bluuebunny/crossref_metadata_2025 huggingface dataset:
Incorporation of Radioactive L-Cystine by Normal and Leukemic Leukocytes in Vivo
Authors: WEISBERGER AUSTIN S. , LEVINE BENNETT
Published: October 2019

Methods for studying the in vivo incorporation of radioactive amino acids by leukocytes are described. The incorporation of S35 L-cystine into normal and leukemic leukocytes was studied and compared with the utilization of S35 L-methionine, S35 sodium sulfate, and C14 sodium formate.

Radioactive L-cystine is rapidly absorbed and rapidly incorporated into the proteins of leukocytes. There are distinctive differences in rates and levels of utilization of S35 L-cystine by normal leukocytes and by leukocytes of acute and chronic leukemia.

The leukocytes of acute leukemia and of chronic myeloid leukemia have a greater avidity for L-cystine than normal leukocytes. This rapid utilization of an amino acid by immature leukocytes is not limited to L-cystine and is probably a manifestations of the rapid metabolic turnover characteristic of immature cells in general. However, in view of the observations on the specificity of L-cysteine and related compounds in modifying the leukotoxic effects of nitrogen mustard, the avidity of immature leukocytes for L-cystine may have greater significance than the utilization of other metabolites.

The data indicate that the life span of normal leukocytes is approximately 13 days. The life span of leukocytes in chronic myeloid leukemia is slightly less than that of normal leukocytes whereas the life span of acute leukemic leukocytes is much shorter than that of normal leukocytes. The life span of lymphocytes in chronic lymphatic leukemia appears to be much longer than that of normal leukocytes. Although L-cystine and L-methionine are readily utilized its acute leukemia, there is marked impairment in the utilization of sodium sulfate in vivo. By comparison, normal leukocytes readily utilize sodium sulfate in vivo. This discrepancy may be due to the rapid turnover of sulfur in acute leukemia or to an intrinsic metabolic defect of acute leukemic leukocytes.
Observations on the New Rh Agglutinin Anti-f
Authors: JONES ALAN RICHARDSON , STEINBERG ARTHUR G. , ALLEN FRED H. , DIAMOND LOUIS K. , KRIETE BERTRAND
Published: October 2019

1. An example of the new anti-Rh serum, anti-f, is described.

2. It is shown that anti-c and anti-e sera may contain anti-f.

3. The significance of the antigen f is discussed in terms of its genetics and practical importance.
Iron Metabolism in Steatorrhea
Authors: BADENOCH J. , CALLENDER SHEILA T.
Published: October 2019

The absorption and utilization of radioactive iron has been studied in sixteen patients with steatorrhea and a control group of twelve patients with hypochromic anemia without steatorrhea and three subjects without anemia.

The patients with idiopathic steatorrhea showed a poor absorption of the test dose given by mouth. This is in keeping with their poor clinical response to oral treatment with iron.

There is some evidence to suggest that poor absorption in such patients may not be the sole factor involved in the development of the anemia.
Observations on the Thrombocytopenia Due to Hypersensitivity to Quinidine
Authors: BARKHAM PERCY , TOCANTINS LEANDRO M.
Published: October 2019

Studies are reported on a patient who developed thrombocytopenic purpura following quinidine therapy. Quinidine caused lysis of the patient’s platelets in vitro within 40 minutes and complete inhibition of clot retraction. Platelet lysis preceded platelet agglutination. Increasing the concentration of the sodium citrate used as anticoagulant inhibited the in vitro action of the quinidine. No thrombocytolytic effect of the patient’s serum with quinidine could be shown when tested against normal blood.

A test dose of quinidine administered orally after the patient had recovered from the thrombocytopenia produced a profound decrease in the number of platelets within 90 minutes, associated with a prolongation of the bleeding time and disappearance of clot retraction.

The in vitro hypersensitivity to quinidine persisted for at least four weeks after recovery from the purpura; after six weeks it could no longer be demonstrated.
Proaccelerin Deficiency Following Irradiation with Therapeutic Doses of Radiophosphorus
Authors: VETTER HERBERT , VINAZZER HELMUT
Published: October 2019

In six out of seven polycythemic patients treated with radiophosphorus a significant decrease of the proaccelerin content of the plasma was observed.

This diminution began to become detectable by means of semiquantitative estimations on the second day after the radioactive isotope was given and lasted for three to sixteen days.

The degree of the deficiency seemed to depend on the amount of radiophosphorus administered, only the highest dose having been capable of impairing the coagulation severely.
Immunopancytopenia Associated with Incomplete Cold Hemagglutinins in a Case of Primary Atypical Pneumonia
Authors: MOESCHLIN S. , SIEGENTHALER W. , GASSER C. , HÄSSIG A.
Published: October 2019

In a case of primary atypical pneumonia (blood grounp O), incomplete antibodies of the cold agglutinin type with a very wide thermal spectrum (35 to 37 C.) led to a severe immunopancytopenia in the form of severe hemolytic anemia, agranulocytosis, and thrombocytopenia.

It is obvious that hemolysis was not induced by cold agglutinins which were active 1:1024 at 4 C. and only 1:32 at 22 C. The indirect cold antibodies which were probably responsible for the hemolysis which occurred were thermolabile (1:16 at 22 C., 1:8 at 37 C.) and could be demonstrated only in the antiglobulin plasma test. During the acute phase, too, a hemolysin of the Dacie type active for normal erythrocytes, but even more active for trypsinized cells, at 37 C. and at a pH of 6.8 and 7.6 could be demonstrated. The severe hemolysis could be stopped by ACTH treatment but the titer of the complete and incomplete cold antibodies showed no distinct change and the direct and indirect Coombs test remained positive for a long time. However, there was a distinct diminution of the thermal amplitude of the incomplete antibodies.

