Top

Journal of Experimental & Clinical Cancer Research

Published in:

Open Access 01-12-2021 | Pancreatic Cancer | Research

C4b-binding protein α-chain enhances antitumor immunity by facilitating the accumulation of tumor-infiltrating lymphocytes in the tumor microenvironment in pancreatic cancer

Authors: Kosuke Sasaki, Shigetsugu Takano, Satoshi Tomizawa, Yoji Miyahara, Katsunori Furukawa, Tsukasa Takayashiki, Satoshi Kuboki, Mamoru Takada, Masayuki Ohtsuka

Published in: Journal of Experimental & Clinical Cancer Research | Issue 1/2021

Abstract

Background

Recent studies indicate that complement plays pivotal roles in promoting or suppressing cancer progression. We have previously identified C4b-binding protein α-chain (C4BPA) as a serum biomarker for the early detection of pancreatic ductal adenocarcinoma (PDAC). However, its mechanism of action remains unclear. Here, we elucidated the functional roles of C4BPA in PDAC cells and the tumor microenvironment.

Methods

We assessed stromal C4BPA, the C4BPA binding partner CD40, and the number of CD8⁺ tumor-infiltrating lymphocytes in resected human PDAC tissues via immunohistochemical staining. The biological functions of C4BPA were investigated in peripheral blood mononuclear cells (PBMCs) and human PDAC cell lines. Mouse C4BPA (mC4BPA) peptide, which is composed of 30 amino acids from the C-terminus and binds to CD40, was designed for further in vitro and in vivo experiments. In a preclinical experiment, we assessed the efficacy of gemcitabine plus nab-paclitaxel (GnP), dual immune checkpoint blockades (ICBs), and mC4BPA peptide in a mouse orthotopic transplantation model.

Results

Immunohistochemical analysis revealed that high stromal C4BPA and CD40 was associated with favorable PDAC prognosis (P=0.0005). Stromal C4BPA strongly correlated with the number of CD8⁺ tumor-infiltrating lymphocytes (P=0.001). In in vitro experiments, flow cytometry revealed that recombinant human C4BPA (rhC4BPA) stimulation increased CD4⁺ and CD8⁺ T cell numbers in PBMCs. rhC4BPA also promoted the proliferation of CD40-expressing PDAC cells. By contrast, combined treatment with gemcitabine and rhC4BPA increased PDAC cell apoptosis rate. mC4BPA peptide increased the number of murine T lymphocytes in vitro and the number of CD8⁺ tumor-infiltrating lymphocytes surrounding PDAC tumors in vivo. In a preclinical study, GnP/ICBs/mC4BPA peptide treatment, but not GnP treatment, led to the accumulation of a greater number of CD8⁺ T cells in the periphery of PDAC tumors and to greater tumor regression than did control treatment.

Conclusions

These findings demonstrate that the combination of GnP therapy with C4BPA inhibits PDAC progression by promoting antitumor T cell accumulation in the tumor microenvironment.

Available only for authorised users

Siegel RL, Miller KD, Jemal A. Cancer statistics, 2020. CA Cancer J Clin. 2020;70:7–30.PubMedCrossRef

Ryan DP, Hong TS, Bardeesy N. Pancreatic adenocarcinoma. N Engl J Med. 2014;371:1039–49.PubMedCrossRef

Clark CE, Beatty GL, Vonderheide RH. Immunosurveillance of pancreatic adenocarcinoma: insights from genetically engineered mouse models of cancer. Cancer Lett. 2009;279:1–7.PubMedCrossRef

Feig C, Gopinathan A, Neesse A, Chan DS, Cook N, Tuveson DA. The pancreas cancer microenvironment. Clin Cancer Res. 2012;18:4266–76.PubMedPubMedCentralCrossRef

Sogawa K, Takano S, Iida F, Satoh M, Tsuchida S, Kawashima Y, et al. Identification of a novel serum biomarker for pancreatic cancer, C4b-binding protein a-chain (C4BPA) by quantitative proteomic analysis using tandem mass tags. Br J Cancer. 2016;115:949–56.PubMedPubMedCentralCrossRef

