Anna Maria Barbui

Prevention of hepatitis B virus recurrence after liver transplantation in cirrhotic patients treated with lamivudine and passive immunoprophylaxis

BACKGROUND/AIMSnTreatment with hepatitis B virus immune globulins (HBIG) or lamivudine has reduced the rate of hepatitis B recurrence after liver transplantation to approximately 50%.nnnMETHODSnTo further decrease hepatitis B recurrence, 33 hepatitis B virus (HBV)-related cirrhotic patients were treated with lamivudine before liver transplantation and with lamivudine together with low-dose HBIG (46 500 IU the first month followed by 5,000 lU/monthly) after surgery.nnnRESULTSnWhile on lamivudine, serum HBV DNA level decreased significantly in all patients and in 11 (33%) the Child-Pugh score improved. Twenty-six patients were transplanted. Among the 25 who survived for longer than 12 months, only one (4%) experienced a hepatitis B recurrence over an average follow-up of 31 months, a rate significantly lower (P = 0.0002) than the 50% recurrence rate among a historical control group of 12 patients. However, low-level HBV replication was detected sporadically throughout the follow-up in 64% of patients.nnnCONCLUSIONSnOver the medium-term, combined prophylaxis with lamivudine and HBIG significantly decreases the risk of hepatitis B recurrence after liver transplantation. Though low-level HBV infection recurred in two thirds of patients, the pathogenic expression of HBV was prevented.

Genetic PTX3 Deficiency and Aspergillosis in Stem-Cell Transplantation

BACKGROUNDnThe soluble pattern-recognition receptor known as long pentraxin 3 (PTX3) has a nonredundant role in antifungal immunity. The contribution of single-nucleotide polymorphisms (SNPs) in PTX3 to the development of invasive aspergillosis is unknown.nnnMETHODSnWe screened an initial cohort of 268 patients undergoing hematopoietic stem-cell transplantation (HSCT) and their donors for PTX3 SNPs modifying the risk of invasive aspergillosis. The analysis was also performed in a multicenter study involving 107 patients with invasive aspergillosis and 223 matched controls. The functional consequences of PTX3 SNPs were investigated in vitro and in lung specimens from transplant recipients.nnnRESULTSnReceipt of a transplant from a donor with a homozygous haplotype (h2/h2) in PTX3 was associated with an increased risk of infection, in both the discovery study (cumulative incidence, 37% vs. 15%; adjusted hazard ratio, 3.08; P=0.003) and the confirmation study (adjusted odds ratio, 2.78; P=0.03), as well as with defective expression of PTX3. Functionally, PTX3 deficiency in h2/h2 neutrophils, presumably due to messenger RNA instability, led to impaired phagocytosis and clearance of the fungus.nnnCONCLUSIONSnGenetic deficiency of PTX3 affects the antifungal capacity of neutrophils and may contribute to the risk of invasive aspergillosis in patients treated with HSCT. (Funded by the European Society of Clinical Microbiology and Infectious Diseases and others.).

Treatment of recurrent hepatitis C in liver transplants: efficacy of a six versus a twelve month course of interferon alfa 2b with ribavirin ☆

BACKGROUND/AIMSnInterferon (IFN) with ribavirin combination therapy (CT) was proposed for the treatment of hepatitis C recurring in liver transplants. We assessed the efficacy of two protocols of CT in transplanted patients with recurrent severe hepatitis C virus (HCV) hepatitis.nnnMETHODSnFifty-seven patients (68% genotype 1b) were treated with IFN alfa-2b 3 million units three times weekly and oral ribavirin 800mg/die for 6 or 12 months. Study end-points were the end of treatment (ETVR) and the 12-month post-therapy sustained virologic response (SVR; negative HCV-RNA).nnnRESULTSnETVR was induced in 9/27 (33%) and in 7/30 patients (23%) treated, respectively, for 6 and 12 months (P=0.4); a SVR was induced in six (22%) of the former and five (17%) of the latter (P=0.4). HCV genotype non-1 patients responded better than genotype 1 (SVR: 43% in genotype non-1 versus 12% in genotype 1, P: 0.02). In ETV responders the hepatitis activity index improved by >2 points in biopsies taken after therapy compared to pre-therapy biopsies. Anemia and leukopenia required reduction of therapy in 51% of the patients.nnnCONCLUSIONSnCT is efficacious in controlling HCV disease in about 20% of transplants with recurrent hepatitis C. Six months of therapy are as efficacious as 12 months.

