Gianfranco Catalano

The Management of Membranous Glomerulopathy in Allogeneic Stem Cells Transplantation: Updated Literature

BACKGROUND membranous glomerulopathy (MG) is an immunomediated disorder which accounts for the most common cause of nephrotic syndrome (NS) following allogeneic hematopoietic stem cell transplantation (HSCT). OBJECTIVE AND METHODS to provide an update on the issue by reviewing pertinent literature on the MEDLINE database. RESULTS sixty-nine post allogenic HSCT patients (42 male) with MG were identified. The median age was 43 (5 to 68) years. Time interval from allogenic HSCT to MG diagnosis ranged from 3 to 134 months (median 17). Most MG patients had a history of acute (70%) or chronic (84%) graft versus host disease (GVHD). Corticosteroids and cyclosporine were the most common therapeutic agents used in this setting; alternative therapies, including rituximab, were given to a lower number of patients. Outcome data were available in 64 out of 69 MG patients; 38 (59%) and 18 (28%) patients achieved a complete and a partial response respectively, whereas treatment failure was recorded in the remaining 8 (13%). CONCLUSION MG after allogenic HSCT appears to be associated with a sub clinical or overt cGVHD, which follows the discontinuation of immunosuppressive prophylaxis. Although a standard therapeutic approach has not been established, the application of available measures can induce favorable response in more than 80% of affected patients, but treatment-failure and progressive deterioration of the renal function may occur in about one fifth of cases.

Spontaneous apoptosis and proliferation detected by BCL-2 and CD71 proteins are important progression indicators within ZAP-70 negative chronic lymphocytic leukemia

In chronic lymphocytic leukemia (CLL), inhibition of spontaneous apoptosis determines a worse prognosis and increasing evidences show that disease progression relies also upon cycling CLL cells. We investigated bcl-2, as measure of apoptosis, and CD71, as measure of proliferation, by flow cytometry in 265 patients with CLL. Combining bcl-2 with CD71 values, we defined three subgroups: (1) bcl2 − CD71−; (2) bcl2 + CD71+; and (3) bcl2 + CD71− or bcl2− CD71+. Both a shorter progression-free survival (PFS) and overall survival (OS) were observed in ZAP-70+ (p < 0.00001) and in patients with bcl2 + CD71+ (p < 0.00001 and p = 0.02). The patients with discordant in bcl2 + CD71− and bcl2− CD71+ showed an intermediate outcome. Noteworthy, patients with bcl2 + CD71+ showed a shorter PFS within ZAP-70 negative subgroup (p = 0.00009). In multivariate analysis of PFS, age (p = 0.005), beta-2 microglobulin (B2-M) (p = 0.003), bcl-2 (p = 0.004), CD49d (p = 0.001), and ZAP-70 (p < 0.001) resulted to be significant prognostic factors. The independent prognostic significance of B2-M (p = 0.009) and bcl-2 (p = 0.03) was confirmed within ZAP-70 negative patients. Bcl-2 and CD71 can be considered as interesting progression indicators, which should be validated in an independent cohort of patients, to take timely therapeutic decisions in CLL.

High-dose ascorbate and arsenic trioxide selectively kill acute myeloid leukemia and acute promyelocytic leukemia blasts in vitro

The use of high-dose ascorbate (ASC) for the treatment of human cancer has been attempted several decades ago and has been recently revived by several in vitro and in vivo studies in solid tumors. We tested the cytotoxic effects of ASC, alone or in combination with arsenic trioxide (ATO) in acute myeloid leukemia (AML) and acute promyelocytic leukemia (APL). Leukemic cell lines and primary blasts from AML and APL patients were treated with graded concentrations of ASC, alone or in association with standard concentration (1 μM) of ATO. The ASC/ATO combination killed myeloid blasts, including leukemic CD34+ cells, while sparing CD34+ progenitors obtained from normal cord blood and bone marrow. Actually, approximately one-third (11/36) of primary AML cases were highly sensitive to the ASC/ATO combination. The mechanism of cell killing appeared to be related to increased oxidative stress and overproduction of ROS in a non-quantitative fashion, which resulted in induction of apoptosis. These effects were reverted by the addition of the antioxidant N-Acetyl-Cysteine (NAC). In the APL NB4 model, ASC induced direct degradation of the PML and PML/RARA proteins via caspase activation, while the transcriptional repressor DAXX was recruited in re-constituted PML nuclear bodies. Our findings encourage the design of pilot studies to explore the potential clinical benefit of ASC alone or in combination with ATO in advanced AML and APL.

