Infectious complications among individuals with clinical monoclonal B-cell lymphocytosis (MBL): a cohort study of newly diagnosed cases compared to controls.

Although the risk of progression from monoclonal B-cell lymphocytosis (MBL) to chronic lymphocytic leukemia (CLL) has been well characterized, it is unknown whether other common complications associated with CLL, such as increased risk of infection, occurs in individuals with MBL. We used the Mayo CLL database to identify cohorts of individuals with newly diagnosed MBL (n=154) or newly diagnosed CLL (n=174) who resided within 50 miles of Mayo Clinic. A cohort of 689 adult patients seen for a general medical examination who resided within 50 miles of Mayo clinic and who enrolled in a case-control study of non-Hodgkin lymphoma (NHL) was used as a comparison cohort. Hospitalization with infection was more common among individuals with MBL (25/154; 16.2%), and CLL (32/174; 18.4%) than controls (18/689; 2.6%). On pooled multivariable Cox proportional hazards analysis of all 1017 patients (controls, MBL and CLL), male sex (hazards ratio (HR)=2.3; P=0.002), major co-morbid health problems (HR=1.7, P=0.04), the presence of CLL (HR=3.2, P<0.001), treatment for progressive CLL (HR=2.4, P=0.001) and the presence of MBL (HR=3.0, P=0.001) were independently associated with risk of hospitalization for infection. These results suggest the risk of serious infection in clinical MBL is substantially greater than the risk of progression requiring treatment.

Immunophenotypic and gene expression analysis of monoclonal B-cell lymphocytosis shows biologic characteristics associated with good prognosis CLL.

Monoclonal B-cell lymphocytosis (MBL) is a hematologic condition wherein small B-cell clones can be detected in the blood of asymptomatic individuals. Most MBL have an immunophenotype similar to chronic lymphocytic leukemia (CLL), and 'CLL-like' MBL is a precursor to CLL. We used flow cytometry to identify MBL from unaffected members of CLL kindreds. We identified 101 MBL cases from 622 study subjects; of these, 82 individuals with MBL were further characterized. In all, 91 unique MBL clones were detected: 73 CLL-like MBL (CD5(+)CD20(dim)sIg(dim)), 11 atypical MBL (CD5(+)CD20(+)sIg(+)) and 7 CD5(neg) MBL (CD5(neg)CD20(+)sIg(neg)). Extended immunophenotypic characterization of these MBL subtypes was performed, and significant differences in cell surface expression of CD23, CD49d, CD79b and FMC-7 were observed among the groups. Markers of risk in CLL such as CD38, ZAP70 and CD49d were infrequently expressed in CLL-like MBL, but were expressed in the majority of atypical MBL. Interphase cytogenetics was performed in 35 MBL cases, and del 13q14 was most common (22/30 CLL-like MBL cases). Gene expression analysis using oligonucleotide arrays was performed on seven CLL-like MBL, and showed activation of B-cell receptor associated pathways. Our findings underscore the diversity of MBL subtypes and further clarify the relationship between MBL and other lymphoproliferative disorders.

Cyclophosphamide, bortezomib and dexamethasone induction for newly diagnosed multiple myeloma: high response rates in a phase II clinical trial.

We have studied a three-drug combination with cyclophosphamide, bortezomib and dexamethasone (CyBorD) on a 28-day cycle in the treatment of newly diagnosed multiple myeloma (MM) patients to assess response and toxicity. The primary endpoint of response was evaluated after four cycles. Thirty-three newly diagnosed, symptomatic patients with MM received bortezomib 1.3 mg/m(2) intravenously on days 1, 4, 8 and 11, cyclophosphamide 300 mg/m(2) orally on days 1, 8, 15 and 22 and dexamethasone 40 mg orally on days 1-4, 9-12 and 17-20 on a 28-day cycle for four cycles. Responses were rapid with a mean 80% decline in the sentinel monoclonal protein at the end of two cycles. The overall intent to treat response rate (>or= partial response) was 88%, with 61% of very good partial response or better (>or=VGPR) and 39% of complete/near complete response (CR/nCR). For the 28 patients who completed all four cycles of therapy, the CR/nCR rate was 46% and VGPR rate was 71%. All patients undergoing stem cell harvest had a successful collection. Twenty-three patients underwent stem cell transplantation (SCT) and are evaluable through day 100 with CR/nCR documented in 70% and >or=VGPR in 74%. In conclusion, CyBorD produces a rapid and profound response in patients with newly diagnosed MM with manageable toxicity.