Kusada-Funakoshi M, Sasaki J, Takada Y, Soji T, Arakawa K. Evidence that C4b-binding protein (proline-rich protein) is synthesized by hepatocytes. Biochem Med Metab Biol. 1991;45:350–4.PubMedCrossRef

Lappin DF, Whaley K. Interferon-induced transcriptional and post-transcriptional modulation of factor H and C4 binding-protein synthesis in human monocytes. Biochem J. 1990;271:767–72.PubMedPubMedCentralCrossRef

Blom AM, Villoutreix BO, Dahlback B. Complement inhibitor C4b-binding protein - friend or foe in the innate immune system? Mol Immunol. 2004;40:1333–46.PubMedCrossRef

Banchereau J, Dubois B, Fayette J, Burdin N, Brière F, Miossec P, et al. Functional CD40 antigen on B cells, dendritic cells and fibroblasts. Adv Exp Med Biol. 1995;378:79–83.PubMedCrossRef

10.

Vonderheide RH. Prospect of targeting the CD40 pathway for cancer therapy. Clin Cancer Res. 2007;13:1083–8.PubMedCrossRef

11.

Lederman S, Yellin MJ, Krichevsky A, Belko J, Lee JJ, Chess L. Identification of a novel surface protein on activated CD4+ T cells that induces contact-dependent B cell differentiation (help). J Exp Med. 1992;175:1091–101.PubMedCrossRef

12.

Croft M, Siegel RM. Beyond TNF: TNF superfamily cytokines as targets for the treatment of rheumatic diseases. Nat Rev Rheumatol. 2017;13:217–33.PubMedPubMedCentralCrossRef

13.

Foy TM, Durie FH, Noelle RJ. The expansive role of CD40 and its ligand, gp39, in immunity. Semin Immunol. 1994;6:259–66.PubMedCrossRef

14.

Splawski JB, Lipsky PE. CD40-mediated regulation of human B-cell responses. Res Immunol. 1994;145:226–34.PubMedCrossRef

15.

Sotomayor EM, Borrello I, Tubb E, Rattis FM, Bien H, Lu Z, et al. Conversion of tumor-specific CD4+ T-cell tolerance to T-cell priming through in vivo ligation of CD40. Nat Med. 1999;5:780–7.PubMedCrossRef

16.

Brahmer JR, Tykodi SS, Chow LQ, Hwu WJ, Topalian SL, Hwu P, et al. Safety and activity of anti-PD-L1 antibody in patients with advanced cancer. N Engl J Med. 2012;366:2455–65.PubMedPubMedCentralCrossRef

17.

Sharma P, Allison JP. The future of immune checkpoint therapy. Science. 2015;348:56–61.PubMedCrossRef

18.

Diehl L, den Boer AT, Schoenberger SP, van der Voort EI, Schumacher TN, Melief CJ, et al. CD40 activation in vivo overcomes peptide-induced peripheral cytotoxic T-lymphocyte tolerance and augments anti-tumor vaccine efficacy. Nat Med. 1999;5:774–9.PubMedCrossRef

19.

French RR, Chan HT, Tutt AL, Glennie MJ. CD40 antibody evokes a cytotoxic T-cell response that eradicates lymphoma and bypasses T-cell help. Nat Med. 1999;5:548–53.PubMedCrossRef

20.

van Mierlo GJ, den Boer AT, Medema JP, van der Voort EI, Fransen MF, Offringa R, et al. CD40 stimulation leads to effective therapy of CD40(-) tumors through induction of strong systemic cytotoxic T lymphocyte immunity. Proc Natl Acad Sci U S A. 2002;99:5561–6.PubMedPubMedCentralCrossRef

21.

Cho HI, Celis E. Optimized peptide vaccines eliciting extensive CD8 T-cell responses with therapeutic antitumor effects. Cancer Res. 2009;69:9012–9.PubMedPubMedCentralCrossRef

22.

Ahonen CL, Wasiuk A, Fuse S, Turk MJ, Ernstoff MS, Suriawinataet AA, et al. Enhanced efficacy and reduced toxicity of multifactorial adjuvants compared with unitary adjuvants as cancer vaccines. Blood. 2008;111:3116–25.PubMedPubMedCentralCrossRef

23.

Nowak AK, Robinson BW, Lake RA. Synergy between chemotherapy and immunotherapy in the treatment of established murine solid tumors. Cancer Res. 2003;63:4490–6.PubMed

24.

