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To design new CARs targeting hepatitis B virus (HBV), we isolated human monoclonal antibodies recognizing the HBV envelope proteins from single B cells of a patient with a resolved infection. HBV-specific memory B cells were isolated by incubating peripheral blood mononuclear cells with biotinylated hepatitis B surface antigen (HBsAg), followed by single-cell flow cytometry-based sorting of live, CD19 + IgG + HBsAg + cells. Amplification and sequencing of immunoglobulin genes from single memory B cells identified variable heavy and light chain sequences. Corresponding immunoglobulin chains were cloned into IgG1 expression vectors and expressed in mammalian cells. Two antibodies named 4D06 and 4D08 were found to be highly specific for HBsAg, recognized a conformational and a linear epitope, respectively, and showed broad reactivity and neutralization capacity against all major HBV genotypes. 4D06 and 4D08 variable chain fragments were cloned into a 2 nd generation CAR format with CD28 and CD3zeta intracellular signaling domains. The new CAR constructs displayed a high functional avidity when expressed on primary human T cells. CAR-grafted T cells proved to be polyfunctional regarding cytokine secretion and killed HBV-positive target cells. Interestingly, background activation of the 4D08-CAR recognizing a linear instead of a conformational epitope was consistently low. In a preclinical model of chronic HBV infection, murine T cells grafted with the 4D06 and the 4D08 CAR showed on target activity indicated by a transient increase in serum transaminases, and a lower number of HBV-positive hepatocytes in the mice treated. This study demonstrates an efficient and fast approach to identifying pathogen-specific monoclonal human antibodies from small donor cell numbers for the subsequent generation of new CARs., Competing Interests: UP is a co-founder, shareholder, and SCG Cell Therapy Pte Ltd board member. UP received personal fees from Abbott, Abbvie, Arbutus, Gilead, GSK, J&J, MSD, Roche, Sanofi, Sobi, and Vaccitech. SR was a founder, has served as an advisor, and has patents licensed to Juno Therapeutics; is a founder of and holds equity in Lyell Immunopharma; and has served on the advisory boards for Adaptive Biotechnologies and Nohla. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Schreiber, Dressler, Loffredo-Verde, Asen, Färber, Wang, Groll, Chakraborty, Kolbe, Kreer, Kosinska, Simon, Urban, Klein, Riddell and Protzer.)

Background & Aims: Current antiviral therapies control but rarely eliminate HBV, leaving chronic HBV carriers at risk of developing hepatocellular carcinoma (HCC). Lacking or dysfunctional virus-specific adaptive immunity prevents control of HBV and allows the virus to persist. Restoring antiviral T-cell immunity could lead to HBV elimination and cure of chronically infected patients., Methods: We constructed bispecific T-cell engager antibodies that are designed to induce antiviral immunity through simultaneous binding of HBV envelope proteins (HBVenv) on infected hepatocytes and CD3 or CD28 on T cells. T-cell engager antibodies were employed in co-cultures with healthy donor lymphocytes and HBV-infected target cells. Activation of the T-cell response was determined by detection of pro-inflammatory cytokines, effector function (by cytotoxicity) and antiviral effects. To study in vivo efficacy, immune-deficient mice were transplanted with HBVenv-positive and -negative hepatoma cells., Results: The 2 T-cell engager antibodies synergistically activated T cells to become polyfunctional effectors that in turn elicited potent antiviral effects by killing infected cells and in addition controlled HBV via non-cytolytic, cytokine-mediated antiviral mechanisms. In vivo in mice, the antibodies attracted T cells specifically to the tumors expressing HBVenv resulting in T-cell activation, tumor infiltration and reduction of tumor burden., Conclusion: This study demonstrates that the administration of HBVenv-targeting T-cell engager antibodies facilitates a robust T-cell redirection towards HBV-positive target cells and provides a feasible and promising approach for the treatment of chronic viral hepatitis and HBV-associated HCC., Lay Summary: T-cell engager antibodies are an interesting, novel therapeutic tool to restore immunity in patients with chronic hepatitis B. As bispecific antibodies, they bind envelope proteins on the surface of the hepatitis B virus (HBV) and CD3 or CD28 on T cells. This way, they induce a potent antiviral and cytotoxic T-cell response that leads to the elimination of HBV-positive cells. These bispecific T-cell engager antibodies are exciting therapeutic candidates for chronic hepatitis B and HBV-associated hepatocellular carcinoma., Competing Interests: Conflict of interest U.P., F.M.,F.B., G.M. and O.Q. are named as inventors on patents WO 2015/036606 held by HMGU and DKFZ and WO 2016/146702 held by HMGU, DKFZ and TUM. U.P. serves as ad hoc advisor for Roche, Gilead, GSK, Merck, Arbutus and Vir Biotech. U.P. is co-founder and share-holder of SCG Cell Therapy who licensed patents WO 2015/036606 and WO 2016/146702. The remaining authors do not disclose a conflict of interest. