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. 2023 Jan 4:12:1049183.
doi: 10.3389/fonc.2022.1049183. eCollection 2022.

A pan-cancer landscape of IGF2BPs and their association with prognosis, stemness and tumor immune microenvironment

Wei Shao  1 , Hui Zhao  2 , Shoudu Zhang  1 , Qian Ding  1 , Yugang Guo  1 , Kaiqi Hou  1 , Yunchao Kan  1 , Fan Deng  3 , Qian Xu  1
Affiliations

A pan-cancer landscape of IGF2BPs and their association with prognosis, stemness and tumor immune microenvironment

Wei Shao et al. Front Oncol. .

Abstract

Background: The human insulin-like growth factor 2 mRNA binding proteins 1-3 (IGF2BP1-3, also called IMP1-3) play essential roles in mRNA regulation, including its splicing, translocation, stability, and translation. However, knowledge regarding the involvement of IGF2BPs in tumor immunity and stemness across cancer types is still lacking.

Methods: In this study, we comprehensively analyzed pan-cancer multi-omic data to determine the correlation of IGF2BPs mRNA and protein expression with various cancer parameters such as mutation frequency, prognostic value, the tumor microenvironment (TME), checkpoint blockade, tumor immune infiltration, stemness and drug sensitivity. Validation of the expression of IGF2BPs in cancer samples and glioma cells were performed by quantitative real-time (qRT)-PCR, and immunofluorescence staining. Investigation of the functional role of IGF2BP3 in glioma stem cells(GSCs) were performed by sphere formation, cytotoxicity, transwell, and wound healing assays.

Results: We found that IGF2BP1 and 3 are either absent or expressed at very low levels in most normal tissues. However, IGF2BP1-3 can be re-expressed in a broad range of cancer types and diverse cancer cell lines, where their expression often correlates with poor prognosis. Immunofluorescence staining and qRT-PCR analyses also showed that the expression of IGF2BP2 and IGF2BP3 were higher in cancer tissues than that in adjacent normal tissues. Moreover, IGF2BPs are associated with TME and stemness in human pan-cancer. Remarkably, IGF2BP3 participated in the maintenance and self-renewal of glioma stem cell (GSCs). Knockdown of IGF2BP3 attenuated GSC and glioma cell proliferation, invasion, and migration.

Conclusions: Our systematic pan-cancer study confirmed the identification of IGF2BPs as therapeutic targets and highlighted the need to study their association with stemness, and the TME, which contribute to the cancer drug-discovery research. Especially, preliminary studies demonstrate the IGF2BP3 as a potential negative regulator of glioma tumorigenesis by modulating stemness.

Keywords: glioma stem cells; insulin-like growth factor 2(IGF2) messenger RNA; pan-cancer; prognosis signature; tumor microenvironment.

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Conflict of interest statement