A severe agranulocytosis (140 granulocytes) developed which could be improved each time by cortisone. However, permanent improvement resulted only after the disappearance of the incomplete antibodies one year later.

A leukopenic factor could be demonstrated by transfusion of 300 cc. blood to a recipient of the same blood group producing a pronounced fall in the granulocytes of from 4200 to 910. Control transfusions from a normal inidividual to the same recipient showed no change and the same is true for the transfusion experiment performed one year later from the now cured patient to the same recipient. Leukocyte agglutination tests were negative, but it is presumed that agglutinins of the incomplete type may have been present. Opsonins could not be demonstrated. The thrombocytopenia never descended below 50,000.

The bone marrow showed a distinct shift to the left, i.e. a predominance of unripe forms. On the basis of our previous experimental work (amidopyrine agranulocytosis) it is believed that the changes in the marrow are brought about by a depletion and exhaustion of the bone marrow due to the enormously increased peripheral destruction of the granulocytes by agglutination rather than to an actual inhibition of the marrow.

The relation of this immunoleukopenia in our previous findings to the occurrence of leukocyte agglutination in agranulocytosis of allergic origin is discussed. Agranulocytosis and leukopenia may be produced on an immunologic basis in many other cases and thus some form of agranulocytosis may be related to the mechanism of erythrocyte destruction in acquired hemolytic anemia and to the agglutination of thrombocytes ins essential thrombocytopenia on an immunologic basis.
Atypical Amyloidosis Associated With Nonthrombocytopenic Purpura and Plasmocytic Hyperplasia of the Bone Marrow
Authors: PROPP SIMON , SCHARFMAN WILLIAM B. , BEEBE RICHARD T. , WRIGHT ARTHUR W.
Published: October 2019

1. Two cases of chronic nonthrombocytopenic purpura of unusual distribution, associated with generalized amyloidosis and plasma cell hyperplasia of the bone marrow, are described in detail with their postmortem findings.

2. Grossly recognizable amyloid was present in the antemortem sternal marrow aspirates of both patients.

3. Skin sections in each patient revealed hemorrhages associated with marked amyloid deposition in the dermis and blood vessel walls. It is postulated that this purpura was of vascular type.

4. The diagnosis of the underlying pathology in both patients was made possible by marrow aspiration. The importance of proper use of marrow particle smears in the diagnosis of plasma cell disturbances in the bone marrow is emphasized.

5. The difficulty in differentiating plasma cell myeloma from other causes of plasma cell hyperplasia of the marrow is clearly documented in this report.
Complement in Hemolytic Disease of the Newborn
Authors: TRIANTAPHYLLOPOULOS DEMETRIOS , GREENWALT TIBOR J.
Published: October 2019

1. The 50 per cent complement levels in thirty-three normal newborn infants and twenty-three infants with D (Rh0)-hemolytic disease were determined.

2. No statistically significant difference in complement values was found between the two groups.

3. In vitro tests with ten anti-D (Rh0) sera demonstrated that there was no fixation of human complement.
Leukemia in Atomic Bomb Survivors
Authors: LANGE ROBERT D. , MOLONEY WILLIAM C. , YAMAWAKI TOKUSO
Published: October 2019

1. Observations on seventy-five established cases of leukemia occurring in people exposed to atomic bomb radiation are presented.

2. These data indicate a great increase in the incidence of leukemia among atomic bomb survivors due to a single massive exposure to ionizing radiation.

3. The leukemogenic effects of radiation are manifested equally in both sexes and at all age levels represented in this series.

4. The striking preponderance of chronic myelogenous leukemia compared to chronic lymphatic leukemia has been noted in exposed individuals but it is pointed out that chronic lymphatic leukemia is comparatively rare among the Japanese.

5. Cases of leukemia are still appearing in atomic bomb survivors. However, since 1950 there has been a steady decline in the number of cases.
Polycythemic Response in Normal Adult Rats to a Nonprotein Plasma Extract from Anemic Rabbits
Authors: BORSOOK HENRY , GRAYBIEL ASHTON , KEIGHLEY GEOFFREY , WINDSOR EMANUEL
Published: October 2019

Experiments are described in which a protein-free extract of plasma obtained from anemic rabbits was injected into healthy young rats. The resulting erythropoietic response was comparable to that induced by hypoxia. The findings suggest that a humoral factor was at work which was capable of disturbing the physiologic mechanisms which establish the equilibrium between red blood cell formation and destruction. The far-reaching significance of these findings is apparent.
Predictors of Long-Term Response to High-Dose Interferon Therapy in Type II Cryoglobulinemia Associated With Hepatitis C Virus Infection
Authors: Casato Milvia , Agnello Vincent , Pucillo Leopoldo P. , Knight Glenn B. , Leoni Marco , Del Vecchio Savino , Mazzilli Cristina , Antonelli Guido , Bonomo Lorenzo
Published: October 2019