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2021 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background: Chronic hepatitis B develops more frequently in countries with high prevalence of helminth infections. The crosstalk between these 2 major liver-residing pathogens, Schistosoma mansoni and hepatitis B virus (HBV), is barely understood., Methods: We used state-of-the-art models for both acute and chronic HBV infection to study the pathogen-crosstalk during the different immune phases of schistosome infection., Results: Although liver pathology caused by schistosome infection was not affected by either acute or chronic HBV infection, S mansoni infection influenced HBV infection outcomes in a phase-dependent manner. Interferon (IFN)-γ secreting, HBV- and schistosome-specific CD8 T cells acted in synergy to reduce HBV-induced pathology during the TH1 phase and chronic phase of schistosomiasis. Consequently, HBV was completely rescued in IFN-γ-deficient or in TH2 phase coinfected mice demonstrating the key role of this cytokine. It is interesting to note that secondary helminth infection on the basis of persistent (chronic) HBV infection increased HBV-specific T-cell frequency and resulted in suppression of virus replication but failed to fully restore T-cell function and eliminate HBV., Conclusions: Thus, schistosome-induced IFN-γ had a prominent antiviral effect that outcompeted immunosuppressive effects of TH2 cytokines, whereas HBV coinfection did not alter schistosome pathogenicity., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Helicobacter pylori chronically colonizes the stomach and is strongly associated with gastric cancer. Its concomitant occurrence with helminths such as schistosomes has been linked to reduced cancer incidence, presumably due to suppression of H. pylori-associated pro-inflammatory responses. However, experimental evidence in support of such a causal link or the mutual interaction of both pathogens is lacking. We investigated the effects of co-infection during the different immune phases of S. mansoni infection. Surprisingly, co-infected mice had increased H. pylori gastric colonization during the interferon gamma (IFNγ) phase of schistosome infection but reduced infiltration of T cells in the stomach due to misdirection of antigen-experienced CXCR3 + T cells to the liver. Unexpectedly, H. pylori co-infection resulted in partial protection from schistosome-induced liver damage. Here, we demonstrate that an increase in fibrosis-protective IL-13Ra2 is associated with H. pylori infection. Thus, our study strongly points to an immunological interaction of anatomically isolated pathogens, eventually resulting in altered disease pathology., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Schistosomiasis is a nontransplacental helminth infection. Chronic infection during pregnancy suppresses allergic airway responses in offspring. We addressed the question whether in utero exposure to chronic schistosome infection (Reg phase) in mice affects B-cell and T-cell development. Therefore, we focused our analyses on T-cell differentiation capacity induced by epigenetic changes in promoter regions of signature cytokines in offspring. Here, we show that naïve T cells from offspring of schistosome infected female mice had a strong capacity to differentiate into T H 1 cells, whereas T H 2 differentiation was impaired. In accordance, reduced levels of histone acetylation of the IL-4 promoter regions were observed in naïve T cells. To conclude, our mouse model revealed distinct epigenetic changes within the naïve T-cell compartment affecting T H 2 and T H 1 cell differentiation in offspring of mothers with chronic helminth infection. These findings could eventually help understand how helminths alter T-cell driven immune responses induced by allergens, bacterial or viral infections, as well as vaccines., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Tendon's natural healing potential is extremely low and inefficient, with significant dysfunction and disability