Clonal non-malignant hematological disorders: unraveling molecular pathogenic mechanisms to develop novel targeted therapeutics

Clonal non-malignant hematological disorders are a heterogeneous group of diseases that are particularly challenging for hematologists. Indeed, most obvious and frequent hematological diseases include a broad spectrum of malignancies, such as leukemias, lymphomas, myeloma, and other myeloproliferative or lymphoproliferative disorders. In recent years, all these diseases have been categorized by the WHO according to a novel classification of myeloid and lymphoid malignancies, which takes in account the outstanding progress in our understanding of molecular defects underlying hematological malignancies. Regardless of a number of novel technologies, hematologists continue to deal daily with conditions where a clear diagnosis of a malignancy is missing: this is the case of several clonal hematological disorders, which are considered bona fide non-malignant. Some myelodysplastic syndromes, chronic T and NK disorders of granular lymphocytes, myelofibrosis, monoclonal gammopathies, monoclonal B-cel lymphocytosis, mastocytosis and paroxysmal nocturnal hemoglobinuria are paradigmatic examples of how clonal disorders are clearly different from cancers, even if they may share with hematological malignancies similar molecular, genetic, epigenetic and immunological processes. Indeed, it is not entirely clear whether in individual conditions such pathogenic mechanisms may represent initial step(s) of malignant transformation, making a bridge between these clonal non-malignant disorders and typical hematological cancers. Some of these non-malignant disorders imply specific pathogenic mechanisms and/or clinical course, and so they have been definitely established with their own biological and clinical identity. However, the obvious question whether some of these clonal non-malignant hematological diseases form some a kind of disease-continuum with their corresponding malignant counterpart is still to be answered

Clinical relevance of intrahepatic hepatitis B virus DNA in HBsAg-negative HBcAb-positive patients undergoing hematopoietic stem cell transplantation for hematological malignancies.

Hepatitis flare-up due to hepatitis B virus (HBV) is a well-recognized complication associated with chemotherapy. Data on the clinical impact of HBV DNA in liver tissue of HB-surface antigen (sAg)-negative patients undergoing immunosuppression are scanty

Molecular Pathogenesis of Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS) are a group of clonal hematologic disorders characterized by inefficient hematopoiesis, hypercellular bone marrow, dysplasia of blood cells and cytopenias. Most patients are diagnosed in their late 60s to early 70s. MDS is a risk factor for the development of acute myeloid leukemia which can occur in 10-15% of patients with MDS. A variety of pathophysiologic mechanisms contributes to the genesis and persistence of MDS including immunologic, epigenetic, cytogenetic and genetic factors. The only potential curative option for MDS is hematopoietic cell transplantation which is suitable for only a few patients. Currently approved therapeutic options for MDS, including lenalidomide, decitabine, and 5-azacytidine, are targeted to improve transfusion requirements and quality of life. Moreover, 5-azacytidine has also been demonstrated to improve survival in some patients with higher risk MDS. New ways to predict which patients will better gain benefit from currently available therapeutic agents are the primary challenges in MDS. In the last 10 years, chromosome scanning and high throughput technologies (single nucleotide polymorphism array genotyping, comparative genomic hybridization, and whole genome/ exome sequencing) have tremendously increased our knowledge of MDS pathogenesis. Indeed, the molecular heterogeneity of MDS supports the idea of different therapeutic approaches which will take into account the diverse morphologic and clinical presentations of MDS patients rather than a restricted therapeutic strategy. This review will summarize the molecular abnormalities in key relevant components of the biology and pathogenesis of MDS and will provide an update on the clinical impact and therapeutic response in MDS patients

Immunological derangement in Hypocellular Myelodysplastic Syndromes

Hypocellular or hypoplastic myelodysplastic syndromes (HMDS) are a distinct subgroup accounting for 10–15% of all MDS patients, that are characterized by the presence of bone marrow (BM) hypocellularity, various degree of dysmyelopoiesis and sometimes abnormal karyotype. Laboratory and clinical evidence suggest that HMDS share several immune-mediated pathogenic mechanisms with acquired idiopathic aplastic anemia (AA). Different immune-mediated mechanisms have been documented in the damage of marrow hematopoietic progenitors occurring in HMDS; they include oligoclonal expansion of cytotoxic T lymphocytes (CTLs), polyclonal expansion of various subtypes of T helper lymphocytes, overexpression of FAS-L and of the TNF–related apoptosis-inducing ligand (TRAIL), underexpression of Flice-like inhibitory protein long isoform (FLIPL) in marrow cells as well as higher release of Th1 cytokines, such as interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α). It has also been documented that some HMDS patients have higher frequency of polymorphisms linked both to high production of proinflammatory cytokines such as TNF-α and transforming growth factor-β and to the inhibition of T-cell mediated immune responses such as interleukin-10, further suggesting that immune-mediated mechanisms similar to those seen in AA patients may also operate in HMDS. Clinically, the strongest evidence for immune–mediated hematopoietic suppression in some HMDS is the response to immunosuppression including mainly cyclosporine, anti-thymocyte globulin and/or cyclosporine, or alemtuzumab. Here we review all these immune mechanisms as well as the influence of this deranged cellular and humoral immunologic mileau on the initiation and possible progression of MDS. All these observations are pivotal not only for a better understanding of MDS pathophysiology, but also for their immediate clinical implications, eventually leading to the identification of MDS patients who may benefit from immunosuppression