We have prospectively studied patients with type II cryoglobulinemia since 1985 to assess the efficacy of treatment with interferon-α at cumulative doses ranging from 234 to 849 MU. In the present study we retrospectively evaluated in this cohort parameters associated with complete response to therapy in 31 consecutive patients with type II cryoglobulinemia associated with hepatitis C virus (HCV) infection. Prevalence of complete response of cryoglobulinemia (disappearance of symptoms and signs of vasculitis and decrease of cryocrit below 10% of the initial value) was 62%, with a median response duration of 33 months and a range of 3 to 100 months. Three patients were putatively cured, as they remained in complete remission for more than 5 years off therapy. Eighteen patients (58%) had liver disease evidenced by histopathology and/or raised transaminase levels. Prevalence of normalization of transaminase levels was 100%, with a median response duration of 36 months. Relapse of hypertransaminasemia occurred in 100% and 8% of patients receiving less than or greater than 621 MU, respectively. By logistic regression analysis, the only pretherapy parameter that associated significantly (P = .0393) with complete response of cryoglobulinemia was the solitary anti-C22 (HCV core) antibody pattern, which was observed in 29% of patients. Association with older age and low cryocrit approached statistical significance (P = .06), while no significant correlations were found with serum IgM levels, duration of disease, HCV genotype, NS5a gene mutations, liver histology, HLA-DR phenotype, or WA cross-idiotype. Complete responses were also associated, on univariate statistical analysis, with low pretherapy HCV viremia. Responses were accompanied by decrease of viremia, of anti-HCV antibody levels and cryocrit. The usefulness of a high dose regimen is underscored by the higher rates of sustained responses of cryoglobulinemia and transaminase levels compared with previous studies.
Pooled Normal Human Polyspecific IgM Contains Neutralizing Anti-Idiotypes to IgG Autoantibodies of Autoimmune Patients and Protects From Experimental Autoimmune Disease
Authors: Hurez Vincent , Kazatchkine Michel D. , Vassilev Tchavdar , Ramanathan Sheela , Pashov Anastas , Basuyaux Bertrand , de Kozak Yvonne , Bellon Blanche , Kaveri Srini V.
Published: October 2019

Normal human serum contains IgM antibodies that regulate the natural autoantibody activity of IgG in autologous serum. In the present study, we show that pooled normal human IgM (IVIgM) purified from plasma of more than 2,500 healthy donors and processed in a similar fashion to that of therapeutic preparations of pooled normal human IgG (IVIg) suppresses activity of IgG autoantibodies purified from the serum of patients with autoimmune diseases in vitro. The inhibitory effect of IVIgM was greater or equivalent to that of IVIg on a molar basis. We show that IVIgM contains anti-idiotypic antibodies directed against idiotypic determinants of autoantibodies, in particular by showing that Sepharose-bound IVIgM selectively retained F(ab′)2 fragments of IgG autoantibodies. The infusion of (Lewis × Brown-Norway) F1 rats with IVIgM protected the animals against experimental autoimmune uveitis induced by immunization with the soluble retinal S antigen, as evidenced by clinical scoring and histopathological analysis. The present findings provide a rationale for considering pooled IgM for immunomodulation of autoimmune disease.
The Plasmacytoma Resistance Gene, Pctr2, Delays the Onset of Tumorigenesis and Resides in the Telomeric Region of Chromosome 4
Authors: Mock Beverly A. , Hartley Janet , Le Tissier Paul , Wax Judith S. , Potter Michael
Published: October 2019

Mouse plasmacytomas share pathogenetic features in common with both multiple myeloma and Burkitt's lymphoma in humans. Susceptibility to plasmacytoma induction by intraperitoneal pristane in mice is controlled by multiple genes. At least two of these genes reside on mouse chromosome 4 in regions of the genome sharing linkage homology with human chromosomes 9p21, 1p32, and 1p36. A series of congenic strains recombinant for regions of mouse chromosome 4 in the vicinity of the Pctr2 predisposition locus were created and typed for their tumor susceptibility/resistance phenotypes. These strains were derived by introgressively backcrossing alleles from resistant DBA/2 mice onto the susceptible BALB/cAnPt background. Six resistant and two susceptible strains were allelotyped for 10 genes and 49 random DNA markers to identify the smallest region of overlap in the resistant strains. These studies have determined that the Pctr2 locus resides in either a 500-kb interval proximal to Nppa, or in a 1- to 2-centiMorgan (cM) interval distal to Nppa. In these congenic strain analyses, the Nppa and Fv1 loci, in addition to genes within about 1 cM of these loci, have been excluded as candidates for the Pctr2 locus. A relevant locus that may reside in this interval is Rep2; it is associated with the efficiency of repairing X-ray induced DNA damage sustained during the G2 phase of the mitotic cycle. The Pctr2 locus acts in a codominant fashion. F1 hybrids between resistant and susceptible congenic strains exhibit a reduced tumor incidence and a significant delay in the onset of tumorigenesis. Identification and eventual cloning of the Pctr2 locus may assist in the identification of genes involved in many types of cancer showing aberrations in human chromosome 1p36.
Failure of Immunologic Purging in Mantle Cell Lymphoma Assessed by Polymerase Chain Reaction Detection of Minimal Residual Disease
Authors: Andersen Niels S. , Donovan John W. , Borus Joseph S. , Poor Christina M. , Neuberg Donna , Aster Jon C. , Nadler Lee M. , Freedman Arnold S. , Gribben John G.
Published: October 2019

