Somekh and colleagues identify CD137, a member of the tumor necrosis factor superfamily, as a novel cause of immunodeficiency associated with a risk of autoimmunity and lymphoid malignancy.

Aberrant activation of Wnt/β-catenin signaling occurs frequently in cancer. However, therapeutic targeting of this pathway is complicated by the role of Wnt in stem cell maintenance and tissue homeostasis. Here, we evaluated antibodies blocking 6 of the 10 human Wnt/Frizzled (FZD) receptors as potential therapeutics. Crystal structures revealed a common binding site for these monoclonal antibodies (mAbs) on FZD, blocking the interaction with the Wnt palmitoleic acid moiety. However, these mAbs displayed gastrointestinal toxicity or poor plasma exposure in vivo. Structure-guided engineering was used to refine the binding of each mAb for FZD receptors, resulting in antibody variants with improved in vivo tolerability and developability. Importantly, the lead variant mAb significantly inhibited tumor growth in the HPAF-II pancreatic tumor xenograft model. Taken together, our data demonstrate that anti-FZD cancer therapeutic antibodies with broad specificity can be finetuned to navigate in vivo exposure and tolerability while driving therapeutic efficacy.antibody therapeutic | Frizzled receptors | Wnt signaling | X-ray crystallography | protein engineering W nt signaling is an evolutionarily conserved signaling cascade that plays a critical role in diverse biological processes, including embryonic development, tissue differentiation, organogenesis, stem cell maintenance, and normal adult tissue homeostasis (1-3). Activation of the well-characterized β-catenindependent canonical pathway is initiated by the binding of secreted Wnt proteins to Frizzled (FZD) receptors and coreceptors such as LRP5 and LRP6 (4-8). To date, 19 human Wnt and 10 FZD receptors have been identified that mediate differential cellular functions (9, 10). FZD receptors interact with Wnt through their N-terminal extracellular cysteine-rich domain (CRD). A Wnt residue, Ser187, (position number according to Xenopus Wnt8) is posttranslationally modified, leading to its palmitoleation, which mediates interactions with the FZD-CRD (site 1); a second site of interaction between FZD and Wnt is located at the opposing end of Wnt (11).Wnt/FZD signaling is essential for normal cell function, but aberrations in the pathway are frequently found in cancers, fibrosis, and degenerative diseases (12,13). Abnormal activation of the Wnt pathway is an essential driver of primary tumor formation and metastasis in multiple cancer types (14-18). Inactivating mutations in E3 ubiquitin ligase RNF43 inhibit the downmodulation of FZD expression on the cell surface and sensitize tumor cells to Wnt-dependent growth. These mutations have been identified in pancreatic, biliary duct, and colorectal cancers (19-21). FZD5 expression is up-regulated in renal cell carcinoma (22), prostate cancer (23), and pancreatic tumors (16); aberrant FZD7 expression is observed in hepatocellular carcinoma and colorectal and triple negative breast cancer (14,24,25); FZD8 is upregulated in acute lymphoblastic leukemia and lung cancer (17, 26); and FZD4 is elevated and drives epithelial-to-mesenchy...

Cell surface antigen discovery is of great interest for biomedical research both for isolation of rare cell populations and therapeutic targeting. We developed a rapid, cost-effective, fully in vitro technology which facilities the simultaneous target discovery and human antibody generation on the surface of virtually any cell population of interest. We apply our technique to human colorectal cancer-initiating cells (CICs) and identify hundreds of unique human antibodies. We characterized the top three antibody candidates targeting these CICs and identify their protein targets as integrin α7 (ITGA7), HLA-A1 and integrin β6 (ITGB6). We demonstrate that these antibodies can be used to isolate self-renewing colorectal CICs, and that the integrin α7 antibody can prospectively identify glioblastoma brain tumor initiating cells as well as human muscle stem cells. We also demonstrate that genetic ablation of integrin β6 impedes colorectal CIC function. The methodology can be readily applied to other cell populations including stem cells, cancer, or immune cells to facilitate the rapid identification of novel targets and simultaneous generation of potent and specific antibodies with therapeutic potential.

Background: Understanding genetic predisposition to cancer is of paramount importance to tailor therapeutic strategies. Several defects in immune checkpoint regulators have been discovered in patients with EBV-induced lymphoma (e.g. CD27, PRKCD, RASGRP1, MAGT1, SH2D1A, ITK). Here, we describe the clinical and immune phenotype of 2 unrelated patients from consanguineous families presenting with a primary immune deficiency (PID) and EBV-associated lymphoproliferation due to biallelic mutations in CD137 (TNFRSF9/4-1BB). Methods: One patient of Turkish origin (P1) and Palestinian origin (P2), respectively were evaluated. Genetic analysis using whole exome sequencing was conducted. Immunological and biochemical assays were performed on primary patient material. Results: Clinical findings included recurrent sinopulmonary and herpes virus infections from childhood on. One patient suffered from auto-immunity (AIHA and auto-immune thrombocytopenia). Abnormal immunoglobulin levels were documented (IgG 413-1670, IgM 105-714, IgA 49-68 mg/dL). Both patients developed EBV-associated lymphoproliferative disorders: P1 developed Burkitt lymphoma and was treated according to NHL-BFM2000 regimen in combination with rituximab. He is currently in remission. P2 had monoclonal EBV-positive lymphoproliferation and was successfully treated with immunosuppressive therapy (e.g. cellcept, glucocorticoids). To shed light on the underlying genetic etiology, we performed whole exome sequencing. A large homozygous deletion in CD137 (c.1_545+1716del) was identified for P1, while P2 harbored a homozygous missense mutation (c.C452T, p.Thr151Met). Impaired anti-CD3 T cell lymphocyte activation and proliferation were observed, amenable to correction upon addition of anti-CD28 monoclonal antibodies. In an attempt to provide definitive proof that the CD137 gene variant causes the activation defect of T-cells, we designed a genetic rescue experiment in patient T cells. Upon retrovirus-mediated recombinant expression of WT CD137, proliferation and activation defects in T cells were restored. Conclusions: In sum, we here show that a genetic defect in the immune checkpoint molecule CD137 causes a new primary immunodeficiency disorder with susceptibility to EBV-induced lymphomagenesis. Disclosures No relevant conflicts of interest to declare.