Prognostic Value of Homotypic Cell Internalization by Nonprofessional Phagocytic Cancer Cells

Abstract

Background: In this study, we investigated the prognostic role of homotypic tumor cell cannibalism in different cancer types.

Methods: The phenomenon of one cell being internalized into another, which we refer to as "cell-in-cell event," was assessed in 416 cases from five head and neck cancer cohorts, as well as one anal and one rectal cancer cohort. The samples were processed into tissue microarrays and immunohistochemically stained for E-cadherin and cleaved caspase-3 to visualize cell membranes and apoptotic cell death.

Results: Cell-in-cell events were found in all of the cohorts. The frequency ranged from 0.7 to 17.3 cell-in-cell events per mm(2). Hardly any apoptotic cells were found within the cell-in-cell structures, although apoptotic cell rates were about 1.6 to two times as high as cell-in-cell rates of the same tissue sample. High numbers of cell-in-cell events showed adverse effects on patients' survival in the head and neck and in the rectal cancer cohorts. In multivariate analysis, high frequency was an adverse prognostic factor for overall survival in patients with head and neck cancer (p = 0.008).

Conclusion: Cell-in-cell events were found to predict patient outcomes in various types of cancer better than apoptosis and proliferation and might therefore be used to guide treatment strategies.

Figure 1

Cell-in-cell structures are found in different tumor tissues. (a) Representative image of an E-cadherin-labeled head and neck squamous cell carcinoma tissue microarray spot with numerous CIC structures and magnifications of indicated regions. (b) Representative images of E-cadherin-labeled head and neck cancer CIC structures. (c) Frequency of CIC structures in different cancer types. (d) Comparative frequency of CIC (A) in the tumor tissue of rectal and anal cancer patients, (B) in low-risk and high-risk HNSCC patients, (C) in the primary tumor of HNSCC tumors and the affected regional lymph nodes, (D) before and after neoadjuvant radiochemotherapy, and (E) in the central tumor area and invasive front of HNSCC. R(C)T: radiochemotherapy or radiotherapy.

Figure 2

Apoptotic and proliferating cell rates. (a) CIC events, apoptotic cells, and proliferating cells in the central tumor area and invasive front of HNSCC. Frequency of (b) apoptotic and (c) proliferating cells in the central tumor area compared to the invasive front of individual HNSCC patients.

Figure 3

Cell-in-cell events compared to apoptotic events and prognostic impact of cell-in-cell events. (a) E-cadherin and cleaved caspase-3-labeled tumor sections containing CIC structures. (b) Frequency of CIC structures compared with the frequency of apoptotic cells in the central tumor area and invasive front of HNSCC. (c) CIC frequency compared to apoptotic cells in pretherapeutic biopsies and in posttherapeutic tumor resections of HNSCC. The shaded area marks tissues containing more CIC/mm² than apoptotic cells/mm². (d) Immunofluorescence staining for E-cadherin and cleaved caspase-3 in the left panel and Ki-67 and CD68 in the right panel. Nuclei were stained with DAPI.

Figure 4

Kaplan-Meier analyses. (a) Kaplan-Meier curves depicting anal and (b) rectal cancer patients with fewer than 10 CIC structures per 10 mm² (blue solid lines) and with equal or more than 10 CIC structures per 10 mm² (green dotted lines). HNSCC patients with CIC structures (green dotted lines) and without CIC structures (blue solid lines) (c) in the central tumor area and (d) from all five HNSCC patient cohorts; from those patients the clinical data, patients' characteristics, and immunohistological data were completely available.