Gencefe Client Article| SERPINB5 is a prognostic biomarker and promotes proliferation, metastasis and (EMT) in lung Adenocarcinoma

The following article is transferred from GENCEFE BIOTECH （Disease Research Newsletter

The most common form of lung cancer is lung adenocarcinoma (LUAD), which is characterized by poor prognosis, recurrence, and easy metastasis. [1, 2] Surgery, chemotherapy, radiotherapy, targeted therapy, immunotherapy, etc. are all clinically effective forms of treatment; however, improving the survival rate of LUAD patients is still a huge challenge. [3, 4] Serine protease inhibitors (SERPINs) are the largest family of protease inhibitors and are related to tumors and the regulation of many important life processes in humans. [5, 6] Studies have shown that SERPINB5 is a tumor suppressor gene that is expressed at abnormally low levels in a variety of tumors, inhibiting the invasion, migration, proliferation and other functions of tumor cells. [7, 8] However, the role of SERPINB5 in LUAD is still controversial.

In July 2023, the team of Director Zhang Lanjun of Sun Yat-sen University Cancer Hospital published an article titled "SERPINB5 is a prognostic biomarker and promotes proliferation, metastasis and epithelial-mesenchymal transition (EMT) in lung adenocarcinoma" in the journal "Thoracic Cancer" [9]. SERPINB5 was found to serve as a potential prognostic biomarker and may become a potential therapeutic target for lung adenocarcinoma.

In this study, small interfering RNA (siRNA) targeting SERPINB5 was designed and synthesized by GenCefe Biotech and successfully achieved the knockdown of SERPINB5.

Research result:

1. Clinical correlation analysis between SERPINB5 and LUAD

By analyzing more than thirty types of cancer in the TCGA database, the researchers found that the expression of SERPINBs was significantly different between different tumor types, among which SERPINB5, SERPINB6 and SERPINB7 were associated with poor prognosis in lung adenocarcinoma. Through further comprehensive analysis of the SERPINB gene family, it was found that SERPINB5 is up-regulated and demethylated in lung adenocarcinoma, and its abnormally high expression is associated with poor prognosis in the TCGA-LUAD cohort. Subsequently, by performing immunohistochemistry on 44 pairs of tumor and normal lung tissue samples, the researchers found that the expression level of SERPINB5 protein in LUAD tissue was higher than that in normal tissue (Figure 1b, c). Analysis of 106 LUAD samples by RT-PCR found that elevated SERPINB5 mRNA expression levels indicated poor prognosis and multivariable regression analysis showed that SERPINB expression was not related to prognostic factors such as age and gender (Fig. 1d, e). Combined with the transcriptome data verification of multiple lung adenocarcinoma GEO cohorts, it was shown that SERPINB5 expression is a potential biomarker for the prognosis of lung adenocarcinoma.

2. Reduced SERPINB5 expression inhibits the proliferation, migration, invasion and EMT of LUAD cells

Changes in mRNA and protein levels were detected by RT-PCR and Western Blot, and the results showed that si-SERPINB5 exhibited good knockdown efficiency (Figure 2a). After SERPINB5 knockdown, the healing ability of cells was significantly reduced (Figure 2b); the migration and invasion abilities of A549 and PC9 cell lines were significantly reduced (Figure 2c, d). Cell proliferation was significantly reduced 48 hours after SERPINB5 knockdown (Figure 2b). Figure 2e,f)

EMT-related vimentin and E-cadherin expression changes in SERPINB5 knockdown cells were detected by immunofluorescence and Western Blot, in which vimentin expression decreased and E-cadherin expression increased (Figure 3), indicating that SERPINB5 can pass through the epithelial EMT regulates cell proliferation, migration and invasion.

3. Up-regulation of SERPINB5 expression promotes the proliferation, migration and invasion of LUAD cells

The stable overexpression of SERPINB5 in A549 and PC9 cells was first verified by qRT-PCR and Western blotting respectively (Fig. 4a). Overexpression of SERPINB5 can promote lung cancer cell migration (Figure 4b), and overexpression of SERPINB5 significantly increased the invasion and migration of LUAD cells (Figure 4c, d). SERPINB5-overexpressing A549 and PC9 cells showed increased proliferation (Fig. 4e, f).

In addition, GSEA (gene set enrichment analysis) results found that there is a significant correlation between SERPINB5 expression and multiple proliferation-related pathways, such as G2M CHECKPOINTS, E2F TARGETS, MTORC1 signaling, MYC TARGETS V1&V2 pathway; and epithelial-mesenchymal transition (EMT ) markers were significantly enriched in the high SERPINB5 subgroup. These results indicate that cells with higher SERPINB5 expression are more likely to proliferate and migrate.

This study found that SERPINB family genes are not only tumor suppressor genes, but also sometimes tumor promoter genes. In addition, SERPINB5 is abnormally upregulated in lung adenocarcinoma, and its abnormally high expression is associated with poor prognosis in LUAD. High expression of SERPINB5 in LUAD is associated with the activation of many tumorigenesis pathways. By evaluating the effect of SERPINB5 on malignant progression in vitro, it was found that SERPINB5 promotes the proliferation, invasion, migration and EMT of lung adenocarcinoma cells. Therefore, SERPINB5 can serve as a potential prognostic biomarker and may become a potential therapeutic target for lung adenocarcinoma.

References:

[1] Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2021; 71: 209–49.

[2] Herbst RS, Morgensztern D, Boshoff C. The biology and management of non-small cell lung cancer. Nature. 2018; 553: 446–54.

[3] Hirsch FR, Scagliotti GV, Mulshine JL, Kwon R, Curran WJ Jr, Wu YL, et al. Lung cancer: current therapies and new targeted treatments. Lancet. 2017; 389: 299–311.

[4] Castellanos E, Feld E, Horn L. Driven by mutations: the predictive value of mutation subtype in EGFR-mutated non-small cell lung cancer. J Thorac Oncol. 2017; 12: 612–23.

[5] Kellici TF, Pilka ES, Bodkin MJ. Small-molecule modulators of serine protease inhibitor proteins (serpins). Drug Discov Today. 2021; 26: 442–54.

[6] Gettins PGW. Serpin structure, mechanism, and function. Chem Rev. 2002; 102: 4751–804.

[7] Chen EI, Yates JR. Maspin and tumor metastasis. IUBMB Life. 2006; 58: 25–9.

[8] Lonardo F, Li X, Kaplun A, Soubani A, Sethi S, Gadgeel S, et al. The natural tumor suppressor protein maspin and potential application in non small cell lung cancer. Curr Pharm Des. 2010; 16: 1877–81.

[9] He, Xiaotian, et al. SERPINB5 is a prognostic biomarker and promotes proliferation, metastasis and epithelial‐mesenchymal transition (EMT) in lung adenocarcinoma.?Thoracic Cancer?14.23 (2023): 2275-2287.

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