Surgically removed formalin-fixed tumour samples were obtained from 65 patients (35 males and 30 females with a median age at diagnosis of 55 years) treated with the histopathological diagnosis of an SGC according to the WHO classification (Barnes et al, 2005). All patients received surgery and in selected cases of postoperative radiation therapy. Epidermal growth factor receptor-targeted therapy was not applied. The study cohort consisted of ACC (n=25), mucoepidermoid carcinoma (n=10), myoepithelial carcinoma (n=8), acinic cell carcinoma (n=12) and adenocarcinoma ex pleomorphic adenoma (n=10).

Deoxyribonucleic acid was isolated from microdissected tumour areas using the QIAamp DNA Mini Kit (Qiagen GmbH, Hilden, Germany).

Epidermal growth factor exons 18, 19, 20 and 21 were amplified by nested PCR using primers described previously (Eberhard et al, 2005). Polymerase chain reaction products were purified with the QIAquick PCR Purification Kit (Qiagen GmbH). Genomic sequencing was performed using fluorescent dye-terminator chemistry and the primer M13F: 5′-TGTAAAACGACGGCCAGT-3′ (MWG Biotech, Martinsried, Germany). The reported mutations were scored as reproducible, having been identified in two independent PCR amplifications and by M13 sequencing in both directions (M13R: 5′-CAGGAAACAGCTATGACC-3′).

The numbering of the EGFR nucleotide and amino acid positions was according RefSeq sequences NM 005228.3 and NP 005219.2, respectively.

This study was conducted in accordance with the principles of the Declaration of Helsinki, as adopted by the 29th World Medical Assembly, Helsinki, Finland.

Sequencing analysis of exons 18–21 of the EGFR tyrosine kinase domain in 65 SGCs revealed two cases (3.1, 95% exact confidence limit: 0.5–1.2) with a heterozygous exon 19 deletion mutation. The two exon 19 E746_A750 deletions (both of them were variant c.2235_2249del15) were identified in a mucoepidermoid carcinoma of the left parotid gland grade 2 (54-year-old female patient, smoker), and in an ACC of solid type of the right parotid gland (69-year-old male patient, non-smoker). The female patient with the mucoepidermoid carcinoma was treated with surgery and postoperative radiation, the male patient with ACC by surgical tumour resection alone. Both patients had a favourable course of their cancer disease with no manifestation of recurrences (follow-up time 24 months).

In exon 20, an SNP c.2607 G>A (p.Q787Q) was detected in 29 out of 65 tumour samples (44.6%, 95% exact confidence limit: 32.5–57.4).

Somatic mutations in the EGFR kinase domain are found in a subset of lung adenocarcinomas and are associated with sensitivity to TKI (point mutations G719A/C in exon 18, L858R and L861Q in exon 21 and in-frame deletions at position 745 of exon 19). Three kinase domain mutations are associated with drug resistance: D761Y in exon 19, T790M in exon 20 and an exon 20 insertion (D770N771insNPG). The exon 19 deletions and L858R in exon 21 represent 85–90% of EGFR mutations in non-small cell lung cancer. In two recent trials in advanced non-small cell lung cancer, therapy with gefitinib administered in a genotype-directed fashion to patients harbouring EGFR mutations resulted in very favourable clinical outcomes with good tolerance (Sequist et al, 2008; Tamura et al, 2008).

Given the comparable embryonal tissue origin between lung adenocarcinoma and SGCs and the potential for genotype-directed trials of EGFR inhibitors, we undertook a comprehensive mutational analysis of EGFR in SGCs. We found EGFR mutations in the tyrosine kinase domain to be a rare event in SGCs of Caucasian origin. Whereas approximately 10–30% of lung carcinoma contain EGFR mutations (Mitsudomi and Yatabe, 2007), they seem to be seldom acquired in the cancers of other organs (EGFR mutation database:http://www.cityofhope.org/cmdl/egfr%5Fdb/).

Both of the mutations detected in our cohort were found in histopathological types (mucoepidermoid carcinoma and ACC), which also occur in the lung. In a recently published study, in two out of five (40%) of mucoepidermoid carcinoma of lung (which originate from minor salivary glands of the tracheobrochial tree), the L858R mutation was detected and all five patients well responded to TKI therapy independent of the mutation status (Han et al, 2008). In contrast, in another study with 24 cases of adenoid cystic and mucoepidermoid salivary gland-type lung carcinomas, EGFR protein expression was found in 91%, but no mutation was detected in exons 18–21 of the EGFR gene (Macarenco et al, 2008). In head-and-neck cancers, in which about 10% of cases showed responses to gefitinib, no mutations were noted in a US study, but 7% of tumours of Asian origin were found to harbour such mutations (Cohen et al, 2005; Lee et al, 2005).

In conclusion, drug-sensitising EGFR mutations are rather rare in SGCs and seem to be mainly found in a few mucoepidermoid and ACC. Taking into consideration the encouraging relative high number of responding SGC patients (particularly ACC patients with poor prognosis) who achieved stable disease after cetuximab treatment in the phase II trials and the apparently low incidence of drug-sensitising EGFR mutations, it would appear that also other biological factors than EGFR tyrosine kinase mutations might influence the therapy response. The underlying mechanisms of response to targeted therapies are complex and still not fully elucidated. Investigating those mechanisms and the mutation status of proteins in the cascade downstream of EGFR might identify further predictive markers for EGFR-targeted therapy in SGCs and would allow the selection of patients for a successful treatment.

We thank K Tunnat and G Fiedler from HELIOS Erfurt and A Reck from the University of Regensburg for excellent technical assistance.