Open Access

CD103+ T Cells May Be a Useful Biomarker in Borrmann Type 4 Gastric Cancer

MORI TAKUYA
TANAKA HIROAKI
DEGUCHI SOTA
MIKI YUICHIRO
YOSHII MAMI
TAMURA TATSURO
TOYOKAWA TAKAHIRO
LEE SHIGERU
MUGURUMA KAZUYA
  &  
OHIRA MASAICHI
Cancer Diagnosis & Prognosis May-June; 2(3): 384-390 DOI: 10.21873/cdp.10121
Received 07 December 2021 | Revised 21 March 2023 | Accepted 18 February 2022
Corresponding author
Hiroaki Tanaka, Department of Gastroenterological Surgery, Osaka City University Graduate School of Medicine, Asahimachi 1-4-3, Abenoku, Osaka, Japan. Tel: +81 666453838 hiroakitan@med.osaka-cu.ac.jp

Abstract

Background/Aim: Recently, several studies have reported that CD103+ T cells are associated with antitumor immunity in gastric cancer (GC). However, the significance of CD103+ T cells in Borrmann type 4 GC remains unclear. The aim of this study is to assess the association of CD103+ T cells with type 4 GC. Materials and Methods: Tissue samples obtained from surgically resected specimens of patients with type 4 GC were collected, and immunohistochemical staining was performed to detect the presence of CD103+ T cells. Results: A total of 46 patients were analyzed. In some patients, high CD103 expression was observed, and patients with high CD103 expression tended to have a better prognosis than those with low CD103 expression. In particular, for patients who receive doublet chemotherapy after surgery, high CD103 expression was associated with a good prognosis. Conclusion: CD103+ T cells may be a prognostic marker in type 4 GC.
Keywords: gastric cancer, type 4, diffuse type, CD103

Gastric cancer (GC) is classified into six types using Borrmann’s classification or two types using Laurén’s classification: the diffuse and intestinal type (1). In particular, Borrmann type 4 GC, including diffuse type of GC, is a type of advanced GC with extremely high malignancy and accounts for nearly 10% of all GC cases in Asia (2). Type 4 GC exhibits unique characteristics such as extensive fibrosis with sparse tumor cell infiltration in the stroma and is characterized by lesions that diffusely infiltrate the gastric wall, accompanied by a higher rate of lymph node metastasis and peritoneal metastasis (3). Type 4 GC is frequently resistant to treatment such as chemotherapy, and the prognosis is worse than with other types of GC (4,5).

The tumor immune microenvironment has attracted increasing attention because of the development of immunotherapy (6). Cytotoxic T lymphocytes, particularly CD8+ T cells, play an important role in immunosurveillance against cancer (7). Although infiltration of CD8+ T cells is associated with favorable prognosis in GC (8,9), Pernot et al. reported that a higher number of CD8+ T cells in diffuse-type GC is not correlated with prognosis (10). Thus, the significance of the tumor immune microenvironment in type 4 GC remains unclear.

Recently, tissue-resident memory T cells have attracted scientific attention. These cells are defined by expression of CD103, whose ligand is the epithelial cell marker E-cadherin (11). CD103 plays an important role in immunity against several cancers and expresses PD-1, CTLA-4 inhibitory receptors (12,13). Previous studies have shown that high infiltration of CD103+ T cells is associated with a better prognosis (14-16), suggesting that CD103+ T cells are a potential predictive biomarker. We also previously reported that CD103+ T cells are associated with a good prognosis in GC and that CD103 is expressed at higher levels in patients with the differentiated type than in those with the undifferentiated type (17). On the other hand, high expression of CD103 was found even in some patients with type 4 GC. The aim of this study was to assess the clinical significance of CD103+ T cells in type 4 GC.

Materials and Methods

Patients. A total of 1,147 patients were diagnosed with GC and underwent initial gastrectomy at Osaka City University Hospital between 2007 and 2017. Among them, 46 patients (4.0%) with type 4 GC were included in our analysis. The inclusion criteria for patients were as follows: 1) type 4 lesions were diagnosed by preoperative detection and postoperative histopathology; 2) the depth of the tumor was pathological T3 or higher; 3) patients did not receive neoadjuvant chemotherapy. Pathological TNM staging was diagnosed histologically based on the 7th Edition of the Union for International Cancer Control TNM classification, and the histological type was determined based on the 14th Edition of the Japanese Classification of Gastric Cancer.