To assess the clinical significance of minimal residual disease (MRD) detection by polymerase chain reaction (PCR) we analyzed samples from 26 patients with mantle cell lymphoma (MCL) who had undergone bone marrow transplantation (BMT) at the Dana-Farber Cancer Institute. The BCL-1/IgH translocation and clonally rearranged Ig heavy chain genes (IgH) provided molecular markers for detection and follow-up of MRD by polymerase chain reaction (PCR) amplification in 19 of the 26 (73%) MCL patients studied. IgH gene sequencing analysis showed somatic mutations in MCL that are characteristic of an antigen driven process suggesting that, in MCL, the final malignant transformation occurs in a mature B cell. Of the 19 patients with a PCR amplifiable marker, 17 underwent autologous, 1 an allogeneic, and 1 a syngeneic bone marrow transplantation (BMT). All patients had PCR-detectable MRD in the bone marrow (BM) at the time of BMT, irrespective of any history of histological BM involvement. In contrast to other B-cell malignancies, we found that immunological purging with complement-mediated lysis eradicated PCR-detectable MCL in only two patients. Moreover reinfusion of MRD was associated with a poor outcome. More than half of the patients undergoing autologous BMT had relapsed by the time of restaging at 2 years after autologous BMT. In four MCL patients in whom no residual lymphoma was reinfused, including the allogeneic and the syngeneic BMT, only one patient relapsed. Persistence of MRD detection after BMT was also associated with a high probability of relapse, although one patient did not have PCR-detectable MRD in peripheral blood or BM before relapse at nodal sites. We conclude that PCR amplification of disease-specific markers is a feasible and sensitive method to assess MRD and its clinical significance in patients with MCL. Moreover, PCR amplification provides a tool to evaluate modifications of purging and stem cell collection procedures that may be required for the management of this otherwise incurable disease.
Retroviral-Mediated Transfer of the Green Fluorescent Protein Gene Into Murine Hematopoietic Cells Facilitates Scoring and Selection of Transduced Progenitors In Vitro and Identification of Genetically Modified Cells In Vivo
Authors: Persons Derek A. , Allay James A. , Allay Esther R. , Smeyne Richard J. , Ashmun Richard A. , Sorrentino Brian P. , Nienhuis Arthur W.
Published: October 2019

We have investigated the utility of the green fluorescent protein (GFP) to serve as a marker to assess retroviral gene transfer into hematopoietic cells and as a tool to identify and enrich for cells expressing high levels of the vector-encoded transcript. GFP, by virtue of a naturally occurring chromophore encoded in its primary sequence, displays autonomous fluorescence, thus eliminating the need for antibody or cytochemical staining to detect its expression. A bicistronic murine stem cell virus (MSCV)-based retroviral vector was constructed containing the GFP cDNA and a mutant, human dihydrofolate reductase gene. High-titer, ecotropic retroviral producer cells free of replication competent virus were generated and used to transduce murine bone marrow cells by cocultivation. Within 24 hours after completion of the transduction procedure, a high proportion (40% to 70%) of the marrow cells were intensely fluorescent compared to mock-transduced cells or cells transduced with a control retrovirus. Erythroid and myeloid hematopoietic colonies derived from GFP-transduced marrow were easily scored for retroviral gene transfer by direct in situ fluorescence microscopy. Clonogenic progenitors expressing increased levels of antifolate drug resistance could be enriched from the GFP-transduced marrow population by fluorescence activated cell sorting of cells expressing high levels of GFP. In vivo, splenic hematopoietic colonies and peripheral blood cells from animals transplanted with GFP-transduced marrow displayed intense fluorescence. These results show that GFP is an excellent marker for scoring and tracking gene-modified hematopoietic cells and for allowing rapid selection and enrichment of transduced cells expressing high levels of the transgene.
CD40 Ligand Induces an Antileukemia Immune Response In Vivo
Authors: Dilloo Dagmar , Brown Michael , Roskrow Marie , Zhong Wanyung , Holladay Martha , Holden William , Brenner Malcolm
Published: October 2019

Leukemia cells may express tumor specific antigens in association with Class I and II major histocompatability complex (MHC) molecules. However, lack of expression of conventional costimulator molecules means that these cells tend to induce specific T-cell anergy rather than activation. CD40 ligand (CD40L) is a costimulator molecule that directly activates T cells and may promote antigen presentation by CD40-expressing cells, which include professional antigen presenting cells and B-acute lymphoblastic leukemia (ALL) cells from many patients. We determined whether transgenic expression of CD40L could enhance an antileukemia immune response using a CD40+ murine lymphoblastic (A20) leukemia and a CD40− myeloblastic (WEHI-3) leukemia in a tumor treatment model. Injection of otherwise nonimmunogenic A20 cells in the presence of CD40L induced an immune response active against preexisting A20 tumor at a distant site. Moreover, concomitant local secretion of transgenic interleukin-2 (IL-2) further amplified the antileukemic response induced and increased protection against preexisting tumor. In ex vivo studies, CD40 activation of A20 cells enhances the antigen presenting potential of A20 cells by upregulating expression of B7.1 (CD80), Class I and II MHC molecules, and increases expression of fas antigens. The importance of CD40 activation to the resulting antitumor response is further emphasized by the failure of transgenic CD40L to protect against the CD40− WEHI myeloblastic leukemia. Depletion studies showed the protective effects against A20 cells to be mediated by a combination of CD4+ and CD8+ T lymphocytes and by natural killer (NK) cells. These results suggest a means by which CD40+ leukemia cells may be rendered immunogenic in vivo.
Allelotype Analysis in the Evolution of Chronic Myelocytic Leukemia
Authors: Mori Naoki , Morosetti Roberta , Lee Stephen , Spira Susanne , Ben-Yehuda Dina , Schiller Gary , Landolfi Raffaele , Mizoguchi Hideaki , Koeffler H. Phillip
Published: October 2019

