Development of SERS nanoprobes for multiplex receptors imaging in cancerous tissues

One of the modern challenges in oncology is the fine molecular characterization of cancer for diagnosis, monitoring and treatment [1]. Nanoprobes based on Surface-Enhanced Raman scattering (SERS) are emerging as powerful alternative to fluorescent labels in biomedicine. SERS nanoprobes offer higher spectral specificity than fluorescent labels in combination with a small vibrationnal bandwidth allowing higher multiplexing capabilities. Therefore, multiple molecular information about a tumor can be obtained simultaneously from a single sample. SERS nanoprobes are composed of a metallic nanoparticle (NP) covered by a Raman-active molecule, and functionalized by a targeting molecule able to provide specificity toward a particular biological target. The role of the metallic nanoparticle is to enhance the Raman signal of the Raman-active molecule to be able to detect small amount of nanoprobes in the sample. SERS nanoprobes combined to Raman micro-imaging can be used to localize and/or quantify the biological target through the unique and intense spectral signature of the Raman-active molecule. In this work, we developed two targeting SERS nanoprobes to evaluate simultaneously the expression of membrane receptors in cancerous tissues slides. We focused on two receptors of interest : Folate Receptors (FRs) and Sialic Acid. Overexpression of FRs is observed in several cancers (ovary, lung, breast, etc.) and has been linked to poor overall survival, to an increase in the tumor agressivness or in the resistance to the treatment. Sialic acid overexpression is closely related to the metastatic phenotype of the tumor. Therefore, obtaining simultaneous information about the expression status of those receptors is relevant for phenotyping tumors. In our case, SERS nanoprobes are both composed of a bimetallic gold@silver core@shell nanoparticle (Au@Ag NP). These nanoparticles provide superior SERS enhancement than traditionnal gold nanoparticles while providing highly tunable optical properties by controlling the shell thickness. Moreover, the reproducibility of the synthesis is greatly increased compared to silver nanoparticles. Both SERS nanoprobes were used independently firstly on cell lines and then on model tumors to confirm their targeting ability. The microscopic resolution offered by micro-Raman imaging allows to follow really accurately the tumor borders and to highlight the overexpression of those receptor in tumors. Also we discuss the simultaneous detection of both nanoprobes on the same tissue sample.