Honeychurch J, Glennie MJ, Johnson PW, Illidge TM. Anti-CD40 monoclonal antibody therapy in combination with irradiation results in a CD8 T-cell-dependent immunity to B-cell lymphoma. Blood. 2003;102:1449–57.PubMedCrossRef

25.

Byrne KT, Vonderheide RH. CD40 stimulation obviates innate sensors and drives T cell immunity in cancer. Cell Rep. 2016;15:2719–32.PubMedPubMedCentralCrossRef

26.

Rech AJ, Dada H, Kotzin JJ, Henao-Mejia J, Minn AJ, Twyman-Saint Victor C, et al. Radiotherapy and CD40 activation separately augment immunity to checkpoint blockade in cancer. Cancer Res. 2018;78:4282–91.PubMedPubMedCentralCrossRef

27.

Carpenter EL, Mick R, Rüter J, Vonderheide RH. Activation of human B cells by the agonist CD40 antibody CP-870,893 and augmentation with simultaneous toll-like receptor 9 stimulation. J Transl Med. 2019;7:93.CrossRef

28.

Winograd R, Byrne KT, Evans RA, Odorizzi PM, Meyer AR, Bajor DL, et al. Induction of T-cell immunity overcomes complete resistance to PD-1 and CTLA-4 blockade and improves survival in pancreatic carcinoma. Cancer Immunol Res. 2015;3:399–411.PubMedPubMedCentralCrossRef

29.

Li J, Byrne KT, Yan F, Yamazoe T, Chen Z, Baslan T, et al. Tumor cell-intrinsic factors underlie heterogeneity of immune cell infiltration and response to immunotherapy. Immunity. 2018;49:178–93.PubMedPubMedCentralCrossRef

30.

Brodeur SR, Angelini F, Bacharier LB, Blom AM, Mizoguchi E, Fujiwara H, et al. C4b-binding protein (C4BP) activates B cells through the CD40 receptor. Immunity. 2003;18:837–48.PubMedCrossRef

31.

Michel NA, Zirlik A, Wolf D. CD40L and its receptors in atherothrombosis-An update. Front Cardiovasc Med. 2017;4:40.PubMedPubMedCentralCrossRef

32.

Schoenberger SP, Toes RE, van der Voort EI, Offringa R, Melief CJ. T-cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions. Nature. 1998;393:480–3.PubMedCrossRef

33.

Lefrançois L, Altman JD, Williams K, Olson S. Soluble antigen and CD40 triggering are sufficient to induce primary and memory cytotoxic T cells. J Immunol. 2000;164:725–32.PubMedCrossRef

34.

Hofmeyer T, Schmelz S, Degiacomi MT, Dal Peraro M, Daneschdar M, Scrima A, et al. Arranged sevenfold: structural insights into the C-terminal oligomerization domain of human C4b-binding protein. J Mol Biol. 2013;425:1302–17.PubMedCrossRef

35.

Draper SJ, Moore AC, Goodman AL, Long CA, Holder AA, Gilbert SC, et al. Effective induction of high-titer antibodies by viral vector vaccines. Nat Med. 2008;14:819–21.PubMedPubMedCentralCrossRef

36.

Ithier MC, Parobchak N, Yadava S, Cheng J, Wang B, Rosen T. Fetal lung C4BPA induces p100 processing in human placenta. Sci Rep. 2019;9:5519.PubMedPubMedCentralCrossRef

37.

Karamitopoulou E. Tumour microenvironment of pancreatic cancer: immune landscape is dictated by molecular and histopathological features. Br J Cancer. 2019;121:5–14.PubMedPubMedCentralCrossRef

38.

Shimazaki R, Takano S, Satoh M, Takada M, Miyahara Y, Sasaki K, et al. Complement factor B regulates cellular senescence and is associated with poor prognosis in pancreatic cancer. Cell Oncol (Dordr). 2021. https://doi.org/10.1007/s13402-021-00614-z.

39.

Wang Y, Sun SN, Liu Q, Yu YY, Guo J, Wang K, et al. Autocrine complement inhibits IL10-dependent T-cell-mediated antitumor immunity to promote tumor progression. Cancer Discov. 2016;6:1022–35.PubMedPubMedCentralCrossRef

40.

Ermert D, Blom AM. C4b-binding protein: The good, the bad and the deadly. Novel functions of an old friend. Immunol Lett. 2016;169:82–92.PubMedCrossRef

41.

Caux C, Massacrier C, Vanbervliet B, Dubois B, Van Kooten C, Durand I, et al. Activation of human dendritic cells through CD40 cross-linking. J Exp Med. 1994;180:1263–72.PubMedCrossRef

42.

Ma DY, Clark EA. The role of CD40 and CD154/CD40L in dendritic cells. Semin Immunol. 2009;21:265–72.PubMedPubMedCentralCrossRef

43.

Vonderheide RH, Glennie MJ. Agonistic CD40 antibodies and cancer therapy. Clin Cancer Res. 2013;19:1035–43.PubMedPubMedCentralCrossRef

44.

Beatty GL, Chiorean EG, Fishman MP, Saboury B, Teitelbaum UR, Sun W, et al. CD40 agonists alter tumor stroma and show efficacy against pancreatic carcinoma in mice and humans. Science. 2011;331:1612–6.PubMedPubMedCentralCrossRef

45.

Beatty GL, Torigian DA, Chiorean EG, Saboury B, Brothers A, Alavi A, et al. A phase I study of an agonist CD40 monoclonal antibody (CP-870,893) in combination with gemcitabine in patients with advanced pancreatic ductal adenocarcinoma. Clin Cancer Res. 2013;19:6286–95.PubMedCrossRef

46.

Morrison AH, Diamond MS, Hay CA, Byrne KT, Vonderheide RH. Sufficiency of CD40 activation and immune checkpoint blockade for T cell priming and tumor immunity. Proc Natl Acad Sci U S A. 2020;117:8022–31.PubMedPubMedCentralCrossRef

Title: C4b-binding protein α-chain enhances antitumor immunity by facilitating the accumulation of tumor-infiltrating lymphocytes in the tumor microenvironment in pancreatic cancer
Authors: Kosuke Sasaki
Shigetsugu Takano
Satoshi Tomizawa
Yoji Miyahara
Katsunori Furukawa
Tsukasa Takayashiki
Satoshi Kuboki
Mamoru Takada
Masayuki Ohtsuka
Publication date: 01-12-2021
Publisher: BioMed Central
Keywords: Pancreatic Cancer
Pancreatic Cancer
Published in: Journal of Experimental & Clinical Cancer Research / Issue 1/2021
Electronic ISSN: 1756-9966
DOI: https://doi.org/10.1186/s13046-021-02019-0

Webinar | 19-02-2024 | 17:30 (CET)

Keynote webinar | Spotlight on antibody–drug conjugates in cancer

Watch now

Antibody–drug conjugates (ADCs) are novel agents that have shown promise across multiple tumor types. Explore the current landscape of ADCs in breast and lung cancer with our experts, and gain insights into the mechanism of action, key clinical trials data, existing challenges, and future directions.

Dr. Véronique Diéras

Prof. Fabrice Barlesi

Developed by: Springer Medicine

Keynote webinar | Spotlight on medication adherence

Springer Medicine

Abstract

Background

Methods

Results

Conclusions

Please log in to get access to this content

Other articles of this Issue 1/2021

RNA m6A demethylase FTO-mediated epigenetic up-regulation of LINC00022 promotes tumorigenesis in esophageal squamous cell carcinoma

The role of the tumor microbe microenvironment in the tumor immune microenvironment: bystander, activator, or inhibitor?

Correction to: Knockdown of FOXO3a induces epithelial-mesenchymal transition and promotes metastasis of pancreatic ductal adenocarcinoma by activation of the β-catenin/TCF4 pathway through SPRY2

Group III phospholipase A2 downregulation attenuated survival and metastasis in ovarian cancer and promotes chemo-sensitization

RNA-binding protein IMP3 is a novel regulator of MEK1/ERK signaling pathway in the progression of colorectal Cancer through the stabilization of MEKK1 mRNA

Efficacy and mechanism of the combination of PARP and CDK4/6 inhibitors in the treatment of triple-negative breast cancer

Keynote webinar | Spotlight on antibody–drug conjugates in cancer