J Liver Cancer.  2022 Mar;22(1):40-50. 10.17998/jlc.2022.03.06.

The effects of immune checkpoint modulators on the clinical course of patients with resectable hepatocellular carcinoma

• An J
• Kang HJ
• Yu E
• Lee HC
• Shim JH

• KMID: 2528295
• DOI: http://doi.org/10.17998/jlc.2022.03.06

Abstract

Background
/Aim: Immune checkpoint proteins regulating T-cell mediated anti-tumor immunity have been reported to affect clinical outcomes in multiple malignancies. This study aimed to investigate the prognostic effect of histological expression of immune checkpoint proteins in patients with resected hepatocellular carcinoma (HCC).
Methods
A total of 221 patients with HCC who underwent curative resection were included. Expression of programmed-cell death ligand-1 (PD-L1) in tumor cells (tPD-L1) and tumor infiltrating mononuclear cells (TIMCs) (iPD-L1), programmed-cell death-1 in TIMCs (iPD-1), and cytotoxic T lymphocyte antigen-4 in TIMCs (iCTLA-4) were measured immunohistochemically.
Results
Histo-positivity for iCTLA-4, iPD-1, iPD-L1, and tPD-L1 was 32.1%, 42.5%, 35.3%, and 14.9%, respectively. Multivariate logistic analyses revealed that male sex and tumor >5 cm were variables related to iCTLA-4 positivity (odds ratio [OR], 0.46 and 1.94, respectively; P<0.05). Poor differentiation was related to PD-L1 expression in both tumor cells and TIMCs (OR, 2.88 and 3.46, respectively; P<0.05). Microvascular invasion was significantly associated only with iPD-L1 (OR, 2.24; P<0.05). In time-dependent outcome analyses, expression of immune checkpoint proteins in TIMCs (i.e., iCTLA-4, iPD-1, and iPD-L1) was significantly related to longer overall survival and non-cancer-related survival (all P<0.05), but not to time-to-recurrence or cancer-specific deaths. Concurrent activation of the PD-1:PD-L1 and CTLA-4 pathways predicted improved outcomes in terms of overall survival and non-cancer related survival (P=0.06 and P=0.03, respectively).
Conclusions
Immune checkpoint proteins upregulated in TIMCs in HCC tissues have individual and additive effects in prolonging the survival of patients, specifically in terms of survival not related to cancer recurrence.

Keyword

Liver neoplasms; Prognosis; CTLA-4; PD-L1; PD-1

Figure

• 

  Figure 1 Presence of immune checkpoint proteins in formalin-fixed paraffin-embedded samples. Positive staining of each molecule is shown as follows (×400): (A) cytotoxic T lymphocyte antigen-4 in tumor-infiltrating mononuclear cells (TIMCs), (B) programmed-cell death-1 in TIMCs, (C) programmed-cell death ligand-1 (PD-L1) in TIMCs, and (D) PD-L1 in tumor cells.

• 

  Figure 2 Associations between presence of immune checkpoint proteins and overall survival. Enhanced expression of immune checkpoint molecules in tumor-infiltrating mononuclear cells (A: iCTLA-4, B: iPD-1, and C: iPD-L1) was significantly associated with longer survival of hepatocellular carcinoma patients, whereas tumoral PD-L1 (D: tPD-L1) had no prognostic significance. iCTLA-4, cytotoxic T lymphocyte antigen-4 in tumor-infiltrating mononuclear cells; iPD-1, programmed-cell death-1 in tumor-infiltrating mononuclear cells; iPD-L1, programmed-cell death ligand-1 in tumor-infiltrating mononuclear cells; tPD-L1, programmed-cell death ligand-1 in tumor cells.

• 

  Figure 3 Expression of immune checkpoint proteins and time-to-recurrence. None of the immune checkpoint proteins was significantly associated with time-to-recurrence in the HCC patients (all P >0.05). (A) iCTLA4, (B) iPD-1, (C) iPD-L1, and (D) tPD-L1. HCC, hepatocellular carcinoma; iCTLA-4, cytotoxic T lymphocyte antigen-4 in tumor-infiltrating mononuclear cells; iPD-1, programmed-cell death-1 in tumor-infiltrating mononuclear cells; iPD-L1, programmed-cell death ligand-1 in tumor-infiltrating mononuclear cells; tPD-L1, programmed-cell death ligand-1 in tumor cells.

