Expression of Human Kallikrein Protein and mRNA in Maxillofacial Cysts and Tumours

Non-inflammatory odontogenic cysts and tumours vary in clinical appearance and growth potential and can cause widespread destruction and deformation of the face. Proteases may play a role in the differing pathogenesis of these cysts and tumours. Human kallikrein proteins (KLKs) are a group of 15 serine proteases implicated in a variety of signalling and regulatory roles including as tumour suppressors in breast and gastric cancer.  KLK over-expression is associated with cancer/neogenesis.  KLK’s are used as biomarkers for identifying and monitoring progression of disease, most notably KLK 3 which is known as prostate specific antigen.  This study aimed to evaluate common neoplasms and cysts of odontogenic origin for the presence of KLK 3, 4, 5, 9 and 11.  Secondarily, we evaluated ameloblastoma tissue for expression of KLK 1-15 mRNA.

Archived paraffin embedded tissue samples were obtained from the Division of Oral Pathology at Western University.  Sixty (n=60) lesions were investigated including lateral periodontal cysts (n=9), dentigerous cysts (n=10), KOTs (n=11) and ameloblastomas (n=11) as well as nasopalatine duct cysts (n=9, non-odontogenic cystic control) and odontomas (n=10, neoplasm control).  These samples were cut and assessed for the presence of KLKs using a standard immunostaining technique utilizing antibodies for KLK 3, 4, 5, 9 & 11. Analysis of epithelial immunostaining was performed utilizing a scoring system assessing staining intensity as well as proportion of cells stained. Results were analyzed using the Kruskal-Wallis and Dunn’s multiple comparison tests.  KLK 1-15 mRNA expression was evaluated in ameloblastoma tissue using nine (n=9) pooled tumour samples, which were analyzed using reverse transcription-polymerase chain reaction (RT-PCR).   

Immunostaining revealed the presence of KLK 3, 4, 9 & 11 in all tissue types studied.  KLK 5 was present only in KOT’s (p<0.001).  Greater KLK 3 staining was present in ameloblastomas (p<0.01) and KOTs (p<0.05) than in the odontogenic control.  KLK 9 exhibited greater staining in KOTs (p<0.05) and dentigerous cysts (p<0.05) than the odontogenic control.  KLK 11 had greater staining in the ameloblastomas (p<0.05) than in non-odontogenic cystic controls. Expression of KLK 1, 4, 7, 8, 10 & 12 mRNA was found in pooled ameloblastoma tissue.   Expression of KLK 7 and 10 mRNA was greater than that of KLK 1, 4, 8 & 12 in the ameloblastoma tissue. KLK 2, 3, 5, 6, 9, 11, 13, 14 & 15 mRNA were not identified in ameloblastoma tissue.        

For the first time, KLK 3, 4, 5, 9 & 11 have been identified in maxillofacial cysts and tumours. KLK 3 is present in significantly greater amounts in benign neoplasms of odontogenic origin, the ameloblastoma and KOT, when compared to the odontogenic hamartoma. KLK 11 is present in significantly greater amounts in the benign neoplasm (ameloblastoma) compared to non-odontogenic cystic control (nasopalatine duct cyst). Interestingly, KLK 3 and 11 mRNA was not identified in the ameloblastoma using RT-PCR, suggesting that KLK 3 and 11 protein may be recruited, sequestered or have a long half life in benign neoplasms of odontogenic origin.  As expected, KLK 5, which is known to be involved in the process of keratinization, is present only in the KOT. Identification of KLK 4 mRNA in ameloblastoma tissue was also expected; it is implicated in normal amelogenesis.  With further investigation, it may be possible to use KLK’s as biomarkers for identification and monitoring these lesions.  Additionally, if the physiologic role of KLK’s in neoplasia can be identified, they may provide a target for minimally invasive therapeutics. 

References:

Darling MR et al. Human Kallikrein 8 Expression in Salivary Gland Tumors.  Head and Neck Pathol 2008 2:169–174.

Sotiropoulou G et al. Functional Roles of Human Kallikrein-related Peptidases. J Biol Chem 2009 Nov 27;284(48):32989-9.