Positive selection of CD34+ cells by immunoadsorption: factors affecting the final yield and hematopoietic recovery in patients with hematological malignancies and solid tumors

There is a progressive increase in the use of selected hematopoietic progenitor cells after myeloablative therapy in patients affected by malignancies. Our goal was to determine which blood parameters, in the starting cell population, influence the concentration of CD34+ progenitors and the removal of unwanted cells in the final product. Also, we evaluated the hematopoietic recovery and toxicity associated with peripheral blood stem cell infusion. We retrospectively reviewed 53 procedures of positive selection of CD34+ cells, performed with the Ceprate SC immunoadsorption system, in 47 paticnts affected by various hematologic malignancies and solid tumors. An increased percentage of CD34+ cells in the starting fraction was associated both with the final purity and enrichment of CD34+ cells and with a decreased percentage of CD3+ and CD19+ cells in the final product. A low platelet count before selection had a borderlinc influence on the recovery of CD34+ cells. Forty patients received a median of 5 x 10(6) CD34+ cells per kg; the absolute neutrophil count (ANC) reached 0.5 x 10(9)/l in a median of 10 days whereas a PLT count above 20 x 10(9)/l was observed in 14 days. The reinfusion of selected CD34+ cells, containing a very low amount of dymethylsulfoxide. was well tolerated and no adverse reactions were observed. Autologous transplantation with selected CD34+ cells is a safe and well-tolerated procedure in patients affected by hematologic malignancies and solid tumors. Positive selection of CD34+ cells seems to be related to the quality of the apheresis products, particularly to the initial CD34+ cell and PLT content.

PYRROLO[1,2-b][1,2,5]BENZOTHIADIAZEPINES (PBTDs) induce apoptosis in K562 cells

BackgroundThe objective of this study was to gain insight into the molecular mechanism of induced cell death (apoptosis) by PYRROLO [1,2-b][1,2,5]BENZOTHIADIAZEPINES (PBTDs) series compounds, using human (K562) cells as a model.MethodsWe focused our attention on some members of the PBTDs family to test their potential apoptotic activity in K562 cells. Important apoptotic activity was demonstrated, as evidenced by the concentration and percentage of cell death quantified by measuring PI-uptake by flow cytometry, and DNA fragmentation analyzed by agarose gel electrophoresis, generating a characteristic ladder pattern of discontinuous DNA fragments. The expression of Bcl-2 family was tested using western blotting and transfection method.ResultsPBTDs-mediated suppression of K562 cell proliferation was induced by apoptosis characterized by the appearance of DNA fragmentation and was associated with the poly(ADP-ribose)polymerase (PARP) cleavage. PBTD-1 and -3 treatment resulted in caspase-3 activation through down-regulation of Bcl-2 and up-regulation of Bax. Furthermore, we used K562/vector and K562/bcl-2 cells, which were generated by transfection of the cDNA of the Bcl-2 gene. As compared with K562/vector, K562/Bcl-2 cells exhibited a 4-fold greater expression of Bcl-2. Treatment with 10 μM PBTD-1 and -3 for 24 h produced morphological features of apoptosis and DNA fragmentation in K562/vector cells, respectively. In contrast, PBTD-1 and -3-induced caspase-3 activation and apoptosis were inhibited in K562/Bcl-2. Furthermore, Bcl-2 overexpressing cells exhibited less cytocrome c release during PBTDs-induced apoptosis.ConclusionThese results indicate that PBTDs effectively induce apoptosis of K562 leukemia cells through the activation of caspase cascades. In addition, these findings indicate that Bcl-2 inhibits PBTD-1 and -3 induced-apoptosis via a mechanism that interferes with cytocrome c release, and the activity of caspase-3, which is involved in the execution of apoptosis.

The Prevention of Oral Mucositis in Patients with Blood Cancers: Current Concepts and Emerging Landscapes

BACKGROUND The prevention of oral mucositis (OM) in the management of hematological malignancies continues to represent an unmet clinical need. Addressing this issue has major clinical implications as OM can also greatly impair patients quality of life. OBJECTIVES To review currently available measures and investigational agents to prevent OM in hematological patients. METHODS we searched for OM and related issues using Medline; the abstract books of the most important hematological and oncological meetings were also reviewed. RESULTS/CONCLUSIONS Many agents targeting different mechanisms of mucosal damage have been applied in order to prevent OM; most of them have failed or its efficacy has not been fully demonstrated. Palifermin is the first pharmaceutical/biological agent approved for the prevention of OM; its use is currently restricted to patients who have received radiotherapy-containing conditioning regimens prior to autologous hematopoietic stem cell transplantation. No clear benefit by this agent has been demonstrated outside of this specific setting and its application should be limited to clinical trials. Other interventions, such as other growth factors and non mitogenic measures are under investigation or in development and their application in the hematological setting is expected in the short term.

