Evaluation of hypoxic cell radio-sensitizers in terms of radio-sensitizing and repair-inhibiting potential. Dependency on p53 status of tumor cells and the effects on intratumor quiescent cells.
BACKGROUND
Intratumor quiescent (Q) cells and p53-mutated tumor cells are more difficult to control than intratumor proliferating (P) cells and p53 wild-type tumor cells, respectively. The usefulness of 3 hypoxic cell radio-sensitizers was compared in terms of a radio-sensitizing effect under aerobic and hypoxic conditions and a repair-inhibiting effect following irradiation on both Q and total (P + Q) cell populations in solid tumors. The dependency of these effects on the p53 status of tumor cells was also examined using tumor cell lines with identical genetic backgrounds except for their p53 status.


METHODS
Human head and neck squamous cell carcinoma cells transfected with mutant TP53 (SAS/mp53) or with neo vector as a control (SAS/neo) were inoculated subcutaneously into both the hind legs of Balb/cA nude mice. The nude mice bearing the tumors and C3H/He mice bearing SCC VII tumors received 5-bromo-2'-deoxyuridine (BrdU) continuously to label all the P cells in the tumors. Tumor-bearing mice received gamma-ray irradiation while alive or following tumor clamping after being administered no drug, nimorazole, SR-2514 or misonidazole, or received no drug, nimorazole, SR-2514 or misonidazole straight after gamma-ray irradiation. For the group irradiated after receiving the drug, the tumors were excised immediately following irradiation, while for the group irradiated before receiving the drug, the tumors were excised 24 h after irradiation. The excised tumors were minced and trypsinized. The tumor cell suspensions thus obtained were incubated with cytochalasin-B (a cytokinesis blocker), and the micronucleus (MN) frequency in the cells without BrdU labelling (= quiescent (Q) cells) was determined using immunofluorescence staining for BrdU. Meanwhile, the MN frequency in the total tumor cell population was determined from the tumors that had not been pretreated with BrdU. The clonogenic cell survival was also determined in the mice given no BrdU.


RESULTS
Both the radio-sensitizing effects under aerobic and hypoxic conditions and the repair-inhibiting effects following gamma-ray irradiation increased in the following order: nimorazole < SR-2514 < misonidazole in both total and Q cells in these 3 tumors. Both effects were more marked in the Q cells and p53-mutated tumors than in the total cells and p53-wild tumors, respectively.


CONCLUSIONS
In terms of controlling radio-resistant Q tumor cells and p53-mutated tumor cells, the combination of radio-sensitizers and conventional radiotherapy is promising both for radio-sensitization and for repair-inhibition, but further study of the toxicity to normal tissues is needed.
Masunaga
Shin-ichiro
S
Radiation Oncology Research Laboratory, Research Reactor Institute, Kyoto University, 2-1010, Asashiro-nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494, Japan. smasuna@rri.kyoto-u.ac.jp
Uto
Yoshihiro
Y
Nagasawa
Hideko
H
Hori
Hitoshi
H
Nagata
Kenji
K
Suzuki
Minoru
M
Kinashi
Yuko
Y
Ono
Koji
K
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000235:genetics / Q000532:radiotherapy*; D015687:Cell Hypoxia / Q000502:physiology; D003131:Combined Modality Therapy; D004260:DNA Repair / Q000235:genetics; D004307:Dose-Response Relationship, Radiation; D005260:Female; D005720:Gamma Rays; D016158:Genes, p53 / Q000502:physiology*; D006258:Head and Neck Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000532:radiotherapy*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008920:Misonidazole / Q000494:pharmacology; D009554:Nimorazole / Q000494:pharmacology; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D023041:Xenograft Model Antitumor Assays
D000970:Antineoplastic Agents; D011838:Radiation-Sensitizing Agents; D009554:Nimorazole; D008920:Misonidazole