The suppression mechanism of Tregs remains an intensely investigated topic. As our focus has shifted toward a model centered on indirect inhibition of DCs, a universally applicable effector mechanism controlled by the transcription factor forkhead box P3 (Foxp3) expression has not been found. Here, we report that Foxp3 blocked the transcription of ER Ca2+-release channel ryanodine receptor 2 (RyR2). Reduced RyR2 shut down basal Ca2+ oscillation in Tregs, which reduced m-calpain activities that are needed for T cells to disengage from DCs, suggesting a persistent blockage of DC antigen presentation. RyR2 deficiency rendered the CD4+ T cell pool immune suppressive and caused it to behave in the same manner as Foxp3+ Tregs in viral infection, asthma, hypersensitivity, colitis, and tumor development. In the absence of Foxp3, Ryr2-deficient CD4+ T cells rescued the systemic autoimmunity associated with scurfy mice. Therefore, Foxp3-mediated Ca2+ signaling inhibition may be a central effector mechanism of Treg immune suppression.
Xiaobo Wang, Shuang Geng, Junchen Meng, Ning Kang, Xinyi Liu, Yanni Xu, Huiyun Lyu, Ying Xu, Xun Xu, Xinrong Song, Bin Zhang, Xin Wang, Nuerdida Nuerbulati, Ze Zhang, Di Zhai, Xin Mao, Ruya Sun, Xiaoting Wang, Ruiwu Wang, Jie Guo, S.R. Wayne Chen, Xuyu Zhou, Tie Xia, Hai Qi, Xiaoyu Hu, Yan Shi
Cell therapies such as tumor-infiltrating lymphocyte (TIL) therapy have shown promise in the treatment of patients with refractory solid tumors, with improvement in response rates and durability of responses nevertheless sought. To identify targets capable of enhancing the antitumor activity of T cell therapies, large-scale in vitro and in vivo clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 screens were performed, with the SOCS1 gene identified as a top T cell–enhancing target. In murine CD8+ T cell–therapy models, SOCS1 served as a critical checkpoint in restraining the accumulation of central memory T cells in lymphoid organs as well as intermediate (Texint) and effector (Texeff) exhausted T cell subsets derived from progenitor exhausted T cells (Texprog) in tumors. A comprehensive CRISPR tiling screen of the SOCS1-coding region identified sgRNAs targeting the SH2 domain of SOCS1 as the most potent, with an sgRNA with minimal off-target cut sites used to manufacture KSQ-001, an engineered TIL therapy with SOCS1 inactivated by CRISPR/Cas9. KSQ-001 possessed increased responsiveness to cytokine signals and enhanced in vivo antitumor function in mouse models. These data demonstrate the use of CRISPR/Cas9 screens in the rational design of T cell therapies.
Michael R. Schlabach, Sharon Lin, Zachary R. Collester, Christopher Wrocklage, Sol Shenker, Conor Calnan, Tianlei Xu, Hugh S. Gannon, Leila J. Williams, Frank Thompson, Paul R. Dunbar, Robert A. LaMothe, Tracy E. Garrett, Nicholas Colletti, Anja F. Hohmann, Noah J. Tubo, Caroline P. Bullock, Isabelle Le Mercier, Katri Sofjan, Jason J. Merkin, Sean Keegan, Gregory V. Kryukov, Caroline Dugopolski, Frank Stegmeier, Karrie Wong, Fiona A. Sharp, Louise Cadzow, Micah J. Benson
Activation of TGF-β signaling serves as an extrinsic resistance mechanism that limits the potential for radiotherapy. Bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) antagonizes TGF-β signaling and is implicated in cancer progression. However, the molecular mechanisms of BAMBI regulation in immune cells and its impact on antitumor immunity after radiation have not been established. Here, we show that ionizing radiation (IR) specifically reduces BAMBI expression in immunosuppressive myeloid-derived suppressor cells (MDSCs) in both murine models and humans. Mechanistically, YTH N6-methyladenosine RNA-binding protein F2 (YTHDF2) directly binds and degrades Bambi transcripts in an N6-methyladenosine–dependent (m6A-dependent) manner, and this relies on NF-κB signaling. BAMBI suppresses the tumor-infiltrating capacity and suppression function of MDSCs via inhibiting TGF-β signaling. Adeno-associated viral delivery of Bambi (AAV-Bambi) to the tumor microenvironment boosts the antitumor effects of radiotherapy and radioimmunotherapy combinations. Intriguingly, combination of AAV-Bambi and IR not only improves local tumor control, but also suppresses distant metastasis, further supporting its clinical translation potential. Our findings uncover a surprising role of BAMBI in myeloid cells, unveiling a potential therapeutic strategy for overcoming extrinsic radioresistance.