Immunohistochemistry. Formalin-fixed, paraffin-embedded samples were sliced into 4-μm sections. Nonspecific binding was blocked using nonspecific staining blocking reagent (Dako, Denmark). The sections were reacted with rabbit monoclonal anti-CD103 antibody [clone: EPR4166 (2); 1/1000; Abcam, Cambridge, UK], or mouse anti-CD8 antibody (clone: C8/144; 1/250; Dako), or rabbit anti-CD4 antibody (clone: EPR6855; 1/250; Abcam). Sections were incubated with secondary antibody for 10 min at room temperature, and then were visualized using 3-3’-diamino-benzidine for 5 min and counterstained with hematoxylin.

Evaluation of immunohistochemical staining. Tumor sections stained with each antibody were scanned at ×200 magnification, and the five high-power fields were randomly captured for each tumor region. The average of five fields was calculated. Positive T cells in each section were counted manually by two observers who were blinded to the clinical outcome. All microscopic images were imported from the digital photo filing system DP-73 (Olympus, Tokyo, Japan). We divided the patients into two groups according to the median range.

Statistical analysis. Statistical analysis was performed with JMP 14 (SAS Institute, Cary, NC, USA). The Mann–Whitney test was used to assess the associations between the expression of CD103 and clinicopathological features. Overall survival (OS) and progression-free survival (PFS) curves were calculated with the Kaplan-Meier method, and significant differences in survival were determined using the log-rank test. PFS is defined as the time between the day of surgery and disease progression. For all other experiments, data were compared using the two-tailed Student’s t-test. p-Values of <0.05 were considered statistically significant.

Results

Patient characteristics. In total, 46 patients with type 4 GC (19 females and 27 males; median age 70 years) were analyzed. According to histological type, undifferentiated types were observed in 43 patients (93.5%) [10 of poorly differentiated type (solid type), 28 of poorly differentiated type (non-solid type), 3 of signet-ring cell type, and 2 of mucinous type]. Four patients were stage II, 18 patients were stage III, and 24 patients were stage IV [2 had liver metastasis, 12 had peritoneal metastasis (P1), 12 had only intraoperative peritoneal lavage cytology positivity (P0CY1)]. Forty patients received adjuvant chemotherapy (26 of monotherapy, 14 of doublet chemotherapy).

The presence of CD103+ cells in type 4 GC. Although CD103 staining was low in most patients, some patients with high infiltration of CD103+ T cells were found (Figure 1). The median range of CD103+ T cells per area was 12.7 (5.75-20.1). No significant difference was found in clinicopathological features between high and low CD103 expression (Table I).

Survival analysis. Kaplan-Meier analysis showed that high CD103 expression tended to predict better prognosis (Figure 2A; p=0.052). On the other hand, no significant difference was found for CD8 or CD4 (Figure 2C-F). Considering each stage, high CD103 expression predicted a better prognosis than low CD103 expression for patients with stage IV (Figure 3B; p=0.003). For patients with P1 or P0CY1, high CD103 expression tended to predict a better prognosis than low CD103 expression (Figure 3C and D; p=0.054 and p=0.077, respectively). When considering patients who received adjuvant chemotherapy, no significant difference in prognosis was found for patients who received monotherapy (Figure 4A and 4B). On the other hand, both OS and PFS in high CD103 expression were longer than in low CD103 expression for patients who received doublet chemotherapy (Figure 4C and 4D; p<0.001 and p=0.063, respectively).

Discussion

In this study, we showed that patients with type 4 GC who had high CD103 expression tended to have a better prognosis than those who were low CD103 expression. In particular, for patients with stage IV or patients who receive doublet adjuvant chemotherapy, high CD103 expression was associated with a good prognosis.

