSP is associated with neuronal intranuclear inclusions comprised of TDP-43 protein, we developed a cellular model whereby proteostatic stress results in the formation of insoluble intranuclear TDP-43 aggregates. Consistent with a loss of nuclear VCP function, cells harboring MSP variants or cells treated with VCP inhibitor exhibited decreased clearance of insoluble intranuclear TDP-43 aggregates. Moreover, we identified four compounds that activate VCP primarily by increasing D2 ATPase activity whereby pharmacologic VCP activation appears to enhance clearance of insoluble intranuclear TDP-43 aggregate. Our findings suggest that VCP function is important for nuclear protein homeostasis, that impaired nuclear proteostasis may contribute to MSP, and that VCP activation may be potential therapeutic by virtue of enhancing the clearance of intranuclear protein aggregates.
Jessica M. Phan, Benjamin C. Creekmore, Aivi T. Nguyen, Darya D. Bershadskaya, Nabil F. Darwich, Carolyn N. Mann, Edward B. Lee
Intratumoral regulatory T cells (Tregs) are key mediators of cancer immunotherapy resistance, including anti-PD-(L)1 immune checkpoint blockade (ICB). The mechanisms driving Treg infiltration into the tumor microenvironment (TME) and the consequence on CD8+ T cell exhaustion remains elusive. Herein, we report that heat shock protein gp96 (GRP94) is indispensable for Treg tumor infiltration, primarily through gp96’s roles in chaperoning integrins. Among various gp96-dependent integrins, we found that only LFA-1 (αL integrin) but not αV, CD103 (αE) or β7 integrin was required for Treg tumor homing. Loss of Treg infiltration into the TME by genetically deleting gp96/LFA-1 potently induces rejection of multiple ICB-resistant murine cancer models in a CD8+ T cell-dependent manner without loss of self-tolerance. Moreover, gp96 deletion impeded Treg activation primarily by suppressing IL-2/STAT5 signaling, which also contributes to tumor regression. By competing for intratumoral IL-2, Tregs prevent activation of CD8+ tumor-infiltrating lymphocytes (TILs), drive TOX induction and induce bona fide CD8+ T cell exhaustion. By contrast, Treg ablation leads to striking CD8+ T cell activation without TOX induction, demonstrating clear uncoupling of the two processes. Our study reveals that the gp96/LFA-1 axis plays a fundamental role in Treg biology and suggests that Treg-specific gp96/LFA-1 targeting represents a valuable strategy for cancer immunotherapy without inflicting autoinflammatory conditions.
Lei Zhou, Maria Velegraki, Yi Wang, J K Mandula, Yuzhou Chang, Weiwei Liu, No-Joon Song, Hyunwoo Kwon, Tong Xiao, Chelsea Bolyard, Feng Hong, Gang Xin, Qin Ma, Mark P. Rubinstein, Haitao Wen, Zihai Li
No posts were found with this tag.