Fausto Adami

The chemokine receptor CXCR3 is expressed on malignant B cells and mediates chemotaxis

B- and T-cell recirculation is crucial for the function of the immune system, with the control of cell migration being mainly mediated by several chemokines and their receptors. In this study, we investigated the expression and function of CXCR3 on normal and malignant B cells from 65 patients with chronic lymphoproliferative disorders (CLDs). Although CXCR3 is lacking on CD5(+) and CD5(-) B cells from healthy subjects, it is expressed on leukemic B lymphocytes from all (31/31) patients with chronic lymphocytic leukemia (CLL). The presence of CXCR3 was heterogeneous in other B-cell disorders, being expressed in 2 of 7 patients with mantle cell lymphoma (MCL), 4 of 12 patients with hairy cell leukemia (HCL), and 11 of 15 patients with other subtypes of non-Hodgkins lymphomas (NHLs). Chemotaxis assay shows that normal B cells from healthy subjects do not migrate in response to IFN-inducible protein 10 (IP-10) and IFN-gamma-induced monokine (Mig). In contrast, a definite migration in response to IP-10 and Mig has been observed in all malignant B cells from patients with CLL, but not in patients with HCL or MCL (1/7 cases tested). Neoplastic B cells from other NHLs showed a heterogenous pattern. The migration elicited by IP-10 and Mig was inhibited by blocking CXCR3. No effect of IP-10 and Mig chemokines was observed on the cytosolic calcium concentration in malignant B cells. The data reported here demonstrate that CXCR3 is expressed on malignant B cells from CLDs, particularly in patients with CLL, and represents a fully functional receptor involved in chemotaxis of malignant B lymphocytes.

New pathogenetic insights into the sarcoid granuloma.

Although the most important enigma of sarcoidosis, ie, its etiology, remains an unsolved problem, the past few years have seen remarkable advances in understanding general immunologic and molecular aspects of the mechanisms leading to granuloma formation in sarcoidosis. It is now clear that during the sarcoid inflammatory process several cytokines are secreted at sites of disease activity; in addition, high affinity receptors for cytokines participating in the granuloma development have been recently identified. This article provides a detailed description of recent data that have clarified cellular interactions governing the dynamics of granuloma formation in sarcoidosis. Together, these new results provide important insights that can refocus efforts at developing immunotherapeutic methods of inhibiting cytokine production at sites of granuloma formation.

Cobalamin Deficiency: Clinical Picture and Radiological Findings

Vitamin B12 deficiency causes a wide range of hematological, gastrointestinal, psychiatric and neurological disorders. Hematological presentation of cobalamin deficiency ranges from the incidental increase of mean corpuscular volume and neutrophil hypersegmentation to symptoms due to severe anemia, such as angor, dyspnea on exertion, fatigue or symptoms related to congestive heart failure, such as ankle edema, orthopnea and nocturia. Neuropsychiatric symptoms may precede hematologic signs and are represented by myelopathy, neuropathy, dementia and, less often, optic nerve atrophy. The spinal cord manifestation, subacute combined degeneration (SCD), is characterized by symmetric dysesthesia, disturbance of position sense and spastic paraparesis or tetraparesis. The most consistent MRI finding is a symmetrical abnormally increased T2 signal intensity confined to posterior or posterior and lateral columns in the cervical and thoracic spinal cord. Isolated peripheral neuropathy is less frequent, but likely overlooked. Vitamin B12 deficiency has been correlated negatively with cognitive functioning in healthy elderly subjects. Symptoms include slow mentation, memory impairment, attention deficits and dementia. Optic neuropathy occurs occasionally in adult patient. It is characterized by symmetric, painless and progressive visual loss. Parenteral replacement therapy should be started soon after the vitamin deficiency has been established.

Regulation of alveolar macrophage-T cell interactions during Th1-type sarcoid inflammatory process

The accessory function of antigen-presenting cells depends on the presence of a number of costimulatory molecules, including members of the B7 family (CD80 and CD86) and the CD5 coligand CD72. The aim of this study was to evaluate the regulation of T cell-antigen-presenting cell costimulatory pathways in the lung of patients with a typical Th1-type reaction, i.e., sarcoidosis. Although normal alveolar macrophages (AMs) did not bear or bore low levels of costimulatory molecules, AMs from sarcoid patients with CD4 T-cell alveolitis upmodulated CD80, CD86, and CD72 and expressed high levels of interleukin (IL)-15; lymphocytes accounting for T-cell alveolitis expressed Th1-type cytokines [interferon (IFN)-γ and/or IL-2] and bore high levels of CD5 and CD28 but not of CD152 molecules. In vitro stimulation of AMs with Th1-related cytokines (IL-15 and IFN-γ) upregulated the expression of CD80 and CD86 molecules. However, stimulation with IL-15 induced the expression of Th1-type cytokines (IFN-γ) and CD28 on sarcoid T cells, suggesting a role for this macrophage-derived cytokine in the activation of the sarcoid T-cell pool. The hypothesis that CD80 and CD86 molecules regulate the sarcoid T-cell response was confirmed by the evidence that AMs induced a strong proliferation of T cells that was inhibited by pretreatment with CD80 and CD86 monoclonal antibodies. To account for these data, it is proposed that locally released cytokines provide AMs with accessory properties that contribute to the development of sarcoid T-cell alveolitis.

