Phenotypic correction of primary Fanconi anemia T cells with retroviral vectors as a diagnostic tool.
OBJECTIVE
The aim of this study was to develop a rapid laboratory procedure that is capable of subtyping Fanconi anemia (FA) complementation groups FA-A, FA-C, FA-G, and FA-nonACG patients from a small amount of peripheral blood.


METHODS
For this test, primary peripheral blood-derived FA T cells were transduced with oncoretroviral vectors that expressed FANCA, FANCC, or FANCG cDNA. We achieved a high efficiency of gene transfer into primary FA T cells by using the fibronectin fragment CH296 during transduction. Transduced cells were analyzed for correction of the characteristic DNA cross-linker hypersensitivity by cell survival or by metaphase analyses.


RESULTS
Retroviral vectors containing the cDNA for FA-A, FA-C, and FA-G, the most frequent complementation groups in North America, allowed rapid identification of the defective gene by complementation of primary T cells from 12 FA patients.


CONCLUSIONS
Phenotypic correction of FA T cells using retroviral vectors can be used successfully to determine the FA complementation group immediately after diagnosis of the disease.
Hanenberg
Helmut
H
Klinik für Pädiatrische Hämatologie und Onkologie, Zentrum für Kinderheilkunde, Heinrich-Heine-Universität, Düsseldorf, Germany. Helmut.Hanenberg@uni-duesseldorf.de
Batish
Sat Dev
SD
Pollok
Karen E
KE
Vieten
Lydia
L
Verlander
Peter C
PC
Leurs
Cordula
C
Cooper
Ryan J
RJ
Göttsche
Kerstin
K
Haneline
Laura
L
Clapp
D Wade
DW
Lobitz
Stephan
S
Williams
David A
DA
Auerbach
Arleen D
AD
D016475:3T3 Cells; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology; D002460:Cell Line; D002461:Cell Line, Transformed; D002470:Cell Survival; D004268:DNA-Binding Proteins; D005006:Ethnicity; D005199:Fanconi Anemia / Q000097:blood / Q000175:diagnosis / Q000235:genetics* / Q000276:immunology*; D052217:Fanconi Anemia Complementation Group A Protein; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D006367:HeLa Cells; D004854:Herpesvirus 4, Human; D006801:Humans; D016130:Immunophenotyping; D008677:Metaphase; D051379:Mice; D016685:Mitomycin / Q000494:pharmacology; D011506:Proteins / Q000235:genetics*; D011994:Recombinant Proteins / Q000378:metabolism; D012190:Retroviridae; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology* / Q000473:pathology; D014162:Transfection
D004268:DNA-Binding Proteins; C498471:FANCA protein, human; C120277:Fanca protein, mouse; D052217:Fanconi Anemia Complementation Group A Protein; D011506:Proteins; D011994:Recombinant Proteins; D016685:Mitomycin