CD103 binds the cell surface marker E-cadherin and labels tissue-resident memory T cells. We previously reported that CD103 expression is higher in differentiated type GC than in undifferentiated type GC (17). On the other hand, in this study, high infiltration of CD103+ T cells was observed in some patients with type 4 GC, which has reduced E-cadherin (18). Thus, the expression of CD103 may be influenced by factors other than E-cadherin.

CD103 expression is associated with a good prognosis in several cancers including GC (15-17). In particular, CD8+ CD103+ T cells exhibit higher levels of cytotoxic molecules such as interferon-γ and Granzyme-B than CD8+CD103 T cells (15), indicating that CD103 is an important marker in antitumor immunity. In general, tumor-infiltrating lymphocytes, especially cytotoxic CD8+ T cells, are correlated with an improved outcome in several cancers (9,19,20), and predict the response to chemotherapy in breast cancer (21,22). However, our results showed that CD8 expression was not correlated with prognosis in type 4 GC, as in previous reports (10,23). On the other hand, high CD103 expression was associated with an improved prognosis even in stage IV patients. Furthermore, CD103 expression had an impact on prognosis in patients who received doublet chemotherapy. Several studies revealed the influence of the immune microenvironment on the efficacy of chemotherapy (24,25). Involvement of immunogenic cell death is considered. Although the association between CD103+ T cells and the efficacy of chemotherapy remain unclear, we hypothesize that chemotherapy induces immunogenic cell death, and the immunogenic cell death may be associated with the activation of CD103+ T cells. Accordingly, we consider that more attention should be paid to CD103+ T cells than CD8+ T cells in type 4 GC.

Our study has several limitations. First, this was a retrospective study with a small sample size. Second, these results were evaluated only with immunohistochemistry, and thus, intratumoral heterogeneity could have affected the results. Third, patients in this study were treated between 2007 and 2017, and changes in chemotherapy over the last 10 years could have affected the prognosis.

In conclusion, we herein confirmed that high CD103 expression is associated with a good prognosis even in type 4 GC although CD103 expression was low in most patients. Our results indicate that CD103+ T cells may be useful as a prognostic factor in type 4 GC.

Conflicts of Interest

The Authors declare no conflicts of interest.

Authors’ Contributions

TM acquired, analyzed and interpreted the data, and wrote the manuscript. HT made substantial contributions to the conception and design of the study, interrupted the data and revised the manuscript. SD, YM, MY, TTa, TTo, SL, and KM acquired and analyzed the data. MO contributed to the conception and design of the study, interrupted the data and revised the manuscript.

Acknowledgements

All experimental procedures were approved by the Osaka City University ethics committee (approval No. 4092), and all patients provided informed consent for collection and analysis of their specimens.