Identification of a potential topoisomerase II “hotspot” DNA region in the DEK gene in two t(6;9)-positive therapy-related myeloid neoplasms

Dear Editor, DEK/NUP214 is considered the unique product of the translocation t(6;9)(p22;q34), first identified in 1976 and described in 1992 by von Lindern [1]. Relatively rare (1–5 % of adult AML and <1% of childhood AML), it has been reported also in AML secondary to MDS and in therapy-related myeloid neoplasms (t-MN), defines a high-risk group [2], and is frequently associated to FLT3-ITDmutation (80%) [3]. We analyzed the DEK and NUP214 breakpoint sequences in two patients who developed chemotherapy-resistant t-AML after cytotoxic treatment for breast cancer. Clinical history is described in supplementary material. The sample karyotypes were defined as follows: 46,XX)[12],t(6;9)(p22;q34)[8] for UPN1 and 46,XX[2],t(6;9)(p22;q34) [13], +13[5] for UPN2 (Supplementary Figure 2). Screening for gene alterations [4] detected DEK-NUP214 fusion gene, JAK2 V617F mutation, and FLT3-ITD mutation in both the patients. DNA was amplified by a two-step long template PCR (LTP) according manufacturer’s instructions (TAKARA Biotechnology, Dalian Biotechnology, Dalian). Five forward and eight reverse primers were designed to cover 17.4-kb spanning DEK intron 8 (genomic coordinates ch6: 18224099–18265054) and NUP214 intron 17 (genomic coordinates ch9:134000948–134110057) (Supplementary Figure 1 and Supplementary Table 1). PCR products were analyzed by direct Sanger sequencing, using the ABI Genetic Analyzer 3500 platform (Applied Biosystems Inc., Foster City, CA, USA); junction sequences were aligned using the BLAST program. The genomic DEK-NUP214 junction regions of UPN1 and UPN2 are shown in Fig. 1a, b, respectively. Several reports suggest that drugs interfering with DNA remodeling by topoisomerase II (TOP2) can mediate the formation of specific chromosomal translocation breakpoints. Chemotherapy agents targeting TOP2 enzymes increase the steady-state levels of cleavage complexes by preventing relegation of the transient TOP2 break (DSB), eventually leading to apoptosis and cell death [5]. This increases the chances of DNA being repaired by non-homologous end joining (NHEJ) leading to chromosomal translocations [6, 7]. In studying patients with multiple sclerosis who developed acute promyelocytic leukemia (APL) after mitoxantrone (MTZ) treatment, we described the presence of “hot-spot” target sequences in the PML gene, representing a potential TOP2 cleavage site [8]. Here, we report for the first time an analysis of the breakpoint sequence in tumor cells from two patients affected by t-MN with a t(6;9) translocation. UPN1 Electronic supplementary material The online version of this article (doi:10.1007/s00277-016-2843-3) contains supplementary material, which is available to authorized users.

Decitabine as salvage therapy for primary induction failure of acute myeloid leukemia

Primary induction failure (PIF) in patients with acute myeloid leukemia (AML) is defined by the persistence of more than 5% of blasts [1] after 1–2 cycles of intensive chemotherapy (ICT) [1,2]. PIF...

PML/RARa inhibits PTEN expression in hematopoietic cells by competing with PU.1 transcriptional activity.

Acute promyelocitic leukemia (APL) is characterized by the pathognomonic presence in leukemic blasts of the hybrid protein PML/RARA, that acts as a transcriptional repressor impairing the expression of genes that are critical to myeloid differentiation. Here, we show that primary blasts from APL patients express lower levels of the oncosuppressor protein PTEN, as compared to blast cells from other AML subtypes or normal bone marrow, and demonstrate that PML-RARA directly inhibits PTEN expression. We show that All-Trans Retinoic Acid (ATRA) triggers in APL cells an active chromatin status at the core regulatory region of the PTEN promoter, that allows the binding of the myeloid-regulating transcription factor PU.1, and, in turn, the transcriptional induction of PTEN. ATRA, via PML/RARA degradation, also promotes PTEN nuclear re-localization and decreases expression of the PTEN target Aurora A kinase. In conclusion, our findings support the notion that PTEN is one of the primary targets of PML/RARA in APL

A case of SRSF2 mutation in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is characterized by extremely variable clinical course indicating substantial differences in the biology of the disease. Molecular characterization provides new insights useful for treatment decision making. We report on a patient diagnosed with CLL, whose disease was characterized by episodes of rapid progression and disease stabilization, and in which a SRSF2 gene mutation was identified in the absence of other commonly known mutations of CLL. To the best of our knowledge this is the first case of SRSF2 gene mutation ever reported in CLL.