Chromosomal translocations t(4;14), t(11;14) and proliferation rate stratify patients with mature plasma cell myelomas into groups with different survival probabilities: a molecular epidemiologic study on tissue microarrays.

Plasma cell myelomas (PMs) exhibit clinical and molecular heterogeneity. To date, morphology and immunohistochemistry on bone marrow trephines are of limited value to stratify patients into different prognostic categories. However, some chromosomal translocations are of prognostic and/or of predictive importance in PMs. In this study, the prognostic significance of morphology, CyclinD1 expression, proliferation index (Mib1) and presence of the translocations FGFR3/IgH [t(4;14)] and CCND1/IgH [t(11;14)] are compared in 119 patients with PM. Immunohistochemistry and fluorescence in situ hybridization analysis were carried out on a tissue microarray containing bone marrow trephines. Hundred and one PMs showed a mature morphology whereas 10 were immature. All but one PM carrying a translocation showed a mature morphology. Patients with a t(4;14) (12%) had a statistically significant shorter 1-year survival (P=0.004), whereas those with a t(11;14) (21%) had a trend towards a better clinical outcome. CyclinD1 protein expression was not significantly associated with survival. Besides the t(4;14), an immature morphology (P<0.001) and a proliferation index (Mib1) of more than 10% (P=0.002) were associated with a significantly worse outcome. A high occurrence of strong CyclinD1 protein expression in the tumor cells was predictive of either a t(11;14) or of a low level amplification of the CCND1 gene, suggesting that different molecular mechanisms may have lead to an over-expression of the CyclinD1 protein in PMs. These findings demonstrate that a high proliferation rate and translocations involving the IgH locus can stratify mature PMs into groups with distinct survival probabilities.

Ultrasonic decalcification offers new perspectives for rapid FISH, DNA, and RT-PCR analysis in bone marrow trephines.

The requisite analyses on bone marrow biopsies are increasing: Molecular analyses such as fluorescence in situ hybridization (FISH), polymerase chain reaction (PCR), and reverse transcriptase (RT)-PCR are demanded in addition to morphology and immunohistochemistry to improve diagnostic accuracy. Moreover, analysis of certain molecular prognostic or predictive biomarkers is increasingly mandatory in the assessment of hematologic diseases. In some circumstances, only formalin fixed, bone-containing tissue is available for molecular analysis. Because various fixation and decalcification procedures can impair DNA and RNA quality, there is an urgent need for standardized decalcification protocols which allow FISH and PCR analysis. In this study we developed a routinely applicable decalcification protocol to optimize the molecular analysis method although preserving morphology and immunohistochemical results. Therefore, we compared 2 different approaches including ultrasonic decalcification versus nonultrasonic procedures and ethylenediaminetetraacetate-based reagents versus acid-based ones. In our hands, the combined use of ultrasound and ethylenediaminetetraacetate-based reagents permits successful interphase FISH, PCR, and RT-PCR analysis whereas concomitantly preserving morphology and antigeneicity.