Abstract P6-14-16: Claudin-9 and its subcoat anchorage proteins ZO-1 and ZO-3 in breast cancer, the clinical and therapeutic significance
Authors: Zhuang Xinguo , Jiang Wen G. , Davies Eleri , Xu Bing , Martin Tracey A. 
Published: March 2023

Abstract
Introduction. Claudin-9 (CLDN9), a member of the claudin protein family, is thought to play a role in the control of tight junctions (TJ) in various cell types. CLDN9 protein needs intracellular subcoat proteins, primarily the Zonula Occluden (ZO)-1 and ZO-3 to anchor to the cytoskeleton in order to form TJ with other proteins, including occludin. In the body, CLDN9 is highly expressed in endocrine tissues but is relatively low in mammary tissues. The role of CLDN9 is not well understood in cancer. In the present study, we have for the first time examined the pattern of CLDN9 expression at protein and transcript levels in breast cancer and explored its clinical and therapeutic implications. Methods. CLDN9 protein and CLDN9 transcript in fresh frozen human mammary tissues were evaluated by immunohistochemistry and quantitative transcript analyses and, together with the ZO family members and occludin in our database, correlated with clinical indicators. Assessment of expression in a range of cell lines was also determined. The levels of CLDN9 transcript, was also assessed against the therapeutic responses of the patients to chemotherapies by using a dataset from TCGA database. Results. Breast tumour tissues had high levels of CLDN9 transcript in tumours versus normal tissues. However, high grade breast tumours had significantly lower levels than low grade (p=0.02 and p=0.003, grade-2 and 3 vs grade-1 respectively). Patients with metastasis also had significantly lower levels than those without (p=0.0075). Patients who died of breast cancer had higher levels of the transcript than those who survived, although this was not significant. CLDN9 expression was significantly correlated with ZO-1 (r=0.20, p&lt; 0.001) and ZO-3 (r=0.179, p&lt; 0.01), but not ZO-2 in our cohort. There was significant correlation with another key TJ molecule, occludin (r=0.236, p=0.11). CLDN9, together with ZO-1 and ZO-3 significantly linked to the survival of patients (1444.6 months for the low expressing group versus 1138.2 months for the high expression group, p=0.013). The expression profile of the CLDN9/ZO1/ZO3 complex also indicated potential as an independent prognostic indicator (p=0.004, HR=2.033). The prognostic value was highly applicable to non-triple negative breast cancer patients. CLDN9 transcript also appeared to have a significant impact on treatment responses; patients who were sensitive to chemotherapies had a significantly lower levels of CLDN9 transcript than those who were resistant to treatment (p&lt; 0.000001). In human cell lines, expression levels of CLDN-9 in ER (+) breast cancer cell lines, MDAMB361, MCF7 and BT474 were high; in a ER(-) breast cancer cell line MDAMB231, the expression level of CLDN-9 was lower. Conclusion. In conclusion, CLDN9 expression was differentially expressed in human breast cancer cells lines and appeared to reflect changes in ER status. It was apparent from levels of CLDN9 in a breast cancer cohort that expression was associated with grade and metastatic disease. Of significant notice was the finding that expression of CLDN9 in an expression profile with ZO1 and ZO3 has potential as a prognostic indicator, particularly in non-triple negative patients and in those who are chemotherapy resistant.
Citation Format: Xinguo Zhuang, Wen G. Jiang, Eleri Davies, Bing Xu, Tracey A. Martin. Claudin-9 and its subcoat anchorage proteins ZO-1 and ZO-3 in breast cancer, the clinical and therapeutic significance [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P6-14-16.