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Dual targeting of PD-L1 and PD-L2 by PCED1B-AS1 via sponging hsa-miR-194-5p induces immunosuppression in hepatocellular carcinoma

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PD-L1 and PD-L2 are PD-1 ligands (PD-Ls). PD-Ls over-expression is associated with poor prognosis in hepatocellular carcinoma (HCC). However, little is known about how PD-Ls expression is regulated. Here, we investigated the involvement of lncRNA-microRNA network in the regulation of PD-Ls in HCC.


The expression of PD-Ls, PCED1B-AS1 and hsa-miR-194-5p was measured in 45 pairs of HCC samples. The interaction between PCED1B-AS1 and hsa-miR-194-5p was measured by microRNA pull down and in vitro binding assay. The effects of PCED1B-AS1 knockdown and over-expression on hsa-miR-194-5p and PD-Ls expression were investigated in HCC cell lines. Immunosuppression was evaluated in co-culture of HCC cell line and human T cells. Exosomes were isolated from HCC cells and their effects on receipt cells were investigated. Tumor behaviors were evaluated by in vitro and in vivo assays.


PD-L1 expression was highly correlated with PD-L2 expression in HCC. PCED1B-AS1 and hsa-miR-194-5p expression was up-regulated in HCC. PCED1B-AS1 was positively correlated with PD-Ls but negatively correlated hsa-miR-194-5p in HCC. These correlations were cross-validated by TCGA-LIHC dataset. PCED1B-AS1 interacted with hsa-mir-194-5p which inhibited PD-Ls expression. PCED1B-AS1 enhanced the expression of PD-Ls via sponging hsa-mir-194-5p. PCED1B-AS1-induced PD-Ls-mediated immunosuppression in co-cultured T cells. HCC cells released PCED1B-AS1 containing exosomes and the exosomal PCED1B-AS1 enhanced PD-Ls expression in receipt HCC cells while inhibited receipt T cells and macrophages. Blood exosomal PCED1B-AS1 was correlated with HCC PD-Ls expression. Finally, PCED1B-AS1 promoted cell proliferation, colony formation and in vivo tumor formation in xenografted nude mice while inhibited apoptosis.


PCED1B-AS1 enhances the expression and function of PD-Ls via sponging hsa-miR-194-5p to induce immunosuppression in HCC.

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  1. Villanueva A. Hepatocellular carcinoma. N Engl J Med 2019;380(15):1450–1462

    Article  CAS  Google Scholar 

  2. Alves RC, Alves D, Guz B, Matos C, Viana M, Harriz M, et al. Advanced hepatocellular carcinoma. Review of targeted molecular drugs. Ann Hepatol 2011;10(1):21–27

    Article  Google Scholar 

  3. Ribas A, Wolchok JD. Cancer immunotherapy using checkpoint blockade. Science 2018;359(6382):1350

    Article  CAS  Google Scholar 

  4. Freeman GJ, Long AJ, Iwai Y, Bourque K, Chernova T, Nishimura H, et al. Engagement of the PD-1 immunoinhibitory receptor by a novel B7 family member leads to negative regulation of lymphocyte activation. J Exp Med 2000;192(7):1027–1034

    Article  CAS  Google Scholar 

  5. Larsen TV, Hussmann D, Niesen AL. PD-L1 and PD-L2 expression correlated genes in non-small-cell lung cancer. Cancer Commun 2019;3:39

    Google Scholar 

  6. Yearley JH, Gibson C, Yu N, Moon C, Murphy E, Juco J, et al. PD-L2 expression in human tumors: relevance to anti-PD-1 therapy in cancer. Clin Cancer Res 2017;23(12):3158–3167

    Article  CAS  Google Scholar 

  7. 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(3):971–979

    Article  CAS  Google Scholar 

  8. Jung HI, Jeong D, Ji S, Ahn TS, Bae SH, Chin S, et al. Overexpression of PD-L1 and PD-L2 Is associated with poor prognosis in patients with hepatocellular carcinoma. Cancer Res Treat 2017;49(1):246–254

    Article  CAS  Google Scholar 

  9. Yang HY, Zhou XX, Sun LJ, Mao YL. Correlation between PD-L2 expression and clinical outcome in solid cancer patients: a meta-analysis. Front Oncol 2019;13:9

