Integration of Pig-a, micronucleus, chromosome aberration, and Comet assay endpoints in a 28-day rodent toxicity study with 4-nitroquinoline-1-oxide.
As part of the Stage III Pig-a multilaboratory validation trial, we examined the induction of CD59-negative reticulocytes and total red blood cells (RET(CD59-) and RBC(CD59-) , respectively) in male Sprague Dawley(®) rats treated with 4-nitroquinoline-1-oxide (4NQO), for 28 consecutive days by oral gavage, at doses of 1.25, 2.50, 3.75, 5.00, and 7.50 mg kg(-1) day(-1) (the high dose group was sacrificed on Day 15 due to excessive morbidity/mortality). Animals also were evaluated for: micronucleated reticulocytes (mnRET) by flow cytometry; DNA damage in peripheral blood, liver, and stomach using the Comet assay; and chromosome aberrations (CAb) in peripheral blood lymphocytes (PBL). All endpoints were analyzed at two or more timepoints where possible. Mortality, body and organ weights, food consumption, and clinical pathology also were evaluated, and demonstrated that the maximum tolerated dose was achieved at 5.00 mg kg(-1) day(-1) . The largest increases observed for the genetic toxicology endpoints (fold-increase compared to control, where significant; all at 5.00 mg kg(-1) day(-1) on Day 29) were: RET(CD59-) (21X), RBC(CD59-) (9.0X), and mnRET (2.0X). In contrast, no significant increases were observed for the CAb or Comet response, in any tissue analyzed, at any timepoint. Because 4NQO is a well known mutagen, clastogen, and carcinogen, the lack of response for these latter endpoints was unexpected. These results emphasize the extreme care that must betaken in dose and endpoint selection when incorporating genotoxicity endpoints into routine toxicity studies as has been recommended or is under consideration by various regulatory and industrial bodies.
Covance Laboratories, Vienna, Virginia, USA. leon.stankowski@bioreliance.com
Leon F
LF
Stankowski
Daniel J
DJ
Roberts
Hepei
H
Chen
Timothy
T
Lawlor
Marie
M
McKeon
Hemalatha
H
Murli
Ajit
A
Thakur
Yong
Y
Xu
D015112:4-Nitroquinoline-1-oxide / Q000633:toxicity*; D000818:Animals; D001921:Brain / Q000187:drug effects / Q000648:ultrastructure; D018957:CD59 Antigens / Q000235:genetics; D002138:Calibration; D002869:Chromosome Aberrations / Q000139:chemically induced*; D020552:Comet Assay / Q000379:methods / Q000592:standards; D003627:Data Interpretation, Statistical; D004305:Dose-Response Relationship, Drug; D023381:Endpoint Determination; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007753:Laboratories / Q000592:standards; D008099:Liver / Q000187:drug effects / Q000648:ultrastructure; D008297:Male; D008565:Membrane Proteins / Q000235:genetics*; D015162:Micronucleus Tests / Q000379:methods / Q000592:standards; D009152:Mutagenicity Tests / Q000379:methods / Q000592:standards; D009153:Mutagens / Q000633:toxicity*; D009929:Organ Size / Q000187:drug effects; D009928:Organ Specificity; D051381:Rats; D017207:Rats, Sprague-Dawley; D012015:Reference Standards; D015203:Reproducibility of Results; D012156:Reticulocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D018570:Risk Assessment; D013270:Stomach / Q000187:drug effects / Q000648:ultrastructure; D013997:Time Factors
D018957:CD59 Antigens; D008565:Membrane Proteins; D009153:Mutagens; C080086:phosphatidylinositol glycan-class A protein; D015112:4-Nitroquinoline-1-oxide