Isolation in the allogeneic transplant environment: how protective is it?

Aggressive infection control measures that include isolating patients within protective hospital environments have become a standard practice during allogeneic stem cell transplantation. A wide range of interventions includes the management of ventilation systems, BMT unit construction and cleaning, isolation and barrier precautions, interactions with health-care workers and visitors, skin and oral care, infection surveillance, and the prevention of specific nosocomial and seasonal infections. However, many of these practices have not been definitively proven to provide patients the intended benefit of decreased infection rates or improved survival. Furthermore, each intervention comes with a financial and social cost. With institutional cost containment efforts and recent trials suggesting that patients may be safely cared for in the outpatient environment after allogeneic transplantation, many widely held practices in managing the transplant environment are being reconsidered. With changing practices, transplant teams are encouraged to review local patterns of infections and associated complications and communicate regularly with infection control committees for guidance on the evolution of isolation needs for the immunosuppressed patient.

Toxic megacolon: a life-threatening complication of high-dose therapy and autologous stem cell transplantation among patients with AL amyloidosis.

AL amyloidosis is a plasma cell disorder in which tissue deposition of immunoglobulin light chains leads to organ dysfunction. Recent reports of high-dose therapy with autologous stem cell transplantation for amyloidosis suggest higher response rates and extended survival compared to those seen with conventional chemotherapy. However, substantial treatment-related toxicity has been observed. This case series describes our institutional experience with autologous transplantation in four patients with amyloidosis with an emphasis on unique gastrointestinal toxicities, including toxic megacolon.

Marrow transplants from unrelated donors for patients with aplastic anemia: minimum effective dose of total body irradiation.

Patients with aplastic anemia who do not have suitably HLA-matched, related donors generally receive immunosuppressive treatment as first-line therapy and are considered for transplantation from an unrelated donor only if they fail to respond to immunosuppressive treatment. In this setting, rates of transplantation-related morbidity and mortality have been high. We conducted a prospective study to determine the minimal dose of total body irradiation (TBI) sufficient to achieve sustained engraftment when it is used in combination with 3 cycles of 30 mg/kg of antithymocyte globulin (ATG) and 4 cycles of 50 mg/kg of cyclophosphamide (CY). We also wanted to determine the tolerability and toxicity of the regimen. The starting dosage of TBI was 3 x 200 cGy given over 2 days following CY/ATG. The TBI dose was to be escalated in increments of 200 cGy if graft failure occurred in the absence of prohibitive toxicity, and de-escalated for toxicity in the absence of graft failure. Twenty-one female and 29 male patients aged 1.3 to 46.5 years (median age, 14.4 years) underwent transplantation at 14 medical centers. The time interval from diagnosis to transplantation was 2.8 to 264 months (median, 14.5 months). All patients had been transfused multiple times and all had received 1 to 11 courses (median, 4 courses) of immunosuppressive treatment and other modalities of treatment. In 38 cases, the donors were HLA-A, -B and -DR phenotypically matched with the patients, and, in 12 cases, the donor phenotype differed from that of the recipient by 1 HLA antigen. Recipients of mismatched transplants were considered separately for TBI dose modification, and this study is still ongoing. Seven patients did not tolerate ATG and were prepared with 6 x 200 cGy of TBI plus 120 mg/kg of CY. Of the HLA-matched recipients prepared with CY/ATG/TBI, all 20 who received 3 x 200 or 2 x 200 cGy of TBI achieved engraftment, and 10 are alive. Of the 13 patients who received 1 x 200 cGy of TBI, 1 failed to engraft, and 8 are alive. Each of 10 patients who received an HLA-nonidentical transplant achieved engraftment, and 3 of 6 who were given 3 x 200 cGy of TBI, and 4 of 4 who were given 2 x 200 cGy are alive. Pulmonary toxicity occurred in 8 of 30 patients who were given 3 x 200 or 2 x 200 cGy of TBI concurrently with ATG and CY at 200 mg/kg, and in 2 of 13 patients who received 1 x 200 cGy of TBI, a pattern that suggests a decrease in toxicity with TBI dose de-escalation. Overall, the highest probability of survival (73%) was observed among patients who underwent transplantation within 1 year of diagnosis, compared with patients who underwent transplantation after a longer period of disease. In addition, younger patients (aged < or = 20 years) were more likely to survive than older patients (aged > 20 years). Thus, for patients with an HLA-matched, unrelated donor, a TBI dose of 200 cGy (in combination with CY/ATG) was sufficient to allow for engraftment without inducing prohibitive toxicity. As in previous studies, patient age and pretransplantation disease duration remain important prognostic factors.

