O6-Methylguanine-DNA methyltransferase status in neuroendocrine tumours: prognostic relevance and association with response to alkylating agents

Abstract

Background: O(6)-Methylguanine-DNA methyltransferase (MGMT) loss of expression has been suggested to be predictive of response to temozolomide in neuroendocrine tumours (NETs), but so far, only limited data are available. We evaluated the prognostic and predictive value of MGMT status, assessed by two molecular methods and immunohistochemistry, in a large series of NETs of different origins.

Methods: A total of 107 patients, including 53 treated by alkylants (temozolomide, dacarbazine or streptozotocin), were retrospectively studied. In each case, we used methyl-specific PCR (MS-PCR) and pyrosequencing for evaluation of promoter methylation and immunohistochemistry for evaluation of protein status.

Results: MGMT promoter methylation was detected in 12 out of 99 (12%) interpretable cases by MS-PCR and in 24 out of 99 (24%) by pyrosequencing. O(6)-Methylguanine-DNA methyltransferase loss of expression was observed in 29 out of 89 (33%) interpretable cases. Status of MGMT was not correlated with overall survival (OS) from diagnosis. Progression-free survival and OS from first alkylant use (temozolomide, dacarbazine and streptozotocin) were higher in patients with MGMT protein loss (respectively, 20.2 vs 7.6 months, P<0.001 and 105 vs 34 months, P=0.006) or MGMT promoter methylation assessed by pyrosequencing (respectively, 26.4 vs 10.8 months, P=0.002 and 77 vs 43 months, P=0.026).

Conclusions: Our results suggest that MGMT status is associated with response to alkylant-based chemotherapy in NETs.

Figure 1

Immunostaining for MGMT protein. Representative examples of strong positive nuclear staining of tumour cells in a pancreatic NET (A) and of unambiguous negative staining of an ileal NET (B) are shown. Note in B the nuclear staining of lymphoid cells (L) and endothelial cells (arrows), which serve as internal positive controls (B). (C) An example of non-interpretable staining in an ileal NET: despite a very faint positivity observed in some tumour cell nuclei (open arrowheads), the absence of any positive internal control, especially in endothelial cells (arrows), prevents the definitive interpretation of the case. In D, an example of highly heterogeneous pancreatic NET is shown, with two distinct tumour cell populations, one with a faint nuclear labelling (arrows) and the other with no detectable labelling. Immunoperoxidase with nuclear counterstaining using Mayer's haematoxylin. Original magnifications: A, × 120; B, × 350; C, × 380; D, × 240; scale bar=50 μm.

Figure 2

Progression-free survival (A and B) and overall survival (C and D) from treatment initiation in patients treated by alkylating agents. In A and C, patients are compared with MGMT promoter methylation in tumour tissue, assessed by pyrosequencing (group 1: patients with methylated MGMT; group 2: patients with unmethylated MGMT). In B and D, patients are compared according to MGMT protein expression, assessed by immunohistochemistry (group 1: patients with MGMT-negative tumours; group 2: patients with MGMT-positive tumours).

Figure 3

Hypermethylated phenotype in NETs. In A, the distribution of tumours according to the number of methylated genes detected in tumour tissue is given. Open bars represent cases with methylated MGMT; solid bars represent cases with non-methylated MGMT. In B, overall survival curves from diagnosis are compared between patients with tumours containing ⩽3 methylated genes (group 1) and those with tumours containing >3 methylated genes (group 2). The difference is statistically significant (log-rank test, P=0.017). In C, overall survival curves from diagnosis are compared for the subset of patients with tumours containing >3 methylated genes according to their MGMT status (group 1: methylated MGMT, group 2: non-methylated MGMT). The difference is not statistically significant.