    Google Scholar 

  10. Quinn JJ, Chang HY. Unique features of long non-coding RNA biogenesis and function. Nat Rev Genet 2016;17(1):47–62

    Article  CAS  Google Scholar 

  11. Cesana M, Cacchiarelli D, Legnini I, Santini T, Sthandier O, Chinappi M, et al. A long noncoding RNA controls muscle differentiation by functioning as a competing endogenous RNA. Cell 2011;147(2):358–369

    Article  CAS  Google Scholar 

  12. Zhou X, Gao Q, Wang J, Zhang X, Liu K, Duan Z. Linc-RNA-RoR acts as a “sponge” against mediation of the differentiation of endometrial cancer stem cells by microRNA-145. Gynecol Oncol 2014;133(2):333–339

    Article  CAS  Google Scholar 

  13. Wu XS, Wang F, Li HF, Hu YP, Jiang L, Zhang F, et al. LncRNA-PAGBC acts as a microRNA sponge and promotes gallbladder tumorigenesis. EMBO Rep 2017;18(10):1837–1853

    Article  CAS  Google Scholar 

  14. Xu S, Wang Q, Kang Y, Liu J, Yin Y, Liu L, et al. Long noncoding RNAs control the modulation of immune checkpoint molecules in cancer. Cancer Immunol Res 2020;8:937–951

    Article  CAS  Google Scholar 

  15. Ding J, Li X, Hu H. TarPmiR: a new approach for microRNA target site prediction. Bioinformatics 2016;32(18):2768–2775

    Article  CAS  Google Scholar 

  16. Kruger J, Rehmsmeier M. RNAhybrid: microRNA target prediction easy, fast and flexible. Nucleic Acids Res 2006;34(Web server issue):W451–W454

    Article  Google Scholar 

  17. Paliard X, de Waal MR, Yssel H, Blanchard D, Chretien I, Abrams J, et al. Simultaneous production of IL-2, IL-4, and IFN-gamma by activated human CD4+ and CD8+ T cell clones. J Immunol 1988;141(3):849–855

    CAS  PubMed  Google Scholar 

  18. Boyman O, Sprent J. The role of interleukin-2 during homeostasis and activation of the immune system. Nat Rev Immunol 2012;12(3):180–190

    Article  CAS  Google Scholar 

  19. Yang JH, Yu D, Liu XSBJ, Changyong E, Yu S. LncRNA PCED1B-AS1 activates the proliferation and restricts the apoptosis of glioma through cooperating with miR-194-5p/PCED1B axis. J Cell Biochem 2020;121:1823–1833

  20. Hou JW, Zhao RC, Xia WY, Chang CW, You Y, Hsu JM, et al. PD-L1-mediated gasdermin C expression switches apoptosis to pyroptosis in cancer cells and facilitates tumour necrosis. Nat Cell Biol 2020;22:1264–1275

  21. Azuma T, Yao S, Zhu G, Flies AS, Flies SJ, Chen L. B7-H1 is a ubiquitous antiapoptotic receptor on cancer cells. Blood 2008;111(7):3635–3643

    Article  CAS  Google Scholar 

  22. Clark CA, Gupta HB, Sareddy G, Pandeswara S, Lao S, Yuan B, et al. Tumor-intrinsic PD-L1 signals regulate cell growth, pathogenesis, and autophagy in ovarian cancer and melanoma. Cancer Res 2016;76(23):6964–6974

    Article  CAS  Google Scholar 

  23. Yao Z, Zhang Q, Guo F, Guo S, Yang B, Liu B, et al. Long noncoding RNA PCED1B-AS1 promotes the Warburg effect and tumorigenesis by upregulating HIF-1alpha in glioblastoma. Cell Transplant 2020;29:963689720906777

    Article  Google Scholar 

  24. Yau T, Park JW, Finn RS, Cheng AL, Mathurin P, Edeline J, et al. CheckMate 459: a randomized, multi-center phase III study of nivolumab (NIVO) vs sorafenib (SOR) as first-line (1L) treatment in patients (pts) with advanced hepatocellular carcinoma (aHCC). Ann Oncol 2019;30:874

    Article  Google Scholar 

  25. Kudo M. Immuno-oncology therapy for hepatocellular carcinoma: current status and ongoing trials. Liver Cancer 2019;8(4):221–238

    Article  Google Scholar 

  26. Dorand RD, Nthale J, Myers JT, Barkauskas DS, Avril S, Chirieleison SM, et al. Cdk5 disruption attenuates tumor PD-L1 expression and promotes antitumor immunity. Science 2016;353(6297):399–403

