A novel ACE2 decoy for both neutralization of SARS-CoV-2 variants and killing of infected cells

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to millions of infections and deaths worldwide. As this virus evolves rapidly, there is a high need for treatment options that can win the race against new emerging variants of concern. Here, we describe a novel immunotherapeutic drug based on the SARS-CoV-2 entry receptor ACE2 and provide experimental evidence that it cannot only be used for (i) neutralization of SARS-CoV-2 in vitro and in SARS-CoV-2-infected animal models but also for (ii) clearance of virus-infected cells. For the latter purpose, we equipped the ACE2 decoy with an epitope tag. Thereby, we converted it to an adapter molecule, which we successfully applied in the modular platforms UniMAB and UniCAR for retargeting of either unmodified or universal chimeric antigen receptor-modified immune effector cells. Our results pave the way for a clinical application of this novel ACE2 decoy, which will clearly improve COVID-19 treatment

A Small Step, a Giant Leap: Somatic Hypermutation of a Single Amino Acid Leads to Anti-La Autoreactivity

The anti-La mab 312B, which was established by hybridoma technology from human-La transgenic mice after adoptive transfer of anti-human La T cells, immunoprecipitates both native eukaryotic human and murine La protein. Therefore, it represents a true anti-La autoantibody. During maturation, the anti-La mab 312B acquired somatic hypermutations (SHMs) which resulted in the replacement of four aa in the complementarity determining regions (CDR) and seven aa in the framework regions. The recombinant derivative of the anti-La mab 312B in which all the SHMs were corrected to the germline sequence failed to recognize the La antigen. We therefore wanted to learn which SHM(s) is (are) responsible for anti-La autoreactivity. Humanization of the 312B ab by grafting its CDR regions to a human Ig backbone confirms that the CDR sequences are mainly responsible for anti-La autoreactivity. Finally, we identified that a single amino acid replacement (D > Y) in the germline sequence of the CDR3 region of the heavy chain of the anti-La mab 312B is sufficient for anti-La autoreactivity

T Cell Mediated Conversion of a Non-Anti-La Reactive B Cell to an Autoreactive Anti-La B Cell by Somatic Hypermutation

Since the first description of nuclear autoantigens in the late 1960s and early 1970s, researchers, including ourselves, have found it difficult to establish monoclonal antibodies (mabs) against nuclear antigens, including the La/SS-B (Sjögrens’ syndrome associated antigen B) autoantigen. To date, only a few anti-La mabs have been derived by conventional hybridoma technology; however, those anti-La mabs were not bona fide autoantibodies as they recognize either human La specific, cryptic, or post-translationally modified epitopes which are not accessible on native mouse La protein. Herein, we present a series of novel murine anti-La mabs including truly autoreactive ones. These mabs were elicited from a human La transgenic animal through adoptive transfer of T cells from non-transgenic mice immunized with human La antigen. Detailed epitope and paratope analyses experimentally confirm the hypothesis that somatic hypermutations that occur during T cell dependent maturation can lead to autoreactivity to the nuclear La/SS-B autoantigen

Validation of CD98hc as a Therapeutic Target for a Combination of Radiation and Immunotherapies in Head and Neck Squamous Cell Carcinoma

Most patients with head and neck squamous cell carcinomas (HNSCC) are diagnosed at a locally advanced stage and show heterogeneous treatment responses. Low SLC3A2 (solute carrier family 3 member 2) mRNA and protein (CD98hc) expression levels are associated with higher locoregional control in HNSCC patients treated with primary radiochemotherapy or postoperative radiochemotherapy, suggesting that CD98hc could be a target for HNSCC radiosensitization. One of the targeted strategies for tumor radiosensitization is precision immunotherapy, e.g., the use of chimeric antigen receptor (CAR) T cells. This study aimed to define the potential clinical value of new treatment approaches combining conventional radiotherapy with CD98hc-targeted immunotherapy. To address this question, we analyzed the antitumor activity of the combination of fractionated irradiation and switchable universal CAR (UniCAR) system against radioresistant HNSCC cells in 3D culture. CD98hc-redirected UniCAR T cells showed the ability to destroy radioresistant HNSCC spheroids. Also, the infiltration rate of the UniCAR T cells was enhanced in the presence of the CD98hc target module. Furthermore, sequential treatment with fractionated irradiation followed by CD98hc-redirected UniCAR T treatment showed a synergistic effect. Taken together, our obtained data underline the improved antitumor effect of the combination of radiotherapy with CD98hc-targeted immunotherapy. Such a combination presents an attractive approach for the treatment of high-risk HNSCC patients

Targeting Acute Myeloid Leukemia Using the RevCAR Platform: A Programmable, Switchable and Combinatorial Strategy