To elucidate the genetic events that may play important roles in the progression of chronic myelocytic leukemia (CML), we performed allelotype analysis in 30 patients with CML as the disease transformed to accelerated phase or blast crisis (21 myeloid and 9 lymphoid cases). DNAs were extracted from slides of bone marrow smears or from freshly isolated bone marrow mononuclear cells. The DNAs from the same individuals in both chronic phase and either blast crisis or accelerated phase were analyzed at 82 microsatellite markers, which mapped to each of the autosomal arms except the short arms of the acrocentric chromosomes. Loss of heterozygosity (LOH) on at least one locus was observed in 21 of the 30 cases (70%) as the disease progressed. Frequent allelic loss of ≥ 20% of the informative cases was observed on chromosome arms 1p (35%), 7p (21%), 19p (20%), and 20q (29%). Allelic losses were also analyzed according to phenotypes. LOH of ≥ 20% was detected on 1p (29%), 18p (20%), and 20q (27%) in myeloid blast crisis, and on 1p (50%), 4p (25%), 7p (43%), 9p (29%), 18q (25%), 19p (43%), and 20q (33%) in lymphoid blast crisis. Serial cytogenetic information was available for most of our cases with LOH on these arms, and only one case had loss of both chromosomes 9 and 20. Fractional allelic loss, calculated for each sample as the total number of chromosomal arms lost/total number of arms with information, showed a median value of 0.06 and a mean of 0.098 (range 0 to 0.60). These results suggest that tumor suppressor genes especially on 1p, 7p, 19p, and 20q probably have an important role in the progression to blast crisis of CML.
High-Dose Chemotherapy With Autologous Bone Marrow Rescue in Children With Poor-Risk Burkitt's Lymphoma: A Report From the European Lymphoma Bone Marrow Transplantation Registry
Authors: Ladenstein Ruth , Pearce Rachel , Hartmann Olivier , Patte Catherine , Goldstone Tony , Philip Thierry
Published: October 2019

To evaluate the role of high-dose chemotherapy (HDC) followed by autologous bone marrow transplantation (ABMT) in children with poor-prognosis Burkitt's lymphoma, the European Lymphoma BMT registry was critically reviewed. Between February 1979 and July 1991, a selected group of 89 children (78 boys and 11 girls) were considered as ABMT candidates in 12 European cancer centers for the following reasons: poor initial response (PIR) to first-line chemotherapy in 28 patients, primary refractory disease (PRD) in nine patients, sensitive relapse (SR) in 38 patients, and resistant relapse (RR) in 14 patients. The median age at ABMT was 8.2 years (range, 2.8 to 16.2 years). Thus, this report reflects data for patients surviving the salvage attempt deemed appropriate for HDC/ABMT and who then actually underwent the transplant procedure. The median follow-up period after HDC/ABMT was 4.3 years (range, 2 to 12 years). The prognosis was dismal for PRD patients and those with RR, ie, all patients died within 1 year. The 5-year event-free survival (EFS) was 56.6% (P < .0001) for patients in partial remission (PR) and 48.7% (P = .002) for patients with SR. The toxic death rate was 11.1%. Continuous complete remissions (CRs) in 39.4% of these otherwise incurable children highlight the fact that HDC/ABMT was an effective complementary procedure after conventional-dose chemotherapy protocols used during the given period. In addition, these data show that patients with PRD or RR clearly had no advantage from this aggressive and cost-intensive procedure. It has to be considered that the need for HDC/ABMT has greatly diminished in parallel with the improvement in survival using the modern intensive pulsed CCT of current protocols. To further rescue patients failing to respond to modern protocols, new approaches appear necessary, ie, combinations of HDC with antibody-targeted therapy plus allogeneic BMT for the additional benefits of the potential graft-versus-lymphoma effect.
Peripheral Blood Progenitor Cell Mobilization Using Stem Cell Factor in Combination With Filgrastim in Breast Cancer Patients
Authors: Glaspy J.A. , Shpall E.J. , LeMaistre C.F. , Briddell R.A. , Menchaca D.M. , Turner S.A. , Lill M. , Chap L. , Jones R. , Wiers M.D. , Sheridan W.P. , McNiece I.K.
Published: October 2019

The safety and optimal dose and schedule of stem cell factor (SCF ) administered in combination with filgrastim for the mobilization of peripheral blood progenitor cells (PBPCs) was determined in 215 patients with high-risk breast cancer. Patients received either filgrastim alone (10 μg/kg/d for 7 days) or the combination of 10 μg/kg/d filgrastim and 5 to 30 μg/kg/d SCF for either 7, 10, or 13 days. SCF patients were premedicated with antiallergy prophylaxis. Leukapheresis was performed on the final 3 days of cytokine therapy and, after high-dose chemotherapy and infusion of PBPCs, patients received 10 μg/kg/d filgrastim until absolute neutrophil count recovery. The median number of CD34+ cells collected was greater for patients receiving the combination of filgrastim and SCF, at doses greater than 10 μg/kg/d, than for those receiving filgrastim alone (7.7 v 3.2 × 106/kg, P < .05). There were significantly (P < .05) more CD34+ cells harvested for the 20 μg/kg/d SCF (median, 7.9 × 106/kg) and 25 μg/kg/d SCF (median, 13.6 × 106/kg) 7-day combination groups than for the filgrastim alone patients (median, 3.2 × 106/kg). The duration of administration of SCF and filgrastim (7, 10, or 13 days) did not significantly affect CD34+ cell yield. Treatment groups mobilized with filgrastim alone or with the cytokine combination had similar hematopoietic engraftment and overall survival after PBPC infusion. In conclusion, the results of this study indicate that SCF therapy enhances CD34+ cell yield and is associated with manageable levels of toxicity when combined with filgrastim for PBPC mobilization. The combination of 20 μg/kg/d SCF and 10 μg/kg/d filgrastim with daily apheresis beginning on day 5 was selected as the optimal dose and schedule for the mobilization of PBPCs.
Overexpression of Human Stem Cell Factor Impairs Melanocyte, Mast Cell, and Thymocyte Development: A Role for Receptor Tyrosine Kinase-Mediated Mitogen Activated Protein Kinase Activation in Cell Differentiation
Authors: Kapur Reuben , Everett Eric T. , Uffman Josh , McAndrews-Hill Monica , Cooper Ryan , Ryder John , Vik Terry , Williams David A.
Published: October 2019

