Utilization of fish for constitutional and acquired chromosomal abnormalities for diagnostic and prognostic purposes

Abstract

Fluorescence in-situ hybridization (FISH) is a useful molecular cytogenetics technique for counting chromosomes and identifying specific chromosomal sequences of interest. FISH probe targets include centromeres, single loci, subtelomeres, and telomeres, using DNA or peptide nucleic acid (PNA) probes. FISH probes were used to determine chromosome number, copy loss or gain, and signal size in four studies involving acquired chromosomal changes in malignancy, mosaicism, and aneuploidy in pre-and postnatal constitutional abnormalities. In the first series of experiments, subtelomere probes for 5p and 5q were used to evaluate paraffin embedded patient samples for a 5q deletion and compare that to pathological and clinical characteristics of small cell lung carcinoma (SCLC). The correlation between del(5q) and spindle cell morphology was found to be significant (p<0.025), but of unknown relevance. In the second study, FISH was used in conjunction with microarray analysis to define the karyotype of a patient with a Pallister-Killian (PKS) phenotype, a tissue-limited mosaic condition. Her karyotype was determined to be 46,XX,dup(12)(p11.2p13.2),trp(12) (p13.2pter)[5]/46,XX[15], with the abnormal cell line remaining at 25% from 6 to 19 months of age. PNA telomere length studies were performed on this same patient to determine if there was a difference between the normal and abnormal cells lines, shorter telomeres explaining loss of abnormal cells, however, no difference was found. In the third series of studies, samples from pregnancy losses that failed to grow in culture were investigated using FISH. Two of the five culture failure samples identified a mosaic trisomy 9 female and a mosaic tetraploid female using FISH probes in interphase cells. In the fourth and final study, PNA FISH probes were used to assess the difference between telomere lengths in newborns with trisomy 21 and normal chromosomes. Telomere lengths in cells with trisomy 21 were significantly shorter than those with normal chromosomes in both metaphase (p<0.05) and interphase (p<0.01) cells. Cell senescence with shortened telomeres, would correlate with shorter lifespan of individuals with trisomy 21. Overall, FISH is an important tool that can enhance the diagnostic capabilities of conventional cytogenetics testing, particularly when used in conjunction with traditional karyotyping. FISH offers a more rapid alternative when turn around times are critical and allows for more specific resulting, particularly in cases where an abnormality of interest is unable to be visualized within a karyotype, specimens are nonliving, or the study of interphase cells is necessary.