Clinical translation of novel immunotherapeutic strategies such as chimeric antigen receptor (CAR) T-cells in acute myeloid leukemia (AML) is still at an early stage. Major challenges include immune escape and disease relapse demanding for further improvements in CAR design. To overcome such hurdles, we have invented the switchable, flexible and programmable adaptor Reverse (Rev) CAR platform. This consists of T-cells engineered with RevCARs that are primarily inactive as they express an extracellular short peptide epitope incapable of recognizing surface antigens. RevCAR T-cells can be redirected to tumor antigens and controlled by bispecific antibodies cross-linking RevCAR T- and tumor cells resulting in tumor lysis. Remarkably, the RevCAR platform enables combinatorial tumor targeting following Boolean logic gates. We herein show for the first time the applicability of the RevCAR platform to target myeloid malignancies like AML. Applying in vitro and in vivo models, we have proven that AML cell lines as well as patient-derived AML blasts were efficiently killed by redirected RevCAR T-cells targeting CD33 and CD123 in a flexible manner. Furthermore, by targeting both antigens, a Boolean AND gate logic targeting could be achieved using the RevCAR platform. These accomplishments pave the way towards an improved and personalized immunotherapy for AML patients

UniCAR T cell immunotherapy enables efficient elimination of radioresistant cancer cells

Induction or selection of radioresistant cancer (stem) cells following standard radiotherapy is presumably one of the major causes for recurrence of metastatic disease. One possibility to prevent tumor relapse is the application of targeted immunotherapies including, e.g., chimeric antigen receptor (CAR) T cells. In light of long-term remissions, it is highly relevant to clarify whether radioresistant cancer cells are susceptible to CAR T cell-mediated killing. To answer this question, we evaluated the anti-tumor activity of the switchable universal chimeric antigen receptor (UniCAR) system against highly radioresistant head and neck squamous cell carcinoma cells both in vitro and in vivo. Following specific UniCAR T cell engagement via EGFR or CD98 target modules, T cell effector mechanisms were induced including secretion of pro-inflammatory cytokines, up-regulation of granzyme B and perforin, as well as T cell proliferation. CD98- or EGFR-redirected UniCAR T cells further possess the capability to efficiently lyse radioresistant tumor cells. Observed anti-tumor effects were comparable to those against the radiosensitive parental cell lines. Finally, redirected UniCAR T cells significantly inhibited the growth of radioresistant cancer cells in immunodeficient mice. Taken together, our obtained data underline that the UniCAR system is able to overcome radioresistance. Thus, it represents an attractive technology for the development of combined radioimmunotherapeutic approaches that might improve the outcome of patients with metastatic radioresistant tumor diseases

Pathogen-induced tissue-resident memory T(H)17 (T(RM)17) cells amplify autoimmune kidney disease

Although it is well established that microbial infections predispose to autoimmune diseases, the underlying mechanisms remain poorly understood. After infection, tissue-resident memory T (T-RM) cells persist in peripheral organs and provide immune protection against reinfection. However, whether T-RM cells participate in responses unrelated to the primary infection, such as autoimmune inflammation, is unknown. By using high-dimensional single-cell analysis, we identified CD4(+) T-RM cells with a T(H)17 signature (termed T(RM)17 cells) in kidneys of patients with ANCA-associated glomerulonephritis. Experimental models demonstrated that renal T(RM)17 cells were induced by pathogens infecting the kidney, such as Staphylococcus aureus, Candida albicans, and uropathogenic Escherichia coli, and persisted after the clearance of infections. Upon induction of experimental glomerulonephritis, these kidney T(RM)17 cells rapidly responded to local proinflammatory cytokines by producing IL-17A and thereby exacerbate renal pathology. Thus, our data show that pathogen-induced T(RM)17 cells have a previously unrecognized function in aggravating autoimmune disease.erman Research Foundation (DFG) SFB1192 SFB1286 SFBTR57 Deutsche Nierenstiftung Deutsche Gesellschaft fur Nephrologie Werner Otto Stiftung eMed Consortia "Fibromap" from the Federal Ministry of Education and Research Comision Nacional de Investigacion Cientifica y Tecnologica (CONICYT) AFB 170004 Conicyt/FONDEQUIP/EQM140016 Else Kroner-Fresenius Foundatio

Pseudorapidity densities of charged particles with transverse momentum thresholds in pp collisions at √ s = 5.02 and 13 TeV

The pseudorapidity density of charged particles with minimum transverse momentum (pT) thresholds of 0.15, 0.5, 1, and 2 GeV/c is measured in pp collisions at the center of mass energies of √s=5.02 and 13 TeV with the ALICE detector. The study is carried out for inelastic collisions with at least one primary charged particle having a pseudorapidity (η) within 0.8pT larger than the corresponding threshold. In addition, measurements without pT-thresholds are performed for inelastic and nonsingle-diffractive events as well as for inelastic events with at least one charged particle having |η|2GeV/c), highlighting the importance of such measurements for tuning event generators. The new measurements agree within uncertainties with results from the ATLAS and CMS experiments obtained at √s=13TeV.