Cytogenetic Analysis of Multiple Myeloma Identifies Cytogenetic Alterations Implicated in Disease Complexity and Progression

Background: Multiple myeloma is the second most common blood borne neoplasia, accounting for nearly 10% of all diagnosed hematologic malignancies and has a disproportionately high incidence in elderly populations. Here we explored copy number variations using the high fidelity CytoScan HD arrays to develop a detailed map of copy number variations and identify novel mediators of disease progression. The results from CytoScan HD microarrays provide a detailed view of the entire genome with a resolution up to 25kb. Furthermore, 750,000 single-nucleotide polymorphisms are included and the array provides information about loss of heterozygosity and uniparental disomy.

Materials and methods: CytoScan HD arrays were performed on 97 myeloma patient samples to identify cytogenetic regions important to the development and progression of the disease. Gene expression profiles from 351 patients were analyzed to identify genes with a change in gene expression of 1.5 fold or more. Data from CytoScan and gene expression arrays was combined to perform chromosomal positional enrichment analysis to identify cytogenetic driver lesions, or lesions that provide a small, but significant growth and survival advantage to the cell. Furthermore, Kaplan-Meier, log-rank test and Hazard ratio analyses were performed to identify gene within the driver lesions that have a significant impact on survival when dysregulated.

Results: The results from the CytoScan HD analysis closely mirrored what has been shown by FISH and SNP arrays, with gains to the odd numbered chromosomes, specifically 3, 5, 7, 9, 11, 15 and 17 as well as losses to chromosomes 1p and 13. Interestingly, we identified gains to a small region within chromosome 8p, contrary to published reports demonstrating a large scale loss of this region. We identified numerous genes within this region that are important for survival and their overexpression resulted in a decreased progression free survival. For example, Cathepsin B (CTSB) is encoded for in chromosome 8p22-p21 with an increased gene expression of at least 1.5 fold over normal controls, among others. Furthermore, Cathepsin B, a cysteine protease, has been linked to cancer of the ileum, suggesting that a similar role may be present within myeloma.

We then integrated the 97 copy number profiles results with 351 myeloma gene expression profiles to identify cytogenetic driver lesions in myeloma important for disease development, progression and poor clinical outcome. Chromosomal positional enrichment analysis was employed to identify global myeloma cytogenetic driver aneuploidies as well as develop unique cytogenetic copy number profiles. Our results identified portions of chromosomes 1q, 3, 8p, 9, 13q and 16q, among others, as important driver lesions with changes to these regions providing growth advantages to the cell. Furthermore, our analysis identified five unique cytogenetic classifications based on common cytogenetic lesions. We continue to explore these driver regions to identify lesions important for the oncogenic properties of the larger regions.

Conclusion: The data presented here represents a novel and highly sensitive approach for the identification of novel copy number variations and driver lesions. Furthermore, correlations between copy number variations and gene expression arrays identified novel targets important for disease progression and patient survival. CytoScan HD arrays in conjunction with gene expression analysis provided a high resolution image of important cytogenetic lesions in myeloma and identified potentially important therapeutic targets for drug development. Further work is needed to validate our findings and determine the therapeutic efficacy of the identified targets.