Summary
Multiple functions of CD38 need exploring to expand clinical application of anti‐CD38 antibodies in multiple myeloma (MM). We investigated membrane dynamics of MM cells and subsequent events ...when CD38 is targeted by therapeutic antibodies. Human MM cells (BF01) were co‐cultured in vitro with therapeutic antibody (or control immunoglobulin G) and analysed using gene expression profiling. Microvesicles from antibody‐exposed cells were analysed for differential gene and microRNA (miRNA) expression, and for phenotypic characterisation. Exposure of BF01 cells to anti‐CD38 antibody resulted in CD38 membrane redistribution, upregulation of metabolism‐related genes and downregulation of genes involved in cell cycle processes. Microvesicles derived from antibody‐exposed cells showed increased CD73 and CD39 expression, presence of programmed death‐ligand 1 and significant up‐/down‐modulation of miRNAs. Microvesicles accumulated around immunoglobulin Fc receptor‐positive (FcR+) cells. Upon internalisation, natural killer cells displayed significantly increased expression of genes related to activation and immune response, and downregulation of genes involved in the cell cycle. Cells may use microvesicles to transmit signals distally as part of a survival strategy. Microvesicles are equipped on their surface with enzymatic machinery leading to production of tolerogenic adenosine. Further, they are internalised in FcR+ cells with significant functional modifications. These observations have relevance for improving anti‐CD38 therapeutic antibodies through targeting this mechanism and its sequelae.
Extracellular adenosine (ADO), generated from ATP or ADP through the concerted action of the ectoenzymes CD39 and CD73, elicits autocrine and paracrine effects mediated by type 1 purinergic ...receptors. We have tested whether the expression of CD39 and CD73 by chronic lymphocytic leukemia (CLL) cells activates an adenosinergic axis affecting growth and survival. By immunohistochemistry, CD39 is widely expressed in CLL lymph nodes, whereas CD73 is restricted to proliferation centers. CD73 expression is highest on Ki-67+ CLL cells, adjacent to T lymphocytes, and is further localized to perivascular areas. CD39+/CD73+ CLL cells generate ADO from ADP in a time- and concentration-dependent manner. In peripheral blood, CD73 expression occurs in 97/299 (32%) CLL patients and pairs with CD38 and ZAP-70 expression. CD73-generated extracellular ADO activates type 1 purinergic A2A receptors that are constitutively expressed by CLL cells and that are further elevated in proliferating neoplastic cells. Activation of the ADO receptors increases cytoplasmic cAMP levels, inhibiting chemotaxis and limiting spontaneous drug-induced apoptosis of CLL cells. These data are consistent with the existence of an autocrine adenosinergic loop, and support engraftment of leukemic cells in growth-favorable niches, while simultaneously protecting from the action of chemotherapeutic agents.
In vivo use of monoclonal antibodies (mAbs) has become a mainstay of routine clinical practice in the treatment of various human diseases. A number of molecules can serve as targets, according to the ...condition being treated. Now entering human clinical trials, CD38 molecule is a particularly attractive target because of its peculiar pattern of expression and its twin role as receptor and ectoenzyme. This review provides a range of analytical perspectives on the current progress in and challenges to anti-CD38 mAb therapy. We present a synopsis of the evidence available on CD38, particularly in myeloma and chronic lymphocytic leukemia (CLL). Our aim is to make the data from basic science helpful and accessible to a diverse clinical audience and, at the same time, to improve its potential for in vivo use. The topics covered include tissue distribution and signal implementation by mAb ligation and the possibility of increasing cell density on target cells by exploiting information about the molecule's regulation in combination with drugs approved for in vivo use. Also analyzed is the behavior of CD38 as an enzyme: CD38 is a component of a pathway leading to the production of adenosine in the tumor microenvironment, thus inducing local anergy. Consequently, not only might CD38 be a prime target for mAb-mediated therapy, but its functional block may contribute to general improvement in cancer immunotherapy and outcomes.