Stem cell factor (SCF) is synthesized as both soluble (S) and membrane-associated (MA) proteins. Indirect insight into the function of MA and S isoforms of SCF has come from studies performed in Steel (Sl) mutant mice. However, the physiologic role(s) of these two isoforms remain unknown. In an attempt to better understand the in vivo role of c-kit/SCF interactions on various cell lineages, transgenic mice were generated that overexpress MA isoform of human SCF (hSCF). In murine cells, hSCF behaves as an antagonist to normal SCF function, due to interference with the interaction between endogenous murine SCF and its receptor, c-kit, encoded by the dominant white spotting (W) gene. Mice expressing the hSCF transgene display a variety of phenotypic abnormalities, which are accentuated when combined with W alleles. Here we show that mice homozygous for the hSCF transgene demonstrate a coat color deficiency seen in some mice homozygous for mild W alleles. Specifically, homozygous hSCF transgenic mice (hSCF220) display a pronounced forehead blaze, with additional white spots over the cervical region, as well as a very large belly spot. Doubly heterozygous animals that carry both a mutated W allele and the hSCF transgene also display an unusual pigment defect and a dramatic reduction in the number of dermal mast cells. Furthermore, overexpression of MA hSCF in the thymus results in abnormal thymocyte differentiation and proliferation, which is associated with reduced mitogen activated protein (MAP) kinase activation. Thus, MAP kinase activation by a receptor tyrosine kinase, such as c-kit, may be critical for the differentiation of thymocytes in vivo.
Abnormal Expression of Mouse Mast Cell Protease 5 Gene in Cultured Mast Cells Derived From Mutant mi/mi Mice
Authors: Morii Eiichi , Jippo Tomoko , Tsujimura Tohru , Hashimoto Koji , Kim Dae-Ki , Lee Young-Mi , Ogihara Hideki , Tsujino Kumiko , Kim Hyung-Min , Kitamura Yukihiko
Published: October 2019

Mast cells contain a lot of mast cell-specific proteases. We have reported that the expression of mouse mast cell protease 6 (MMCP-6) is remarkably reduced in both cultured mast cells (CMCs) and skin mast cells of mi/mi mutant mice. In the present study, we found that the expression of MMCP-5 was reduced in CMCs but not in skin mast cells of mi/mi mice, and we compared the regulation mechanisms of MMCP-5 with those of MMCP-6. The mi locus encodes a member of the basic-helix-loop-helix-leucine zipper (bHLH-Zip) protein family of transcription factors (hereafter called MITF ). The consensus sequence recognized and bound by bHLH-Zip transcription factors is CANNTG. The overexpression of the normal (+) MITF but not of mi-MITF normalized the poor expression of the MMCP-5 gene in mi/mi CMCs, indicating the involvement of +-MITF in transactivation of the MMCP-5 gene. Although +-MITF directly bound CANNTG motifs in the promoter region of the MMCP-6 gene and transactivated it, the binding of +-MITF to the CAGTTG motif in the promoter region of the MMCP-5 gene was not detectable. The +-MITF appeared to regulate the transactivation of the MMCP-5 gene indirectly. Moreover, addition of stem cell factor to the medium normalized the expression of the MMCP-5 but not of the MMCP-6 gene in mi/mi CMCs. Despite the significant reduction of both MMCP-5 and MMCP-6 expressions in mi/mi CMCs, their regulation mechanisms appeared to be different.
A Leu117 → Trp Mutation Within the RGD-Peptide Cross-Linking Region of β3 Results in Glanzmann Thrombasthenia by Preventing αIIbβ3 Export to the Platelet Surface
Authors: Basani Ramesh B. , Brown Deborah L. , Vilaire Gaston , Bennett Joel S. , Poncz Mortimer
Published: October 2019

We report a case of Glanzmann thrombasthenia in a Pakistani child whose platelets express less than 10% of the normal amount of αIIbβ3 on their surface. Single-stranded conformation polymorphism analysis of the exons of the patient's αIIb and β3 genes showed an abnormality in exon 4 of the β3 gene. Direct sequence analysis showed that the patient was homozygous for a T → G nucleotide substitution in this exon, resulting in the replacement of a highly conserved Leu at position 117 with Trp. Heterologous expression of αIIbβ3 containing the β3 mutation in COS-1 cells confirmed the pathogenicity of the Leu117 → Trp substitution and showed that it resulted in the intracellular retention of malfolded αIIbβ3 heterodimers. Additional site-directed mutagenesis at position 117 indicated that, although the smaller hydrophobic amino acid Val could be substituted for the wild-type Leu, the larger hydrophobic amino acids Trp and Phe or the charged amino acids Asp and Lys were not tolerated. These studies indicate that Leu117 in β3 plays a critical role in attaining the correct folded conformation of αIIbβ3. These studies also suggest that the hydrophobic side chain of Leu117 is likely folded into the interior of β3, where it serves to stabilize internal packing of the protein and determines its overall shape.
Molecular Characterization of the t(8; 13)(p11;q12) Translocation Associated With an Atypical Myeloproliferative Disorder: Evidence for Three Discrete Loci Involved in Myeloid Leukemias on 8p11
Authors: Still Ivan H. , Chernova Olga , Hurd David , Stone Richard M. , Cowell John K.
Published: October 2019

