Osteoclasts (OCs) are bone-resorbing cells that are formed from hematopoietic precursors. OCs ordinarily maintain bone homeostasis, but they can also cause major pathology in autoimmune and inflammatory diseases. Under homeostatic conditions, receptor activator of nuclear factor kappa-B (RANK) ligand on osteoblasts drives OC differentiation by interaction with its receptor RANK on OC precursors. During chronic immune activation, RANK ligand on activated immune cells likewise drives pathogenic OC differentiation. We here report that the related TNF family member CD70 and its receptor CD27 can also mediate cross-talk between immune cells and OC precursors. We identified CD27 on a rare population (0.3%) of B220 − c-Kitlow cells in the mouse bone marrow (BM) that are highly enriched for osteoclastogenic potential. We dissected this population into CD27 high common precursors of OC, dendritic cells (DCs) and macrophages and CD27 low/neg downstream precursors that could differentiate into OC and macrophages, but not DC. In a recombinant mouse model of chronic immune activation, sustained CD27/CD70 interactions caused an accumulation of OC precursors and a reduction in OC activity. These events were due to a CD27/CD70-dependent inhibition of OC differentiation from the OC precursors by BM-infiltrating, CD70 + -activated immune cells. DC numbers in BM and spleen were increased, suggesting a skewing of the OC precursors toward DC differentiation. The impediment in OC differentiation culminated in a high trabecular bone mass pathology. Mice additionally presented anemia, leukopenia, and splenomegaly. Thus, under conditions of constitutive CD70 expression reflecting chronic immune activation, the CD27/CD70 system inhibits OC differentiation and favors DC differentiation.hematopoiesis | osteoimmunology | costimulation | TNF receptor family

Epstein-Barr virus (EBV) latency proteins EBNA1, LMP1, LMP2, and BARF1 are expressed in tumor cells of nasopharyngeal carcinoma (NPC). IgG and IgA antibody responses to these non-self tumor antigens were analyzed in NPC patients (n=125) and regional controls (n=100) by three approaches, focusing on the putative LMP1, LMP2 extracellular domains. Despite abundant IgG and IgA antibody responses to lytic antigens and EBNA1, patients had low titer (1:25-1:100) IgG to LMP1 (81.2%), LMP2 (95.6%), and BARF1 (84.8%), while immunoblot showed such reactivity in 24.2%, 12.5%, and 12.5% at 1:50 dilution, respectively. Few IgA responses were detected, except for EBNA1. Controls only showed IgG to EBNA1. ELISA using peptides from different domains of LMP1, LMP2, and BARF1 also yielded mostly negative results. When existing, low level IgG to intracellular C-terminus of LMP1 (62.9%) prevailed. Rabbit immunization with peptides representing extracellular (loop) domains yielded loop-specific antibodies serving as positive control. Importantly, these rabbit antibodies stained specifically extracellular domains of LMP1 and LMP2 on viable cells and mediated complement-driven cytolysis. Rabbit anti-LMP1 loop-1 and -3 killed 50.4% and 59.4% of X50/7 and 35.0% and 35.9% of RAJI cells, respectively, and 22% of both lines were lysed by anti-LMP2 loop-2 or -5 antibodies. This demonstrates that (extracellular domains of) EBV-encoded tumor antigens are marginally immunogenic for humoral immune responses. However, peptide-specific immunization may generate such antibodies, which can mediate cell killing via complement activation. This opens options for peptide-based tumor vaccination in patients carrying EBV latency type II tumors such as NPC.

The absorption, efflux and transport properties of two of the most commonly used tyrosine kinase inhibitors (TKIs), Erlotinib (E)

Acute myeloid leukemia (AML) is a high-risk disease with a poor prognosis, particularly in elderly patients. Because current AML treatment relies primarily on untargeted therapies with severe side effects that limit patient eligibility, identification of novel therapeutic AML targets is highly desired. We recently described AT1413, an antibody produced by donor B cells of a patient with AML cured after allogeneic hematopoietic stem cell transplantation. AT1413 binds CD43s, a unique sialylated epitope on CD43, which is weakly expressed on normal myeloid cells and overexpressed on AML cells. Because of its selectivity for AML cells, we considered CD43s as a target for a bispecific T-cell-engaging antibody (bTCE) and generated a bTCE by coupling AT1413 to two T-cell-targeting fragments using chemo-enzymatic linkage. In vitro, AT1413 bTCE efficiently induced T-cell-mediated cytotoxicity toward different AML cell lines and patient-derived AML blasts, whereas endothelial cells with low binding capacity for AT1413 remained unaffected. In the presence of AML cells, AT1413 bTCE induced upregulation of T-cell activation markers, cytokine release, and T-cell proliferation. AT1413 bTCE was also effective in vivo. Mice either coinjected with human peripheral blood mononuclear cells or engrafted with human hematopoietic stem cells [human immune system (HIS) mice] were inoculated with an AML cell line or patient-derived primary AML blasts. AT1413 bTCE treatment strongly inhibited tumor growth and, in HIS mice, had minimal effects on normal human hematopoietic cells. Taken together, our results indicate that CD43s is a promising target for T-cell-engaging antibodies and that AT1413 holds therapeutic potential in a bTCE-format. Significance: These findings offer preclinical evidence for the therapeutic potential of a bTCE antibody that targets a sialylated epitope on CD43 in AML.

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