• 

  Figure 4 (A) Effect of combined expression of the PD-1/PD-L1 and CTLA-4 pathways on overall survival. The patients were divided into the following three groups based on the expression of immune checkpoint proteins: activation of both pathways (PD-1:PD-L1 & CTLA-4; group 1), activation of only one of the two pathways (group 2), and activation of neither pathway (group 3). The difference between the overall survival of the three groups was marginally significant (P=0.06). When further analyzed, a significant difference was observed for non-cancer-related survival (C), but not for cancer-specific survival (B). PD-1, programmed-cell death-1; PD-L1, programmed-cell death ligand-1; CTLA-4, cytotoxic T lymphocyte antigen-4.

Reference

References

1. Dunn GP, Old LJ, Schreiber RD. The immunobiology of cancer immunosurveillance and immunoediting. Immunity. 2004; 21:137–148.

2. Schreiber RD, Old LJ, Smyth MJ. Cancer immunoediting: integrating immunity’s roles in cancer suppression and promotion. Science. 2011; 331:1565–1570.

3. Pardoll DM. The blockade of immune checkpoints in cancer immunotherapy. Nat Rev Cancer. 2012; 12:252–264.

4. Buchbinder EI, Desai A. CTLA-4 and PD-1 pathways: similarities, differences, and implications of their inhibition. Am J Clin Oncol. 2016; 39:98–106.

5. Fife BT, Bluestone JA. Control of peripheral T-cell tolerance and autoimmunity via the CTLA-4 and PD-1 pathways. Immunol Rev. 2008; 224:166–182.

6. Krummel MF, Allison JP. CD28 and CTLA-4 have opposing effects on the response of T cells to stimulation. J Exp Med. 1995; 182:459–465.

7. Fife BT, Pauken KE, Eagar TN, Obu T, Wu J, Tang Q, et al. Interactions between PD-1 and PD-L1 promote tolerance by blocking the TCR-induced stop signal. Nat Immunol. 2009; 10:1185–1192.

8. Keir ME, Butte MJ, Freeman GJ, Sharpe AH. PD-1 and its ligands in tolerance and immunity. Annu Rev Immunol. 2008; 26:677–704.

9. Boonstra A, Woltman AM, Janssen HL. Immunology of hepatitis B and hepatitis C virus infections. Best Pract Res Clin Gastroenterol. 2008; 22:1049–1061.

10. Ye B, Liu X, Li X, Kong H, Tian L, Chen Y. T-cell exhaustion in chronic hepatitis B infection: current knowledge and clinical significance. Cell Death Dis. 2015; 6:e1694.

11. Bertoletti A, Maini MK. Protection or damage: a dual role for the virus-specific cytotoxic T lymphocyte response in hepatitis B and C infection? Curr Opin Microbiol. 2000; 3:387–392.

12. Kassel R, Cruise MW, Iezzoni JC, Taylor NA, Pruett TL, Hahn YS. Chronically inflamed livers up-regulate expression of inhibitory B7 family members. Hepatology. 2009; 50:1625–1637.

13. Wu P, Wu D, Li L, Chai Y, Huang J. PD-L1 and survival in solid tumors: a meta-analysis. PLoS One. 2015; 10:e0131403.

14. Vassilakopoulou M, Avgeris M, Velcheti V, Kotoula V, Rampias T, Chatzopoulos K, et al. Evaluation of PD-L1 expression and associated tumor-infiltrating lymphocytes in laryngeal squamous cell carcinoma. Clin Cancer Res. 2016; 22:704–713.

15. Choueiri TK, Fay AP, Gray KP, Callea M, Ho TH, Albiges L, et al. PD-L1 expression in nonclear-cell renal cell carcinoma. Ann Oncol. 2014; 25:2178–2184.

16. McLaughlin J, Han G, Schalper KA, Carvajal-Hausdorf D, Pelekanou V, Rehman J, et al. Quantitative assessment of the heterogeneity of PD-L1 expression in non-small-cell lung cancer. JAMA Oncol. 2016; 2:46–54.

