Production of anti-KLK14 polyclonal antibodies

• Grant number: 22/01968-8
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: August 01, 2022
• End date: July 31, 2023
• Field of knowledge: Health Sciences - Dentistry
• Principal Investigator: Katiuchia Uzzun Sales
• Grantee: Bruna Miyoko Ikenaga de Brito
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

Head and neck squamous cell Carcinoma (HNSCC) is a type of Cancer with high incidence and is difficult to treat. Therefore, the discovery of new therapeutic targets for better treatments and, consequently, higher quality of life for these patients is needed. Kallikreins (KLKs) are serine proteases that have roles in both physiological and pathological processes, such as epithelial desquamation and malignant neoplasias. Indeed, a recent study from our laboratory has shown that the relative expression of kallikrein 5 (KLK5) to its inhibitor Lympho-epithelial Kazal-type inhibitor (LEKTI) is an important implement to the histopathologic diagnosis of HNSCC. Among other targets, KLK5 is capable of cleaving and activating kallikrein 14 (KLK14). Thus, aiming to continue our studies, we have hypothesized that KLK5 may activate KLK14 in malignant tumors. It is important to mention that KLK14 is overexpressed in different types of cancers, such as prostate, breast, ovary, and testicle. To this end, it is necessary for us to develop a specific methodology to assess KLK14 expression in tissues. In particular, the relevance of antibodies in biochemical assays for research and diagnosis is explained by the high specificity between antigen and antibody. Indeed, antibody production emerges as a fundamental tool for the study of KLK14 role in carcinomas. Therefore, the production of immune serum in rabbits is in fact an alternative. This proposal presents the development of three polyclonal antibodies: (I) anti-murine KLK14, using the full murine KLK14 protein; (II) anti-murine KLK14, using a fragment of the murine KLK14 protein; and (III) anti-human KLK14, using the full human KLK14 protein. Methodologically, the production of the aforementioned anti-KLK14 antibodies will follow the following steps: (I) DNA cloning in pGEX4T-1 vector; (II) production of the full KLK14 protein or its fragment, and (III) production of serum immune using rabbits immunization. At last, this proposal is justified by the absence of available specific antibodies on the market combined with the need to study KLK14 in the HNSCC context. Kallikreins (KLKs) are extracellular proteins and, therefore, promising therapeutic targets for developing therapies for this challenging disease.(AU)