PRDM1/BLIMP1 induces cancer immune evasion by modulating the USP22-SPI1-PD-L1 axis in hepatocellular carcinoma cells

In the paper

“Programmed death ligand 1 (PD-L1, namely CD274) is widely expressed in various malignancies”

What the sources say

• Supported

  The Introduction (page not provided) identifies CD274 as PD-L1 and states that it is overexpressed in many cancers. This supports the claim that PD-L1 is widely expressed across malignancies. (LLM, conf=0.92)

  cites [1] Papalexi, E. et al. Characterizing the molecular regulation of inhibitory immune checkpoints with multimodal single-cell screens. Nat. Genet. 53, 322–331 (2021). source
• ?Not checked — no full text of the sourcecites [2] Yu, M. et al. Interferon-gamma induces tumor resistance to anti-PD-1 immunotherapy by promoting YAP phase separation. Mol. cell 81, 1216.e9–1230.e9 (2021). source
• ?Not checked — no full text of the sourcecites [3] Pinato, D. J., Fessas, P., Sapisochin, G. & Marron, T. U. Perspectives on the neoadjuvant use of immunotherapy in hepatocellular carcinoma. Hepatology 74, 483–490 (2021). source
• Supported

  The Discussion states that PD-L1 is constitutively upregulated in various tumors, including melanoma, non-small-cell lung cancer, and head and neck squamous cell carcinoma. The Methods search strategy also identifies CD274 as a term for PD-L1. Page numbers are not available in the supplied passages. (LLM, conf=0.91)

  cites [4] Li, Z. et al. Clinical benefit of neoadjuvant anti-PD-1/PD-L1 utilization among different tumors. MedComm 2, 60–68 (2021). source

In the paper

“induces immune evasion by targeting its ligand, programmed cell death 1 (PD-1) on activated T cells, resulting in T cell exhaustion1–4.”

What the sources say

• Partly supported

  The Introduction (page unavailable) says PD-L1 interacts with the PD-1 receptor on T cells to inhibit T-cell activation, and the Results (page unavailable) say cancer cells up-regulate PD-L1 to escape immune surveillance. Our copy does not specify activated T cells or say that this interaction results in T-cell exhaustion; a search for exhaustion found no supporting passage. (LLM, conf=0.88)

  cites [1] Papalexi, E. et al. Characterizing the molecular regulation of inhibitory immune checkpoints with multimodal single-cell screens. Nat. Genet. 53, 322–331 (2021). sourceExperimental — accuracy not yet measured
• ?Not checked — no full text of the sourcecites [2] Yu, M. et al. Interferon-gamma induces tumor resistance to anti-PD-1 immunotherapy by promoting YAP phase separation. Mol. cell 81, 1216.e9–1230.e9 (2021). source
• ?Not checked — no full text of the sourcecites [3] Pinato, D. J., Fessas, P., Sapisochin, G. & Marron, T. U. Perspectives on the neoadjuvant use of immunotherapy in hepatocellular carcinoma. Hepatology 74, 483–490 (2021). source
• Partly supported

  The Introduction (page not provided) says PD-L1 binds PD-1 on T cells, suppresses their activity, and enhances tumor immune escape. The Discussion (page not provided) identifies activated T cells as expressing PD-1. Neither passage says this interaction results in T-cell exhaustion, so that part of the claim is not supported by our copy. (LLM, conf=0.87)

  cites [4] Li, Z. et al. Clinical benefit of neoadjuvant anti-PD-1/PD-L1 utilization among different tumors. MedComm 2, 60–68 (2021). sourceExperimental — accuracy not yet measured

In the paper

“Moreover, several critical proteins including CMTM4, CMTM6, GSK3β, CSN5, CDK4, CDK6, and palmitoylated B3GNT3 have been shown to affect post-translational PD-L1 stability31–36.”

