B-cell reconstitution after allogeneic SCT impairs minimal residual disease monitoring in children with ALL

Abstract

Minimal residual disease (MRD) detection using quantification of clone-specific Ig or TCR rearrangements before and after transplantation in children with high-risk ALL is an important predictor of outcome. The method and guidelines for its interpretation are very precise to avoid both false-negative and -positive results. In a group of 21 patients following transplantation, we observed detectable MRD positivities in Ig/TCR-based real-time quantitative PCR (RQ-PCR) leading to no further progression of the disease (11 of 100 (11%) total samples). We hypothesized that these positivities were mostly the result of nonspecific amplification despite the application of strict internationally agreed-upon measures. We applied two non-self-specific Ig heavy chain assays and received a similar number of positivities (20 and 15%). Nonspecific products amplified in these RQ-PCR systems differed from specific products in length and sequence. Statistical analysis proved that there was an excellent correlation of this phenomenon with B-cell regeneration in BM as measured by flow cytometry and Ig light chain-j excision circle quantification. We conclude that although Ig/TCR quantification is a reliable method for post transplant MRD detection, isolated positivities in Ig-based RQ-PCR systems at the time of intense B-cell regeneration must be viewed with caution to avoid the wrong indication of treatment.

Loss of B cells and their precursors is the most constant feature of GATA-2 deficiency in childhood myelodysplastic syndrome

“…22 The lowest KREC levels were observed in GATA-2-deficient patients in BM and PB (range BM 25-118; median 25; range PB 25-146, median 25) (Table 3 and Figures 1 and 2). Very low levels of KRECs, together with a decrease in B-…”

“…Thus, the final KREC (TREC) levels in unsorted populations serve as a surrogate marker of developing B (T) lymphocytes, irrespective of their proliferation history. 22 The detection limit was 25 copies/mg DNA. The controls for KREC and TREC analyses in PB consisted of 87 samples (median age 8 years; range 0-18 years) and 5 samples from BM (median age 10.1 years; range 5.2-14.2 years).…”

Improved flow cytometric detection of minimal residual disease in childhood acute lymphoblastic leukemia

“…Consequently, the RQ-PCR data are interpreted in such a way that false-negative MRD results are avoided, but this implies that low amplification levels that potentially do not reflect ALL cells but regenerating B-cells are considered as MRD positivity. 32,43,50 If alternative guidelines for data interpretation, used to prevent false-positive MRD results, were used, the concordance between FCM and PCR increased significantly (from 68 to 85%).…”

Standardized MRD quantification in European ALL trials: Proceedings of the Second International Symposium on MRD assessment in Kiel, Germany, 18–20 September 2008

“…Also, false positivity cannot be completely excluded, as massive regeneration of normal lymphoid progenitors might lead to (very) low levels of nonspecific amplification. 54,55 In addition, the method needs the initial characterization of the leukemic Ig/TCR rearrangements with a panel of different PCR and sequencing reactions and is restricted to experienced molecular hematology laboratories because of the complexity of the analyses (see Table 2). …”