Exp Mol Med.  2000 Jun;32(2):84-87.

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

• Iqbal MP
• Burney IA
• Sultana F
• Mehboobali N
• Siddiqui T

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 1,054 +/- 1,095 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.

Keyword

Dihydrofolate reductase; neutropenia; colony stimulating factor; granulocyte colony stimulating factor; granulocyte-macrophage colony stimulating factor; immunoreactive nonfunctional form

MeSH Terms

Adolescence
Adult
Cell Division/drug effects
Child
Female
Granulocyte Colony-Stimulating Factor/therapeutic use
Granulocyte Colony-Stimulating Factor/pharmacology*
Granulocyte Colony-Stimulating Factor/adverse effects
Granulocyte-Macrophage Colony-Stimulating Factor/therapeutic use
Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology*
Granulocyte-Macrophage Colony-Stimulating Factor/adverse effects
Human
Isoenzymes/metabolism
Isoenzymes/biosynthesis
Leukocyte Count
Leukocytes/pathology
Leukocytes/enzymology
Leukocytes/drug effects
Male
Middle Age
Neoplasms/enzymology
Neoplasms/drug therapy
Neoplasms/blood*
Neutropenia/metabolism*
Neutropenia/chemically induce
Neutropenia/blood
Tetrahydrofolate Dehydrogenase/metabolism*
Tetrahydrofolate Dehydrogenase/biosynthesis