The fission yeast protein p73res2 is an essential component of the mitotic MBF complex and a master regulator of meiosis.

Depending on environmental conditions, Schizosaccharomyces pombe can remain in the stationary phase or enter into either premitotic or premeiotic DNA synthesis. This decision point is known as Start. In the mitotic cell cycle, regulation of G1/S-specific gene expression is dependent upon the MBF (Mlu1 binding factor) complex, known to contain p85cdc10 and p72res1. Here we demonstrate that p73res2 controls cell cycle progression via its participation in the MBF complex, interacting directly with both p85cdc10 and p72res1. In contrast, when cells enter into meiosis, the MBF complex is disrupted, and p73res2 shifts its regulatory function towards the transactivation of genes required for meiotic progression. These observations suggest that p73res2 plays a pivotal role at Start and constitutes an example of a transcription factor involved in the control of both mitotic and meiotic progression.

The Schizosaccharomyces pombe MBF complex requires heterodimerization for entry into S phase.

In Schizosaccharomyces pombe, MBF is a DNA-binding complex suspected to activate the transcription of genes necessary for entry into S phase. The MBF complex contains both p85cdc10 and p72res1/sct1. To obtain a better understanding of how the MBF complex regulates gene expression at the G1/S transition, we have performed a genetic analysis of p72res1. We determined that p72res1 can bind specifically to the cdc22 promoter, when analyzed by gel mobility shift assay, and that the N-terminal 157 amino acids of p72res1 are sufficient for this specific binding. When overexpressed in vivo, a fragment of p72res1 containing this DNA-binding domain could rescue a strain carrying a temperature-sensitive cdc10 allele at the restrictive temperature as well as a strain with a cdc10 null allele. We also determined that the C-terminal region of p72res1 is necessary and sufficient for binding to p85cdc10. Overexpression of the cdc10-binding domain of p72res1 leads to a G1 arrest with a cdc phenotype and a decrease on MBF activity. Overexpression of full-length p72res1 also leads to a growth arrest that can be rescued by overexpression of p85cdc10. These results imply that the MBF activity in vivo is dependent on the interaction of p85cdc10 with p72res1.

Demonstration of separate phosphotyrosyl- and phosphoseryl- histone phosphatase activities in the plasma membranes of a human astrocytoma.

A plasma membrane preparation from a human astrocytoma contained p-nitrophenyl phosphate (pNPP), phosphotyrosyl histone, and phosphoseryl histone hydrolysis activities. The pNPPase and phosphotyrosyl histone phosphatase activities were inhibited by vanadate, whereas the phosphoseryl histone phosphatase activity was not; the latter activity was inhibited by pyrophosphate and nucleoside di- and triphosphates. When the membranes were solubilized by Triton X-100 and the solubilized proteins were subjected to column chromatography on DEAE-Sephadex, Sepharose 6B-C1, and wheat germ agglutinin-Sepharose 4B columns, the pNPPase activity from the phosphoseryl histone phosphatase activity. The results from column chromatography also indicated that there may be multiple phosphotyrosyl and phosphoseryl protein phosphatases in the plasma membranes.

Amplification of the murine leukemia virus Mr 70,000 glycoprotein gene product by human xenografts in athymic mice.

