Expression profiling of formalin-fixed paraffin embedded primary human uveal melanomas using DASL matrices.
OBJECTIVE
Fresh biopsied ocular tumor tissues are difficult to obtain for the purpose of performing microarray experiments on extracted nucleic acids. Present technology allows for extraction of total RNA from formalin-fixed paraffin embedded (FFPE) tissue analyzed by the cDNA mediated Annealing Sectioning and Ligation (DASL) method. We aimed to correlate gene transcript differences between two uveal melanoma (UM) clinical-histopathological parameters (metastasis, cell type).


METHODS
A total of 43 FFPE UM were used. The expression of RPL13a, a ribosomal protein gene, for each sample was used to evaluate the quality of RNA extracted from FFPE tissue. Gene expression values generated from the array were analyzed using the GeneSpring GX software (Agilent). Immunohistochemistry was used in order to validate transcriptional findings at the protein level.


RESULTS
A total of 106 genes were identified with (P < 0.05, Welch ANOVA test) a difference in transcript abundance for the metastasis clinical parameter. Furthermore, we identified 64 genes with a statistically significant (P < 0.05) difference in transcript abundance between the spindle and epithelioid cell types. Each individual sample for both groups (metastasis, cell type) exhibited distinct transcriptional profiles that were separated on a PCA. Positive nuclear immunostaining for LIG4-metastasis, ErbB3-cell type was found to be associated with better patient prognosis and outcome.


CONCLUSIONS
To the best of our knowledge, this is the first time that a successful retrospective analysis has been done with UM FFPE RNA. This data may lead to future customized therapeutic targets, which may improve the now unchanged mortality rate of this particular malignancy.
Di Cesare
Sebastian
S
The Henry C. Witelson Ophthalmic Pathology Laboratory and Registry, McGill University Health Center, 3775 University St., Lyman Duff Building, Room 216, Montreal, QC, H3A 2B4, Canada. sebastian.dicesare@mail.mcgill.ca
Nantel
Andre
A
Marshall
Jean-Claude
JC
Fernandes
Bruno F
BF
Antecka
Emilia
E
Orellana
Maria E
ME
Abourbih
Daniel
D
Saornil
Antonia M
AM
Burnier
Miguel N
MN
D005404:Fixatives / Q000737:chemistry; D005557:Formaldehyde / Q000737:chemistry; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D007150:Immunohistochemistry; D008545:Melanoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016612:Paraffin Embedding; D012334:RNA, Neoplasm / Q000032:analysis; D015203:Reproducibility of Results; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism; D014604:Uveal Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
D005404:Fixatives; D012334:RNA, Neoplasm; C402487:RPL13A protein, human; D012269:Ribosomal Proteins; D005557:Formaldehyde