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G Kvalheim

Publications and source records attributed to G Kvalheim.

68 records · Page 4Linked to original sources
PubMed1988

Epitopes of the Mycobacterium tuberculosis 65-kilodalton protein antigen as recognized by human T cells.

A synthetic peptide approach has been used to identify the epitopes recognized by clonal and polyclonal human T cells reactive to the recombinant mycobacterial 65-kDa protein Ag. Three of the four epitopes identified were recognized as cross-reactive between Mycobacterium tuberculosis and Mycobacterium leprae, although their amino acid sequence in two of three cases was not identical. The peptide (231-245) defining an epitope recognized as specific to the M. tuberculosis complex contains two substitutions compared with the homologous M. leprae region of which one or both are critical to T cell recognition. The reactive T cell clones showed helper/inducer phenotype (CD4+, CD8-), and secrete IL-2, granulocyte-macrophage-CSF, and IFN-gamma upon Ag stimulation. The same clones display cytotoxicity against macrophages pulsed with the relevant peptides or mycobacteria.

Amino Acid Sequence↗
PubMed1988

New indirect approach to the therapeutic use of immunotoxins.

To improve the applicability of immunotoxins (ITs), we have developed a new two-step indirect procedure. The target cells to be killed are first incubated with cell-specific mouse monoclonal antibodies (MAbs). After removal of excess unbound antibody, the cells are incubated in the presence of lactose with an indirect IT, a conjugate of whole abrin and sheep anti-mouse immunoglobulin (SAM) that binds only to cells having primary mouse MAbs on their surfaces. The SAM-abrin IT is affinity purified before use to remove molecules with exposed B-chain-binding sites; it was nontoxic in the absence of the specific mouse MAbs, demonstrating the specificity of the two-step method. We compared the indirect approach, using four different primary MAbs, with the conventional method, in which abrin is coupled directly to the mouse MAbs. In three human cell lines--the melanoma line FEMX, the Burkitt cell line Rael, and the leukemia cell line KM3--the cell kill, measured by a clonogenic assay, was consistently greater with the indirect than with the direct method. In the melanoma and Rael cells, the indirect method gave a higher cell kill than even native abrin. With a mixture of two different antibodies an additive effect was observed with the indirect but not with the direct method. The new approach greatly simplifies the therapeutic application in vitro of ITs, because it permits the use of different primary antibodies, singly or in mixtures, in conjunction with only one or a few general indirect ITs. In efforts to further improve the usefulness of the indirect method, other indirect ITs containing different toxin moieties are being examined. The possibility of employing the indirect principle in vivo is being explored.

Abrin↗
PubMed1988

Survival of breast cancer patients diagnosed during pregnancy or lactation.

This survival study includes 20 breast cancer patients diagnosed during pregnancy and 15 patients diagnosed during the lactation period. The survival rate of these patients is compared with that of ordinary breast cancer patients taking stage of the disease, age and calendar-year at diagnosis into account. The pregnancy group showed a significantly poorer prognosis compared with the control groups. Only 3 out of 20 survived more than 4 years. The tendency of a worse prognosis for the lactation group than for the control group was, however, not significant.

Adult↗
PubMed1988

Immunomagnetic removal of B-lymphoma cells from human bone marrow: a procedure for clinical use.

B-lymphoma cells were purged from human bone marrow by incubating the cell suspension with a cocktail of three different pan-B cell mouse IgG1 monoclonal antibodies, and then with immunobeads charged with sheep anti-mouse antibody, followed by magnetic separation. The primary antibodies used, HD37 (CD19), HD6 (CD22), and HH1 (CD37), bind to a very high percentage of the cells in non-Hodgkin's lymphomas of poor prognosis. The secondary antibody is directed against the Fc portion of the IgG antibodies. In model experiments Burkitt's lymphoma cells (Rael) were admixed to mononuclear bone marrow cells in the ratio 1/9. With a ratio of immunobeads/total antibody-binding B cells of 50/1 in a first treatment cycle and repeating the procedure with the same number of beads in a second cycle, a tumor cell depletion of more than 5 logs was achieved, as judged by a clonogenic assay. The concomitant reduction of CFU-GM and CFU-GEMM was about 20%. The purging procedure has been scaled up to clinical use. Equipment suitable for purging patients' marrow specimens, employing standard transfusion facilities, is described. With this equipment the efficacy of tumor cell removal was the same as in the model experiments, and the whole magnetic separation could be completed in 2 hours.