17. Kim JW, Nam KH, Ahn SH, Park DJ, Kim HH, Kim SH, et al. Prognostic implications of immunosuppressive protein expression in tumors as well as immune cell infiltration within the tumor microenvironment in gastric cancer. Gastric Cancer. 2016; 19:42–52.

18. Fusi A, Festino L, Botti G, Masucci G, Melero I, Lorigan P, et al. PD-L1 expression as a potential predictive biomarker. Lancet Oncol. 2015; 16:1285–1287.

19. Bellmunt J, Mullane SA, Werner L, Fay AP, Callea M, Leow JJ, et al. Association of PD-L1 expression on tumor-infiltrating mononuclear cells and overall survival in patients with urothelial carcinoma. Ann Oncol. 2015; 26:812–817.

20. Umemoto Y, Okano S, Matsumoto Y, Nakagawara H, Matono R, Yoshiya S, et al. Prognostic impact of programmed cell death 1 ligand 1 expression in human leukocyte antigen class I-positive hepatocellular carcinoma after curative hepatectomy. J Gastroenterol. 2015; 50:65–75.

21. Calderaro J, Rousseau B, Amaddeo G, Mercey M, Charpy C, Costentin C, et al. Programmed death ligand 1 expression in hepatocellular carcinoma: relationship with clinical and pathological features. Hepatology. 2016; 64:2038–2046.

22. Shi F, Shi M, Zeng Z, Qi RZ, Liu ZW, Zhang JY, et al. PD-1 and PDL1 upregulation promotes CD8(+) T-cell apoptosis and postoperative recurrence in hepatocellular carcinoma patients. Int J Cancer. 2011; 128:887–896.

23. Onuma AE, Zhang H, Huang H, Williams TM, Noonan A, Tsung A. Immune checkpoint inhibitors in hepatocellular cancer: current understanding on mechanisms of resistance and biomarkers of response to treatment. Gene Expr. 2020; 20:53–65.

24. Willimsky G, Schmidt K, Loddenkemper C, Gellermann J, Blankenstein T. Virus-induced hepatocellular carcinomas cause antigen-specific local tolerance. J Clin Invest. 2013; 123:1032–1043.

25. Gao Q, Wang XY, Qiu SJ, Yamato I, Sho M, Nakajima Y, et al. Overexpression of PD-L1 significantly associates with tumor aggressiveness and postoperative recurrence in human hepatocellular carcinoma. Clin Cancer Res. 2009; 15:971–979.

26. Kim HR, Ha SJ, Hong MH, Heo SJ, Koh YW, Choi EC, et al. PD-L1 expression on immune cells, but not on tumor cells, is a favorable prognostic factor for head and neck cancer patients. Sci Rep. 2016; 6:36956.

27. Muenst S, Soysal SD, Gao F, Obermann EC, Oertli D, Gillanders WE. The presence of programmed death 1 (PD-1)-positive tumor-infiltrating lymphocytes is associated with poor prognosis in human breast cancer. Breast Cancer Res Treat. 2013; 139:667–676.

28. Huang PY, Guo SS, Zhang Y, Lu JB, Chen QY, Tang LQ, et al. Tumor CTLA-4 overexpression predicts poor survival in patients with nasopharyngeal carcinoma. Oncotarget. 2016; 7:13060–13068.

29. Zhang XF, Pan K, Weng DS, Chen CL, Wang QJ, Zhao JJ, et al. Cytotoxic T lymphocyte antigen-4 expression in esophageal carcinoma: implications for prognosis. Oncotarget. 2016; 7:26670–26679.

30. Hato T, Goyal L, Greten TF, Duda DG, Zhu AX. Immune checkpoint blockade in hepatocellular carcinoma: current progress and future directions. Hepatology. 2014; 60:1776–1782.

31. Makarova-Rusher OV, Medina-Echeverz J, Duffy AG, Greten TF. The yin and yang of evasion and immune activation in HCC. J Hepatol. 2015; 62:1420–1429.

32. Breous E, Thimme R. Potential of immunotherapy for hepatocellular carcinoma. J Hepatol. 2011; 54:830–834.

33. Sawada Y, Yoshikawa T, Nobuoka D, Shirakawa H, Kuronuma T, Motomura Y, et al. Phase I trial of a glypican-3-derived peptide vaccine for advanced hepatocellular carcinoma: immunologic evidence and potential for improving overall survival. Clin Cancer Res. 2012; 18:3686–3696.

