RASSF1 alteration has been suggested as a molecular hallmark of pancreatic cancer [
2]. In the attempt to clarify the involvement of
RASSF1 in this setting, we performed a comprehensive analysis of expression, methylation and copy number status of this putative tumor suppressor gene in a relatively large series of primary PDAC, xenografts and PDAC cell lines.
Rassf1a expression was downregulated in about 50 % of samples. However, such downregulation was not correlated with the
RASSF1 methylation status, which indicates that
RASSF1 methylation is not an essential mechanism for regulating protein expression in PDAC. This is the first study investigating RASSF1 isoforms expression in PDAC. RASSF1A, RASSF1B and RASSF1C mRNAs were expressed in all xenografts and cell lines, and, as observed for immunohistochemical data, no correlation between the expression level of the RASSF1 isoforms and methylation of CpG islands was found. However, RASSF1F was preferentially expressed by cell lines lacking methylation at CpG island A (Wilcoxon test,
p = 0.008), in line with the results of Dammann et al. [
2,
3]. This suggests that methylation of CpG island A may affect expression and splicing of RASSF1 transcripts regulated by the same promoter. The lack of correlation between the level of methylation of CpG island A and RASSF1A mRNA level in PDAC xenografts and cell lines further suggests that methylation is not decisive for transcription regulation of RASSF1 isoforms. No correlation was found between methylation and outcome of disease, at variance with other cancer types [
13,
16‐
21]. A consistent number (71/96, 74 %) of
RASSF1 locus alterations was found in our series, including 54 % (50/96) of losses and 22 % (21/96) of gains. The frequency of
RASSF1 loss found is similar to that reported in a previous LOH microsatellite study investigating 82 PDAC xenografts [
26]. Notably, among the 50 samples with loss of
RASSF1 locus, 30 (60 %) had a normal chromosome 3 ploidy, while the remaining 20 (40 %) samples had chromosome 3 polysomy. Previous studies documented frequent chromosomal 3 alterations in pancreatic cancer [
25‐
27,
35,
36], including in particular the allelic loss at 3p21.3 [
24,
25]. Harada et al. observed frequent gains of chromosome 3 by high-density single nucleotide polymorphism array (78 % of cases), associated with losses in the specific region of chromosome 3p21 [
26], this might reflect a propensity for 3p21.3 loss to occur as a secondary event of large 3p deletions, which involves regions coding for other tumor suppressor genes. Interestingly, chromosome 3 poliploidy was associated with a worse prognosis in our series, independently of the status of
RASSF1 locus, while patients with a disomic status of both
CEP3 and
RASSF1 had a similar prognosis compared to patients with disomic
CEP3 and loss of
RASSF1. It has been described that
RASSF1 methylation can have different roles depending on the ploidy status and patient’s age in neuroblastoma [
40]. However, there is no correlation between methylation and ploidy status in our PDAC series. It is possible that other tumor suppressor genes in chromosome 3p may be implicated in the clinical course of these tumors.