This work addresses the question whether CD38, a negative prognostic marker in B-cell chronic lymphocytic leukemia (B-CLL), plays a role in neoplastic B-cell growth and survival. We show that CD38+ ...B-CLL cells bind to murine fibroblasts transfected with the CD31 ligand. The interaction triggers an extensive remodeling of the B-CLL membrane, with relocalization of BCR/CD19 to the CD38/CD31 contact areas, and it also increases cell survival and proliferation. A second event is the up-modulation of the survival receptor CD100, restricted to proliferating cells, and a concomitant decrease of CD72 (low-affinity CD100 ligand and negative regulator of immune responses). The most efficient signals are delivered through sequential interactions between CD38/CD31 and CD100/plexin-B1 (high-affinity CD100 ligand), as inferred by coculture experiments using specific transfectants and blocking monoclonal antibodies (mAbs). The finding that nurselike cells from B-CLL patients express CD31 and plexin-B1, which deliver growth and survival signals to CD38+/CD100+ B-CLL cells, further confirms the model proposed. These findings show that a set of normal receptors and ligands ruling physiologic signaling pathways in B lymphocytes becomes detrimental when expressed in the context of B-CLL cells, ultimately leading to the generation of a tumor reservoir.
Summary
Respiratory syncytial virus (RSV) is the most common cause of hospitalization due to bronchiolitis in infants. Although the mechanisms behind this association are not completely elucidated, ...they appear to involve an excessive immune response causing lung pathology. Understanding the host response to RSV infection may help in the identification of targets for therapeutic intervention. We infected in‐vitro human monocyte‐derived dendritic cells (DCs) with RSV and analysed various aspects of the cellular response. We found that RSV induces in DCs the expression of CD38, an ectoenzyme that catalyses the synthesis of cyclic ADPR (cADPR). Remarkably, CD38 was under the transcriptional control of RSV‐induced type I interferon (IFN). CD38 and a set of IFN‐stimulated genes (ISGs) were inhibited by the anti‐oxidant N‐acetyl cysteine. When CD38‐generated cADPR was restrained by 8‐Br‐cADPR or kuromanin, a flavonoid known to inhibit CD38 enzymatic activity, RSV‐induced type I/III IFNs and ISGs were markedly reduced. Taken together, these results suggest a key role of CD38 in the regulation of anti‐viral responses. Inhibition of CD38 enzymatic activity may represent an encouraging approach to reduce RSV‐induced hyperinflammation and a novel therapeutic option to treat bronchiolitis.
The infection of human dendritic cells (DC) by respiratory syncytial virus induces activation of anti‐viral and proinflammatory pathways mediated by type I IFNs and fostered by the production of reactive oxygen species (ROS). CD38, an ectoenzyme that catalyses the synthesis of cyclic ADPR (cADPR), is under the transcriptional control of RSV‐induced type I interferon (IFN). In RSV‐infected DC, CD38 activity exacerbates the activation of inflammatory and anti‐viral processes, probably through cADPR‐mediated activation of ROS; inhibition of CD38 enzymatic activity may represent a novel approach to reduce RSV‐induced hyperinflammation and airway pathology.
There is a real medical need of new diagnostic tools for the early recognition of invasive Candida infections. We exploited a rather simple and rapid redox methodology to construct a bispecific ...monoclonal antibody (bsmAb) that combines a monoclonal antibody (mAb) directed against 1,3-β-D-glucan, a well-known, pan-fungal diagnostic biomarker, with a mAb recognizing MP65, a major immunogenic mannoprotein secreted by C.albicans and other Candida species. The bsmAb (MP65/bglu mAb) was successfully produced and purified at high yields and proved to bind and reveal simultaneously, with high sensitivity, the β-glucan and MP65 antigens in both purified and native forms. The MP65/bglu mAb is the first bispecific antibody generated against a fungal microorganism and may prove useful for the concurrent detection of different and clinically significant Candida biomarkers in patient sera.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Nicotinamide adenine dinucleotide (NAD⁺) is an essential co-enzyme reported to operate both intra- and extracellularly. In the extracellular space, NAD⁺ can elicit signals by binding purinergic P2 ...receptors or it can serve as the substrate for a chain of ectoenzymes. As a substrate, it is converted to adenosine (ADO) and then taken up by the cells, where it is transformed and reincorporated into the intracellular nucleotide pool. Nucleotide-nucleoside conversion is regulated by membrane-bound ectoenzymes. CD38, the main mammalian enzyme that hydrolyzes NAD⁺, belongs to the ectoenzymatic network generating intracellular Ca(2+)-active metabolites. Within this general framework, the extracellular conversion of NAD⁺ can vary significantly according to the tissue environment or pathological conditions. Accumulating evidence suggests that tumor cells exploit such a network for migrating and homing to protected areas and, even more importantly, for evading the immune response. We report on the experience of this lab to exploit human multiple myeloma (MM), a neoplastic expansion of plasma cells, as a model to investigate these issues. MM cells express high levels of surface CD38 and grow in an environment prevalently represented by closed niches hosted in the bone marrow (BM). An original approach of this study derives from the recent use of the clinical availability of therapeutic anti-CD38 monoclonal antibodies (mAbs) in perturbing tumor viability and enzymatic functions in conditions mimicking what happens in vivo.