Protein Kinase CK2 Protects Multiple Myeloma Cells from ER Stress–Induced Apoptosis and from the Cytotoxic Effect of HSP90 Inhibition through Regulation of the Unfolded Protein Response

Purpose: Protein kinase CK2 promotes multiple myeloma cell growth by regulating critical signaling pathways. CK2 also modulates proper HSP90-dependent client protein folding and maturation by phosphorylating its co-chaperone CDC37. Because the endoplasmic reticulum (ER) stress/unfolded protein response (UPR) is central in myeloma pathogenesis, we tested the hypothesis that the CK2/CDC37/HSP90 axis could be involved in UPR in myeloma cells. Experimental Design: We analyzed CK2 activity upon ER stress, the effects of its inactivation on the UPR pathways and on ER stress–induced apoptosis. The consequences of CK2 plus HSP90 inhibition on myeloma cell growth in vitro and in vivo and CK2 regulation of HSP90-triggered UPR were determined. Results: CK2 partly localized to the ER and ER stress triggered its kinase activity. CK2 inhibition reduced the levels of the ER stress sensors IRE1α and BIP/GRP78, increased phosphorylation of PERK and EIF2α, and enhanced ER stress–induced apoptosis. Simultaneous inactivation of CK2 and HSP90 resulted in a synergic anti-myeloma effect (combination index = 0.291) and in much stronger alterations of the UPR pathways as compared with the single inhibition of the two molecules. Cytotoxicity from HSP90 and CK2 targeting was present in a myeloma microenvironment model, on plasma cells from patients with myeloma and in an in vivo mouse xenograft model. Mechanistically, CK2 inhibition led to a reduction of IRE1α/HSP90/CDC37 complexes in multiple myeloma cells. Conclusions: Our results place CK2 as a novel regulator of the ER stress/UPR cascades and HSP90 function in myeloma cells and offer the groundwork to design novel combination treatments for this disease. Clin Cancer Res; 18(7); 1888–900. ©2012 AACR.

Immune mechanisms in interstitial lung diseases

Several features have long suggested a strict relationship between the functional behavior of the host immune system and the pathobiology of interstitial lung diseases (ILDs). Most clinical manifestations of ILDs are secondary to the direct effect of the accumulation of activated immunocompetent cells in involved tissues, notably T cells, macrophages, or neutrophils. However, it is also believed that pathophysiologic events leading to the persistence and/or progression of the pulmonary disease are the same in most ILDs: the persistence of the causative agents and/ or the imbalance of mechanisms for the removal of in ammatory cells favor a prolongation of in ammatory responses, the formation of pulmonary granulomas, and, in some cases, the development of irreversible ®brosis. Other characteristics which have captured the attention of immunologists are related to the fact that the development of some complications of ILDs, including the progression to ®brosis, are regulated by a complex cascade of interactions among T cells, accessory cells, and cytokines released by relevant immunocompetent cells. Recent data on the complex network of interactions between cells and biologic response modi®ers which control the pulmonary immune system have dramatically expanded our knowledge of the pathogenesis of ILDs. Thanks to the cloning of cytokine genes, in situ hybridization methods, and cytokine receptor characterization, the cascade of interacting extracellular signaling molecules which orchestrate the traf®cking of immune cells accounting for the alveolitis are beginning to be clari®ed. In this paper, we will mainly summarize the basic aspects of pathogenetic mechanisms leading to the recruitment, homing, and activity of in ammation in the respiratory tract of patients with ILDs. In the second part of the paper, we will comment on the role of the immunologic evaluations of pulmonary immunocompetence in speci®c ILDs.