due to hypocellularity and hypovascularity of tendon tissues. The application of stem cells can aid in significantly enhanced repair of tendon rupture; therefore, the main aim of this study is to assess the potential of using periodontal ligament cells (PDL), usually obtained from patients undergoing orthodontic treatment, as a novel cell source for cell-based therapy for tendon injuries in a clinically relevant rat full-size Achilles tendon defect. In addition, the study compares the differences between the healing effects of Achilles tendon-derived cells (AT) versus PDL and, hence, comprises of four experimental groups, native tendon (NT), empty defect (ED), PDL and human AT (hAT). The tendon healing in each group was assessed in the late remodelling phase at 16 weeks after surgery using a combination of methods, including evaluation of gross morphological appearance; various histological and immunohistological stainings; and detailed analyses of cell morphometry. Based on these outcome measures, PDL cell-implanted tendons exhibited not only advanced tissue maturation, less ectopic fibrocartilage formation, more organised collagen fibres, tendon matrix expression corresponding to the final healing stage, and better cell-morphometry parameters when compared with the ED group, but were also very similar to the tendons treated with hAT-derived cells. Taken together, our study clearly demonstrates the feasibility of using PDL cells as a novel cell source for tendon repair and strongly recommends this cell type for the future development of innovative regenerative applications for treatment of different tendon or ligament pathologies.

Aim: Currently there is no effective approach to enhance tendon repair, hence we aimed to identify a suitable cell source for tendon engineering utilizing an established clinically relevant animal model for tendon injury., Materials & Methods: We compared, by in-depth histomorphometric evaluation, the regenerative potential of uncommitted human mesenchymal stem cells (hMSC) and Scleraxis (Scx)-programmed tendon progenitors (hMSC-Scx) in the healing of a full-size of rat Achilles tendon defect., Results: Our analyses clearly demonstrated that implantation of hMSC-Scx, in contrast to hMSC and empty defect, results in smaller diameters, negligible ectopic calcification and advanced cellular organization and matrix maturation in the injured tendons., Conclusion: Scaffold-free delivery of hMSC-Scx aids in enhanced repair in a clinically translatable Achilles tendon injury model.

The continual rise of asthma in industrialised countries stands in strong contrast to the situation in developing lands. According to the modified Hygiene Hypothesis, helminths play a major role in suppressing bystander immune responses to allergens, and both epidemiological and experimental studies suggest that the tropical parasitic trematode Schistosoma mansoni elicits such effects. The focus of this study was to investigate which developmental stages of schistosome infection confer suppression of allergic airway inflammation (AAI) using ovalbumin (OVA) as a model allergen. Moreover, we assessed the functional role and localization of infection-induced CD4(+)Foxp3(+) regulatory T cells (Treg) in mediating such suppressive effects. Therefore, AAI was elicited using OVA/adjuvant sensitizations with subsequent OVA aerosolic challenge and was induced during various stages of infection, as well as after successful anti-helminthic treatment with praziquantel. The role of Treg was determined by specifically depleting Treg in a genetically modified mouse model (DEREG) during schistosome infection. Alterations in AAI were determined by cell infiltration levels into the bronchial system, OVA-specific IgE and Th2 type responses, airway hyper-sensitivity and lung pathology. Our results demonstrate that schistosome infection leads to a suppression of OVA-induced AAI when mice are challenged during the patent phase of infection: production of eggs by fecund female worms. Moreover, this ameliorating effect does not persist after anti-helminthic treatment, and depletion of Treg reverts suppression, resulting in aggravated AAI responses. This is most likely due to a delayed reconstitution of Treg in infected-depleted animals which have strong ongoing immune responses. In summary, we conclude that schistosome-mediated suppression of AAI requires the presence of viable eggs and infection-driven Treg cells. These data provide evidence that helminth derived products could be incorporated into treatment strategies that specifically target suppression of immune responses in AAI by inducing Treg cells.