References

1 Jarvi O & Lauren P On the role of heterotopias of the intestinal epithelium in the pathogenesis of gastric cancer. Acta Pathol Microbiol Scand. 29(1) 26 - 44 1951. PMID: 14902458.
2 Huang JY Wang ZN Lu CY Miao ZF Zhu Z Song YX Xu HM & Xu YY Borrmann type IV gastric cancer should be classified as pT4b disease. J Surg Res. 203(2) 258 - 267 2016. PMID: 27363630. DOI: 10.1016/j.jss.2016.04.026
3 Luo Y Gao P Song Y Sun J Huang X Zhao J Ma B Li Y & Wang Z Clinicopathologic characteristics and prognosis of Borrmann type IV gastric cancer: a meta-analysis. World J Surg Oncol. 14(1) 49 2016. PMID: 26912240. DOI: 10.1186/s12957-016-0805-9
4 Nashimoto A Akazawa K Isobe Y Miyashiro I Katai H Kodera Y Tsujitani S Seto Y Furukawa H Oda I Ono H Tanabe S & Kaminishi M Gastric cancer treated in 2002 in Japan: 2009 annual report of the JGCA nationwide registry. Gastric Cancer. 16(1) 1 - 27 2013. PMID: 22729699. DOI: 10.1007/s10120-012-0163-4
5 Dong RZ Guo JM Zhang ZW Zhou YM & Su Y Prognostic impact and implications of extracapsular lymph node spread in Borrmann type IV gastric cancer. Oncotarget. 8(57) 97593 - 97601 2017. PMID: 29228635. DOI: 10.18632/oncotarget.18400
6 Topalian SL Taube JM Anders RA & Pardoll DM Mechanism-driven biomarkers to guide immune checkpoint blockade in cancer therapy. Nat Rev Cancer. 16(5) 275 - 287 2016. PMID: 27079802. DOI: 10.1038/nrc.2016.36
7 Speiser DE Ho PC & Verdeil G Regulatory circuits of T cell function in cancer. Nat Rev Immunol. 16(10) 599 - 611 2016. PMID: 27526640. DOI: 10.1038/nri.2016.80
8 Kang BW Kim JG Lee IH Bae HI & Seo AN Clinical significance of tumor-infiltrating lymphocytes for gastric cancer in the era of immunology. World J Gastrointest Oncol. 9(7) 293 - 299 2017. PMID: 28808502. DOI: 10.4251/wjgo.v9.i7.293
9 Lee HE Chae SW Lee YJ Kim MA Lee HS Lee BL & Kim WH Prognostic implications of type and density of tumour-infiltrating lymphocytes in gastric cancer. Br J Cancer. 99(10) 1704 - 1711 2008. PMID: 18941457. DOI: 10.1038/sj.bjc.6604738
10 Pernot S Terme M Radosevic-Robin N Castan F Badoual C Marcheteau E Penault-Llorca F Bouche O Bennouna J Francois E Ghiringhelli F De La Fouchardiere C Samalin E Baptiste Bachet J Borg C Boige V Voron T Stanbury T Tartour E Gourgou S Malka D & Taieb J Infiltrating and peripheral immune cell analysis in advanced gastric cancer according to the Lauren classification and its prognostic significance. Gastric Cancer. 23(1) 73 - 81 2020. PMID: 31267360. DOI: 10.1007/s10120-019-00983-3
11 Mueller SN & Mackay LK Tissue-resident memory T cells: local specialists in immune defence. Nat Rev Immunol. 16(2) 79 - 89 2016. PMID: 26688350. DOI: 10.1038/nri.2015.3
12 Mackay LK Rahimpour A Ma JZ Collins N Stock AT Hafon ML Vega-Ramos J Lauzurica P Mueller SN Stefanovic T Tscharke DC Heath WR Inouye M Carbone FR & Gebhardt T The developmental pathway for CD103(+)CD8+ tissue-resident memory T cells of skin. Nat Immunol. 14(12) 1294 - 1301 2013. PMID: 24162776. DOI: 10.1038/ni.2744
13 Webb JR Milne K & Nelson BH PD-1 and CD103 are widely coexpressed on prognostically favorable intraepithelial CD8 T cells in human ovarian cancer. Cancer Immunol Res. 3(8) 926 - 935 2015. PMID: 25957117. DOI: 10.1158/2326-6066.CIR-14-0239
14 Djenidi F Adam J Goubar A Durgeau A Meurice G de Montpréville V Validire P Besse B & Mami-Chouaib F CD8+CD103+ tumor-infiltrating lymphocytes are tumor-specific tissue-resident memory T cells and a prognostic factor for survival in lung cancer patients. J Immunol. 194(7) 3475 - 3486 2015. PMID: 25725111. DOI: 10.4049/jimmunol.1402711