What the sources say

• Supported

  The paper’s Abstract identifies CMTM4 and CMTM6 as regulators of PD-L1 protein stability. This paper is one of citations 31–36, so its support is assessed for CMTM4 and CMTM6, not for every protein in the list. No page number is provided in our copy. (LLM, conf=0.92)

  cites [31] Mezzadra, R. et al. Identification of CMTM6 and CMTM4 as PD-L1 protein regulators. Nature 549, 106–110 (2017). source
• ?Not checked — no full text of the sourcecites [32] Visan, I. CMTM6 controls PD-L1. Nat. Immunol. 18, 1067 (2017). source
• Supported

  This paper supports the CMTM6 portion of the multi-citation claim. Its main text reports accelerated degradation of PD-L1 after CMTM6 knockout, and Extended Data Figure 7 describes selective destabilisation of PD-L1 when CMTM6 is lost. No page number is available in our copy. (LLM, conf=0.90)

  cites [33] Burr, M. L. et al. CMTM6 maintains the expression of PD-L1 and regulates anti-tumour immunity. Nature 549, 101–105 (2017). source
• ?Not checked — no full text of the sourcecites [34] Borden, B. A. et al. The landscape of glycogen synthase kinase-3 beta genomic alterations in cancer. Mol. Cancer Ther. 20, 183–190 (2021). source
• Supported

  This source supports the CSN5 part of the multiply cited list. In Results, under “TNF-α Upregulates CSN5 Expression to Stabilize PD-L1,” CSN5 downregulation shortened PD-L1 protein half-life, while wild-type CSN5 overexpression stabilized PD-L1. (LLM, conf=0.93)

  cites [35] Lim, S. O. et al. Deubiquitination and stabilization of PD-L1 by CSN5. Cancer Cell 30, 925–939 (2016). source
• Partly supported

  The SUMMARY (page not provided) shows that CDK4 affects PD-L1 protein abundance and that CDK4/6 inhibitors increase its stability. It also reports that CDK6 depletion did not increase PD-L1; this paper therefore does not support listing CDK6 as an independently demonstrated regulator. The other listed proteins are not established by this paper’s findings. (LLM, conf=0.89)

  cites [36] Zhang, J. et al. Cyclin D-CDK4 kinase destabilizes PD-L1 via cullin 3-SPOP to control cancer immune surveillance. Nature 553, 91–95 (2018). sourceExperimental — accuracy not yet measured

In the paper

“Various critical transcription factors, including HIF-1α, c-MYC, NF-kB, STAT3, c-JUN, and miR-138-5p have been shown to transcriptionally regulate PD-L1 expression25–30.”

What the sources say

• Partly supported

  The Results section ('TET1 interacts with HIF-1A to regulate PD-L1'; page unavailable) reports that HIF-1A enhanced PD-L1 expression in gastric cancer cells. The copy does not establish PD-L1 regulation by the other listed factors—c-MYC, NF-kB, STAT3, c-JUN, or miR-138-5p—so it supports only the HIF-1α part of the sentence. (LLM, conf=0.88)

  cites [25] Wang, J. et al. LncRNA NUTM2A-AS1 positively modulates TET1 and HIF-1A to enhance gastric cancer tumorigenesis and drug resistance by sponging miR-376a. Cancer Med. 9, 9499–9510 (2020). sourceExperimental — accuracy not yet measured
• Supported

  The cited paper supports the c-MYC portion of the multi-citation claim. In the passage labeled Abstract (no page number provided), it reports that MYC binds the PD-L1 gene promoter and directly regulates its expression. (LLM, conf=0.93)

  cites [26] Casey, S. C. et al. MYC regulates the antitumor immune response through CD47 and PD-L1. Science 352, 227–231 (2016). source
• ?Could not verify

  The Introduction (page unavailable) says IFNγ induces PD-L1, but does not identify the listed factors as its transcriptional regulators. The Results section on TRAF3 depletion discusses NF-κB regulation of MHC-I and reports no change in PD-L1 transcription after TRAF3 knockout. Searches for the listed factors and PD-L1 transcriptional regulation found no passage establishing the attributed claim. (LLM, conf=0.76)

  cites [27] Gu, S. S. et al. Therapeutically increasing MHC-I expression potentiates immune checkpoint blockade. Cancer Discov. 11, 1524–1541 (2021). sourceExperimental — accuracy not yet measured
• Supported