67 kDa laminin receptor (67LR) in normal and neoplastic hematopoietic cells: is its targeting a feasible approach?

The 67 kDa laminin receptor (67LR) is a non-integrin cell surface receptor for laminin (LM) that derives from a 37 kDa precursor (37LRP). 67LR expression is increased in neoplastic cells and correlates with an enhanced invasive and metastatic potentialin many human solid tumors, recommending this receptor as a new promising target for cancer therapy. This is supported by in vivo studies showing that 67LR downregulation reduces tumour cell proliferation and tumour formation by inducing apoptosis. 67LR association with the anti-apoptotic protein PED/PEA-15 activates a signal transduction pathway, leading to cell proliferation and resistance to apoptosis. However, the main function of 67LR is to enhance tumor cell adhesion to the LM of basement membranes and cell migration, two crucial events in the metastasis cascade.Thus, inhibition of 67LR binding to LM has been proved to be a feasible approach to block metastatic cancer cell spread. Despite accumulating evidences on 67LR overexpression in hematologic malignancies, 67LR role in these diseases has not been clearly defined. Here, we review 67LR expression and function in normal and malignant hematopoietic cells, 67LR role and prognostic impact in hematological malignancies and first attempts in targeting its activity

Urokinase type plasminogen activator receptor (uPAR) as a new therapeutic target in cancer

The urokinase (uPA)-type plasminogen activator receptor (uPAR) is a GPIanchored receptor that focuses urokinase (uPA) proteolytic activity on the cell surface. uPAR also regulates cell adhesion, migration and proliferation, protects from apoptosis and contributes to epithelial mesenchymal transition (EMT), independently of uPA enzymatic activity. Indeed, uPAR interacts with beta1, beta2 and beta3 integrins, thus regulating their activities. uPAR cross-talks with receptor tyrosine kinases through integrins and regulates cancer cell dormancy, proliferation and angiogenesis. Moreover, uPAR mediates uPA-dependent cell migration and chemotaxis induced by fMet-Leu-Phe (fMLF), through its association with fMLF-receptors (fMLFRs). Further, uPAR is an adhesion receptor because it binds vitronectin (VN), a component of provisional extracellular matrix. High uPAR expression predicts for more aggressive disease in several cancer types for its ability to increase invasion and metastasis. In fact, uPAR has been hypothesized to be the link between tumor cell dormancy and proliferation that usually precedes the onset of metastasis. Thus, inhibiting uPAR could be a feasible approach to affect tumor growth and metastasis. Here, we review the more recent advances in the development of uPAR-targeted anti-cancer therapeutic agents suitable for further optimization or ready for the evaluation in early clinical trials

Mean reticolocyte hemoglobin content index plays a key role to identify children who are carriers of β –thalassemia

Reticulocyte (r) and red blood cell (RBC) indices provide reliable parameters for screening and monitoring iron deficiency anemia (IDA) patients and β-thalassemia trait (BTT) carriers. The aim of this study is to identify a simple method for use to distinguish β-thalassemia trait carriers from IDA and to evaluate the correlation between BTT genetic mutation and MCV values and new discrimination index for the detection of β-thalassemia trait (DI-BTT). We analyzed CHr, MCHCr, MCVr, RBC, mean cellular hemoglobin concentration (MCHC) and mean cellular volume (MCV) indices among a pediatric population of IDA patients (n=90), βthalassemia trait carriers (n=72) and normal controls (NC) (n=131). Furthermore, to distinguish IDA patients from β-thalassemia trait carriers we evaluated clinical utility of new DI for the detection BTTcarriers, using the following polynomial: (RBC × MCHC × 50/MCV)/CHr. We found that CHr, MCVr and DI-BTT mean values were significantly different between βthalassemia trait carriers and IDA patients. CHr, MCVr and DI-BTT plotting curves showed exclusive distribution in β-thalassemia trait carriers. Moreover, DI-BTT was very accurate in differentiating β-thalassemia trait carriers from IDA patients. All BTT patients showed a heterozygous mutation of the β-globin gene including CD39, IVS1.110, IVS1.6 and IVS2.745, IVS2.1 and IVS1.1. The highest MCV values were displayed by those carrying the IVS1.6 mutation. Conclusions: The simultaneous measurement and plotting of CHr and MCVr indices, as well as the DI-BTT allow to distinguish β-thalassemia carriers from IDA patients