Serum levels of tumour necrosis factor-α in patients with B-cell chronic lymphocytic leukaemia

Serum levels of tumour necrosis factor-alpha (TNF-alpha) have been evaluated in the peripheral blood of 91 patients with B-cell chronic lymphocytic leukaemia (B-CLL), and have been correlated with the clinical stage (according to Rais staging system) and relevant haematological and immunological data. Increased values were detected, compared to 36 normal age-matched controls (36 pg/ml +/- 5 versus 0.11 pg/ml +/- 0.08; P < 0.05). An increase of TNF-alpha serum levels was observed in all stages including stage 0, with a progressive increase in relation to the stage of the disease. A significant relationship between serum TNF-alpha levels and the number of circulating monocytes (P < 0.002) and an inverse correlation with the level of the haemoglobin (P < 0.001) was established, as defined by the Pearsons correlation test. In contrast, no correlation was observed between TNF-alpha serum levels and the other parameters taken into account, including the white blood cell and platelet counts, the absolute number of peripheral blood (PB) lymphocytes, CD5+ B lymphocytes, CD57+ lymphocytes, serum levels of lactic dehydrogenase, total serum immunoglobulins and the serum levels of IgG, IgA and IgM. These data suggest that, in addition to the B-CLL neoplastic cells, the PB monocytes may be involved in the release of TNF-alpha.

Multiple myeloma plasma cells show different chemokine receptor profiles at sites of disease activity

Chemokines and their receptors play a pivotal role in the regulation of B‐lymphocyte trafficking. This study was aimed at investigating the pattern of chemokine receptor expression, including CCR1 to CCR3, CCR5 to CCR7, CXCR1 to CXCR5, and the migration ability of multiple myeloma (MM) plasma cells (PC). PC were recovered from the bone marrow (BM) of 29 MM patients, extramedullary sites of 10 patients and the BM of five controls. Flow cytometry analysis showed that the receptors mainly expressed on malignant BM PC were represented by CXCR4 (70% of patients), CCR1 (25%), CCR2 (25%), CCR5 (17%) and CXCR3 (20%), while other receptors were commonly lacking. The analysis performed on extramedullary (peripheral blood and pleural effusion) malignant PC demonstrated that the most represented receptors were CXCR4 (100%), CCR2 (66%) and CXCR1 (60%). The migratory capability of malignant PC at resting conditions identified three groups of patients with different migration (low, intermediate and high). As CXCR4 was the relevant chemokine receptor expressed by MM PC, its ligand CXCL12 induced their migration. These data suggest that malignant PC from MM display different chemokine receptor profiles and that CXCR4 is fully functional and might play a role in the spreading of the disease.

Protein Kinase CK2 Inhibition Down Modulates the NF-κB and STAT3 Survival Pathways, Enhances the Cellular Proteotoxic Stress and Synergistically Boosts the Cytotoxic Effect of Bortezomib on Multiple Myeloma and Mantle Cell Lymphoma Cells

CK2 is a pivotal pro-survival protein kinase in multiple myeloma that may likely impinge on bortezomib-regulated cellular pathways. In the present study, we investigated CK2 expression in multiple myeloma and mantle cell lymphoma, two bortezomib-responsive B cell tumors, as well as its involvement in bortezomib-induced cytotoxicity and signaling cascades potentially mediating bortezomib resistance. In both tumors, CK2 expression correlated with that of its activated targets NF-κB and STAT3 transcription factors. Bortezomib-induced proliferation arrest and apoptosis were significantly amplified by the simultaneous inhibition of CK2 with two inhibitors (CX-4945 and K27) in multiple myeloma and mantle cell lymphoma cell lines, in a model of multiple myeloma bone marrow microenvironment and in cells isolated from patients. CK2 inhibition empowered bortezomib-triggered mitochondrial-dependent cell death. Phosphorylation of NF-κB p65 on Ser529 (a CK2 target site) and rise of the levels of the endoplasmic reticulum stress kinase/endoribonuclease Ire1α were markedly reduced upon CK2 inhibition, as were STAT3 phospho Ser727 levels. On the contrary, CK2 inhibition increased phospho Ser51 eIF2α levels and enhanced the bortezomib-dependent accumulation of poly-ubiquitylated proteins and of the proteotoxic stress-associated chaperone Hsp70. Our data suggest that CK2 over expression in multiple myeloma and mantle cell lymphoma cells might sustain survival signaling cascades and can antagonize bortezomib-induced apoptosis at different levels. CK2 inhibitors could be useful in bortezomib-based combination therapies.

Human myeloma cells express the CD38 ligand CD31

Multiple myeloma (MM) plasma cells (PC) are CD38+. A ligand for CD38 is the adhesion molecule CD31. By flow cytometry and immunocytochemistry we have investigated whether malignant PC co‐express CD38 and CD31. All 68 patients studied were CD38+. 14/14 monoclonal gammopathies of undetermined significance (MGUS) and 39/39 plasmacytic MM patients co‐expressed CD38 and CD31 at high density. Only 1/11 plasmablastic MM and 1/4 plasma cell leukaemias (PCL) expressed CD31. These data indicated that PC malignancies co‐expressed high levels of both CD38 and its ligand CD31, with the exception of plasmablastic MM and PCL.