Novel DNA sequence variants in the hHR21 DNA repair gene in radiosensitive cancer patients.
OBJECTIVE
Radiation therapy is an important treatment modality for oncology patients. DNA sequence variants have so far been identified in only a few genes in radiosensitive cancer patients. Patients known to be clinically radiosensitive were tested for mutation of a gene involved in DNA double-strand break repair and sister chromatid cohesion--hHR21.


METHODS
Clinically radiation-sensitive patients were accrued to the study after giving informed consent. Blood samples were obtained and lymphoblastoid cell lines established. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to amplify the hHR21 gene, and the DNA product was sequenced to identify any genetic abnormalities. Northern blot analysis, cell survival, and growth assays were performed on control cells and cells with hHR21 variants, and a restriction digest assay was developed to screen for carriers of a detected gene variant.


RESULTS
The DNA sequence of the hHR21 gene was determined in 19 radiation-sensitive cancer patients. In 6 of the 19 patients, a thymidine (T) to cytosine (C) transition was detected at position 1440 of the hHR21 open reading frame (T1440C). This variant did not alter the amino acid sequence and was likely to be a polymorphism. One patient with a particularly severe radiation reaction had a second sequence variant immediately adjacent to the first. This was a guanine (G) to adenine (A) transition (G1441A), resulting in a change of the amino acid sequence (glycine --> arginine) in a portion of the protein conserved in evolution. This suggests that this DNA alteration may be biologically significant. Restriction digest with the HpaII enzyme confirmed the presence of both sequence variants on the same allele.


CONCLUSIONS
We describe the first two DNA sequence variants ever found in the hHR21 gene, in patients with clinical radiation hypersensitivity. Although no direct evidence for the involvement of hHR21 alterations in the radiosensitivity of the cancer patients examined has been demonstrated, the possibility exists that homozygous mutations or other mutations of this gene could contribute to radiosensitivity. A simple test is described that could be applied to screening for these variants in relevant populations.
Severin
D M
DM
Peter MacCallum Cancer Institute, East Melbourne, Victoria, Australia.
Leong
T
T
Cassidy
B
B
Elsaleh
H
H
Peters
L
L
Venter
D
D
Southey
M
M
McKay
M
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D015152:Blotting, Northern; D018797:Cell Cycle Proteins; D002461:Cell Line, Transformed / Q000528:radiation effects; D004252:DNA Mutational Analysis; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins; D004307:Dose-Response Relationship, Radiation; D020022:Genetic Predisposition to Disease; D005820:Genetic Testing; D006801:Humans; D008214:Lymphocytes / Q000737:chemistry / Q000528:radiation effects; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D009363:Neoplasm Proteins / Q000235:genetics* / Q000502:physiology; D009369:Neoplasms / Q000532:radiotherapy; D009687:Nuclear Proteins / Q000235:genetics* / Q000502:physiology; D016366:Open Reading Frames; D010750:Phosphoproteins / Q000235:genetics* / Q000502:physiology; D012150:Polymorphism, Restriction Fragment Length; D011832:Radiation Injuries / Q000209:etiology / Q000235:genetics*; D011836:Radiation Tolerance / Q000235:genetics*; D011878:Radiotherapy / Q000009:adverse effects*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; C429022:RAD21 protein, human