Krüppel-like factor 2 (KLF2) is a transcription factor with significant roles in development, maturation, differentiation, and proliferation of several cell types. In immune cells, KLF2 regulates maturation and trafficking of lymphocytes and monocytes. KLF2 participates in regulation of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway. Although pulmonary arterial hypertension (PAH) related to KLF2 genetic variant has been suggested, genetic role of KLF2 associated with immune dysregulation has not been described. We identified a family whose members suffered from lymphopenia, autoimmunity, and malignancy. Whole exome sequencing revealed a KLF2 p.(Glu318Argfs*87) mutation disrupting the highly conserved zinc finger domain. We show a reduced amount of KLF2 protein, defective nuclear localization and altered protein-protein interactome. The phenotypically variable positive cases presented with B and T cell lymphopenia and abnormalities in B and T cell maturation including low naive T cell counts and low CD27+IgD-IgM- switched memory B cells. KLF2 target gene (CD62L) expression was affected. Although the percentage of (CD25+FOXP3+, CD25+CD127-) regulatory T cells (Treg) was high, the naive Treg cells (CD45RA+) were absent. Serum IgG1 levels were low and findings in one case were consistent with common variable immunodeficiency (CVID). Transcription of NF-κβ pathway genes and p65/RelA phosphorylation were not significantly affected. Inflammasome activity, transcription of genes related with JAK/STAT pathway and interferon signature were also comparable to controls. Evidence of PAH was not found. In conclusion, KLF2 variant may be associated with familial immune dysregulation. Although the KLF2 deficient family members in our study suffered from lymphopenia, autoimmunity or malignancy, additional study cohorts are required to confirm our observations.
Copyright © 2022 Pernaa, Keskitalo, Chowdhury, Nissinen, Glumoff, Keski-Filppula, Junttila, Eklund, Santaniemi, Siitonen, Seppänen, Vähäsalo, Varjosalo, Åström and Hautala.

CD73 is widely expressed on immune cells playing a critical role in immunomodulatory functions including cell adhesion and migration, as a costimulatory molecule for T cells and in production of adenosine. The function of CD73 expressed on B cells has not been fully characterized. Mupadolimab is an anti-human CD73 antibody that activates B cells. We evaluated the characteristics of this antibody and its effects on immune cells in vitro and in vivo.
Mupadolimab binding to CD73, inhibition of CD73 enzymatic activity, and effects on lymphocyte activation were evaluated in vitro by measuring changes in immunophenotype by flow cytometry. Cryogenic-transmission electron microscopy was used to determine epitope binding. Effects on human B cells in vivo were evaluated in immunodeficient NSG-SGM3 mice immunized with SARS-CoV-2 and influenza viral antigens. Safety and immune effects were evaluated in the completed dose escalation portion of a phase 1 trial conducted in patients with cancer.
Mupadolimab binds to a unique epitope on CD73POS B cells resulting in their activation and differentiation through B cell receptor signaling pathways. Mupadolimab induces expression of CD69, CD83, CD86 and MHC class II on B cells along with morphological transformation into plasmablasts and expression of CD27, CD38 and CD138. These effects are independent of adenosine. Mupadolimab binds to the N-terminal of CD73 in the closed position and competitively inhibits substrate binding. Mupadolimab enhanced antigen specific antibody response to SARS-CoV-2 spike protein and influenza hemagglutinin in humanized mouse models. Mupadolimab was evaluated as a monotherapy in a phase 1 trial (NCT03454451) in 34 patients with advanced cancer and demonstrated binding to CD73POS circulating cells and transient reduction in the number of B cells, with return of CD73NEG B cells with memory phenotype. No dose-limiting toxicities or changes in serum immunoglobulins were seen.
Mupadolimab activates B cells and stimulates the production of antigen specific antibodies. The effects in patients with cancer suggest that activated, CD69POS B cells redistribute to lymphoid tissues. Minor tumor regression was observed in several patients. These results support further investigation of mupadolimab as an immunotherapy for cancer and its potential use as a vaccine adjuvant.
NCT03454451.
© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.

h4>ABSTRACT/h4> COVID-19 is a global pandemic that has resulted in over 800,000 deaths. Robust humoral anti-viral immune responses have the potential to generate a diverse set of neutralizing antibodies to eliminate viruses and protect against re-infection, transmission, and the evolution of mutations that escape targeted therapeutics. CD73 is present on the majority of human B cells and a subset of T cells where it plays a role in lymphocyte activation and migration. CD73 also functions as an ectoenzyme that converts AMP into adenosine, which can be immunosuppressive. Here we report on CPI-006, a humanized FcγR binding-deficient IgG1 anti-CD73 antibody that blocks CD73 enzymatic activity and directly activates CD73 POS B cells, inducing differentiation into plasmablasts, immunoglobulin class switching, and antibody secretion independent of adenosine. Immunophenotypic analysis of peripheral blood from advanced cancer patients receiving CPI-006 revealed evidence of B cell activation, clonal expansion, and development of memory B cells. These immune effects suggested that CPI-006 may be effective at enhancing the magnitude, diversity, and duration of humoral and cellular responses to viruses such as SARS-CoV-2. We have therefore initiated a Phase 1, single-dose, dose-escalation trial in hospitalized patients with mild to moderate COVID-19. The objectives of this trial are to evaluate the safety of CPI-006 in COVID-19 patients and to determine effects of CPI-006 on anti-SARS-CoV-2 antibody responses and the development of memory B cell and T cells. Ten patients have been enrolled in the trial receiving doses of 0.3 mg/kg or 1.0 mg/kg. All evaluable patients had low pre-treatment serum levels of anti-viral antibodies to the SARS-CoV-2 trimeric spike protein and its receptor binding domain, independent of the duration of their COVID-19 related symptoms prior to enrollment. Anti-viral antibody responses were induced 7 days after CPI-006 treatment and titers continued to rise past Day 56. Increases in the frequency of memory B cells and effector/memory T cells were observed 28 days after treatment. These preliminary results suggest that CPI-006 activates B cells and may enhance and prolong anti-SARS-CoV-2 antibody responses in patients with COVID-19. This approach may be useful for treating COVID-19 or as an adjuvant to enhance the efficacy of vaccines.