15 Ganesan AP Clarke J Wood O Garrido-Martin EM Chee SJ Mellows T Samaniego-Castruita D Singh D Seumois G Alzetani A Woo E Friedmann PS King EV Thomas GJ Sanchez-Elsner T Vijayanand P & Ottensmeier CH Tissue-resident memory features are linked to the magnitude of cytotoxic T cell responses in human lung cancer. Nat Immunol. 18(8) 940 - 950 2017. PMID: 28628092. DOI: 10.1038/ni.3775
16 Webb JR Milne K Watson P Deleeuw RJ & Nelson BH Tumor-infiltrating lymphocytes expressing the tissue resident memory marker CD103 are associated with increased survival in high-grade serous ovarian cancer. Clin Cancer Res. 20(2) 434 - 444 2014. PMID: 24190978. DOI: 10.1158/1078-0432.CCR-13-1877
17 Mori T Tanaka H Suzuki S Deguchi S Yamakoshi Y Yoshii M Miki Y Tamura T Toyokawa T Lee S Muguruma K Wanibuchi H & Ohira M Tertiary lymphoid structures show infiltration of effective tumor-resident T cells in gastric cancer. Cancer Sci. 112(5) 1746 - 1757 2021. PMID: 33735485. DOI: 10.1111/cas.14888
18 Shiozaki H Tahara H Oka H Miyata M Kobayashi K Tamura S Iihara K Doki Y Hirano S & Takeichi M Expression of immunoreactive E-cadherin adhesion molecules in human cancers. Am J Pathol. 139(1) 17 - 23 1991. PMID: 1713020.
19 Pagès F Berger A Camus M Sanchez-Cabo F Costes A Molidor R Mlecnik B Kirilovsky A Nilsson M Damotte D Meatchi T Bruneval P Cugnenc PH Trajanoski Z Fridman WH & Galon J Effector memory T cells, early metastasis, and survival in colorectal cancer. N Engl J Med. 353(25) 2654 - 2666 2005. PMID: 16371631. DOI: 10.1056/NEJMoa051424
20 Piersma SJ Jordanova ES van Poelgeest MI Kwappenberg KM van der Hulst JM Drijfhout JW Melief CJ Kenter GG Fleuren GJ Offringa R & van der Burg SH High number of intraepithelial CD8+ tumor-infiltrating lymphocytes is associated with the absence of lymph node metastases in patients with large early-stage cervical cancer. Cancer Res. 67(1) 354 - 361 2007. PMID: 17210718. DOI: 10.1158/0008-5472.CAN-06-3388
21 Denkert C von Minckwitz G Darb-Esfahani S Lederer B Heppner BI Weber KE Budczies J Huober J Klauschen F Furlanetto J Schmitt WD Blohmer JU Karn T Pfitzner BM Kümmel S Engels K Schneeweiss A Hartmann A Noske A Fasching PA Jackisch C van Mackelenbergh M Sinn P Schem C Hanusch C Untch M & Loibl S Tumour-infiltrating lymphocytes and prognosis in different subtypes of breast cancer: a pooled analysis of 3771 patients treated with neoadjuvant therapy. Lancet Oncol. 19(1) 40 - 50 2018. PMID: 29233559. DOI: 10.1016/S1470-2045(17)30904-X
22 Park YH Lal S Lee JE Choi YL Wen J Ram S Ding Y Lee SH Powell E Lee SK Yu JH Ching KA Nam JY Kim SW Nam SJ Kim JY Cho SY Park S Kim J Hwang S Kim YJ Bonato V Fernandez D Deng S Wang S Shin H Kang ES Park WY Rejto PA Bienkowska J & Kan Z Chemotherapy induces dynamic immune responses in breast cancers that impact treatment outcome. Nat Commun. 11(1) 6175 2020. PMID: 33268821. DOI: 10.1038/s41467-020-19933-0
23 Li R Zhang H Cao Y Liu X Chen Y Qi Y Wang J Yu K Lin C Liu H He H Li H Chen L Shen Z Qin J Zhang W Sun Y & Xu J Lauren classification identifies distinct prognostic value and functional status of intratumoral CD8+ T cells in gastric cancer. Cancer Immunol Immunother. 69(7) 1327 - 1336 2020. PMID: 32200421. DOI: 10.1007/s00262-020-02550-7
24 Wang J Lin C Li H Li R Wu Y Liu H Zhang H He H Zhang W & Xu J Tumor-infiltrating γδT cells predict prognosis and adjuvant chemotherapeutic benefit in patients with gastric cancer. Oncoimmunology. 6(11) e1353858 2017. PMID: 29147601. DOI: 10.1080/2162402X.2017.1353858
25 Zitvogel L Kepp O & Kroemer G Immune parameters affecting the efficacy of chemotherapeutic regimens. Nat Rev Clin Oncol. 8(3) 151 - 160 2011. PMID: 21364688. DOI: 10.1038/nrclinonc.2010.223