34. Duffy AG, Ulahannan SV, Makorova-Rusher O, Rahma O, Wedemeyer H, Pratt D, et al. Tremelimumab in combination with ablation in patients with advanced hepatocellular carcinoma. J Hepatol. 2017; 66:545–551.

35. Shimizu K, Kotera Y, Aruga A, Takeshita N, Katagiri S, Ariizumi S, et al. Postoperative dendritic cell vaccine plus activated T-cell transfer improves the survival of patients with invasive hepatocellular carcinoma. Hum Vaccin Immunother. 2014; 10:970–976.

36. Abou-Alfa GK, Chan SL, Kudo M, Lau G, Kelley RK, Furuse J, et al. Phase 3 randomized, open-label, multicenter study of tremelimumab (T) and durvalumab (D) as first-line therapy in patients (pts) with unresectable hepatocellular carcinoma (uHCC): HIMALAYA. J Clin Oncol. 2022; 40(Suppl 4):379.

37. Finn RS, Qin S, Ikeda M, Galle PR, Ducreux M, Kim TY, et al. Atezolizumab plus bevacizumab in unresectable hepatocellular carcinoma. N Engl J Med. 2020; 382:1894–1905.

38. Droeser RA, Hirt C, Viehl CT, Frey DM, Nebiker C, Huber X, et al. Clinical impact of programmed cell death ligand 1 expression in colorectal cancer. Eur J Cancer. 2013; 49:2233–2242.

39. Wimberly H, Brown JR, Schalper K, Haack H, Silver MR, Nixon C, et al. PD-L1 expression correlates with tumor-infiltrating lymphocytes and response to neoadjuvant chemotherapy in breast cancer. Cancer Immunol Res. 2015; 3:326–332.

40. Taube JM, Klein A, Brahmer JR, Xu H, Pan X, Kim JH, et al. Association of PD-1, PD-1 ligands, and other features of the tumor immune microenvironment with response to anti-PD-1 therapy. Clin Cancer Res. 2014; 20:5064–5074.

41. Taube JM, Anders RA, Young GD, Xu H, Sharma R, McMiller TL, et al. Colocalization of inflammatory response with B7-h1 expression in human melanocytic lesions supports an adaptive resistance mechanism of immune escape. Sci Transl Med. 2012; 4:127ra137.

42. Yu H, Yang J, Jiao S, Li Y, Zhang W, Wang J. Cytotoxic T lymphocyte antigen 4 expression in human breast cancer: implications for prognosis. Cancer Immunol Immunother. 2015; 64:853–860.

43. Jiang Y, Li Y, Zhu B. T-cell exhaustion in the tumor microenvironment. Cell Death Dis. 2015; 6:e1792.

44. Duraiswamy J, Kaluza KM, Freeman GJ, Coukos G. Dual blockade of PD-1 and CTLA-4 combined with tumor vaccine effectively restores T-cell rejection function in tumors. Cancer Res. 2013; 73:3591–3603.

45. Hino R, Kabashima K, Kato Y, Yagi H, Nakamura M, Honjo T, et al. Tumor cell expression of programmed cell death-1 ligand 1 is a prognostic factor for malignant melanoma. Cancer. 2010; 116:1757–1766.

46. Baptista MZ, Sarian LO, Derchain SF, Pinto GA, Vassallo J. Prognostic significance of PD-L1 and PD-L2 in breast cancer. Hum Pathol. 2016; 47:78–84.

47. Salgado R, Denkert C, Demaria S, Sirtaine N, Klauschen F, Pruneri G, et al. The evaluation of tumor-infiltrating lymphocytes (TILs) in breast cancer: recommendations by an International TILs Working Group 2014. Ann Oncol. 2015; 26:259–271.

48. Noman MZ, Desantis G, Janji B, Hasmim M, Karray S, Dessen P, et al. PD-L1 is a novel direct target of HIF-1α, and its blockade under hypoxia enhanced MDSC-mediated T cell activation. J Exp Med. 2014; 211:781–790.

49. Prieto J, Melero I, Sangro B. Immunological landscape and immunotherapy of hepatocellular carcinoma. Nat Rev Gastroenterol Hepatol. 2015; 12:681–700.