Genetic Modification of Human T Cells with CD20: A Strategy to Purify and Lyse Transduced Cells with Anti-CD20 Antibodies

A retroviral vector has been constructed that contains the human CD20 cDNA under the control of the Moloney murine leukemia virus (Mo-MuLV) LTR. Freshly isolated mononuclear cells are infected for three consecutive days in the presence of PHA and hrlL-2, and a mean 15.9% of the cells (range, 6.5 to 31.7%) acquire a CD3+CD20+ phenotype. Transduced T lymphocytes grow and expand in vitro for up to 3 weeks like mock-infected cells and, as observed for the T lymphoblastoid CEM cell line, CD20 expression is maintained for several months with no change in the growth curve of the cells. CD20-expressing CEM and fresh T lymphocytes can be positively immunoselected on columns using different anti-CD20 antibodies. Exposure to monoclonal chimeric anti-CD20 IgG1(kappa) Rituximab antibody (Roche), in the presence of complement, results in effective and rapid killing of the transduced CD3+CD20+ human T cells in vitro. This approach represents a new and alternative method to gene manipulation with suicide genes for the production of drug-responsive T cell populations, a crucial step for the future management of graft-versus-host disease in bone marrow transplant patients.

Risk Factors for the Development of Secondary Malignancy After High-Dose Chemotherapy and Autograft, With or Without Rituximab: A 20-Year Retrospective Follow-Up Study in Patients With Lymphoma

PURPOSEnHigh-dose chemotherapy with peripheral blood progenitor cell (PBPC) autograft is effective in high-risk lymphoma, particularly with the addition of rituximab; however, it is associated with risk of secondary malignancy. These issues have been addressed in a series of 1,347 patients with lymphoma treated with a high-dose sequential (HDS) program.nnnPATIENTS AND METHODSnA total of 1,024 patients with B-cell lymphoma, 234 patients with Hodgkins lymphoma, and 89 patients with T-cell lymphoma were treated with HDS between 1985 and 2005 at 11 Gruppo Italiano Terapie Innovative Linfomi centers. HDS was given as salvage treatment to 707 patients (52%); 655 patients (49%) received a modified HDS, with high-dose cytarabine and two consecutive PBPC harvests. Rituximab-supplemented HDS was given to 523 patients (39%).nnnRESULTSnAt a median follow-up of 7 years, the median overall survival (OS) was 16.2 years; in B-cell lymphoma the OS was significantly superior with rituximab HDS compared to HDS alone. The cumulative incidence at 5 and 10 years of secondary myelodysplasia/acute leukemia (sMDS/AL) were 3.09% and 4.52%, respectively, that of solid tumors were 2.54% and 6.79%, respectively. Factors associated with sMDS/AL were male sex and use of the second harvest PBPC for the graft; factors found to be associated with solid tumor were advanced age, post-HDS radiotherapy, and rituximab addition to HDS. Despite the increased risk of solid tumors, rituximab addition to HDS was still associated with survival advantages.nnnCONCLUSIONnThis analysis has relevant implications for the design and use of intensive chemoimmunotherapy with autograft. In addition, it offers useful insights toward the understanding and prevention of tumor development.

Negative selection of peripheral blood stem cells to support a tandem autologous transplantation programme in multiple myeloma

Summary. We recently described a two‐step negative selection procedure whereby peripheral blood stem cells (PBSCs) were efficiently purged of contaminating neoplastic cells by a combination of monoclonal antibodies. Here, we report 60 newly diagnosed multiple myeloma (MM) patients treated with a double transplant programme and randomized to receive either unmanipulated or in vitro purged PBSCs. We demonstrated that this technique is feasible and safe without significant loss of either CD34+ or CD3+ cells. Haematological engraftment and immunological reconstitution were rapid without treatment‐related mortality. Using polymerase chain reaction (PCR), we compared the level of minimal residual disease (MRD) in PBSC before and after in vitro purging and in vivo after transplant. A median of one tumour cell per 102 normal cells (range 101−105) was seen in the unmanipulated aphereses with a 3–4 log reduction after manipulation in vitro. However, despite this tumour debulking, all patients remained PCR positive in vivo. At 3u2003years, the estimated event‐free survival was 40% in the control arm and 72% in the experimental arm (Pu2003=u20030·05), whereas the estimated overall survival was 83% in both arms. This suggests that autologous transplantation using efficiently purged PBSCs can be performed safely, but confirms the need for innovative protocols for MRD eradication in vivo.