A reciprocal chromosome translocation between 13q12 and 8p11 is the consistent cytogenetic abnormality seen in a nonspecific myeloproliferative disorder that is associated with T-cell leukemia/lymphoma and peripheral blood eosinophilia. Detailed molecular analyses of the translocation breakpoints associated with this rearrangement have not been reported to date. We have now generated somatic cell hybrids from a newly described patient with this specific structural rearrangement and analyzed the breakpoints on the derivative chromosomes. We have shown that the breakpoint on chromosome 13 lies within a 300- to 500-kb region defined by the KIAA177 gene and D13S1123 marker. In addition, we have identified a 1.2-Mb YAC, 959A4, that crosses the translocation breakpoint on the short arm of chromosome 8 in this patient. The location of this breakpoint in 8p11 is distinct from the t(8; 16) and t(8; 22) translocations associated with M4/M5 myeloid leukemias, and suggests that three distinct loci located within 8p11 are involved in the pathogenesis of myeloid neoplasias.
Prognostic Significance of Bax Protein Expression in Diffuse Aggressive Non-Hodgkin's Lymphoma
Authors: Gascoyne Randy D. , Krajewska Maryla , Krajewski Stanislaw , Connors Joseph M. , Reed John C.
Published: October 2019

Bax is a proapoptotic member of the Bcl-2 protein family. The incidence and prognostic significance of Bax protein expression in diffuse non-Hodgkin's lymphomas with a large cell component (DLCL) was determined by an immunohistochemical method by using paraffin-embedded tumors from a cohort of patients treated uniformly with combination chemotherapy (n = 139). All patients were between 16 and 70 years of age and had advanced stage disease of diffuse large cell type (diffuse mixed, diffuse large cell, immunoblastic, or anaplastic large cell). Paraffin sections from diagnostic biopsies were successfully immunostained for Bax in 113 cases. Of these, 7 (6%) tumors were scored as Bax immunonegative (<1% Bax-stained tumor cells), 42 (37%) as low (1% to 10%), 9 (8%) as low-intermediate (11% to 30%), 25 (22%) as high-intermediate (31% to 70%), and 30 specimens (27%) as high for Bax expression (<70%). Of the 7 Bax-immunonegative lymphomas, all also scored low (≤10% immunostained tumor cells) for Bcl-2 expression, whereas 78 of the 106 (74%) Bax-immunopositive tumors had low Bcl-2 expression. By itself, Bax expression was not of prognostic significance in univariate analysis, although there was a clear trend for patients with Bax-immunonegative lymphomas (n = 7) to relapse sooner and to die faster than patients whose tumors contained Bax-immunopositive malignant cells (n = 106; 8-year overall survival 29% versus 55%; P = .06). When combined with Bcl-2 immunostaining data, Bax provided additional prognostic information. Among patients with Bcl-2 low-expressing DLCLs, for example, Bax immunonegativity was associated with lower 8-year relapse-free survival (RFS; 29% v 61%; P < .01) and lower 8-year overall survival (OS; 29% v 63%; P < .05), suggesting that absence of Bax protein connotes a more aggressive phenotype when Bcl-2 protein is also not expressed at high levels. In contrast, low Bax expression was associated with improved 8-year disease-free survival (52% v 16%; P < .02), RFS (47% v 11%; P < .02), and OS (64% v 11%; P < .01) in patients whose tumors expressed Bcl-2 at high levels, suggesting that the combination of high levels of Bax and Bcl-2 expression is more deleterious than high levels of Bcl-2 expression alone. Bax expression failed to provide additional prognostic information beyond Bcl-2 expression in multivariate analysis that included the clinical International Prognostic Index factors (age, stage, lactate dehydrogenase, performance status, and number of extranodal sites) and immunophenotype. Taken together, the results suggest that Bax expression is not a major prognostic marker in DLCL. However, the interactions of the Bcl-2 and Bax expression data with respect to clinical outcome may shed new insights into the biological significance of Bcl-2/Bax protein heterodimerization.
Induction of Vascular Endothelial Growth Factor by Hypoxia Is Modulated by a Phosphatidylinositol 3-Kinase/Akt Signaling Pathway in Ha-ras-Transformed Cells Through a Hypoxia Inducible Factor-1 Transcriptional Element
Authors: Mazure Nathalie M. , Chen Eunice Y. , Laderoute Keith R. , Giaccia Amato J.
Published: October 2019

Tumor angiogenesis, the development of new blood vessels, is a highly regulated process that is controlled genetically by alterations in oncogene and tumor suppressor gene expression and physiologically by the tumor microenvironment. Previous studies indicate that the angiogenic switch in Ras-transformed cells may be physiologically promoted by the tumor microenvironment through the induction of the angiogenic mitogen, vascular endothelial growth factor (VEGF). In this report, we show Ras-transformed cells do not use the downstream effectors c-Raf-1 or mitogen activated protein kinases (MAPK) in signaling VEGF induction by hypoxia as overexpression of kinase-defective alleles of these genes does not inhibit VEGF induction under low oxygen conditions. In contrast to the c-Raf-1/MAP kinase pathway, hypoxia increases phosphatidylinositol 3-kinase (PI 3-kinase) activity in a Ras-dependent manner, and inhibition of PI 3-kinase activity genetically and pharmacologically results in inhibition of VEGF induction. We propose that hypoxia modulates VEGF induction in Ras-transformed cells through the activation of a stress inducible PI 3-kinase/Akt pathway and the hypoxia inducible factor-1 (HIF-1) transcriptional response element.
Signal Transduction in Human Hematopoietic Cells by Vascular Endothelial Growth Factor Related Protein, a Novel Ligand for the FLT4 Receptor
Authors: Wang Jian-Feng , Ganju Ramesh K. , Liu Zhong-Ying , Avraham Hava , Avraham Shalom , Groopman Jerome E.
Published: October 2019

