Interphase Chromosome Profiling (ICP) Assay in Peripheral Blood As a Viable Alternative to Bone Marrow Aspirate in the Initial Genetic Screening of Myelodysplastic Syndrome

Myelodysplastic syndrome (MDS) is a common adult malignancy and cytogenetics plays an important role in the diagnosis and classification for management of the disease. Both peripheral blood (PB) and bone marrow (BM) samples are routinely used in the investigation of MDS cases. However, the failure rate for peripheral blood for classical chromosome studies is very high and a common practice in those scenarios is to reflex to FISH testing using a set of probes designed to detect the most common abnormalities involving chromosomes 5, 7, 8 and 20 (MDS-FISH). While FISH is very valuable when less than 20 or no metaphase cells are available for analysis, several studies in the literature clearly indicate that in MDS with normal karyotype results, adding MDS-FISH has no additional clinical value and does not detect any additional cytogenetic abnormalities. Therefore, classical cytogenetics remains the primary testing for MDS. While bone marrow samples have a higher success rate in producing informative karyotype results, one large prospective study suggests that use of PB samples in the initial screening of patients with various anemias and suspected MDS, utilizing FISH probes if necessary, when the karyotype studies are unsuccessful, is valid and justified (Am J Clin Pathol 2011;135:915-920). According to this study, such noninvasive approaches will be very useful in the initial screening and would avoid or at least postpone the bone marrow aspirate in a significant number of patients resulting in significant cost savings.

Interphase chromosome Profiling (ICP) is a new novel molecular cytogenetic technology which is capable of producing a complete molecular karyotype from interphase nuclei of any tissue without the need for tissue culture. ICP has been demonstrated to be a failure proof technology with greater sensitivity to detection of large and/or cryptic cytogenetic aberrations than standard cytogenetics and FISH, and can also characterize marker chromosomes and chromosomal material of unknown origin in unbalanced rearrangements. (Cytogenet Genome Res 2014;142:226, Abstract #22). Besides the common deletions and monosomies involving the chromosomes 5, 7 and 20; and trisomy 8, several balanced translocations have been identified in MDS which cannot be detected by the current MDS-FISH panel. Additionally, other abnormalities present in both simple and complex karyotypes will be missed by MDS-FISH. ICP has been developed to detect most of these changes in both BM and PB samples.

We have tested the utility of ICP on eight peripheral blood and two bone marrow samples from 10 MDS cases. Two of these had no karyotype results. Four had a normal karyotype and the other four had a normal MDS-FISH. Both BM samples had a normal karyotype. One had a history of AML with trisomy 8, but was not detected in the current relapsed sample. Two cases had additional clinical suspicion of CLL or Multiple Myeloma in addition to MDS.

ICP was successful in all ten samples and the normal molecular karyotype is concordant with either the cytogenetics or FISH result. In the two failed cytogenetic cases and no FISH study, ICP showed a normal result. The guidelines for establishing the clonality when utilizing the ICP technology is in progress (manuscript in preparation). In the ICP validation studies referenced above, any abnormality present in four or more cells in a 20 cell analysis is considered positive and was always corroborated by either cytogenetics or FISH studies. However, in couple of patients, the clonal abnormality identified by FISH studies, was present in only two to three cells in the ICP study. At present, the clinical significance of these "minor" clones identified by ICP analyses is not clear. Large prospective studies are needed to answer this question. We have identified such minor clonal abnormalities in two of the ten cases in three to four cells. These abnormalities include marker chromosomes and potential balanced translocations involving chromosomes 1, 2, 12, and 14. As mentioned above, current MDS-FISH will not detect these minor clones but identified by the ICP technology and may be clinically significant.

Our results clearly demonstrate that ICP is a very sensitive technology and may be considered and adopted as the preferred method of choice in the initial screening of MDS patients especially on peripheral blood samples.