Phenomenon of Oligoclonality in Adult Patients with Acute Lymphoblastic Leukemia

Minimal residual disease (MRD) in adult patients with acute lymphoblastic leukemia (ALL) can be analyzed by flow cytometry, fluorescent in situ hybridization (FISH) if there are chromosomal abnormalities, polymerase chain reaction (PCR) for chimerical transcripts and immunoglobulin heavy chain (IgH) and T-cell receptor gamma (TCRG) genes rearrangements. IgH and TCRG rearrangements are the for monitoring of MRD, because it is possible to find specific clonal tumor marker in 80–90% patients. According to the data of different research groups phenomenon of oligoclonality (more than one clonal rearrangement) is identified in 16–80% cases depending on how much molecular markers are used (Scrideli CA, 2001; Gameiro P, 2002; Thorsten Raff, 2006; Csinady E, 2009; Katsibardi K, 2011).

We studied bone marrow samples from 59 patients with new diagnosed B-lineage ALL (43 patients), T-lineage ALL (15 patients) and one patient with bilinear B-T-ALL. Ph-positive ALL was diagnosed in 6 patients, t(4;11) – in one patient. Patients were treated by standard ALL-protocols based on 8 weeks induction remission, 3–5 consolidation courses and maintenance treatment during 2 years. Patients with Ph-positive B-ALL were treated by the same chemotherapy with imanitinib and one patient with mature B-ALL was treated by NHL-BFM-90 chemotherapy. Detection of IgH and TCRG genes rearrangements were carried out by polymerase chain reaction using standard family-specific primers. Synthesis of patient specific primers was based on defined nucleotide sequence of clonal PCR-product. Monitoring MRD was realized by nested PCR using patient specific primers.

IgH and/or TCRG genes rearrangements were detected in 50 of 59 patients (84,7%). IgH complete rearrangements were found in 31of 43 patients (72,1%) with B-lineage ALL, IgH incomplete rearrangements were detected in 12 of 43 patient (27,9%) with B-lineage ALL and in one of 15 patient with T-lineage ALL. TCRG rearrangements were found in 13 of 15 patients (86,7%) with T-lineage ALL and in 24 of 43 patients (55,8%) with B-lineage ALL. Seventeen (34%) of 50 patients had one clonal marker. So, phenomenon of oligoclonality was revealed in 33 patients (66%). Two clonal rearrangements were detected in 23 cases, 3 rearrangements were detected in 7 cases and 3 patients had 4 rearrangements. In 3 of 27 patients, in whom all markers were followed, monitoring of MRD showed different pattern of tumor subclones elimination. Monitoring of MRD in patients who had also chromosomal abnormalities such as t(9;22), t(4;11) was carried out by FISH and PCR in Real-time. In one case the patient had 3 IgH rearrangements and chromosomal abnormality t(4;11) at the diagnosis. Later at various stages of chemotherapy we observed a different clearance of IgH rearrangements, however, when cytogenetic remission was achieved and MLL-AF4 expression was absent there were two of three IgH markers. This fact confirms the presence of several tumor subclones, their different clearance during the treatment and suggests persistence of tumor in this patient.

In ALL patients with several subclones MRD monitoring should carried out with all markers, as we proved different subclone clearance suggesting different cells sensitivity to cytostatic drugs.

No relevant conflicts of interest to declare.