Nucleotide-metabolizing ectoenzymes are endowed with an extracellular catalytic domain, which is involved in regulating the extracellular nucleotide/nucleoside balance. The tumor microenvironment ...contains high levels of adenosine (ADO) generated by this enzymatic network, thus promoting tumor growth by inhibiting anti-tumor immune responses. ADO inhibition in melanoma murine models limits tumor metastases and restores anti-tumor immune responses. This work investigates the expression and function of ectoenzymes in primary human melanoma cell lines. All of latter cells expressed CD38, CD39, CD73, and CD203a/PC-1, and produced ADO from AMP and NAD(+ )T cell proliferation. Accordingly, phosphorylation of S6 ribosomal protein, p38 and Stat1 was lower in activated memory cells than in naïve CD4(+) T lymphocytes. Melanoma cells also inhibited proliferation of naïve, memory and -to a lesser extent- of effector CD8(+) T cells. These different inhibitory effects correlated with distinct patterns of expression of the ADO receptor A2a and A2b. These results show that primary human melanoma cell lines suppress in vitro T cell proliferation through an adenosinergic pathway in which CD38 and CD73 play a prominent role.
This study investigates the mechanism(s) underlying the immunoregulatory activities of placenta-derived human amnion epithelial cells (hAEC). The working hypothesis is that NAD
and ATP, along with ...ectoenzymes involved in their metabolism, play a significant role in hAEC-mediated immune regulation. Proof of principle of the hypothesis was obtained by analyzing the interactions between hAEC and the main human leukocyte populations. The results obtained indicate that hAEC constitutively express a unique combination of functional ectoenzymes, driving the production of adenosine (ADO) via canonical (CD39, CD73) and alternative (CD38, CD203a/PC-1, CD73) pathways. Further, the picture is completed by the observation that hAEC express A1, A2a, and A2b ADO receptors as well as ADO deaminase, the enzyme involved in ADO catabolism. The contribution of the purinergic mediator to immunomodulation was confirmed by exposing in vitro different immune effector cells to the action of primary hAECs. B cells showed an enhanced proliferation and diminished spontaneous apoptosis when in contact with hAEC. T cell proliferation was partially inhibited by hAEC through ADO production, as confirmed by using specific ectoenzyme inhibitors. Further, hAEC induced an expansion of both T and B regulatory cells. Last, hAEC inhibited NK cell proliferation. However, the involvement of ADO-producing ectoenzymes is less apparent in this context. In conclusion, hAEC exert different in vitro immunoregulatory effects, per se, as a result of interactions with different populations of immune effector cells. These results support the view that hAEC are instrumental for regenerative medicine as well as in therapeutic applications for immune-related diseases.
It is known that multiple myeloma (MM) cells express CD38 and that a recently developed human anti-CD38 monoclonal antibody Daratumumab mediates myeloma killing. However, the expression of CD38 and ...other functionally related ectoenzymes within the MM bone niche and the potential effects of Daratumumab on bone cells are still unknown. This study firstly defines by flow cytometry and immunohistochemistry the expression of CD38 by bone marrow cells in a cohort of patients with MM and indolent monoclonal gammopathies. Results indicate that only plasma cells expressed CD38 at high level within the bone niche. In addition, the flow cytometry analysis shows that CD38 was also expressed by monocytes and early osteoclast progenitors but not by osteoblasts and mature osteoclasts. Indeed, CD38 was lost during
osteoclastogenesis. Consistently, we found that Daratumumab reacted with CD38 expressed on monocytes and its binding inhibited
osteoclastogenesis and bone resorption activity from bone marrow total mononuclear cells of MM patients, targeting early osteoclast progenitors. The inhibitory effect was not observed from purified CD14
cells, suggesting an indirect inhibitory effect of Daratumumab. Interestingly,
retinoic acid treatment increased the inhibitory effect of Daratumumab on osteoclast formation. These observations provide a rationale for the use of an anti-CD38 antibody-based approach as treatment for multiple myeloma-induced osteoclastogenesis.
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