Animals↗
PubMed1987

Elimination of B-lymphoma cells from human bone marrow: model experiments using monodisperse magnetic particles coated with primary monoclonal antibodies.

Conditions for removing B-lymphoma cells from human bone marrow using "immunobeads" (IBs) were investigated. The IBs were prepared by coupling monoclonal antibodies directly to a new type of monodisperse magnetic polymer particles (M 450). Two monoclonal immunoglobulin M antibodies, AB-1 (CD 19), a B-cell-specific antibody, and AB-4, an HLA-DR-specific antibody, were used. The IBs were incubated with Rael Burkitt lymphoma cells admixed to fresh, mononuclear human bone marrow cells. After incubation for 30 min at 4 degrees C, the IBs were removed using cobalt samarium magnets. The number of remaining clonogenic tumor cells was assayed by the Courtenay and Mills soft agar procedure, and the clonogenic capacity of the bone marrow progenitor cells was measured by granulocyte-monocyte and granulocyte-erythroid-monocyte-megakaryocyte assays. With a ratio of tumor cells to normal bone marrow cells of 0.1 or 0.01 and a ratio of immunobeads to tumor cells in excess of 75, a tumor cell depletion of more than 3 logs was achieved with the AB-4 IBs and slightly less with the AB-1 beads. After two consecutive cycles of purification with the AB-4 beads, no colonies were found, corresponding to more than 6 logs of purification. In the case of the AB-1 beads, 4 to 5 logs of purification were achieved. The concomitant reduction in clonogenic bone marrow progenitor cells was only 30 to 40%. Flow cytometric studies showed that the tumor cell population contained appreciable proportions of cells binding only small amounts of the antibodies used. The results indicate that the IB procedure is highly efficient and capable of removing tumor cells expressing low levels of antigen. Compared to other purging methods in use the procedure described seems to offer several advantages with respect to efficacy, speed, and simplicity. By the use of a panel of suitable antibodies the new immunobead procedure may be potentially useful in autologous bone marrow transplantation of B-lymphomas and non-T-leukemias with poor prognosis.

Antibodies, Monoclonal↗
PubMed1987

Removal of cells from a malignant B-cell line from bone marrow with immunomagnetic beads and with complement and immunoglobulin switch variant mediated cytolysis.

In this report we describe the generation of complement (C') fixing IgG2b CD19 and CD22 monoclonal antibodies (mAbs) by the isolation of immunoglobulin (Ig) class switch variants using a simple and efficient method for the selection of spontaneously mutating hybridomas. The aim of this study was to compare the efficacy of C'-mediated cytolysis vs immunomagnetic (IB) depletion of tumor cells from mixtures of malignant B cells and normal bone marrow. In a clonogenic assay employing the B-cell lines Namalwa and OCI.LY1, we found that the use of immunomagnetic beads warranted a highly efficient tumor cell removal independent of the Ig isotype of the mAbs used. Elimination of up to 4 log was achieved using a cocktail consisting of CD19, CD20, CD22 and CD37 B-cell mAbs. A less efficient killing of 2 log was obtained by C' lysis using IgM mAbs, while only about 1 log tumor cell elimination was obtained using IgG2b mAbs. Immunomagnetic purging, besides being more effective than C'-mediated cytolysis, is easier to handle and more rapid.

Antibodies, Monoclonal↗
PubMed1987

Depletion of T lymphocytes from human bone marrow. Use of magnetic monosized polymer microspheres coated with T-lymphocyte-specific monoclonal antibodies.

