A validated real-time quantitative PCR approach shows a correlation between tumor burden and successful ex vivo purging in follicular lymphoma patients.
OBJECTIVE
Purging procedures are increasingly used to provide stem cell collections devoid of contaminating tumor cells. In follicle center lymphoma (FCL), most approaches eradicate polymerase chain reaction (PCR);-detectable disease in only a fraction of harvests undergoing ex vivo manipulation. In this study we evaluated whether there is a relationship between tumor burden of stem cell harvests and successful clearance of PCR-detectable disease following ex vivo manipulation.


METHODS
To address this issue, we developed a real-time PCR approach for quantitative measurement of tumor contamination using the bcl-2 rearrangement. Real-time PCR was used to evaluate the relationship between tumor burden of stem-cell harvests and purging effectiveness in PCR(+) samples derived from 10 FCL patients. Ex vivo purging was performed using the MaxSep cell separator (Baxter Immunotherapy, Deerfield, IL, USA).


RESULTS
Our real-time PCR method proved effective, sensitive, accurate, and reproducible. Four collections were successfully cleared of minimal residual disease (MRD) whereas six remained PCR(+). Real-time PCR showed that the four collections successfully cleared of MRD had a prepurging tumor burden significantly lower than those remaining PCR(+) (p = 0.04).


CONCLUSIONS
This study provides the first evidence that evaluation of tumor burden in stem-cell harvests by real-time PCR can predict the effectiveness of therapeutic intervention in non-Hodgkin's lymphoma. Based on these findings, we foresee a more widespread use of this technique to evaluate the impact of different therapeutic approaches in FCL.
Ladetto
M
M
Divisione Universitaria di Ematologia, Azienda Ospedaliera S. Giovanni Battista, Torino, Italy. marco.ladetto@unito.it
Sametti
S
S
Donovan
J W
JW
Ferrero
D
D
Astolfi
M
M
Mitterer
M
M
Ricca
I
I
Drandi
D
D
Corradini
P
P
Coser
P
P
Pileri
A
A
Gribben
J G
JG
Tarella
C
C
D000293:Adolescent; D000328:Adult; D001483:Base Sequence; D001781:Blood Component Removal / Q000379:methods*; D015321:Gene Rearrangement; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D008224:Lymphoma, Follicular / Q000097:blood* / Q000628:therapy; D008875:Middle Aged; D008969:Molecular Sequence Data; D018365:Neoplasm, Residual / Q000097:blood; D016133:Polymerase Chain Reaction / Q000379:methods*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014178:Translocation, Genetic; D014182:Transplantation, Autologous
D019253:Proto-Oncogene Proteins c-bcl-2; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases