Artificial Intelligence-based Digital Pathology Assessment of CD44s Expression in Breast Cancer

Association with Clinicopathological Features and Survival Outcomes

Authors

DOI:

https://doi.org/10.14500/aro.12248

Keywords:

Artificial intelligence, Breast cancer stem cells, Breast cancer, CD44s, Immunohistochemistry, QuantCenter

Abstract

Breast cancer (BC) exhibits considerable molecular and clinical heterogeneity, complicating prognostic evaluation. The cluster of differentiation 44 standard (CD44s) isoform has been proposed as a prognostic marker in various cancers; however, its role in BC remains unclear. This study evaluated CD44s expression in BC tissues and its association with clinicopathological features and survival outcomes using an artificial intelligence (AI)-based digital pathology scoring method. A retrospective analysis of 98 BC tissue samples is conducted, with CD44s cell membrane protein expression assessed through both manual and AI based immunohistochemical (IHC) scoring. Statistical analyses included Pearson’s chi-square test, Kaplan-Meier (log-rank), and Cox regression. CD44s expression was observed in 65.31% of patients. No significant associations are found between CD44s expression and clinicopathological characteristics, including age, tumor size, lymph node metastasis, histological grade, lymphovascular invasion (LVI), or hormone receptor status (all p > 0.05). Survival analysis reveals no significant association between CD44s expression and overall survival (OS, p = 0.1345) or progression-free survival (p = 0.0669). While CD44s expression is prevalent in BC samples, it is not an independent prognostic factor; LVI is the only significant predictor of OS (p = 0.036). Finally, the moderate agreement between AI and manual scoring (Cohen’s Kappa = 0.4337, p < 0.0001) supports the potential of AI-assisted methods for biomarker quantification, warranting further validation in larger cohorts.

Downloads

Download data is not yet available.

Author Biographies

Avan S. Mohammed, Department of Biology, Faculty of Science, University of Duhok, Duhok, Kurdistan Region – F.R. Iraq

Avan S. Mohammed is an Assistant Lecturer at the Ministry of Education, Kurdistan Region, Iraq. She received both her B.Sc. and M.Sc. degrees in Biology - Molecular biology from the University of Duhok and is currently pursuing her Ph.D. in the Department of Biology, College of Science, University of Duhok. Her research interests include molecular biology, human genetics, hematology, and cancer stem cells.

Ramadhan T. Othman, Department of Medicine, College of Medicine, University of Duhok, Duhok, Kurdistan Region – F.R. Iraq

Ramadhan T. Othman is an Assistant Professor at the Department of Clinical Oncology, College of Medicine, University of Duhok. He obtained his MBChB in Medicine, followed by an M.Sc. and Ph.D. in Clinical Oncology from the University of Nottingham, UK. His research interests are in cancer treatment resistance, clinical outcomes, and molecular biology. Dr. Othman is a member of the European Society for Medical Oncology (ESMO) and the American Society of Clinical Oncology (ASCO).

Rafil T. Yaqo, Department of Pathology, College of Medicine, University of Duhok, Duhok, Kurdistan Region – F.R. Iraq

Rafil T. Yaqo is an Assistant Professor at the Department of Pathology, College of Medicine, University of Duhok. He earned his MBChB and FIBMS (Path) from the University of Duhok. He also obtained his certification from the Iraqi Board of Medical Specialties at the Mosul Medical Training Centre, Department of Pathology, Northern Iraq. His research interests are in histopathology, diagnostic pathology, and laboratory medicine. Dr. Yaqo is a member of the Kurdistan Syndicate of Medical Doctors and is currently practicing as a specialty doctor in the Histopathology Department at William Harvey Hospital, UK.

References

Abraham, B.K., Fritz, P., McClellan, M., Hauptvogel, P., Athelogou, M., and Brauch, H., 2005. Prevalence of CD44+/CD24−/low cells in breast cancer may not be associated with clinical outcome but may favour distant metastasis. Clinical Cancer Research, 11(3), pp.1154-1159. DOI: https://doi.org/10.1158/1078-0432.1154.11.3

Acs, B., Pelekanou, V., Bai, Y., Martinez-Morilla, S., Toki, M., Leung, S.C., Nielsen, T.O., and Rimm, D.L., 2019. Ki67 reproducibility using digital image analysis: An inter-platform and inter-operator study. Laboratory Investigation, 99(1), pp.107-117. DOI: https://doi.org/10.1038/s41374-018-0123-7

Aeffner, F., Adissu, H.A., Boyle, M.C., Cardiff, R.D., Hagendorn, E., Hoenerhoff, M.J., Klopfleisch, R., Newbigging, S., Schaudien, D., Turner, O., and Wilson, K., 2018. Digital microscopy, image analysis, and virtual slide repository. ILAR journal, 59(1), pp.66-79. DOI: https://doi.org/10.1093/ilar/ily007

Al-Hajj, M., Wicha, M.S., Benito-Hernandez, A., Morrison, S.J., and Clarke, M.F., 2003. Prospective identification of tumorigenic breast cancer cells. Proceedings of the National Academy of Sciences, 100(7), pp.3983-3988. DOI: https://doi.org/10.1073/pnas.0530291100

Barnard, M.E., Boeke, C.E., and Tamimi, R.M., 2015. Established breast cancer risk factors and risk of intrinsic tumour subtypes. Biochimica Biophysica Acta (BBA)-Reviews on Cancer, 1856(1), pp.73-85. DOI: https://doi.org/10.1016/j.bbcan.2015.06.002

Bei, Y., Cheng, N., Chen, T., Shu, Y., Yang, Y., Yang, N., Zhou, X., Liu, B., Wei, J., Liu, Q., Zheng, W., Zhang, W., Su, H., Zhu, W., Ji, J., and Shen, P., 2020. CDK5 inhibition abrogates TNBC stem‐cell property and enhances anti‐PD‐1 therapy. Advanced Science, 7(22), p.2001417. DOI: https://doi.org/10.1002/advs.202001417

Braun, M., Piasecka, D., Bobrowski, M., Kordek, R., Sadej, R., and Romanska, H.M., 2020. A ‘Real-Life’experience on automated digital image analysis of FGFR2 immunohistochemistry in breast cancer. Diagnostics(Basel), 10(12), p.1060. DOI: https://doi.org/10.3390/diagnostics10121060

Bray, F., Laversanne, M., Sung, H., Ferlay, J., Siegel, R.L., Soerjomataram, I., and Jemal, A., 2024. Global cancer statistics 2022: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA: A Cancer Journal for Clinicians, 74(3), pp.229-263. DOI: https://doi.org/10.3322/caac.21834

Brown, R.L., Reinke, L.M., Damerow, M.S., Perez, D., Chodosh, L.A., Yang, J., and Cheng, C., 2011. CD44 splice isoform switching in human and mouse epithelium is essential for epithelial-mesenchymal transition and breast cancer progression. The Journal of Clinical Investigation, 121(3), pp.1064-1074. DOI: https://doi.org/10.1172/JCI44540

Bustreo, S., Osella-Abate, S., Cassoni, P., Donadio, M., Airoldi, M., Pedani, F., Papotti, M., Sapino, A., and Castellano, I., 2016. Optimal Ki67 cut-off for luminal breast cancer prognostic evaluation: A large case series study with a long-term follow-up. Breast Cancer Research and Treatment, 157, pp.363-371. DOI: https://doi.org/10.1007/s10549-016-3817-9

Cho, Y., Lee, H.W., Kang, H.G., Kim, H.Y., Kim, S.J., and Chun, K.H., 2015. Cleaved CD44 intracellular domain supports activation of stemness factors and promotes tumorigenesis of breast cancer. Oncotarget, 6(11), pp.8709-8721. DOI: https://doi.org/10.18632/oncotarget.3325

Clark, G.C., Hampton, J.D., Koblinski, J.E., Quinn, B., Mahmoodi, S., Metcalf, O., Guo, C., Peterson, E., Fisher, P.B., Farrell, N.P., Wang, X.Y., and Mikkelsen, R.B., 2022. Radiation induces ESCRT pathway dependent CD44v3+ extracellular vesicle production stimulating pro-tumor fibroblast activity in breast cancer. Frontiers in Oncology, 12, p.913656. DOI: https://doi.org/10.3389/fonc.2022.913656

Dai, X., Li, T., Bai, Z., Yang, Y., Liu, X., Zhan, J., and Shi, B., 2015. Breast cancer intrinsic subtype classification, clinical use and future trends. American Journal of Cancer Research, 5(10), pp.2929-2943. DOI: https://doi.org/10.1371/journal.pone.0124964

Elston, C.W., and Ellis, I.O., 1991. Pathological prognostic factors in breast cancer. I. The value of histological grade in breast cancer: Experience from a large study with long‐term follow‐up. Histopathology, 19(5), pp.403-410. DOI: https://doi.org/10.1111/j.1365-2559.1991.tb00229.x

Giuliano, A.E., Edge, S.B., and Hortobagyi, G.N., 2018. Eighth edition of the AJCC cancer staging manual: Breast cancer. Annals of Surgical Oncology, 25, pp.1783-1785. DOI: https://doi.org/10.1245/s10434-018-6486-6

Gong, Y., Sun, X., Huo, L., Wiley, E.L., and Rao, M.S., 2005. Expression of cell adhesion molecules, CD44s and E‐cadherin, and microvessel density in invasive micropapillary carcinoma of the breast. Histopathology, 46(1), pp.24-30. DOI: https://doi.org/10.1111/j.1365-2559.2004.01981.x

Goodison, S., Urquidi, V., and Tarin, D., 1999. CD44 cell adhesion molecules. Molecular Pathology, 52(4), pp.189-196. DOI: https://doi.org/10.1136/mp.52.4.189

Gu, J., Chen, D., Li, Z., Yang, Y., Ma, Z., and Huang, G., 2022. Prognosis assessment of CD44+/CD24− in breast cancer patients: A systematic review and meta-analysis. Archives of Gynecology and Obstetrics, 306(4), pp.1147-1160. DOI: https://doi.org/10.1007/s00404-022-06402-w

Guo, Q., Liu, Y., He, Y., Du, Y., Zhang, G., Yang, C., and Gao, F., 2021. CD44 activation state regulated by the CD44v10 isoform determines breast cancer proliferation. Oncology Reports, 45(4), p.7. DOI: https://doi.org/10.3892/or.2021.7958

Herrera-Gayol, A., and Jothy, S., 1999. Adhesion proteins in the biology of breast cancer: Contribution of CD44. Experimental and Molecular Pathology, 66(2), pp.149-156. DOI: https://doi.org/10.1006/exmp.1999.2251

Hu, J., Li, G., Zhang, P., Zhuang, X., and Hu, G., 2017. A CD44v+ subpopulation of breast cancer stem-like cells with enhanced lung metastasis capacity. Cell Death Disease, 8(3), p.e2679. DOI: https://doi.org/10.1038/cddis.2017.72

Lee, K., Kruper, L., Dieli-Conwright, C.M., and Mortimer, J.E., 2019. The impact of obesity on breast cancer diagnosis and treatment. Current Oncology Reports, 21, p.41. DOI: https://doi.org/10.1007/s11912-019-0787-1

Lee, S.J., Go, J., Ahn, B.S., Ahn, J.H., Kim, J.Y., Park, H.S., Kim, S.I., Park, B.W., and Park, S., 2023. Lymphovascular invasion is an independent prognostic factor in breast cancer irrespective of axillary node metastasis and molecular subtypes. Frontiers in Oncology, 13, p.1269971. DOI: https://doi.org/10.3389/fonc.2023.1269971

Liu, Y., Han, D., Parwani, A.V., and Li, Z., 2023. Applications of artificial intelligence in breast pathology. Archives of Pathology and Laboratory Medicine, 147(9), pp.1003-1013. DOI: https://doi.org/10.5858/arpa.2022-0457-RA

Lopez, J.I., Camenisch, T.D., Stevens, M.V., Sands, B.J., McDonald, J., and Schroeder, J.A., 2005. CD44 attenuates metastatic invasion during breast cancer progression. Cancer Research, 65(15), pp.6755-6763. DOI: https://doi.org/10.1158/0008-5472.CAN-05-0863

McCaffrey, C., Jahangir, C., Murphy, C., Burke, C., Gallagher, W.M., and Rahman, A., 2024. Artificial intelligence in digital histopathology for predicting patient prognosis and treatment efficacy in breast cancer. Expert Review of Molecular Diagnostics, 24(5), pp.363-377. DOI: https://doi.org/10.1080/14737159.2024.2346545

Mohamed, S.Y., Kaf, R.M., Ahmed, M.M., Elwan, A., Ashour, H.R., and Ibrahim, A., 2019. The prognostic value of cancer stem cell markers (Notch1, ALDH1, and CD44) in primary colorectal carcinoma. Journal of Gastrointestinal Cancer, 50, pp.824-837. DOI: https://doi.org/10.1007/s12029-018-0156-6