  This paper supports the STAT3 portion of the list of regulators cited collectively as references 25–30. In “STAT3 activation directly drives PD-L1 expression in NKTL,” the authors report increased STAT3 occupancy at the PD-L1 promoter and link it to increased transcriptional activity and PD-L1 expression. No page number is available in our copy. (LLM, conf=0.91)

  cites [28] Song, T. L. et al. Oncogenic activation of the STAT3 pathway drives PD-L1 expression in natural killer/T-cell lymphoma. Blood 132, 1146–1158 (2018). source
• Supported

  The Results section, “JUN controls pro-fibrotic and immune-checkpoint genes” (page not provided), reports JUN binding at a PD-L1 enhancer and increased PD-L1 RNA and protein after JUN overexpression in lung fibroblasts. This supports the c-JUN portion of the multi-citation sentence; the paper is not needed to support the other factors listed. (LLM, conf=0.91)

  cites [29] Cui, L. et al. Activation of JUN in fibroblasts promotes pro-fibrotic programme and modulates protective immunity. Nat. Commun. 11, 2795 (2020). source
• Partly supported

  The Results section reports that miR-138-5p targets and down-regulates PD-L1, supporting its inclusion as a regulator. But the same section reports no significant change in PD-L1 gene expression despite lower protein expression, while the Introduction describes microRNAs as acting through mRNA degradation or translation inhibition. The paper therefore does not support calling miR-138-5p a transcription factor or saying it transcriptionally regulates PD-L1; it does not establish the other listed factors. (LLM, conf=0.88)

  cites [30] Song, N. et al. MicroRNA-138-5p suppresses non-small cell lung cancer cells by targeting PD-L1/PD-1 to regulate tumor microenvironment. Front. Cell Dev. Biol. 8, 540 (2020). sourceExperimental — accuracy not yet measured

In the paper

“Supporting this view, a recent study revealed that USP22 could regulate PD-L1 expression in a direct and indirect way37.”

What the sources say

• Supported

  The Conclusions explicitly describe two routes by which USP22 regulates PD-L1 protein levels: direct deubiquitination of PD-L1 and an indirect route through the USP22–CSN5–PD-L1 axis. The Background states the same distinction, supporting the citing sentence’s paraphrase. (LLM, conf=0.95)

  cites [37] Wang, Y. et al. The deubiquitinase USP22 regulates PD-L1 degradation in human cancer cells. Cell Commun. Signal. 18, 112 (2020). source

In the paper

“Infiltrated CD8+ T cells and the activity (GZMB+) of infiltrated CD8+ T cells was obviously increased, and infiltrated CD8+ T cell exhaustion was significantly reduced in the sgPrdm1 groups (Fig. 2g–i).”

In the paper

“Further, site-directed mutagenesis abolished the increased promoter activity in Hep3B cells with PRDM1 overexpression and the decreased promoter activity in PRDM1-knockout Huh7 cells (Fig. 5n, o).”

In the paper

“The results revealed that Prdm1 overexpression in immune-competent mice inhibited tumor proliferation and that Prdm1 knockout contributed to tumor proliferation (Supplementary Fig. 3i–n).”

In the paper

“Meanwhile, patients with high tumoral BLIMP1 protein expression showed significant tumor shrinkage after treatment, whereas patients with low tumoral BLIMP1 protein expression showed tumor progression after treatment (Supplementary Fig. 9c, d).”

In the paper

“We noticed that CD8+ cytotoxic T cell subpopulations were predominant in patient 1, who showed low expression of PRDM1 in tumor cells, whereas patient 2 showed a higher percentage of CD8+LAG3+ exhausted T cells and CD4+FOXP3+ Tregs, supporting the notion that PRDM1 overexpression potentiates T cell exhaustion (Fig. 8e).”

