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Background.Although influenza vaccination of healthcare workers reduces influenza-like illness and overall mortality among patients, national rates of vaccination for healthcare providers are unacceptably low. We report the implementation of a new mandatory vaccination policy by means of a streamlined electronic enrollment and vaccination tracking system at the National Institutes of Health (NIH) Clinical Center.Objective.To evaluate the outcome of a new mandatory staff influenza vaccination program.Methods.A new hospital policy endorsed by all the component NIH institutes and the Clinical Center departments mandated that employees who have patient contact either be vaccinated annually against influenza or sign a declination specifying the reason(s) for refusal. Those who fail to comply would be required to appear before the Medical Executive Committee to explain their rationale. We collected in a database the names of all physician and nonphysician staff who had patient contact. When a staff member either was vaccinated or declined vaccination, a simple system of badge scanning and bar-coded data entry captured essential data. The database was continuously updated, and it provided a list of noncompliant employees with whom to follow up.Results.By February 12, 2009, all 2,754 identified patient-care employees either were vaccinated or formally declined vaccination. Among those, 2,424 (88%) were vaccinated either at the NIH or elsewhere, 36 (1.3%) reported medical contraindications, and 294 (10.7%) declined vaccination for other reasons. Among the 294 employees without medical contraindications who declined, the most frequent reason given for declination was concern about side effects.Conclusions.Implementation of a novel vaccination tracking process and a hospital policy requiring influenza vaccination or declination yielded dramatic improvement in healthcare worker vaccination rates and likely will result in increased patient safety in our hospital.

Rituximab is a monoclonal antibody to the CD20 antigen expressed on B lymphocytes. Several recent reports suggest that rituximab may be used to treat complications of SCT including GVHD and EBV reactivation. The long term consequences on IgG recovery aren’t well studied. This is a retrospective analysis of allogeneic pediatric SCT patients treated with rituximab at Loma Linda University from 2004 to 2013. We treated 23 patients with rituximab following allogeneic SCT (11 MUD, 7 MSD, 3 Cord) for post-transplant EBV reactivation alone (4), cGVHD alone (4), cGVHD in combination with thrombocytopenia (9), plus hemolysis (4) and EBV reactivation (2). Most patients required IVIG monthly (18) or more frequently (4) to maintain serum IgG levels >500mg/dl. We examined the length of time from end of rituximab therapy to recovery of endogenous IgG production. The effect on recovery of IgG production of cGVHD therapy, EBV infection, age, type of stem cell graft, and number of doses of rituximab were evaluated. Those treated for cGVHD plus any other indication (n=19) received a median of 11 doses of rituximab (range 4 to >30). Among these patients 11 of 14 whose cGVHD resolved and came off immune suppression recovered the ability to produce IgG completely (9) or partially (2, defined as decreasing frequency of IVIG infusions to maintain IgG>500mg/dl). Median time to recovery was 14 months (range 1-50 months) after last dose of rituximab. For patients who continued to require immune suppression for cGVHD only 1 of 5 recovered IgG production 13 months after last dose of rituximab. The remaining 4 patients required monthly or greater IVIG for >3 years after the last dose of rituximab (all still requiring IVIG). There were 4 patients treated with 2-4 doses of rituximab for EBV reactivation early after SCT, and 1 treated with 11 doses for cGVHD and EBV. Only 1 of these 5 has completely recovered IgG production. The 4 other patients remain dependent on IVIG a median of >39 months since end of rituximab therapy. We found no relationship between age of patient or duration of rituximab therapy on the time interval between end of rituximab therapy and onset of endogenous IgG production. Recipients of MUD grafts had a non-significant increase in failure to recover endogenous IgG production, but when the impact of therapy for cGVHD was taken into account the type of stem cell graft lost significance. Ability to recover endogenous IgG production following treatment with rituximab after SCT depends on successful treatment of cGVHD and in this group of patients requires about 1 year from end of therapy. Among those treated with rituximab whose cGVHD does not resolve and in those with early EBV reactivation, loss of IgG production persists at least several years, and this may be unrelated to rituximab therapy. Disclosures Off Label Use: rituximab for cGVHD.