    Article  CAS  Google Scholar 

  27. Casey SC, Tong L, Li YL, Do R, Walz S, Fitzgerald KN, et al. MYC regulates the antitumor immune response through CD47 and PD-L1. Science 2016;352(6282):227–231

    Article  CAS  Google Scholar 

  28. Mezzadra R, Sun C, Jae LT, Gomez-Eerland R, de Vries E, Wu W, et al. Identification of CMTM6 and CMTM4 as PD-L1 protein regulators. Nature 2017;549(7670):106

    Article  CAS  Google Scholar 

  29. Kataoka K, Shiraishi Y, Takeda Y, Sakata S, Matsumoto M, Nagano S, et al. Aberrant PD-L1 expression through 3′-UTR disruption in multiple cancers. Nature 2016;534(7607):402

    Article  CAS  Google Scholar 

  30. Li M, Cui J, Niu W, Huang J, Feng T, Sun B, et al. Long non-coding PCED1B-AS1 regulates macrophage apoptosis and autophagy by sponging miR-155 in active tuberculosis. Biochem Biophys Res Commun 2019;509(3):803–839

    Article  CAS  Google Scholar 

  31. Zhou H, Su J, Hu X, Zhou C, Li H, Chen Z, et al. Glia-to-neuron conversion by CRISPR–CasRx alleviates symptoms of neurological disease in mice. Cell 2020;181(3):590 e16–603 e16

    Article  Google Scholar 

  32. Herbst RS, Soria JC, Kowanetz M, Fine GD, Hamid O, Gordon MS, et al. Predictive correlates of response to the anti-PD-L1 antibody MPDL3280A in cancer patients. Nature 2014;515(7528):563

    Article  CAS  Google Scholar 

  33. Hewson C, Morris KV. Form and function of exosome-associated long non-coding RNAs in cancer. Curr Top Microbiol 2016;394:41–56

    Google Scholar 

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The work was supported by project of Shanghai JiaDing District Health Commission (2017-KY-02).

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Correspondence to Caifeng Liu or Bin Wu.

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Fei Fan, Keji Chen, Xiaoliang Lu, Aijun Li, Caifeng Liu, Bin Wu have no conflicts of interest to declare.

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The study was approved by the Review Boards of Shanghai Eastern Hepatobiliary Surgery Hospital (Shanghai, China).

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Written informed consent was obtained from each patient.

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Supplementary Figure 1. Expression data and correlations in TCGA LIHC dataset. (A) Expression levels (TPM) of indicated lncRNAs in TCGA LIHC dataset. (B) Scatter plots showing the positive correlation of PCED1B-AS1 with PD-L1 (CD274) or PD-L2 (PDCD1LG2) in tumor tissues from TCGA LIHC dataset. (C) Scatter plots showing the positive correlation of PCED1B-AS1 with PD-L1 (CD274) or PD-L2 (PDCD1LG2) in normal control tissues from TCGA LIHC dataset. (D) Western blots showing PD-L1 and PD-L2 protein levels in Huh-7 cells after knockdown of indicated lncRNAs (TIFF 1870 kb)


Supplementary Figure 2. Correlations of hsa-mir-194-5p with PCED1B-AS1 and PD-Ls in TCGA LIHC dataset. (A) Workflow of predicting PD-L1 targeting microRNAs, PD-L2 targeting microRNAs and PCED1B-AS1 targeting microRNAs using TarPmiR and RNAhybrid. Scatter plots showing the negative correlations of hsa-mir-194-5p with PD-L1 (CD274) (B), PD-L2 (PDCD1LG2) (C) or PCED1B-AS1 (D) in tumor tissues from TCGA LIHC dataset (TIFF 625 kb)

Supplementary Table 1. List of PD-Ls correlated lncRNAs in TCGA LIHC dataset (XLSX 12 kb)

Supplementary Table 2. List of microRNA prediction by TarPmiR and RNAhybrid (XLSX 26 kb)

Supplementary Table 3. Clinico-pathological characteristics of patients (DOCX 16 kb)

Supplementary Table 4. Multivarite Analysis of Factors Associated With Survival (DOCX 13 kb)

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Fan, F., Chen, K., Lu, X. et al. Dual targeting of PD-L1 and PD-L2 by PCED1B-AS1 via sponging hsa-miR-194-5p induces immunosuppression in hepatocellular carcinoma. Hepatol Int 15, 444–458 (2021).

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