THE MUTATION FREQUENCIES IN HPRT LOCUS AS A GOLD
BIOMARKER TO EVALUATE THE SECONDARY NEOPLASMS
Pei
XH, Gu LJ, Zhao JC, Shen L
Shanghai
Institute for Pediatric Research, Shanghai, China
Objective: To investigate the risk
of secondary neoplasms by chemotherapy, we evaluated the somatic cell
mutation frequencies (Mfs) at hprt gene locus.
Methods: The Mfs were measured with
the T-lymphocytes cloning assay in age-match 30 healthy children and 20
treated patients by chemotherapy. Either fresh or carefully thrawed
cryopreserved mononuclicus cells (MNs) were counted and primed by seeding
106 cells/ml in culture medium containing PHA and incubated at
37°
C for 24-40 h. After priming the cells were counted and plated into 96-well
microtitre plates at the cell number 1,2,4,8 cells/well in non-6TG culture
medium (non selection) for determining the none cloning efficiency (CE);
and at 2×104 cell/well
in 10-5 6-TG (selection) for determining the selection CE. There
was 200ul growth culture medium containing IL-2 as T-cell growth factor,
PHA and 1×104 irradiated
accessory cells in every well. The microtiter dishes were incubated for
10-14 days in incubtator. Wells were scored for colony growth by use of an
inverted phase contrast microscope.
Results: The geometric mean Mfs for
patients was 7.5×10-6
and 1.3×10-6 for
age-match controls.
Conclusion:
In our investigation, the hprt mutation frequencies were measured as a
“gold standard”. The result shows that Mfs for patients were significantly
higher than controls. It suggested that the hprt locus be used as
biomarkers for evaluating the secondary neoplasms risk of chemotherapy
patients.