A new technique for depletion of T cells from bone marrow is presented. Bone marrow cells (BMC) were rosetted with magnetic monosized polystyrene microspheres coated with monoclonal antibodies (MAbs) specific for T cell CD2 and CD3 antigens. Rosetted T cells were subsequently removed from non-T cells with the aid of a magnet. This immunomagnetic separation procedure was carried out in less than 40 min and reproducibly removed T cells, leaving a maximum of 0.025% sheep-red-blood-cell (SRBC) rosette-forming cells and less than 0.02% T cells as detected by a T cell limiting dilution assay. The efficacy of the depletion procedure was further shown by flow cytometry data, by effective removal of cells from a T cell line added to the BMC prior to immunomagnetic separation, and by abrogation of interleukin 2 (IL-2)-producing capacity in T-cell-depleted BMC (BMC-T). The T cell depletion procedure provided a 43-74% recovery of non-T cells present in the Isopaque-Ficoll-isolated bone marrow mononuclear cell fraction and did not disturb the growth potential of stem cells, as assayed by hematopoietic stem cell assays.

Antibodies, Monoclonal↗
PubMed1986

Mycobacterium bovis BCG-induced human T-cell clones from BCG-vaccinated healthy subjects: antigen specificity and lymphokine production.

A total of 121 human T-cell clones were raised from nine Mycobacterium bovis BCG-vaccinated healthy individuals. Three clones were autoreactive, 74 responded to BCG in the presence of antigen-presenting cells, and the others required in addition exogenous interleukin 2. Only one clone was CD8+ CD4-, and the rest were CD4+ CD8-. Testing with a panel of mycobacteria suggested that the clones were recognizing epitopes of varied specificity. Out of 44 clones tested, 15 were specific to BCG and Mycobacterium tuberculosis, 22 showed limited cross-reactivity, and 8 were broadly cross-reactive. None of the 22 BCG responder clones could differentiate between Danish, French, Prague, and Moreau strains of BCG. BCG and M. tuberculosis H37Rv also paralleled very closely; however, 6 of 18 BCG- and M. tuberculosis H37Rv-responding clones did not proliferate to Mycobacterium africanum. BCG- and M. tuberculosis H37Rv-specific as well as cross-reactive T-cell clones could be induced to produce interleukin 2, gamma interferon, and granulocyte-macrophage colony-stimulating factor activity.

BCG Vaccine↗
PubMed1985

Intra-arterial infusion of mitomycin C in treatment of breast cancer: occurrence of skin necrosis in irradiated patients.

Eight previously irradiated breast cancer patients with local recurrences were treated with intra-arterial infusions of 8 mg/m2 mitomycin C given at 3-week intervals. The mean time interval between radiotherapy and intra-arterial chemotherapy was 38 months (range 2-60). In five cases a temporary reduction in tumour size was observed. However, in 3 of the 8 patients severe local pain, starting immediately after the third course of treatment, was followed 4 weeks later by the development of deep necrotic ulcers of the chest wall. These cases are reported here and discussed in relation to the results of previous studies.

Adult↗
PubMed1985

Determination of warfarin in human plasma by high performance liquid chromatography and photodiode array detector.

A simple liquid chromatographic method for the determination of warfarin in human plasma is described. The method involves precipitation of plasma proteins with acetonitril. No further processing of samples is required. The supernatant was analyzed on a short (10 cm) 3-microns reversed-phase column eluted with 23% acetonitril in 100 mM ammonium formate, pH 3.5. The method takes advantage of the fact that warfarin dissolved in this mobile phase has a characteristic absorbance spectrum with distinct peaks at 271, 281, and 305 nm. Using an online photodiode array detector, the UV spectrum could be recorded during analysis without interrupting the flow of the mobile phase. This spectral information improves identification possibilities and evaluation of the purity of the chromatographic peaks. Warfarin was separated from other UV-absorbing compounds in plasma in less than 3 min, and there was no interference from numerous drugs given to patients. The standard curve (305 nm) was linear in the concentration range observed after oral intake of the single dose of warfarin and for a time corresponding to several half-lives. The detection limit of the method was about 0.1 microgram/ml when the absorbance was recorded at 305 nm. At this wavelength, the solvent front was small relative to that observed at lower wavelengths. The precision of the method, given as coefficient of variation, was 6.4%. The method was used for the determination of plasma half-lives of R- and S-warfarin in humans.

Chromatography, High Pressure Liquid↗
PubMed1984

Phase I study of the plant protein ricin.

