Increased levels of multiple forms of dihydrofolate reductase in peripheral blood leucocytes of cancer patients receiving haematopoietic colony-stimulating factors: Interim analysis

M. Perwaiz Iqbal, Ikram A. Burney, Fakhra Sultana, Naseema Mehboobali, Tariq Siddiqui

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Abstract

The precise mechanism whereby granulocytes proliferate when haematopoietic colony stimulating factors (CSFs) are used in neutropenic cancer patients is poorly understood. The purpose of this study was to investigate whether these cytokines bring about leucocyte proliferation by increasing the levels of multiple forms of dihydrofolate reductase (DHFR). Blood samples were collected from 36 cancer patients (25 males and 11 females) with chemotherapy-induced neutropenia. One sample of blood from each patient was obtained before therapy either with CSF, such as granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF) or with placebo, and another one at the time of resolution of neutropenia. Peripheral blood leucocytes in these blood samples were counted, separated and lysed. From lysates, cytoplasmic samples were prepared and analyzed for active DHFR by a methotrexate-binding assay and for total immunoreactive DHFR by an enzyme linked immunosorbent assay. The increase in total leucocyte count (TLC) was most prominent (P < 0.005) in the CSF group and less so (P < 0.05) in the placebo group. The mean ± SD concentration values of active DHFR before and after stimulation with GM-CSF found were to be 0.34 ± 0.4 ng/mg protein and 0.99 ± 0.82 ng/mg protein, respectively, and in the group treated with G-CSF, 0.24 ± 0.32 ng/mg protein and 1.18 ± 2.4 ng/mg protein, respectively. This increase in active DHFR after stimulation with CSF was statistically significant (P < 0.05). Similarly concentration values of immunoreactive but nonfunctional form of DHFR (IRE) were 110 ± 97 ng/mg protein and 605 ± 475 ng/mg protein before and after stimulation with GM-CSF, and 115 ± 165 ng/mg protein and 1054 ± 1095 ng/mg protein before and after stimulation with G-CSF. This increase in concentration of IRE after stimulation with GM-CSF or G-CSF was statistically significant (P < 0.005). In the control group, there was an increase in the concentration of both active DHFR and IRE after treatment with placebo. However, this was not statistically significant. Resolution of neutropenia was quicker in the groups treated with CSF compared to the control group. Results of this study indicate that colony stimulating factors (G-CSF and GM-CSF) induce white cell proliferation by increasing the levels of multiple forms of DHFR.

Original languageEnglish
Pages (from-to)84-87
Number of pages4
JournalExperimental and Molecular Medicine
Volume32
Issue number2
Publication statusPublished - Jun 30 2000

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Colony-Stimulating Factors
Tetrahydrofolate Dehydrogenase
Factor analysis
Statistical Factor Analysis
Leukocytes
Blood
Granulocyte Colony-Stimulating Factor
Granulocyte-Macrophage Colony-Stimulating Factor
Neoplasms
Proteins
Neutropenia
Placebos
Assays
Immunosorbents
Control Groups
Chemotherapy
Cell proliferation
Leukocyte Count
Granulocytes
Methotrexate

Keywords

  • Colony stimulating factor
  • Dihydrofolate reductase
  • Granulocyte colony stimulating factor
  • Granulocyte-macrophage colony stimulating factor
  • Immunoreactive nonfunctional form
  • Neutropenia

ASJC Scopus subject areas

  • Biochemistry
  • Genetics

Cite this

Increased levels of multiple forms of dihydrofolate reductase in peripheral blood leucocytes of cancer patients receiving haematopoietic colony-stimulating factors : Interim analysis. / Iqbal, M. Perwaiz; Burney, Ikram A.; Sultana, Fakhra; Mehboobali, Naseema; Siddiqui, Tariq.

In: Experimental and Molecular Medicine, Vol. 32, No. 2, 30.06.2000, p. 84-87.

Research output: Contribution to journalArticle

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abstract = "The precise mechanism whereby granulocytes proliferate when haematopoietic colony stimulating factors (CSFs) are used in neutropenic cancer patients is poorly understood. The purpose of this study was to investigate whether these cytokines bring about leucocyte proliferation by increasing the levels of multiple forms of dihydrofolate reductase (DHFR). Blood samples were collected from 36 cancer patients (25 males and 11 females) with chemotherapy-induced neutropenia. One sample of blood from each patient was obtained before therapy either with CSF, such as granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF) or with placebo, and another one at the time of resolution of neutropenia. Peripheral blood leucocytes in these blood samples were counted, separated and lysed. From lysates, cytoplasmic samples were prepared and analyzed for active DHFR by a methotrexate-binding assay and for total immunoreactive DHFR by an enzyme linked immunosorbent assay. The increase in total leucocyte count (TLC) was most prominent (P < 0.005) in the CSF group and less so (P < 0.05) in the placebo group. The mean ± SD concentration values of active DHFR before and after stimulation with GM-CSF found were to be 0.34 ± 0.4 ng/mg protein and 0.99 ± 0.82 ng/mg protein, respectively, and in the group treated with G-CSF, 0.24 ± 0.32 ng/mg protein and 1.18 ± 2.4 ng/mg protein, respectively. This increase in active DHFR after stimulation with CSF was statistically significant (P < 0.05). Similarly concentration values of immunoreactive but nonfunctional form of DHFR (IRE) were 110 ± 97 ng/mg protein and 605 ± 475 ng/mg protein before and after stimulation with GM-CSF, and 115 ± 165 ng/mg protein and 1054 ± 1095 ng/mg protein before and after stimulation with G-CSF. This increase in concentration of IRE after stimulation with GM-CSF or G-CSF was statistically significant (P < 0.005). In the control group, there was an increase in the concentration of both active DHFR and IRE after treatment with placebo. However, this was not statistically significant. Resolution of neutropenia was quicker in the groups treated with CSF compared to the control group. Results of this study indicate that colony stimulating factors (G-CSF and GM-CSF) induce white cell proliferation by increasing the levels of multiple forms of DHFR.",
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AU - Mehboobali, Naseema

AU - Siddiqui, Tariq

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KW - Dihydrofolate reductase

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KW - Granulocyte-macrophage colony stimulating factor

KW - Immunoreactive nonfunctional form

KW - Neutropenia

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