SIMPLIFIED STRATEGIES FOR MINIMAL RESIDUAL DISEASE DETECTION IN B-CELL PRECURSOR ACUTE LYMPHOBLASTIC-LEUKEMIA

We have developed a simplified fluorescent runoff (FluRO) based IgH PC R strategy in order to facilitate follow-up of large numbers of B-cell precursor (BCP) acute lymphoblastic leukaemias (ALL) in a routine mol ecular diagnostic laboratory. DNA samples from 26 BCP-ALL and one B-ce ll line were amplified using IgH FR1 and FR2 consensus primers and ana lysed in parallel either by ethidium bromide non-denaturing PAGE or, a fter rendering the PCR products fluorescent with an internal JH consen sus primer, by high-resolution analysis on an automated fragment analy ser. The latter led to a minimum of one log increase in sensitivity of detection in 62% of alleles from 19 samples (16/28 in FR1: 11/15 in F R2) tested in parallel on log DNA dilutions, and to at least a 10(-2) level of sensitivity of detection in 15/19. The improved resolution al lowed an approximate 20% increase in the number of clonal alleles dete cted, and consequently doubled incidence of oligoclonality (6/26; 23%) . Using strategies, 6/17 (35%) of children analysed prospectively show ed residual IgH positivity in the post induction complete remission bo ne marrow sample. Both early deaths occurred within this subgroup of p atients and of the three of four surviving patients tested, two remain ed positive 2-3 months later, Although this simplified strategy Is, as expected, less sensitive than anti-V-D-J junction specific strategies , it enables detection of a category of 'slow-remitters' which map hav e prognostic significance at a stage where therapeutic decisions are t aken.