Using Chaperonin Containing Tcp1 As A Marker To Track Clinically Relevant Circulating Tumor Cells.

Abstract Current methods for assessing circulating tumor cells (CTC) in blood focus on capture and enumeration with epithelial markers that provide little information about the aggressive and invasive potential of CTC. As a result, valuable knowledge that improves monitoring of cancer recurrence and metastasis is lacking. To provide actionable information on CTC, we developed a CTC identification method based on detection of the second subunit of Chaperonin Containing TCP1 (CCT2). CCT is a macromolecular protein folding complex composed of eight subunits (CCT1-8) that drives expression of many tumorigenic factors. Bioinformatic analysis in multiple cancers, such as breast cancer, revealed increased gene expression in tumor tissue compared to normal tissue for CCT1-8 (p< 0.0001), with the largest differences observed in CCT2 and CCT3. Genomic alteration of CCT2 inversely correlated with cancer patient survival, and depletion of CCT2 prevented growth of murine breast tumors. In a correlative study of blood and tumor tissues from metastatic breast cancer patients, high CTC counts positively correlated with detectable CCT2 in tumor tissues. These results support CCT2 as a possible marker for CTC. To determine this, antibodies to CCT2 were validated for use in the CellSearch System (CSS), currently the only FDA approved CTC enrichment platform, and tested with breast cancer cells spiked into healthy human blood. Recovery using CCT2 for identification, led to increased collection up to 6% of spiked breast cancer cells that would have been lost by standard CTC classification using cytokeratin (CK). With prognostic thresholds around 3-5 CTC/tube of blood, increased detection of even 1-2 more CTC can be significant for assessing patients' risks. Analysis in an open software program called ACCEPT (Automated CTC Classification, Enumeration and PhenoTyping) confirmed that the majority of recovered spiked cancer cells expressed CCT2. Flow cytometry analysis of spiked cancer cells confirmed that CCT2+ enriched cells expressed cancer markers CD44 or estrogen receptor (ER). These results show that the addition of CCT2 staining to a CTC detection protocol has positive implications for enumeration of CTC from blood, such as increased detection of CTC with low expression of epithelial markers. This experiment was repeated using small cell lung cancer cell lines with similar results. The combined bioinformatics, histologic and experimental results indicate that identification of CCT2-expressing CTC can be incorporated into CTC detection platforms to monitor those patients with a heightened risk for cancer recurrence or metastasis. Citation Format: Amanda Cox, Ana Martini, Eunkyung Lee, Rebecca Moroose, Amr Khaled, Annette R. Khaled. Using chaperonin containing TCP1 as a marker to track clinically relevant circulating tumor cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2601.