In the paper

“Flow cytometry analysis and IF staining confirmed that co-treatment with Prdm1 knockout and PD-1 displayed comparable percentages of infiltrated CD8+ T cells, activities (GZMB+) of infiltrated CD8+ T cells, and exhaustion of infiltrated CD8+ T cells compared with those in Prdm1 knockout alone (Fig. 6j and Supplementary Fig. 7d–f).”

In the paper

“Our preclinical animal experiments including subcutaneous tumor models, orthotopic tumor models, and the DEN-induced HCC model, validated the synergistic effect of Prdm1 overexpression and PD-1 mAb treatment, in terms of a distinct reduction in tumor size/number and extended overall survival, which was attributed to the reinforcement of TILs in the tumors.”

In the paper

“Here, we found that PRDM1-overexpressing HCC cells upregulated PD-L1 expression and inhibited T cell-mediated anti-tumor immunity, thus dampening its role as a tumor growth suppressor in immunocompetent mice.”

In the paper

“Moreover, BLIMP1 and PD-L1 levels were negatively correlated with CD8+ T cells, and the activity (GZMB+) of infiltrated CD8+ T cells (Fig. 7d).”

In the paper

“Ultimately, as expected, patient 2 with high tumoral PRDM1 expression showed significant tumor shrinkage after treatment, whereas patient 1 with low tumoral PRDM1 expression showed tumor progression after treatment.”

In the paper

“Immune regulatory parameters such as positive regulation of immune response, response to interferon-gamma, and lymphocyte activation, were significantly enriched. Regulation of the cell death pathway was also significantly enriched, indicating that PRDM1 might suppress the proliferation ability of HCC cell lines by mediating tumor cell death (Fig. 8f).”

In the paper

“These results indicate that USP22 interacts with SPI1 and enhances SPI1 stability through deubiquitination.”

In the paper

“Thus, Prdm1 fails to impede tumor proliferation in immunocompetent mice but impairs tumor progression in immunodeficient mice.”

In the paper

“Thus, above results confirmed that Prdm1 may promote tumor immune evasion by driving PD-L1 upregulation and neutralizing the anti-tumor efficacy of Prdm1 overexpression.”

In the paper

“Taken together, SPI1 is a downstream effector of PRDM1 and directly binds the PD-L1 promoter to enhance its mRNA expression.”

In the paper

“Each specimen was histologically and pathologically examined and graded by two experienced pathologists.”

In the paper

“Gating strategies used for flow cytometry staining was provided in Supplementary Fig. 10.”

In the paper

“Uncropped and unprocessed scans of blots are included in a Source Data file.”

In the paper

“Tumor sizes were calculated by the volume formula π/6(L × W × H) [L: the length (longest dimension); W: width (shorter dimension, parallel to the mouse body); H: height (diameter of tumor perpendicular to the length and width)], which is confirmed to be the best formula represented the actual volume of a wide spectrum of tumor shapes and sizes38.”

What the sources say

• ?Not checked — no full text of the sourcecites [38] Tomayko, M. M. & Reynolds, C. P. Determination of subcutaneous tumor size in athymic (nude) mice. Cancer Chemother. Pharmacol. 24, 148–154 (1989). source

In the paper

“All the animal studies were approved by the Institutional Animal Care and Use Committee of Nanjing Medical University and conducted according to protocols approved by the Ethical Committee of Nanjing Medical university.”

In the paper

“P value was determined by unpaired two-sided Student’s t test (e, g, i, j, n, o) and Kaplan–Meier method (f, h, p) with no correction for multiple comparisons.”

In the paper

“No patient in Cohort 1 and Cohort 2 received any other HCC treatment prior to surgery.”

In the paper

“Collectively, we demonstrate that the PRDM1-USP22-SPI1 axis regulates PD-L1 levels, resulting in infiltrated CD8+ T cell exhaustion.”

In the paper

“Herein, we identified that PRDM1 may upregulate SPI1 expression, thus contributing to PD-L1 expression.”

In the paper

“Our study was approved by the Ethics Committee of the First Affiliated Hospital of Nanjing Medical University, and informed consent was obtained from each patient.”