We have recently identified a novel ligand of the vascular endothelial growth factor (VEGF) family termed VEGF-related protein (VRP), which specifically binds to the FLT4 receptor. To characterize the signaling events after VRP engagement of its cognate receptor in hematopoietic cells, a population of human erythroleukemia (HEL) cells, termed HEL-JW, expressing high levels of FLT4 receptor was isolated. Stimulation of HEL-JW cells with VRP alone and in combination with the c-kit ligand/stem cell factor increased cell growth. VRP induced tyrosine phosphorylation of various proteins, including the FLT4 receptor. Further characterization of these tyrosine phosphorylated molecules revealed that Shc, Grb2, and SOS form a complex with the activated FLT4 receptor. HEL-JW cells also expressed RAFTK, a recently identified member of the focal adhesion kinase family. RAFTK was phosphorylated and activated upon VRP treatment, and there was an enhanced association of this kinase with the adaptor protein Grb2. Furthermore, the c-Jun NH2-terminal kinase (JNK), involved in growth activation and shown to mediate RAFTK signaling in other cell types, was activated by VRP stimulation. We also observed that VRP treatment of HEL-JW cells resulted in the phosphorylation of the cytoskeletal protein paxillin. This treatment resulted in an increased association of paxillin with RAFTK, which was mediated by the C-terminal region of RAFTK. These studies indicate that VRP stimulation induced the formation of a signaling complex at its activated receptor as well as activation of RAFTK. VRP-mediated activation of RAFTK may facilitate signal transduction to the cytoskeleton and downstream to the JNK pathway in FLT4-expressing blood cells.
Deficient Drug Transporter Function of Bone Marrow–Localized and Leukemic Plasma Cells in Multiple Myeloma
Authors: Pilarski Linda M. , Szczepek Agnieszka J. , Belch Andrew R.
Published: October 2019

Although chemotherapy effectively reduces the plasma cell burden in multiple myeloma (MM), the disease recurs. MM includes circulating and bone marrow (BM) localized components. A large majority of circulating CD11b+ MM B cells (81%) express an IgH VDJ rearrangement identical to that of autologous BM plasma cells. Unlike plasma cells, these monoclonal circulating B cells exhibit dye and drug transport activity before and throughout chemotherapy. Drug resistance was measured as the ability to export the fluorescent dye Rhodamine123 (Rh123) or the drug adriamycin, using flow cytometry. The role of P-glycoprotein 170 (P-gp), the multidrug transporter, was defined by cyclosporin A (CsA)-sensitive dye export. Only 8% to 11% of BM-localized plasma cells exported dye with the majority retaining dye, identified as bright staining. Circulating leukemic plasma cells were also unable to export dye and remained Rh123bright. However, 53% of circulating clonotypic MM B cells exhibited CsA-sensitive dye export. BM plasma cells taken before or after initiation of first line chemotherapy were equally unable to export dye. Thus in myeloma, differentiation to the plasma cell stage is accompanied by a loss of P-gp function, although P-gp phenotypic expression is retained. In contrast, for monoclonal gammopathy of undetermined significance (MGUS), 54% of BM-localized plasma cells exported dye, comparable to the 53% of circulating MGUS B cells that also exported dye, suggesting that the apparent defect in P-gp function is unique to myeloma plasma cells. Virtually all BM plasma cells in MM retained the drug adriamycin, consistent with their initial drug sensitivity in vivo, in contrast to circulating MM B cells, or to T cells in BM or blood. Thus, circulating B cells appear to be the predominant drug resistant component of the MM B-lineage hierarchy. This report suggests that successful therapeutic strategies will be those that target circulating B cells. Chemosensitization methods involving inhibition of P-gp are likely to improve depletion of these cells by compromising their ability to exclude drug. This work suggests that circulating clonotypic B cells should be monitored in clinical trials to confirm their depletion and the overall efficacy of novel treatment strategies.
Human Signaling Protein 14-3-3ζ Interacts With Platelet Glycoprotein Ib Subunits Ibα and Ibβ
Authors: Calverley David C. , Kavanagh Terrance J. , Roth Gerald J.
Published: October 2019

The initiation of primary hemostasis is mediated by interaction of the platelet glycoprotein Ib (GPIb) surface receptor and its arterial subendothelial von Willebrand factor (vWF) ligand. The intracellular signaling immediately following GPIb receptor occupancy connecting the adhesive event to platelet activation and aggregation has not been well characterized. The 14-3-3 proteins are a 27- to 30-kD ubiquitous protein family with diverse biologic roles, including functional modulation of several prominent signaling proteins. We used the yeast two-hybrid system and confocal microscopy to characterize the recently described interaction between GPIb and platelet 14-3-3ζ, and provide evidence for the potential signaling role of this protein. Two-hybrid interactions suggest that platelet 14-3-3ζ associates with the cytoplasmic domain of GPIb subunits Ibα and Ibβ in transformed yeast cells. The 14-3-3 interaction with GPIbβ may be partly mediated through the latter's phosphorylated serine 166 residue as its mutagenesis results in 20% to 40% reduced interaction. There was 51% to 59% reduced interaction between GPIb and three 14-3-3ζ deletion mutants compared with full-length 14-3-3ζ, suggesting that either theN-terminal dimerization or membrane-binding domains or more than one noncontiguous 14-3-3ζ element may be required for optimal GPIb interaction. Confocal studies of platelets and a megakaryocyte cell line provided additional evidence for interaction of 14-3-3ζ with GPIbα and GPIbβ. We also found that, similar to the signaling mediators phosphatidylinositol 3-kinase and Src, platelet cytoskeletal 14-3-3ζ content is increased following vWF and ristocetin stimulation. We suggest that platelet 14-3-3ζ interacts with GPIbα and Ibβ, that this interaction may be partly mediated through phosphoserine recognition, and that 14-3-3ζ cytoskeletal translocation may serve as a GPIb post–receptor occupancy signaling event.