The urokinase/urokinase receptor system in mast cells: effects of its functional interaction with fmlf receptors

Mast cell and basophils express the high affinity receptor for IgE (FcRI) and are primary effector cells of allergic disorders. The urokinase (uPA)-mediated plasminogen activation system is involved in physiological and pathological events based on cell migration and tissue remodelling, such as inflammation, wound healing, angiogenesis and metastasis. uPA is a serine protease that binds uPAR, a high affinity glycosyl-phosphatidyl-inositol (GPI)- anchored receptor. uPAR focuses uPA activity at the cell surface and activates intracellular signaling through lateral interactions with integrins, receptor tyrosine kinases and the G-protein-coupled family of fMLF chemotaxis receptors (FPRs). We investigated the expression of the uPAuPAR system and its functional interaction with FPRs in human mast cells (MCs). Differently from basophils, MCs produced uPA that was able to induce their chemotaxis. Indeed, MCs also expressed uPAR, both in the intact and in a cleaved form (DIIDIII-uPAR) that can expose, at the N-terminus, the SRSRY sequence, able to interact with FPRs and to mediate cell chemotaxis. MCs also expressed mRNAs for FPRs that were functionally active; indeed, uPA and a soluble peptide (uPAR84-95), containing the SRSRY chemotactic sequence of uPAR and able to interact with FPRs, were able to induce MCs chemotaxis. Thus, uPA is a potent chemoattractant for MCs acting through the exposure of the chemotactic epitope of uPAR, that is an endogenous ligand for FPRs. The same mechanism could be involved in VEGF-A secretion by human MCs, also induced by uPA and uPAR84-95 stimulation

CD10, BCL6, and MUM1 expression in diffuse large B-cell lymphoma on FNA samples

BACKGROUND: Gene expression profiling has divided diffuse large B-cell lymphoma (DLBCL) into 2 main subgroups: germinal center B (GCB) and non-GCB type. This classification is reproducible by immunohistochemistry using specific antibodies such as CD10, B-cell lymphoma 6 (BCL6), and multiple myeloma oncogene 1 (MUM1). Fine-needle aspiration (FNA) plays an important role in the diagnosis of non-Hodgkin lymphoma, and in some cases FNA may be the only available pathological specimen. The objectives of the current study were to evaluate CD10, BCL6, and MUM1 immunostaining on FNA samples by testing the CD10, BCL6, and MUM1 algorithm on both FNA cell blocks (CB) and conventional smears (CS), evaluating differences in CB and CS immunocytochemical (ICC) performance, and comparing results with histological data. METHODS: Thirty-eight consecutive DLBCL cases diagnosed by FNA were studied. Additional passes were used to prepare CB in 22 cases and CS in 16 cases; the corresponding sections and smears were immunostained using CD10, BCL6, and MUM1 in all cases. The data obtained were compared with histological immunostaining in 24 cases. RESULTS: ICC was successful in 33 cases (18 CB and 15 CS) and not evaluable in 5 cases (4 CB and 1 CS). The CD10-BCL6-MUM1 algorithm subclassified DLBCL as GCB (9 cases) and non-GCB (24 cases). ICC data were confirmed on histologic staining in 24 cases. CONCLUSIONS: CD10, BCL6, and MUM1 ICC staining can be performed on FNA samples. The results herein prove it is reliable both on CB and CS, and is equally effective and comparable to immunohistochemistry data

Prolonged complete hematologic response in relapsed/refractory T-large granular lymphocyte leukemia after bendamustine treatment

T-large granular lymphocyte leukemia (T-LGLL) is a chronic clonal proliferation of effector memory cytotoxic CD3+CD57+CD56- T cells and the current guidelines suggest immunosuppressive therapy as first-line therapy, but the treatment of refractory/relapsed patients is still challenging due to the lack of prospective studies. We describe a series of two refractory/relapsed T-LGLL patients successfully treated with bendamustine, a chemotherapeutic agent largely used for B-cell neoplasms, but poorly investigated for the treatment of T-cell diseases. Complete remission (CR) was achieved in 3 and 6 months, respectively, and maintained for at least 20 months. One patient relapsed after a 20-month CR, but she was responsive to bendamustine therapy again, obtaining a further prolonged CR. Bendamustine as single agent or in combination could be a feasible therapeutic option in refractory/relapsed T-LGLL, especially for elderly patients because of its safety profile