Giovanni D’Arena

Chronic lymphocytic leukemia is marked by profound defects in T-cell function. Programmed death-1 is a receptor involved in tumor-mediated immunosuppression through binding of the PD-L1 ligand. Multiparametric flow cytometry and immunohistochemistry were used to study PD-1/PD-L1 expression. Functional assays were used to determine the involvement of the PD-1/PD-L1 axis in T-cell responses. PD-1 expression by CD4(+) and CD8(+) T lymphocytes was significantly higher in 117 chronic lymphocytic leukemia patients than in 33 donors of a comparable age. CD4(+) and CD8(+) T lymphocytes from chronic lymphocytic leukemia patients displayed increased numbers of effector memory and terminally differentiated cells, respectively, when compared to controls. The number of effector memory CD4(+) and terminally differentiated CD8(+) lymphocytes positively associated with a more advanced stage of disease, treatment requirements and unfavorable genomic aberrations. Furthermore, leukemic lymphocytes expressed higher levels of PD-L1 than circulating B lymphocytes from normal donors. PD-1 and PD-L1 surface expression spiked in proliferating T and B lymphocytes, suggesting that this interaction works efficiently in activated environments. Within chronic lymphocytic leukemia proliferation centers in the lymph node, CD4(+)/PD-1(+) T lymphocytes were found to be in close contact with PD-L1(+) chronic lymphocytic leukemia cells. Lastly, functional experiments using recombinant soluble PD-L1 and blocking antibodies indicated that this axis contributes to the inhibition of IFN-γ production by CD8(+) T cells. These observations suggest that pharmacological manipulation of the PD-1/PD-L1 axis may contribute to restoring T-cell functions in the chronic lymphocytic leukemia microenvironment.

BACKGROUND: Bisphosphonates (BPs) are effective in the prevention and treatment of skeletal-related events (SREs) in patients with symptomatic myeloma who are receiving chemotherapy. Recent data also suggest a possible antineoplastic activity of BPs. Few studies published to date have explored the role of BPs in patients with untreated, asymptomatic myeloma (AM). No data are available on the efficacy of zoledronic acid in these patients. METHODS: The authors conducted a prospective, multicenter, open-label, phase 3, randomized trial comparing the administration of zoledronic acid versus simple observation in patients with AM. One-hundred sixty-three patients were enrolled and randomized (1:1) to receive zoledronic acid (n = 81 patients) or not to receive zoledronic acid (n = 82 patients) for 1 year at a dose of 4 mg monthly as a single, 15-minute, intravenous infusion. RESULTS: After a median follow-up of 64.7 person-months, 44.4% of patients in the zoledronic acid group and 45.1% of the control group progressed to 'symptomatic' myeloma requiring chemotherapy (P = .9307). The median time to progression was 67 months and 59 months for the treatment and control groups, respectively (P = .8312). At progression, SREs were significantly lower in the zoledronic acid-treated group (55.5%) than in the control group (78.3%; P = .041), whereas anemia, renal failure, and extramedullary disease were not statistically different. More frequent adverse events observed in the zoledronic acid-treated group were asymptomatic hypocalcemia and fever. One patient developed reversible osteonecrosis of the jaw. No renal failure caused by zoledronic acid was reported. CONCLUSIONS: The monthly use of zoledronic acid for 1 year in patients with AM reduced the development of SREs at progression but did not influence the natural history of the disease.

