Flow cytometric evaluation of measurable residual disease in chronic lymphocytic leukemia: Where do we stand?

Measurable residual disease (MRD) has emerged as a relevant parameter of response to therapy in chronic lymphocytic leukemia (CLL). Although several methods have been developed, flow cytometry has emerged as the most useful and standardized approach to measure and quantify MRD. The improved sensitivity of MRD measurements has been paralleled by the development of more effective therapeutic strategies for CLL, increasing the applicability of MRD detection in this setting. Chemotherapy and chemoimmunotherapy have firstly demonstrated their ability to obtain a deep MRD. Combined targeted therapies are also demonstrating a high molecular response rate and prospective trials are exploring the role of MRD to guide the duration of treatment in this setting. In this review we briefly summarize what we have learned about MRD with emphasis on its flow cytometric detection.

Red wine activates plasma membrane redox system in human erythrocytes.

In the present study, we report that polyphenols present in red wine obtained by a controlled microvinification process are able to protect human erythrocytes from oxidative stress and to activate Plasma Membrane Redox System (PMRS). Human plasma obtained from healthy subjects was incubated in the presence of whole red wine at a concentration corresponding to 9.13-73 µg/ml gallic acid equivalents to verify the capacity to protect against hypochlorous acid (HOCl)-induced plasma oxidation and to minimize chloramine formation. Red wine reduced hemolysis and chloramine formation induced by HOCl of 40 and 35%, respectively. PMRS present on human erythrocytes transfers electrons from intracellular molecules to extracellular electron acceptors. We demonstrated that whole red wine activated PMRS activity in human erythrocytes isolated from donors in a dose-dependent manner with a maximum at about 70-100 µg/ml gallic acid equivalents. We also showed that red wine increased glutathione (GSH) levels and erythrocytic antioxidant capacity, measured by 2,2-diphenyl-1-picrylhydrazyl (DPPH) quenching assay. Furthermore, we reported that GSH played a crucial role in regulating PMRS activity in erythrocytes. In fact, the effect of iodoacetamide, an alkylating agent that induces depletion of intracellular GSH, was completely counteracted by red wine. Bioactive compounds present in red wine, such as gallic acid, resveratrol, catechin, and quercetin were unable to activate PMRS when tested at the concentrations normally present in aged red wines. On the contrary, the increase of PMRS activity was associated with the anthocyanin fraction, suggesting the capacity of this class of compounds to positively modulate PMRS enzymatic activity.

ABT-737 resistance in B-cells isolated from chronic lymphocytic leukemia patients and leukemia cell lines is overcome by the pleiotropic kinase inhibitor quercetin through Mcl-1 down-regulation.

Chronic lymphocytic leukemia (CLL) is the most frequent form of leukemia in adult population and despite numerous studies, it is considered an incurable disease. Since CLL is characterized by overexpression of pro-survival Bcl-2 family members, treatments with their antagonists, such as ABT-737, represent a promising new therapeutic strategy. ABT-737 is a BH3 mimetic agent which binds Bcl-2, Bcl-XL and Bcl-w with high affinity, while weakly interacts with Mcl-1 and Bfl-1. Previous studies demonstrated that quercetin, a flavonoid naturally present in food and beverages, was able to sensitize B-cells isolated from CLL patients to apoptosis when associated with death ligands or fludarabine, through a mechanism involving Mcl-1 down-regulation. Here, we report that the association between ABT-737 and quercetin synergistically induces apoptosis in B-cells and in five leukemic cell lines (Combination Index <1). Peripheral blood mononuclear cell from healthy donors were not affected by quercetin treatment. The molecular pathways triggered by quercetin have been investigated in HPB-ALL cells, characterized by the highest resistance to both ABT-737 and quercetin when applied as single molecules, but highly sensitivity to the co-treatment. In this cell line, quercetin down-regulated Mcl-1 through the inhibition of PI3K/Akt signaling pathway, leading to Mcl-1 instability. The same mechanism was confirmed in B-cells. These results may open new clinical perspectives based on a translational approach in CLL therapy.

Short-term thalidomide incorporated into double autologous stem-cell transplantation improves outcomes in comparison with double autotransplantation for multiple myeloma.

PURPOSE: To assess potential benefits with thalidomide incorporated into double autologous stem-cell transplantation (ASCT) for younger patients with newly diagnosed multiple myeloma (MM). PATIENTS AND METHODS: One hundred thirty-five patients who received thalidomide from induction until the second ASCT were retrospectively analyzed in comparison with an equal number of pair mates treated with double ASCT not including thalidomide. RESULTS: On an intention-to-treat basis, the addition of thalidomide to double ASCT effected a significant improvement in the rate (68% v 49%; P = .001) and duration (62% v 33% at 4 years; P < .001) of at least very good partial response (VGPR), time to progression (TTP; 61% v 41% at 4 years; P < .001) and progression-free survival (PFS; 51% v 31% at 4 years; P = .001). A trend was also noted for extended overall survival (OS) among thalidomide-treated patients (69% at 5 years v 53% for the control group), although the difference between the two groups was not statistically significant (P = .07). Benefits with thalidomide in increasing the rate of VGPR or better response, TTP, and PFS were confirmed in a multivariate analysis. Median OS after relapse was 24 months for patients receiving thalidomide added to double ASCT and 25 months for the control group. Overall, 17% of patients discontinued thalidomide, including 8% because of drug-related adverse events. CONCLUSION: In comparison with double ASCT, the addition of first-line thalidomide to double ASCT improved clinical outcomes. Short-term thalidomide was generally well tolerated and had no adverse impact on postrelapse survival.

Visceral leishmaniasis in a patient with common variable immunodeficiency and evans syndrome: clinical remarks.

Visceral Leishmaniasis (VL) is a vector-borne zoonosis endemic in Southern Italy whose usual clinical features include fever, splenomegaly, pancytopenia and hypergammaglobulinemia. The clinical and biochemical picture may be misleading in patients with immunodeficiency diseases hampering the diagnosis. We describe a VL case in a patient whose spleen had been removed and who had Common Variable Immunodeficiency and Evans syndrome.