Liangliang Wang, Wei Si, Xianbin Yu, Andras Piffko, Xiaoyang Dou, Xingchen Ding, Jason Bugno, Kaiting Yang, Chuangyu Wen, Linda Zhang, Dapeng Chen, Xiaona Huang, Jiaai Wang, Ainhoa Arina, Sean Pitroda, Steven J. Chmura, Chuan He, Hua Laura Liang, Ralph Weichselbaum
Acute myeloid leukemia (AML) presents a pressing medical need in that it is largely resistant to standard chemotherapy as well as modern therapeutics such as targeted therapy and immunotherapy, including anti-PD therapy. We demonstrate that Programmed Death-1 Homolog (PD-1H), an immune co-inhibitory molecule is highly expressed in blasts from the bone marrow of AML patients, while normal myeloid cell subsets and T cells have the expression of PD-1H. In studies employing syngeneic and humanized AML mouse models, overexpression of PD-1H promoted the growth of AML cells, mainly by evading T cell-mediated immune responses. Importantly, ablation of AML cell surface PD-1H by antibody blockade or genetic targeting significantly inhibited AML progression by promoting T cell activity. In addition, the genetic deletion of PD-1H from host normal myeloid cells inhibited AML progression as well and the combination of PD-1H blockade with PD-1 blockade conferred a synergistic anti-leukemia effect. Our findings provide the basis for PD-1H as an attractive therapeutic target to treat human AML.
Tae Kon Kim, Xue Han, Qianni Hu, Esten N. Vandsemb, Carly M. Fielder, Junshik Hong, Kwang Woon Kim, Emily F. Mason, R. Skipper Plowman, Jun Wang, Qi Wang, Jian-Ping Zhang, Ti Badri, Miguel F. Sanmamed, Linghua Zheng, Tianxiang Zhang, Jude Alawa, Sang Won Lee, Amer M. Zeidan, Stephanie Halene, Manoj M. Pillai, Namrata S. Chandhok, Jun Lu, Mina L. Xu, Steven D. Gore, Lieping Chen
Sarcoidosis is a disease of unknown etiology in which granulomas form throughout the body and is typically treated with glucocorticoids, but there are no approved steroid-sparing alternatives. Here, we investigated the mechanism of granuloma formation using single-cell RNA-Seq in sarcoidosis patients. We observed that the percentages of triggering receptor expressed on myeloid cells 2–positive (TREM2-positive) macrophages expressing angiotensin-converting enzyme (ACE) and lysozyme, diagnostic makers of sarcoidosis, were increased in cutaneous sarcoidosis granulomas. Macrophages in the sarcoidosis lesion were hypermetabolic, especially in the pentose phosphate pathway (PPP). Expression of the PPP enzymes, such as fructose-1,6-bisphosphatase 1 (FBP1), was elevated in both systemic granuloma lesions and serum of sarcoidosis patients. Granuloma formation was attenuated by the PPP inhibitors in in vitro giant cell and in vivo murine granuloma models. These results suggest that the PPP may be a promising target for developing therapeutics for sarcoidosis.