Because of its relatively indolent clinical course, chronic lymphocytic leukemia (CLL) offers a versatile model for testing novel therapeutic regimens and drug combinations. Nicotinamide is the main NAD(+) precursor and a direct inhibitor of four classes of enzymes, including the sirtuins. SIRT1, the main member of the sirtuin family, inactivates p53 by deacetylating a critical lysine residue. In this study, we showed that CLL cells express high levels of functional SIRT1, which is inhibited by exogenous nicotinamide. This agent blocks proliferation and promotes apoptosis selectively in leukemic cells that express wild-type (wt) p53. Nicotinamide modulates the p53-dependent genes p21, NOXA, BAX, and Mcl-1, indicating an activation of the p53 pathway and of caspase-3. DNA-damaging chemotherapeutics, such as etoposide, activate a functional loop linking SIRT1 and p53 through the induction of miR-34a. When leukemic cells are simultaneously exposed to nicotinamide and etoposide, we observe a significant increase in miR-34a levels with a concomitant inhibition of SIRT1. Furthermore, p53 acetylation levels are higher than with either agent used alone. Overall, treatment with both nicotinamde and etoposide shows strongly synergistic effects in the induction of apoptosis. We therefore concluded that nicotinamide has the dual property of inhibiting SIRT1 through a noncompetitive enzymatic block (p53 independent) and at the same time through miR-34a induction (p53 dependent). These observations suggested the therapeutic potential of nicotinamide, a novel, safe, and inexpensive drug, to be used in addition to chemotherapy for CLL patients with wt p53.

BACKGROUND AND OBJECTIVE: One of the most important potential advantages in the use of human umbilical cord blood (HUCB) for hematopoietic reconstitution after myeloablative therapy seems to be the lower occurrence of acute graft-versus-host-disease (GvHD) in recipients after allogeneic transplantation. Since mature T cells play an important role in GvHD pathogenesis, we tried to verify whether a different immunophenotypic pattern exists between HUCB and peripheral blood (PB) T cells. DESIGN AND METHODS: An immunophenotypic study on 40 HUCB and 40 PB samples from healthy adult volunteers was performed, by means of flow cytometry using a large panel of monoclonal antibodies in double labeling. RESULTS: The absolute lymphocyte count was greater in HUCB (5233 +/- 1808 microL) than in adult PB (1941 +/- 378 microL). Significant differences in percentage were found between cord and adult T-cells, respectively (CD3+: 59.9 +/- 12 vs 74.9 +/- 4.6%), CD3- CD16+ and/or CD56+ natural killer (NK) cells (23.8 +/- 10.1 vs 10.8 +/- 5.3%) and CD3+CD16+ and/or CD56+ cytotoxic T lymphocyte subset (0.3 +/- 0.3 vs 10.7 +/- 4.1%). There was no difference in CD4/CD8 ratio (1.7 +/- 0.5 vs 1.6 +/- 0.2%) between the two groups. In absolute terms, HUCB contained an higher number of all lymphocyte subsets, with the exception of CD3+CD16+ and/or CD56+ T lymphocyte subpopulation, CD3+CD25+ and CD3+HLADR+ activated T-cells. CD38, a marker of activation and immaturity, was present on virtually all cord T cells and on approximately half of the adult T cells. The large majority of HUCB T cells co-expressed CD45RA naive antigen (CD4+CD45RA+: 87.6 +/- 5.2%, CD8+ CD45RA+: 93.5 +/- 7.8%; CD4+CD45RO+: 12.3 +/- 5.2%; CD8+CD45RO+: 6.4 +/- 7.8%) whereas in adult PB T cells an higher number of CD45RO+ memory cells was detected (CD4+CD45RA+: 44.8 +/- 9.6%; CD8+CD45RA: 71.5% +/- 8.1%; CD4+CD45RO+: 55.2 +/- 9.6%; CD8+ CD45RO+: 28.5 +/- 8.1%). Finally, less than 14% of lymphocytes were shown to belong to B lineage in both sources, while, in absolute terms, they were more represented in HUCB with respect to adult PB. INTERPRETATION AND CONCLUSIONS: In the present study we found significant difference between HUCB and adult PB lymphocytes in their immunophenotypic profile. In particular HUCB showed T lymphocytes that appeared to be phenotypically immature. Indeed, as a likely consequence of poor antigenic experience during pregnancy, the majority of HUCB cells were naive, expressing the RA isoform of the CD45 molecule. These findings could justify the previously reported reduced cord blood lymphocyte alloreactivity when allogeneic transplantation is performed and require further functional studies in order to confirm the impairment of HUCB immune system response to alloantigens.