Detection of Karyotype Evolution From Peripheral Blood by Sequential FISH Analyses of Circulating CD34+ Cells In MDS Patients: Results of the Ongoing German Multicenter Prospective Diagnostic Study

Chromosomal aberrations in myelodysplastic syndromes (MDS) play a major role in diagnostics, pathogenesis, prognosis, and, more recently, in treatment allocations. The acquisition of clonal abnormalities in pts with initially normal karyotype, the expansion of an aberrant cell clone with a given anomaly or the occurrence of new abnormalities are called karyotype evolution (KE). A model of stepwise cytogenetic changes is proposed, but only a few systematic studies had focused on this phenomenon as yet. Chromosomal anomalies can be detected by conventional chromosome banding analyses of bone marrow (bm) metaphases and most of them are provable by Fluorescence in Situ Hybridization (FISH) of circulating CD34+ cells from peripheral blood (pb).

To closely follow chromosomal aberrations from pb, to assess karyotype evolution and rare anomalies and to correlate the molecular-cytogenetic results with pb blood counts, bm morphology and treatment modalities. For this purpose, we initiated a German multicenter prospective, non-interventional diagnostic study in October 2008.

Sequential FISH analyses were performed on immunomagnetically enriched circulating CD34+ cells from pb as follows: a “super-panel” (D7/CEP7, EGR1, CEP8, CEP XY, D20, p53, IGH/BCL2, TEL/AML1, RB1, MLL, 1p36/1q25, CSF1R) was used for initial screening, every 12 months during follow-up and in every case of suspected progression. A “standard-panel” (EGR1, D7/CEP7, CEP8, p53, D20, CEP X/Y, TEL/AML1) was performed for analyses at short intervals every 2 months in the 1^st year and every 3 months during the 2^nd year. Results were also correlated with those of conventional banding and FISH analyses performed on bm samples, which typically were collected every 6 months.

As yet, 205 pts have been included in the study. Concerning median age, gender distribution, and MDS subtypes (according to WHO and IPSS) the study cohort was representative for the disease. Up to date, chromosomal aberrations were detectable in 126 pts. (62%) by FISH from pb. Most of them had 1 or 2 anomalies, but there were even pts with 4 or more aberrations. The most common aberrations- as known from other studies based on bm analyses- were del(5q) (64%), aberrations of chromosome 7 (del(7q)/-7) (25%), allelic loss of p53 (12%), +8 (11%), del(20q) (10%) and aberrations of chromosome 21 (del(21q)/-21/+21) (7%). Interestingly, a 12p-deletion was detectable as often as a trisomy 8 (11%), and in most cases as part of complex aberrations. A loss of p53 was significantly associated with aberrations of chromosome 7: 67% of pts with aberrations of chromosome 7 showed an allelic loss of p53 at the same time. In 4 cases, a coexistence of del(7q) and -7 in the same clone was observed.

Within a median observation time of 7 months (2-21), a KE was detectable from pb by FISH in 17 pts (13%) out of 135 with at least 2 sequential analyses. In most cases del(7q)/-7 developed as a new anomaly, and was followed in frequency by aberrations of chromosome 21.

FISH analyses of enriched circulating CD34+ cells are a feasible, less-invasive method to detect chromosomal aberrations in MDS pts and to follow the clone size from peripheral blood. By that method we are able to observe karyotype evolution as a step wise process in detail and to follow even rare abnormalities to learn more about their clinical and prognostic impact.

No relevant conflicts of interest to declare.