Passage of human tumors in athymic mice is accompanied by an increase in serum levels of the Mr 70,000 murine leukemia virus envelope protein, gp70. Elevated levels of gp70 can be detected in tissues of the hematopoietic systems of mice bearing human xenografts, but there is no evidence of synthesis of gp70 in these tissues. By far, the highest concentration of gp70 is in the human xenografts themselves. When assayed for gp70, 8 human xenografts and 12 cell lines established from human xenografts were all positive. In the plasma membrane of the human astrocytoma xenograft, T24, the gp70 was found to be approximately 10% of the total membrane protein. In contrast, the concentration of the Mr 30,000 viral core protein, p30, was 17-fold less. Only trace amounts of complete infectious virus could be detected. A human prostate carcinoma line that had not been grown in the athymic mice was found to have no gp70, but was shown to be able to synthesize gp70 after a single passage in the athymic mice.

An acid phosphatase in the plasma membranes of human astrocytoma showing marked specificity toward phosphotyrosine protein.

The plasma membrane from the human tumor astrocytoma contains an active acid phosphatase activity based on hydrolysis of p-nitrophenyl phosphate. Other acid phosphatase substrates--beta-glycerophosphate, O-phosphorylcholine, and 5'-AMP--are not hydrolyzed significantly. The phosphatase activity is tartrate insensitive and is stimulated by Triton X-100 and EDTA. Of the three known phosphoamino acids, only free O-phosphotyrosine is hydrolyzed by the membrane phosphatase activity. Other acid phosphatases tested from potato, wheat germ, milk, and bovine prostate did not show this degree of specificity. The plasma membrane activity also dephosphorylated phosphotyrosine histone at a much greater rate than did the other acid phosphatases. pH profiles for free O-phosphotyrosine and phosphotyrosine histone showed a shift toward physiological pH, indicating possible physiological significance. Phosphotyrosine histone dephosphorylation activity was nearly 10 times greater than that seen for phosphoserine histone dephosphorylation, and Km values were much lower for phosphotyrosine histone dephosphorylation (0.5 microM vs. 10 microM). Fluoride and zinc significantly inhibited phosphoserine histone dephosphorylation. Vanadate, on the other hand, was a potent inhibitor of phosphotyrosine histone dephosphorylation (50% inhibition at 0.5 microM) but not of phosphoserine histone. ATP stimulated phosphotyrosine histone dephosphorylation (160-250%) but inhibited phosphoserine histone dephosphorylation (95%). These results suggest the existence of a highly specific phosphotyrosine protein phosphatase activity associated with the plasma membrane of human astrocytoma.

Isolation and characterization of plasma membranes from transplantable human astrocytoma, oat cell carcinoma, and melanomas.

Purified plasma membranes were obtained from five transplantable human tumors, a grade IV astrocytoma, an oat cell carcinoma, and three melanomas. Plasma membrane fractions were isolated from tumor homogenates by differential and discontinuous sucrose gradient centrifugation. Determination of enzyme activities indicated that the plasma membranes were enriched 10- to 20-fold with respect to 5'-nucleotidase, nicotinamide adenine dinucleotide glycohydrolase, Mg2+-activated nucleoside triphosphatase, and sialic acid. Specific activities of nearly all the enzymes varied with the individual tumors, even among tumors of the same type, i.e., the melanomas. Electron micrographs of the plasma membrane fractions showed smooth single-membrane vesicles with slight contamination by lysosomes. Therefore, these membranes are suitable for comparative biochemical studies and for the preparation of tumor-specific monoclonal antibodies. Plasma membranes from all five tumors contained very high Mg2+-adenosine triphosphatase (ATPase) activities. The Na+-K+-ATPase was a minor component of the total ATPase of these membranes (less than 30%). The major component was an ATPase exhibiting similar activity toward several nucleoside triphosphates. The activity of such a nucleoside triphosphatase has been correlated with tumorigenicity in cultured liver epithelial cells. The nucleoside triphosphatase of the plasma membranes of astrocytoma and oat cell carcinoma was stimulated from 50 to 1005 by concanavalin A, whereas ATPase of the melanoma plasma membranes was not or only slightly stimulated. The different response to concanavalin A could be due to differences in the ATPase molecules of the individual tumors or to the different environment of the ATPase.