Norovirus infection is a major cause of nonbacterial gastroenteritis and is a self-limited infection in immunocompetent patients. However, in immune compromised patients, norovirus has been reported to cause prolonged infection resulting in complications such as graft versus host disease and sepsis due to mucosal breakdown. Supportive care is the only current therapy for norovirus as attempts to treat norovirus with ribavirin or oral immunoglobulin have been unsuccessful. Nitazoxanide is a thiazolide antimicrobial with broad activity against anaerobic bacteria, protozoa, and a range of viruses in cell culture models. The antiviral activity of nitazoxanide may relate to activation of natural host antiviral defenses, but there is also evidence for a direct-acting antiviral effect on cellular processes possibly through inhibition of virus protein production/maturation and/or assembly. We report our experience treating norovirus gastroenteritis occurring in 12 patients after (9) or prior to (3) HSCT with Nitazoxanide. From November 2012 to July 2013, 12 patients (2 receiving chemotherapy, 1 autologous HSCT and 9 allogeneic HSCT) developed norovirus gastroenteritis. Ages ranged from 1 to 21 years (median 10) diagnoses included ALL/AML (6), aplastic anemia (2), and 1 each for osteopetrosis, Wiskott Aldrich, neuroblastoma, and brain tumor. Norovirus was detected by RNA RT-PCR test of stool performed by Focus Diagnostics, Cypress, Ca. The dose of Nitazoxanide was 100 mg po BID for ages 1 to 4 years, 200 mg po BID for age 4 to 11 years, and 500 mg po BID for greater than 11 years. One patient, 33 months post HSCT with normal immune studies was not treated as his symptoms resolved prior to availability of test results. All other patients clinically responded with improvements in diarrhea, nausea, and abdominal pain within 2-4 days (median 2 days). Three patients were pre-HSCT on chemo/immunotherapy and 8 patients were 1 day to 34 months after HSCT. All of the treated patients were on immune suppression or chemotherapy. Eight allogeneic HSCT patients were on immunosuppression and four of these patients had GVHD at onset of symptoms. Immune suppression included tacrolimus/solumedrol (3), cellcept/solumedrol (2) plus infliximab (1), tacrolimus (1), cyclosporine (1). Three patients were receiving immunotherapy (1), or chemotherapy for solid tumors (2) prior to planned HSCT. Clearance of stool virus was variable. Two of 3 patients treated prior to HSCT became negative on stool study within 3-14 days of treatment (1 unknown duration). Among patients treated after HSCT 5 of 8 had persistent viral shedding, 2 received drug until death (1 adenovirus, 1 congestive heart failure) both were treated greater than 2 months, 3 with GVHD continue to shed virus after 6 months of treatment, and 3 have come off therapy and remain negative for norovirus RNA. One HSCT patient with clinical resolution but persistent viral shedding stopped treatment resulting in re-occurrence of clinical symptoms. This patient responded clinically to reinstitution of therapy within 2 days but continues to shed virus. UGI endoscopy and colonoscopy were performed in 5 patients at the time of clinical infection, all showed inflammation and edema but no GVHD was seen on histology. Peripheral blood CD4 counts among those with persistent viral shedding ranged from Nitazoxinide is effective therapy for norovirus gastroenteritis in immune compromised patients. Therapy needs to be continued until stool RNA studies become negative. Disclosures: Off Label Use: Nitazoxide is a thiazolide antimicrobial that is FDA approved for treatment of cryptosporidium and is known to have broad spectrum activity to bacteria and viral infections. We used it to treat norovirus infections in immune compromised SCT patients.

COX-2 has been implicated in the development of many epithelial cancers, as well as in tumor angiogenesis. COX-2 inhibitors have been shown to have anti-tumor activity in experimental cancer. Little information exists, however, on the expression or role of COX-2 in hematologic malignancies. We have use a variety of immunochemical assays to document expression of COX-2 in human and murine leukemias and hematopoietic cells. The factor-dependent murine cell lines FDCP1 and 32D expressed COX-2 when growing continuously in the presence of IL-3; expression declined markedly when growth factor was removed. FDCP1 cells constitutively expressing bcl-2, pim-1, or bcr-abl had markedly elevated levels of COX-2, and continued to express this enzyme even after removal of growth factor. To assess COX-2 expression in human hematopoietic cells we developed a flow cytometry assay using a FITC-labelled anti-COX-2 MoAb (Cayman). Cells were washed once in serum-free medium, fixed briefly in 1% paraformaldehyde, permeabilized with PBS/0.2% Triton X100, then stained with antibody. Negative control samples were processed similarly but stained with antibody that had been preincubated with immunizing peptide. Specific COX-2 staining was interpreted as the difference between the histograms from blocked versus unblocked anti-COX-2 antibody, as determined by Kolmogorov-Smirnoff analysis. In buffy coat preparations from normal donors, we found constitutive expression of COX-2 in lymphocytes (both B-cells and T-cells). In contrast little or no COX-2 was detected in unstimulated neutrophils or monocytes. In human acute myelogenous leukemia (AML) cell lines we found COX-2 expression to be universal and easily detected. In several cell lines we confirmed the results of our flow cytometry assay with immunoblotting. We further examined 25 cryopreserved samples of human acute leukemia blasts obtained from peripheral blood. COX-2 expression was variable, but universal. Levels generally were less than those seen in immortalized cell lines, and did not correlate with blasts morphology (AML, ALL, APL, AMoL, CML-BT). To determine if COX-2 inhibitors could play a role in the treatment of acute leukemias, we performed cytotoxicity assays using the COX-2 specific inhibitors, celecoxib and NS398. Survival and growth of human AML cell lines were inhibited by both agents. These data demonstrate that 1) a variety of oncogenes can induce expression of COX-2 in hematopoietic cells, 2) clinical human acute leukemias uniformly express COX-2 in circulating blasts, and 3) COX-2 inhibitors are cytotoxic for human leukemia cells. Combination therapies for acute leukemias may evaluate the incorporation of COX-2 inhibitors for added cytotoxic effects or angiogenesis inhibition.

The murine B-cell differentiation antigen Lyb-2 has been shown to be involved in B-lymphocyte activation and has been postulated by some to be related to a receptor for B-cell stimulatory factor I (BSF-1) (H. Yakura et al., J. Immunol. 137 , 1475, 1986). Here we have demonstrated that monoclonal antibody (mAB) to Lyb-2 resembles BSF-1 in its ability to activate small resting B cells and enhancement of surface Ia. Anti-Lyb-2 antibodies bound B cells with very high avidity and were able to induce mobilization of cytosolic-free calcium. Anti-Lyb-2 mAB differs from BSF-1 in that BSF-1 but not anti-Lyb-2 is able to synergize with anti-μ in induction of B-cell proliferation. The relation between Lyb-2 molecules and BSF-1 receptors was tested in assays that measure binding of anti-Lyb-2 or BSF-1 in B cells and were found not to compete with each other. It appears that the two B-cell agonists anti-Lyb-2 and BSF-1 may exert their effects on B cells through different cell surface moieties as well as different intracellular pathways.