Nishimura, R., Osako, T., Okumura, Y., Nakano, M., Ohtsuka, H., Fujisue, M., and Arima, N., 2022. An evaluation of lymphovascular invasion in relation to biology and prognosis according to subtypes in invasive breast cancer. Oncology Letters, 24(2), p.245. DOI: https://doi.org/10.3892/ol.2022.13366

Pati, P., Karkampouna, S., Bonollo, F., Compérat, E., Radić, M., Spahn, M., Martinelli, A., Wartenberg, M., Kruithof-de Julio, M., and Rapsomaniki, M., 2024. Accelerating histopathology workflows with generative AI-based virtually multiplexed tumor profiling. Nature Machine Intelligence, 6(9), pp.1077-1093. DOI: https://doi.org/10.1038/s42256-024-00889-5

Paulsen, I.M.S., Dimke, H., and Frische, S., 2015. A single simple procedure for dewaxing, hydration and heat-induced epitope retrieval (HIER) for immunohistochemistry in formalin fixed paraffin-embedded tissue. European Journal of Histochemistry EJH, 59(4), p.2532. DOI: https://doi.org/10.4081/ejh.2015.2532

Rakha, E.A., Vougas, K., and Tan, P.H., 2022. Digital technology in diagnostic breast pathology and immunohistochemistry. Pathobiology, 89(5), pp.334-342. DOI: https://doi.org/10.1159/000521149

Schmitt, F., Ricardo, S., Vieira, A.F., Dionísio, M.R., and Paredes, J., 2012. Cancer stem cell markers in breast neoplasias: Their relevance and distribution in distinct molecular subtypes. Virchows Archiv, 460, pp.545-553. DOI: https://doi.org/10.1007/s00428-012-1237-8

Senbanjo, L.T., and Chellaiah, M.A., 2017. CD44: A multifunctional cell surface adhesion receptor is a regulator of progression and metastasis of cancer cells. Frontiers in Cell and Developmental Biology, 5, p.18. DOI: https://doi.org/10.3389/fcell.2017.00018

Somal, P.K., Sancheti, S., Sharma, A., Sali, A.P., Chaudhary, D., Goel, A., Dora, T.K., Brar, R., Gulia, A., and Divatia, J., 2023. A clinicopathological analysis of molecular subtypes of breast cancer using immunohistochemical surrogates: A 6-year institutional experience from a tertiary cancer center in north India. South Asian Journal of Cancer, 12(02), pp.104-111. DOI: https://doi.org/10.1055/s-0043-1761942

Song, Y.J., Shin, S.H., Cho, J.S., Park, M.H., Yoon, J.H., and Jegal, Y.J., 2011. The role of lymphovascular invasion as a prognostic factor in patients with lymph node-positive operable invasive breast cancer. Journal of Breast Cancer, 14(3), pp.198-203. DOI: https://doi.org/10.4048/jbc.2011.14.3.198

Steinbichler, T.B., Dudás, J., Skvortsov, S., Ganswindt, U., Riechelmann, H., and Skvortsova, I.I., 2018. December. Therapy resistance mediated by cancer stem cells. In: Seminars in Cancer Biology. Vol. 53. Academic Press, United States, pp.156-167. DOI: https://doi.org/10.1016/j.semcancer.2018.11.006

Turner, K.M., Yeo, S.K., Holm, T.M., Shaughnessy, E., and Guan, J.L., 2021. Heterogeneity within molecular subtypes of breast cancer. American Journal of Physiology Cell Physiology, 321(2), pp.C343-C354. DOI: https://doi.org/10.1152/ajpcell.00109.2021

Vadhan, A., Hou, M.F., Vijayaraghavan, P., Wu, Y.C., Hu, S.C.S., Wang, Y.M., Cheng, T.L., Wang, Y.Y., and Yuan, S.S.F., 2022. CD44 promotes breast cancer metastasis through AKT-mediated downregulation of nuclear FOXA2. Biomedicines, 10(10), p.2488. DOI: https://doi.org/10.3390/biomedicines10102488

Vallejos, C.S., Gómez, H.L., Cruz, W.R., Pinto, J.A., Dyer, R.R., Velarde, R., Suazo, J.F., Neciosup, S.P., León, M., De La Cruz, M.A., and Vigil, C.E., 2010.

