Non-radiolabelled PCR consensus primers and automatic sequencing enable rapid identification of tumor-specific V(H) CDR3 in aggressive B-cell malignancies.
Even though the results of current therapy are improved for B-cell acute lymphoblastic leukemia (B-ALL) and Burkitt's lymphoma (BL), prognosis of relapsed mature B-ALL and BL still remain extremely poor. In this study, we investigated the possibility of applying the use of non-radiolabelled PCR consensus primers and automatic sequencing for the rapid identification of the tumor-specific V(H) CDR3 nucleotide sequence, in mature B-ALL and BL. RNA was extracted from four consecutive, unselected samples from BL cases and three consecutive, unselected samples from mature B-ALL cases. The feasibility of the identification of the tumor-specific V(H) CDR3 nucleotide sequence was then assessed by using non-radiolabelled PCR consensus primers with automatic sequencing. The tumor-specific V(H) CDR3 nucleotide sequence was successfully identified for all seven patients (3 mature B-ALL and BL). The time required was substantially lower than that of the other methods previously published, despite the poor quality of some of the samples. The procedure showed rapidity, reliability and reproducibility. The characteristics of the methodology applied widen the possibility of developing anti-idiotypic therapeutic strategies, even in these B-cell malignancies.
Pession
Andrea
A
Department of Pediatrics, University of Bologna, Ospedale S. Orsola, Via Massarenti 11, 40100 Bologna, Italy. pession@med.unibo.it
Tonelli
Roberto
R
Canderan
Glenda
G
Bendandi
Maurizio
M
Rosolen
Angelo
A
Basso
Katia
K
Basso
Giuseppe
G
Locatelli
Franco
F
Zabalegui
Natalia
N
Paolucci
Guido
G
D001331:Automation; D001483:Base Sequence; D002051:Burkitt Lymphoma / Q000235:genetics* / Q000473:pathology; D002648:Child; D002675:Child, Preschool; D022801:Complementarity Determining Regions / Q000032:analysis / Q000235:genetics*; D016384:Consensus Sequence; D017931:DNA Primers / Q000235:genetics*; D005803:Genes, Immunoglobulin; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000032:analysis / Q000235:genetics*; D007223:Infant; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D022801:Complementarity Determining Regions; D017931:DNA Primers; D007143:Immunoglobulin Heavy Chains