Author HZ was employed by Zhengzhou Revogene Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Expression pattern of IGF2BPs in human cancers. (A-C) Expression pattern of IGF2BPs in diverse normal tissues (data from GTEx). (D-F) Expression pattern of IGF2BPs in GTEx normal, TCGA normal, and TCGA cancer tissues. (G-I) Expression pattern of IGF2BPs in diverse cancer cells (data from CCLE). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
Figure 2
Figure 2
Expression level of IGF2BP2 and IGF2BP3 in COAD, ESCA and STAD cancer tissues. (A, B) IGF2BP2/3 protein expression levels among normal tissue and primary tissue were compared based on the CPTAC dataset. Immunofluorescence of IGF2BP2 (red) in validated cancer tissues and adjacent tissues. Representative immunofluorescence images are presented in validated cancer tissues (C) COAD, (D) ESCA and (E) STAD and adjacent tissues. (F) Immunoreactivity of IGF2BP2 was quantified. (G) The mRNA expression of IGF2BP2 in validated cancer tissues and adjacent tissues by RT-qPCR. Immunofluorescence of IGF2BP3 (green) in validated cancer tissues and adjacent tissues. Representative immunofluorescence images are presented in validated cancer tissues (H) COAD, (I) ESCA and (J) STAD and adjacent tissues. (K) Immunoreactivity of IGF2BP3 was quantified. (L) The mRNA expression of IGF2BP3 in validated cancer tissues and adjacent tissues by RT-qPCR. Data are mean ± SEM (n = 3 per group). *P < 0.05, **P < 0.01, ***P < 0.001.
Figure 3
Figure 3
The relationship between IGF2BPs expression level and overall survival. (A-C) Kaplan–Meier survival curves comparing the high and low expression of IGF2BPs in 33 cancer types. (D) The forest plot of the relationship between IGF2BP1 expression and OS across 13 tumors. (E) The forest plot of the relationship between IGF2BP2 expression and OS across 12 tumors. (F) The forest plot of the relationship between IGF2BP3 expression and OS across 15 tumors.
Figure 4
Figure 4
Correlation analysis between IGF2BPs expression and MMR, DNA methylation level, TMB, and MSI in pan-cancer. (A-C) The Spearman correlation analysis of IGF2BPs expression with expression levels of five MMR genes across cancers. (D-F) The Spearman correlation analysis of IGF2BPs expression with the expression of 4 methyltransferases, Red represents DNMT1, blue represents DNMT2, green represents DNMT3A, and purple represents DNMT3B. (G-I) The correlation analysis between IGF2BPs expression and TMB in pan-cancer. (J-L) The correlation analysis between IGF2BPs expression and MSI in pan-cancer. *P < 0.05, **P < 0.01, ***P < 0.001.
Figure 5
Figure 5
Relationship between the expression of the IGF2BPs and the tumor immune microenvironment.(A) Relationship between the expression of the IGF2BPs and the infiltrating immune sub-types in pan-cancer. C1: wound healing, C2: INF-r dominant, C3: inflammatory, C4: lymphocyte depleted, C5: immunologically quiet, and C6: TGFβ dominant. The correlation matrix of the IGF2BPs expression and (B) the stromal cells, (C) the immune cells, (D) as well as comprehensive scores of 33 cancer types based on the estimation algorithm. ***P < 0.001.
Figure 6
Figure 6
Relationship between IGF2BPs and the expression of tumor-infiltrating immune cell and immune checkpoint inhibitors. (A-C) Heatmap representation of the correlation between IGF2BPs expression and immune cell in pan-cancer. (D, E) Heatmaps represent the association of the IGF2BPs expression with PD-L1 and CTLA4. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
Figure 7
Figure 7
Association of IGF2BPs and tumor immune microenvironment in LGG and GBM based on TCGA. Association of the IGF2BPs expression and the immune infiltrate subtypes in (A) LGG and (B) GBM. (C-E) Association of the IGF2BPs expression and TAMs. Red color represents positive correlation, blue color represents negative correlation, and the deeper the color, the stronger the correlation. **P < 0.01, ***P < 0.001.
Figure 8
Figure 8
IGF2BP3 plays a key role in the maintenance and self-renewal of GSCs. (A–C) The correlation of stemness score and IGF2BPs gene expression. (D, E) GEPIA database showing IGF2BPs expression correlation with stem gene in high-grade glioma tissues compared with that in low-grade glioma tissues. (F) The serum level of IGF2BP3 was significantly increased in the GBM patients. (G) RT-qPCR analysis of IGF2BP3 expression in glioma cell lines (U251 and Hs 683). (H, I) The representative images of 2 established neutrosphere-cultured GSCs and lentiviral transfection efficiency. (J, K) Sphere formation of U251and HS683 after infection with shRNA-IGF2BP3 and shRNA-NT was evaluated by the sphere formation assay. Bar: 100μm. **P < 0.01, ***P < 0.001.
Figure 9
Figure 9
IGF2BP3 knockdown inhibits the cell proliferation, migration, and invasion of GSCs. (A, B) Cell proliferation ability of U251 and HS683 cells transfected with sh-IGF2BP3 was evaluated by CCK8 assay. (C, D) Cell migration capability of U251 and HS683 cells transfected with sh-IGF2BP3 was evaluated by wound healing assays. (E, F) The influence on cell migration and invasion abilities of U251 and HS683 cells transfected with sh-IGF2BP3 was assessed by transwell migration invasion assays. Bar: 100μm. **P < 0.01, ***P < 0.001.
Figure 10
Figure 10
IGF2BP3 knockdown impairs glioma cell proliferation, migration, and invasion. (A, B) Cell proliferation ability of U251 and HS683 cells transfected with sh-IGF2BP3 was evaluated by CCK8 assay. (C, D) Cell migration capability of U251 and HS683 cells transfected with sh-IGF2BP3 was evaluated by wound healing assays. (E, F) The in-fluence on cell migration and invasion abilities of U251 and HS683 cells transfected with sh-IGF2BP3 was assessed by transwell migration invasion assays. Bar: 100μm. *P < 0.05, **P < 0.01, ***P < 0.001.
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