Patterns of BCR/ABL gene rearrangements by interphase fluorescence in situ hybridization (FISH) in BCR/ABL+ leukemias: incidence and underlying genetic abnormalities

Abstract

Interphase fluorescence in situ hybridization (iFISH) is increasingly used for the identification of BCR/ABL gene rearrangements in chronic myeloid leukemia (CML) and acute lymphoblastic leukemia (ALL). In the present study, we have explored the incidence of both typical and atypical iFISH patterns of BCR/ABL gene rearrangements in a series of 168 consecutive BCR/ABL + patients -135 CML, 31 precursor B-ALL and two acute myeloblastic leukemia (AML) cases -and established their underlying genetic alterations through further molecular and chromosome analyses. Two different FISH probes (Vysis Inc., Downers Grove, IL, USA) were used: the LSI BCR/ABL dual color extra signal (ES) and the dual color dual fusion BCR/ABL probe (D-FISH). Our results show that most BCR/ABL + patients (83%, including 88% of all CML, 61% of ALL and one of two AML) displayed typical iFISH patterns of either Major (M) BCR/ABL (87% of CML, 13% of ALL and one of the two AML) or minor (m) BCR/ABL gene rearrangements (1% of all CML and 48% of ALL cases) with the two probes. Further molecular and cytogenetic studies confirmed the presence of such typical rearrangements in all except one of these ALL cases who had coexistence of an MBCR/ABL and an mBCR/ ABL gene rearrangement together with monosomy 9. In the remaining 29 cases (17%), up to five different atypical iFISH patterns were detected with the ES probe. Atypical iFISH patterns were most frequently due to additional numerical changes -most often supernumerary Philadelphia (Ph) chromosome (7%) but also gain or loss of chromosome 9 (1%) or 22 (1%). Deletion of 9q sequences proximal to the breakpoint were also frequently observed with the ES probe (8%). Application of the D-FISH probe showed that in most of these latter cases (5%) deletion of 22q sequences distal to the breakpoint also occurred. The remaining cases with atypical iFISH had cryptic insertion of BCR in 9q34 (1%). Exact interpretation of each iFISH pattern was supported by FISH on metaphases and molecular determination of the BCR breakpoint. In summary, our results indicate that despite the high incidence of typical iFISH patterns of BCR/ABL gene rearrangements, atypical patterns are also found in BCR/ABL + acute leukemias; the precise definition of the alteration present in individual cases is dependent on metaphase studies and molecular definition of the breakpoint.

Atypical fluorescence in situ hybridisation pattern in chronic myeloid leukaemia due to cryptic insertion of BCR at 9q34

“…In contrast, two patients (1%) had a normal karyotype with no Ph chromosome being detected; the BCR/ABL gene rearrangements observed in both cases were due to a cryptic insertion of BCR in 9q34, as confirmed by metaphase FISH. These findings, together with those of Wan et al, 1 would support the notion that, in patients in whom a CML is suspected to show an iFISH pattern consistent with an apparent 9q À and a Ph-negative karyotype, interphase and metaphase FISH investigation of atypical BCR/ABL gene rearrangements should be searched to confirm the presence of cryptic BCR insertion in 9q34. As already mentioned in our previous work, 2 in none of our two cases, neither in the one reported by Wan et al, 1 nuclei carrying a typical MBCR/ABL gene translocation were simultaneously found.…”

“…2 Despite the observation of a normal karyotype, the presence of the BCR/ABL gene fusion on chromosome 9 was confirmed by FISH on metaphase chromosomes, using the two mentioned probes plus a third BCR/ABL dual-color signal fusion (S-FISH) probe (Vysis). 1 In an updated series of 182 CML patients, we have found 15 cases (8%) showing similar patterns with the ES probe: one red, one green and one fused red/green signal (1R1G1F). This is in line with the previously published incidence of this pattern.…”

Derivative chromosome 9 deletions are a significant feature of childhood Philadelphia chromosome positive acute lymphoblastic leukaemia

“…A number of cryptic insertions of either BCR into ABL at 9q34, as reviewed by Primo et al, 24 or rarely ABL into BCR at 22q11, 25 have been reported in CML, with the insertions confirmed by dual fusion FISH. 7,18,[25][26][27][28] An insertion is sometimes seen as the end point of a series of events. Thus, in our case, either the der(9) chromosome which provided ABL sequences to chromosome 22 could have been lost, with subsequent duplication of the normal copy of chromosome 9, or else there could have been a partial duplication of ABL sequences on the der(9), which preceded their insertion into chromosome 22.…”

“…The designs of the SF and the ES probes allow detection of breakpoints involving the m-bcr of the BCR gene as well as the BCR/ABL fusion. 17,18 The interphase FISH patterns expected with each probe are shown in Table 1. For those cases with standard and nonstandard signal patterns observed in interphase, the number and relative size of the signals were confirmed, where possible, on representative metaphase cells from the same samples.…”

Reply to Wan et al

“…This is in line with the previously published incidence of this pattern. 2,3 In most of these 15 cases (n ¼ 13; 7%), the presence of an extensive del(9q) was confirmed by metaphase analyses with both the ES and the D-FISH probes; all of them showed a typical Ph chromosome on conventional cytogenetics. In contrast, two patients (1%) had a normal karyotype with no Ph chromosome being detected; the BCR/ABL gene rearrangements observed in both cases were due to a cryptic insertion of BCR in 9q34, as confirmed by metaphase FISH.…”