A Phase I study was carried out with ricin, a plant toxin acting by inhibiting protein synthesis, on 54 cancer patients with advanced disease. Ricin was given as i.v. bolus injections every two weeks at dose levels ranging from 4.5 to 23 micrograms/sq m of estimated body surface area. Ricin was well tolerated at doses up to 18 to 20 micrograms/sq m. At these levels and at higher levels, flu-like symptoms with fatigue and muscular pain appeared and, in some patients, nausea and vomiting occurred also. No myelo-suppression was seen. Antibodies to ricin were detected in serum after two to three ricin injections. Ricin was eliminated from blood according to first order kinetics. At each dose level, the plasma concentrations, as well as the side effects, showed only minor differences between patients. The highest dose given, 23 micrograms/sq m, gave plasma concentrations twice those found previously to be therapeutically effective in tumor-bearing mice. Of 38 evaluable patients, one patient with lymphoma had a partial response. Stable disease was observed in four patients with renal cancers, in two with soft tissue sarcomas, and in one patient each with mesothelioma, thyroid, and rectal cancer. A dose of 23 micrograms/sq m is recommended for Phase II trials of ricin.

Abrin↗
PubMedDate not supplied

Development of a clinical grade procedure for generation of mRNA transfected dendritic cells from purified frozen CD34(+) blood progenitor cells.

Enriched CD34(+) peripheral blood progenitor cells (PBPC) are frequently used as stem cell support in cancer patients following high dose therapy. Since precursor dendritic cells (DCs) originate from haematopoietic progenitor cells, purified CD34(+) cells might also serve as starting cells for ex- vivo production of DC. In the present study we developed a clinical grade procedure for ex- vivo production of DC derived from enriched CD34(+) cells. Different concentrations of CD34(+) cells were grown in gas-permeable Teflon bags with different serum-free and serum-containing media supplemented with GM-CSF, IL-4, TNF-alpha, SCF, Flt-3L and INF-alpha. Serum-free CellGroSCGM medium for 7 days followed by CellGroDC medium in 7 days gave the same results as serum-containing medium. After incubation the cultured cells containing immature DCs were concentrated and transfected with tumour mRNA from human prostate cancer cell lines employing a highly efficient electroporation procedure. Thawed transfected DCs were able to elicit primary T-cell responses in vitro against antigens encoded by the prostate cancer mRNA as shown by ELISPOT assay using mock-transfected DCs as control. Our results show that frozen enriched CD34(+) cells can be an alternative and efficient source for production of DCs for therapeutic purpose.

Antigens, CD34↗
PubMedDate not supplied

Growth factor receptors in hematopoietic stem cells: EPH family expression in CD34+ and CD133+ cell populations from mobilized peripheral blood.

Cell-surface antigen expression of hematopoietic stem cells has a crucial role in characterizing cell subpopulation with distinct functional properties. The Eph receptors are the largest receptor tyrosine kinase family being involved in processes like vascular remodelling during development and physiological and pathological angiogenesis. Some Eph/Ephrin members are expressed in hematopoietic cells. The ability to isolate purified cell populations co-expressing CD34 and CD133 antigens as most commonly used markers for identification of hematopoietic progenitors has provided the opportunity to identify their surface-receptor profile. As positively expressed CD34 and CD133 cells take place not only in hematopoietic but also in endothelial differentiation, we aimed to define the Eph/Ephrin characteristic of these cells and relate these findings to new therapy strategies. Positive selections of CD34 and CD133 cells from PBPC in lymphoma patients were performed using magnetic beads and AutoMACS (Miltenyi Biotec) device. The purity of isolated cells was tested by flow cytometry. Immunocytochemistry was used to assess the Eph/Ephrin expression profile of positively selected samples. Our study revealed that all samples (10 from CD34+ and 8 from CD133+ cells) expressed one or more of Eph/Ephrin antigens in different proportions. All CD34+ cell samples, and 6 of 8 in the CD133+ cell fraction were strongly immunoreactive for EphA2. EphB2 was strongly expressed in all CD133+ cases, but 50% of the CD34 positive group lacked or weakly expressed this receptor. EphB4 was negative in 9 of 10 CD34+ cases and in all CD133+ cells. Thus, we have shown the surface marker profile of positively selected CD34 and CD133 cells in leukapheresis samples from lymphoma patients with regard to Eph/Ephrin receptors and discussed their biological clinical potential.

AC133 Antigen↗