Satoshi Nakamizo, Yuki Sugiura, Yoshihiro Ishida, Yoko Ueki, Satoru Yonekura, Hideaki Tanizaki, Hiroshi Date, Akihiko Yoshizawa, Teruasa Murata, Kenji Minatoya, Mikako Katagiri, Seitaro Nomura, Issei Komuro, Seishi Ogawa, Saeko Nakajima, Naotomo Kambe, Gyohei Egawa, Kenji Kabashima
The study investigates a mechanistic link if bacterial biofilm mediated host-pathogen interaction leads to immunological complications associated with breast implant illness (BII). Over 10 million women worldwide have breast implants. In recent years, women have described a constellation of immunological symptoms believed to be related to their breast implants. The study included 178 subjects divided in three cohorts. Eighty-six patients reported symptoms consistent with BII. Control group I (non-BII, N=55) included patients with breast implants without BII symptoms but went through explantation of the breast implant. Control group II (normal tissue, N=37) was comprised of women without an implant, whose breast tissue was removed as an unrelated clinically indicated surgical procedures. We report that periprosthetic breast tissue of BII had increased abundance of biofilm and biofilm-derived oxylipin, 10-HOME. S. epidermidis biofilm was observed to be higher in the BII group (73.33%) compared to non-BII group (16.67%, p=0.018) and the normal group (10%, p=0.036). The oxylipin was found to be immunogenic capable of polarizing naïve CD4+ T cells with a resulting Th1 subtype in vitro and in vivo. Consistently, an abundance of CD4+Th1 subtype was observed in the periprosthetic breast tissue and blood of BII subjects. Mice injected with 10-HOME also had increased Th1 subtype in blood akin to BII patients and demonstrated fatigue-like symptoms. The identification of an oxylipin-mediated mechanism of immune activation induced by local bacterial biofilm associated with BII provides insight into the possible pathogenesis of implant-associated immune symptoms of BII.
Imran Khan, Robert E. Minto, Christine Kelley-Patteson, Kanhaiya Singh, Lava Timsina, Lily J. Suh, Ethan Rinne, Bruce W. Van Natta, Colby R. Neumann, Ganesh Mohan, Mary Lester, R. Jason VonDerHaar, Rana German, Natascia Marino, Aladdin H. Hassanein, Gayle M. Gordillo, Mark H. Kaplan, Chandan K. Sen, Marshall E. Kadin, Mithun Sinha
AIOLOS, also known as IKZF3, is a transcription factor highly expressed in the lymphoid lineage and critical for lymphocyte differentiation and development. Here we report nine individuals from three unrelated families carrying AIOLOS variants Q402* or E82K leading to AIOLOS haploinsufficiency through different mechanisms of action. Nonsense mutant Q402* displayed abnormal DNA binding, pericentromeric targeting, post-transcriptional modification, and transcriptome regulation. Structurally, the mutant lacks the AIOLOS zinc finger (ZF) 5-6 dimerization domain, but is still able to homodimerize with WT AIOLOS and negatively regulate DNA binding through ZF1, a previously unrecognized function for this domain. Missense mutant E82K showed overall normal AIOLOS functions; however, by affecting a redefined AIOLOS protein stability domain, it also led to haploinsufficiency. AIOLOS haploinsufficiency patients showed hypogammaglobulinemia, recurrent infections, autoimmunity and allergy, but with incomplete clinical penetrance. Altogether, these data redefine AIOLOS structure-function relationship and expand the spectrum of AIOLOS-associated diseases.
Hye Sun Kuehn, Inga S. Sakovich, Julie E. Niemela, Agustin A. Gil Silva, Jennifer L. Stoddard, Ekaterina A. Polyakova, Ana Esteve-Sole, Svetlana N. Aleshkevich, Tatjana A. Uglova, Mikhail V. Belevtsev, Vladislav R. Vertelko, Tatsiana V. Shman, Aleksandra N. Kupchinskaya, Jolan E. Walter, Thomas A. Fleisher, Luigi D. Notarangelo, Xiao P. Peng, Ottavia Maria Delmonte, Svetlana O. Sharapova, Sergio D. Rosenzweig
BACKGROUND. Systemic administration of Adeno-associated virus (AAV) can trigger life-threatening inflammatory responses including thrombotic microangiopathy (TMA), acute kidney injury due to atypical hemolytic uremic syndrome (aHUS)-like complement activation, immune-mediated myocardial inflammation and hepatic toxicity. METHODS. We describe the kinetics of immune activation following systemic AAV serotype 9 (AAV9) administration in 38 subjects following two distinct prophylactic immunomodulation regimens. Group 1 received corticosteroids and Group 2 received rituximab plus sirolimus in addition to steroids to prevent anti-AAV antibody formation. RESULTS. Group 1 subjects had a rapid increase of immunoglobulin M (IgM) and immunoglobulin G (IgG). Increase in D-dimer, decline in platelet count and complement activation are indicative of TMA. All Group 1 subjects demonstrated activation of both classical and alternative complement pathways indicated by depletion of C4, elevated SC5b-9, Ba, and Bb antigens. Group 2 patients did not have a significant change in IgM or IgG and had minimal complement activation. CONCLUSIONS. This study demonstrates that TMA in the setting of AAV gene therapy is antibody dependent (classical pathway) and amplified by the alternative complement pathway. Critical time points and interventions are identified to allow for management of immune mediated events which impact the safety and efficacy of systemic gene therapy.