Breast cancer classification according to immunohistochemistry markers: Subtypes and association with clinicopathologic variables in a peruvian hospital database. Clinical Breast Cancer, 10(4), pp.294-300. DOI: https://doi.org/10.3816/CBC.2010.n.038

Walcher, L., Kistenmacher, A.K., Suo, H., Kitte, R., Dluczek, S., Strauß, A., Blaudszun, A.R., Yevsa, T., Fricke, S., and Kossatz-Boehlert, U., 2020. Cancer stem cells-origins and biomarkers: Perspectives for targeted personalized therapies. Frontiers in Immunology, 11, p.1280. DOI: https://doi.org/10.3389/fimmu.2020.01280

Wilson, M.M., Weinberg, R.A., Lees, J.A., and Guen, V.J., 2020. Emerging mechanisms by which EMT programs control stemness. Trends in Cancer, 6(9), pp.775-780. DOI: https://doi.org/10.1016/j.trecan.2020.03.011

Winters, S., Martin, C., Murphy, D., and Shokar, N.K., 2017. Breast cancer epidemiology, prevention, and screening. Progress in Molecular Biology and Translational Science, 151, pp.1-32. DOI: https://doi.org/10.1016/bs.pmbts.2017.07.002

Wu, Q., Yang, Y., Wu, S., Li, W., Zhang, N., Dong, X., and Ou, Y., 2015. Evaluation of the correlation of KAI1/CD82, CD44, MMP7 and β-catenin in the prediction of prognosis and metastasis in colorectal carcinoma. Diagnostic Pathology, 10, p.176. DOI: https://doi.org/10.1186/s13000-015-0411-0

Wu, S., Yue, M., Zhang, J., Li, X., Li, Z., Zhang, H., Wang, X., Han, X., Cai, L., Shang, J., Jia, Z., Wang, X., Li, J., and Liu, Y., 2023. The role of artificial intelligence in accurate interpretation of HER2 immunohistochemical scores 0 and 1+ in breast cancer. Modern Pathology, 36(3), p.100054. DOI: https://doi.org/10.1016/j.modpat.2022.100054

Wu, X.J., Li, X.D., Zhang, H., Zhang, X., Ning, Z.H., Yin, Y.M., and Tian, Y., 2015. Clinical significance of CD44s, CD44v3 and CD44v6 in breast cancer. Journal of International Medical Research, 43(2), pp.173-179. DOI: https://doi.org/10.1177/0300060514559793

Xiong, X., Zheng, L.W., Ding, Y., Chen, Y.F., Cai, Y.W., Wang, L.P., Huang, L., Liu, C.C., Shao, Z.M., and Yu, K.D., 2025. Breast cancer: Pathogenesis and treatments. Signal Transduction and Targeted Therapy, 10(1), p.49. DOI: https://doi.org/10.1038/s41392-024-02108-4

Yang, C., Cao, M., Liu, Y., He, Y., Du, Y., Zhang, G., and Gao, F., 2019. Inducible formation of leader cells driven by CD44 switching gives rise to collective invasion and metastases in luminal breast carcinomas. Oncogene, 38(46), pp.7113-7132. DOI: https://doi.org/10.1038/s41388-019-0899-y

Zahwe, M., Bendahhou, K., Eser, S., Mukherji, D., Fouad, H., Fadhil, I., Soerjomataram, I., and Znaor, A., 2025. Current and future burden of female breast cancer in the Middle East and North Africa region using estimates from GLOBOCAN 2022. International Journal of Cancer, 156, pp.2320-2329. DOI: https://doi.org/10.1002/ijc.35325

Zhang, H., Brown, R.L., Wei, Y., Zhao, P., Liu, S., Liu, X., Deng, Y., Hu, X., Zhang, J., Gao, X.D., Kang, Y., Mercurio, A.M., Goel, H.L., and Cheng, C., 2019. CD44 splice isoform switching determines breast cancer stem cell state. Genes and Development, 33(3-4), pp.166-179 DOI: https://doi.org/10.1101/gad.319889.118

Published

2025-06-22

How to Cite

Mohammed, A. S., Othman, R. T. and Yaqo, R. T. (2025) “Artificial Intelligence-based Digital Pathology Assessment of CD44s Expression in Breast Cancer: Association with Clinicopathological Features and Survival Outcomes”, ARO-THE SCIENTIFIC JOURNAL OF KOYA UNIVERSITY, 13(1), pp. 266–274. doi: 10.14500/aro.12248.
Received 2025-05-04
Accepted 2025-06-02
Published 2025-06-22

Similar Articles

<< < 1 2 3 4 5 6 7 8 9 > >> 

You may also start an advanced similarity search for this article.