Targeting cancer stem cells by raising cytotoxic T-cell responses against the stemness protein SOX2

Despite significant advances in medicine, one of the major challenges in oncology remains the frequent recurrence of tumor, even in the absence of clinical signs of neoplasia. Growing evidence indicates that cancer relapse is likely driven by a rare population of cells endowed with stem cell characteristics lurking within treated primary tumor beds and initiating tumors at distant places. Therefore, specific targeting of Cancer Stem Cells (CSCs) could turn a short-term recovery into long-term disease-free survival of many cancer patients.
One of the major properties ascribed to cancer stem cells is the expression of embryonic genes. In our experiments we evaluate the potential of the regulatory transcription factor SOX2 as a target for anti-cancer vaccination. It has been shown that SOX2 together with other transcription factors such as OCT4 and NANOG play an important role in maintaining the stemness of embryonic stem cells and become reactivated in several human cancers.
In a first instance we evaluated the immunogenicity of the SOX2 protein. To that aim we applied a novel immunization technique in which mRNA encoding a given antigenic protein is directly injected into inguinal lymph nodes. Moreover, in our experiments along with antigen-encoding mRNA we co-immunize with mRNA encoding a constitutively active TLR4, CD40L and CD70 ('TriMix'), which was developed at our laboratory and to enhance T-cell stimulation.

By means of in vivo CTL assay, we show that mice immunized with SOX2-TriMix mRNA successfully developed anti-SOX2 immune responses. This has been envisaged by the lysis of SOX2-expressing target cells stained with CFSE and administered intravenously to the recipient mouse. Additionally, we identified candidate groups of peptides that seem to be the most immunogenic out of 86 overlapping SOX2 peptides.
Preliminary ELISA and intracellular cytokine staining experiments performed on spleno-cytes isolated from immunized mice, showed that in response to in vitro re-stimulation these cells secrete cytokines such as IFN-g and IL-2 and exhibit a lytic capacity in vitro against target cells expressing antigen of interest.
Currently performed experi-ments aim to determine specific epitopes within the SOX2 sequence.
Here, SOX2-specific T cells will be subsequently cloned by limiting dilution.
Moreover, in the future we will examine whether SOX2 immunization is safe and does not result in autoimmunity.
Ultimately, we will evaluate the benefits of targeting CSCs versus bulk tumor cells, by evaluating the efficacy of an immunotherapy targeting CSCs through anti-SOX2 immunization versus a TRP2-specific immuno-therapy designed to target target bulk tumor cells.