*3.5. Colorectal Cancer*

Colorectal cancer (CRC) is distinguished by a high inter-patient and intra-tumor heterogeneity that makes the identification of possible target antigens challenging. Hence, targeting a combination of CRC antigens seems to be a more promising approach. A preclinical study developed a VLP from the VP60 capsid proteins of RHDV displaying at the N-terminus peptide of the murine topoisomerase IIα (T.VP60), survivin (S.VP60) or both peptides (TS.VP60) [27]. These two antigens are involved in relevant mechanisms for tumor growth in both murine and human CRC. CRC cell line MC38-OVA was injected subcutaneously on day 0, and vaccination was performed over three consecutive days (days 7, 8, and 9) and monitored for 100 days. Synthetic CpGs (25 μg) were used as an adjuvant. T.VP60, S.VP60, and TS.VP60 vaccination delayed tumor growth; the first two treatments (T.VP60, S.VP60) improved survival by 60%, while TS.VP60 enhanced it by 73%. All mice that survived were tumor free at day 100; these mice received a second challenge of MC38-OVA, and none of the mice developed the tumor with 100% of survival up to the completion of the 200 day trial. The results suggest the development of a systemic memory response and demonstrate that a VLP vaccine with multiple epitopes can be an effective treatment strategy for CRC [27].

Other CRC-associated antigens are under investigation as possible targets of VLP-based vaccination. The Cluster of Differentiation 44 (CD44) is an adhesion molecule in the extracellular matrix that induces tumor cell invasion and metastasis. The variant form CD44v6 is a CRC cell marker and its positiveness is correlated with poor survival in CRC patients. CD44 variants stabilize cystine/glutamate antiporter protein xCT (see below) on the cell surface of several human carcinomas. The presence of xCT itself is an independent predictor of recurrence in CRC patients and is correlated with lymphatic and venous invasion. xCT is associated with CRC cancer cell growth, invasion and metastasis, making xCT a reliable target for the development of a CRC therapeutic vaccine [98]. Recently developed VLP-based vaccines targeting xCT [22,99] would be an ideal candidate for preclinical testing in CRC models.