Zygomycosis in Italy: a Survey of FIMUA-ECMM (Federazione Italiana di Micopatologia Umana ed Animale and European Confederation of Medical Mycology)

Abstract The aims of the study were to analyze the clinical and epidemiological characteristics and treatments for patients who developed zygomycosis enrolled in italy during the european Confederation of medical mycology survey. This prospective multicenter study was performed between 2004 and 2007 at 49 italian Departments. 60 cases of zygomycosis were enrolled: the median age was 59.5 years (range 1-87), with a prevalence of males (70%). The majority of cases were immunocompromised patients (42 cases, 70%), mainly hematological malignancies (37). Among non-immunocompromised (18 cases, 30%), the main category was represented by patients with penetrating trauma (7/18, 39%). The most common sites of infection were sinus (35%) with/without CNS involvement, lung alone (25%), skin (20%), but in 11 cases (18%) dissemination was observed. According to EORTC criteria, the diagnosis of zygomycosis was proven in 46 patients (77%) and in most of them it was made in vivo (40/46 patients, 87%); in the remaining 14 cases (23%) the diagnosis was probable. 51 patients received antifungal therapy and in 30 of them surgical debridement was also performed. The most commonly used antifungal drug was liposomal amphotericin b (L- AmB), administered in 44 patients: 36 of these patients (82%) responded to therapy. Altogether an attributable mortality rate of 32% (19/60) was registered, which was reduced to 18% in patients treated with L-AmB (8/44). Zygomycosis is a rare and aggressive filamentous fungal infection, still associated with a high mortality rate. This study indicates an inversion of this trend, with a better prognosis and significantly lower mortality than that reported in the literature. It is possible that new extensive, aggressive diagnostic and therapeutic procedures, such as the use of L-AmB and surgery, have improved the prognosis of these patients.

Correlation between Cytomegalovirus Infection and Raynaud’s Phenomenon in Lupus nephritis

Relationships between viruses and autoimmune diseases such as systemic lupus erythematosus (SLE) are still elusive. Recent reports demonstrated the association of some viral infections with peculiar clinical events in the general population, such as cytomegalovirus (CMV) with arterial damage and Parvovirus B19 (PV-B19) with hematologic abnormalities. We planned to look for this kind of viral imprinting in SLE, hypothesizing that traces of specific features of some viral infections might be found in some subsets of seropositive SLE patients. In 60 SLE patients recruited at our nephrologic center, serology for CMV, PV-B19, Epstein-Barr virus viral capsid antigen (EBV-VCA), Epstein-Barr nuclear antigen (EBNA) and Epstein-Barr virus early antigen (EBV-EA) was performed. χ2 and ANOVA were employed to compare the frequency and titers of antiviral antibodies in SLE patients with groups of transplant, hemodialysis and blood donor subjects. χ2, Fisher’s test, Bonferroni and Scheffe’s test were employed to compare the different biochemical/clinical features between seropositive and seronegative SLE patients. Univariate and multivariate analysis (logistic regression models) were employed to evaluate the odds ratio (OR) of different risk factors for vascular events (including Raynaud’s phenomenon, deep venous thrombosis) and hematologic abnormalities (including severe anemia, leukopenia and thrombocytopenia). Anti-CMV (82%), anti-PV-B19 (60%), anti-EBV-VCA (92%) and EBV-EA (45%) IgG antibodies were frequent in SLE, with higher prevalence in comparison with the blood donor group and higher titers in comparison with transplant and hemodialysis groups. CMV seropositivity was a highly significant risk factor for Raynaud’s phenomenon (OR +α in univariate and multivariate analysis = 13.51 using a correction of 0.5 in case of a zero event), but not for venous vascular events (OR = 1.31). An increased though not significant risk factor was found for antiphospholipid antibodies (OR = 2.71, p = 0.19), while the presence of nephrotic syndrome during the follow-up was a significant protective factor (OR = 0.15, p = 0.035). There was no significantly increased OR for PV-B19 seropositivity in cases with severe anemia (OR = 2.09, p = 0.29). No significant associations were found with the status of EBV reactivation. In conclusion, our results support the hypothesis that viral infection may imprint the course of SLE leading to specific clinical subsets (i.e. CMV and ‘vascular’ SLE, with more frequent Raynaud’s phenomenon and a less frequent typical histological renal picture responsible for nephrotic syndrome). Further prospective studies are justified to validate these correlations, mainly dealing with associations between acute viral infections and vascular events, thus eventually leading to a better understanding of mutual relationships between viruses and SLE.

