A Comparative Analysis of Molecular Genetic and Conventional Cytogenetic Detection of Diagnostically Important Translocations in More than 400 Cases of Acute Leukemia, Highlighting the Frequency of False-negative Conventional Cytogenetics

Rebecca L. King, MD; Mojdeh Naghashpour, MD, PhD; Christopher D. Watt, MD, PhD; Jennifer J.D. Morrissette, PhD; Adam Bagg, MD


Am J Clin Pathol. 2011;135(6):921-928. 

In This Article


MRP was performed on 494 diagnostic patient specimens during the study. Of these, 52 were excluded for a diagnosis other than AL. Of the remaining 442 cases, acute myeloid leukemia (AML) was diagnosed in 375 (84.8%) and acute lymphoblastic leukemia (ALL) in 67 (15.2%). The HV-7 system was used in 107 specimens and the LTx assay in 335.

Of 442 specimens tested using the MRP assay, 67 (15.2%) were positive for 1 of the detectable translocations Table 2 and Figure 1. When results from the 2 assays were compared, 20 (18.7%) of 107 specimens were positive by the HV-7 and 47 (14.0%) of 335 were positive by the LTx (P = .27). No specimens tested were positive for more than 1 translocation by the molecular assay. Of the 67 MRP+ cases, 19 had additional cytogenetic abnormalities.

Figure 1.

Flow chart highlighting cases detected by MRP but missed by CC (7/57 [12%] false-negative by CC). CC, conventional cytogenetics; MRP, multiplex reverse transcriptase–polymerase chain reaction.

Of the 442 specimens, 20 (4.5%) had inconclusive results from the MRP assay owing to suboptimal RNA quality Figure 2. Six of these inconclusive cases were studied with the HV-7 system and 14 with the LTx. Of the 20 cases with an inconclusive MRP result, CC studies were available for 14 (4 CC not ordered; 2 CC failed). In 5 cases, there was a normal karyotype, and 9 demonstrated a clonal cytogenetic abnormality. Of these 9 cases, 1 revealed an inv(16) by CC, which is detectable by our MRP assay. The remaining 8 cases had abnormalities that are not targeted by our MRP assay. Thus, at least 1 inconclusive MRP case harbored a potentially detectable translocation that was evident by CC (Figure 2); however, this cannot be considered a true false-negative because the RNA quality was suboptimal.

Figure 2.

Flow chart highlighting results of all cases evaluated in this study. Inconclusive MRP are cases in which there was suboptimal RNA quality that resulted in poor amplification of control targets in the MRP assay. CC, conventional cytogenetics; MRP, multiplex reverse transcriptase–polymerase chain reaction.

CC was successfully performed on 330 (78.2%) of 422 cases with valid MRP results available (Figure 2). In the remaining 92 cases (21.8%), CC failed (n = 32) or was not ordered (n = 60).

Of the 330 cases with evaluable CC, 140 (42.4%) had normal cytogenetics and 190 (57.6%) had clonal cytogenetic abnormalities (Figure 2). MRP was negative in 135 (96.4%) of the 140 cases with normal cytogenetics. However, the remaining 5 cases each had a translocation detected by MRP that was missed by CC (Figure 2) Table 3.

Among the 190 cases with cytogenetic abnormalities, there were 50 with translocations identified that were potentially detectable by MRP. Of these, all 50 were also positive by MRP. The remaining 140 cases with clonal cytogenetic abnormalities had genetic aberrations identified that were not targeted by the MRP assay. However, of these, 2 were positive by MRP, again revealing translocations missed by CC (Figure 2) Table 4. Therefore, in total, there were 57 translocations identified that could have been detected by the MRP and CC assays. Of these, 50 were indeed detected by both assays, whereas 7 were missed by CC, yielding a false-negative CC rate of 12% (7/57) (Figure 1).

From this group of 190 cases with abnormal CC, we found 6 AML specimens that had the inv(3)(q21;q26.2) or t(3;3)(q21;q26.2)/RPN1-EVI1 translocation, 1 with a t(6;9) (p23;q34.3)/DEK-NUP214, and 1 with a t(1;22)(p13;q13)/RBM15-MKL1 translocation. Although 9 patients with AML had a translocation involving the 11q23 locus detected by CC, none had a t(9;11)(p22;q23). These 9 cases comprised 6 with t(11;19)(q23;p13.1) and 1 each with t(6;11)(q27;q23), t(11;22)(q23;q13), and t(1;11)(p32;q23).

MRP but not CC was done in 92 cases, and of this group, 10 had a translocation detected by the MRP assay. In 4 of the cases, there was a t(15;17), and the patients were diagnosed with acute promyelocytic leukemia (APL), 1 was a case of AML with t(8;21), and 1 was AML with inv(16). The other 4 were cases of B-cell ALL, 3 with a t(9;22) and 1 with a t(1;19).


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