Stephanie M. Salabarria, Manuela Corti, Kirsten E. Coleman, Megan B. Wichman, Julie A. Berthy, Precilla D'Souza, Cynthia J. Tifft, Roland W. Herzog, Melissa E. Elder, Lawrence R. Shoemaker, Carmen Leon-Astudillo, Fatemeh Tavakkoli, David H. Kirn, Jonathan D. Schwartz, Barry J. Byrne
Although most CD8+ T-cells are equipped to kill infected or transformed cells, a subset may regulate immune responses and preserve self-tolerance. Here we describe a CD8 lineage that is instructed to differentiate into CD8 T regulatory cells (Treg) by a surprisingly restricted set of T-cell receptors (TCR) that recognize MHC-E (mouse Qa-1) and several dominant self-peptides. Recognition and elimination of pathogenic target cells that express these Qa-1self-peptide complexes selectively inhibits pathogenic antibody responses without generalized immune suppression. Immunization with synthetic agonist peptides that mobilize CD8 Treg in vivo efficiently inhibit anti-graft antibody responses and markedly prolong heart and kidney organ graft survival. Definition of TCR-dependent differentiation and target recognition by this lineage of CD8 Treg may open the way to new therapeutic approaches to inhibit pathogenic antibody responses.
Hye-Jung Kim, Hidetoshi Nakagawa, John Y. Choi, Xuchun Che, Andrew Divris, Qingshi Liu, Andrew E. Wight, Hengcheng Zhang, Anis Saad, Zhabiz Solhjou, Christa Deban, Jamil R. Azzi, Harvey Cantor
Even with the prolific clinical use of next-generation cancer therapeutics, many tumors remain unresponsive or become refractory to therapy, creating a medical need. In cancer, DCs are indispensable to T cell activation, so there is a restriction on cytotoxic T cell immunity if DCs are not present in sufficient numbers in the tumor and draining lymph nodes to uptake and present relevant cancer antigens. To address this bottleneck, we developed a Flt3L-based therapeutic named Alb-Flt3L that demonstrated superior pharmacokinetic properties compared to Flt3L, including significantly longer half-life, accumulation in tumor and lymph node, and cross-presenting DCs expansion following a single injection. We demonstrated that Alb-Flt3L, in combination with standard-of-care chemotherapy and radiation therapy, serves as an in situ vaccination strategy capable of engendering polyclonal tumor neoantigen-specific immunity spontaneously. In addition, Alb-Flt3L-mediated tumor control synergized with immune checkpoint blockade delivered as anti-PD-L1. The mechanism of action of Alb-Flt3L treatment revealed a dependency on Batf3, type-I-interferons, and plasmacytoid DCs. Finally, the ability of Alb-Flt3L to expand human DC was explored in humanized mice. We observed significant expansion of human cross-presenting DC subsets, supporting the notion that Alb-Flt3L could be used clinically to modulate human DC populations in future cancer therapeutic regimens.
Brandon Lam, Yu Jui Kung, John Lin, Ssu-Hsueh Tseng, Hsin-Fang Tu, Claire Huang, Brandon Lee, Esteban Velarde, Ya Chea Tsai, Rafael Villasmil, Sung Taek Park, Deyin Xing, Chien-Fu Hung, T.-C. Wu