Occult hepatitis B virus infection in HBsAg negative patients undergoing liver transplantation: Clinical significance

Occult Hepatitis B virus (o‐HBV) infection has been reported in HB surface antigen (HBsAg)‐negative liver donors whose risk of transmitting HBV justifies a specific prophylaxis in liver recipients. The clinical significance of o‐HBV infection in HBsAg‐negative recipients and their need for prophylaxis is unknown. Liver samples collected during surgery from 23 HBsAg‐negative patients (9 liver donors and 14 recipients) and 20 HBsAg‐positive recipients (controls) were studied by polymerase chain reaction with an independent set of primers mapping the core and surface HBV genes. Intrahepatic HBV DNA was detected as core and surface genes in all the HBsAg‐positive recipients, in none of the HBsAg‐negative donors and in 9/14 (64%) of the HBsAg‐negative recipients (2 HCV negative, 7 HCV positive). The intrahepatic amount of HBV was significantly lower in HBsAg‐negative than in HBsAg‐positive livers (median values 1.36 Log10/μg DNA vs. 3.66 Logs, p<0.0001, core gene, and 1.13 vs. 6.21 Logs p<0.0001, surface gene). No HBV DNA was detected in plasma from o‐HBV recipients; one of them tested positive in lymphocytes. No correlation was found between o‐HBV and serologic markers of previous HBV exposure, response to vaccination, acute rejection, hepatitis D and G virus‐infections. None of o‐HBV carriers experienced a de novo hepatitis B after transplantation (median follow‐up: 477 days). Occult HBV is frequent in HBsAg‐negative liver recipients. It is not associated with increased episodes of acute rejection, coinfection with hepatotropic viruses, different responses to HBV vaccination, or the development of de‐novo hepatitis B. In o‐HBV infection a particular virus‐host interaction can explain the low intrahepatic HBV content and the lack of extrahepatic HBV replication, thus justifying the low risk of hepatitis B reactivation, in absence of specific prophylaxis, once the recipient liver is removed. (Liver Transpl 2004;10:356–362.)

The lymphocyte to monocyte ratio improves the IPI‐risk definition of diffuse large B‐cell lymphoma when rituximab is added to chemotherapy

The peripheral blood lymphocyte to monocyte ratio (LMR) at diagnosis can be clinically relevant in patients with diffuse large B‐cell lymphoma (DLBCL). We reviewed the outcome of 1,057 DLBCL patients followed from 1984 to 2012 at four centers. LMR was analyzed as a clinical biomarker by receiver‐operating characteristic (ROC) analysis and Harrells C‐statistics. Patients were characterized by a median age of 61 years, International Prognostic Index (IPI) score of >2 in 39%, and were treated with a rituximab‐containing chemotherapy in 66%. LMR proved strongly predictive for survival in patients treated with rituximab‐based programs, but not in those receiving chemotherapy alone. Additionally, an LMR value of ≤2.6 (as determined by ROC analysis) was associated with a worst performance status, a higher lactate dehydrogenase (LDH) level, an advanced clinical stage, and a higher IPI score (Pu2009=u20090.000). In patients treated with rituximab‐supplemented chemotherapy programs, an LMR value of <2.6 was found in most of the primary refractory patients (75%) which proved as the best cutoff to predict both response and survival (Pu2009=u20090.018). Finally, multivariate analysis and Harrells C‐statistics confirmed the IPI‐independent role of LMR on survival (Pu2009=u20090.0000). In conclusion, LMR is a potent predictor of clinical response and survival in DLBCL treated with rituximab‐containing chemotherapy. Am. J. Hematol. 88:1062–1067, 2013.