Application of SOX4 in preparation of preparation for improving sensitivity of cervical cancer to chemotherapeutic drugs

Abstract

The invention belongs to the technical field of biomedicine, and particularly relates to application of SOX4 in preparation of a preparation for improving sensitivity of cervical cancer to chemotherapeutic drugs, and the amino acid sequence of the SOX4 is as shown in SEQ ID NO.1. An SOX4 knocked-down stable transfection cell line is constructed in a Hela cell, then carboplatin and oxaliplatin are respectively added into the SOX4 knocked-down Hela cell, and the IC50 of the cell to the carboplatin and the oxaliplatin is obviously reduced, which indicates that the sensitivity of the Hela cell to the carboplatin and the oxaliplatin is enhanced by SOX4 knock-down. By reducing the expression of SOX4, the sensitivity of Hela cells to chemotherapeutic drugs such as carboplatin and oxaliplatin is enhanced, so that the treatment effect of the chemotherapeutic drugs such as carboplatin and oxaliplatin is effectively improved. Results of the invention prove that inhibition of SOX4 is beneficial to recovery of chemosensitivity of cervical cancer cells to drugs such as carboplatin and oxaliplatin, and a new treatment strategy is provided for treatment of drug-resistant cervical cancer.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical technology, and particularly relates to an application of SOX4 in the preparation of a preparation for improving the sensitivity of cervical cancer to chemotherapy drugs. Background Art

[0002] Cervical cancer is a common gynecological malignancy. Hela, Caski, C33A, SiHa, and other cervical cancer cell lines are commonly used in scientific research. The characteristics and behavior of cervical cancer cells determine the occurrence, development, and clinical manifestations of cervical cancer. Current clinical treatments for cervical cancer mainly include surgery, radiotherapy, and chemotherapy. Despite progress in screening and treatment, chemotherapy resistance in cervical cancer cells remains a key challenge. The main causes of cancer chemotherapy resistance are as follows: reduced drug uptake or increased drug efflux, inhibition of apoptosis pathways, epigenetic regulation, enhanced tolerance to damaged DNA, or increased DNA repair. This severely limits the efficacy of traditional therapies. Understanding the molecular mechanisms of this resistance is crucial for developing more effective treatment strategies.

[0003] Currently, first-line chemotherapy drugs for cervical cancer include cisplatin, carboplatin, and paclitaxel. Cervical cancer cell resistance to chemotherapy drugs is one of the main causes of treatment failure and tumor recurrence. Drug resistance allows tumor cells to evade the killing effects of drugs, resulting in reduced treatment efficacy or even complete ineffectiveness. Furthermore, drug resistance can lead to accelerated tumor cell proliferation, increased invasiveness, and an increased risk of metastasis, severely impacting patient prognosis.

[0004] To improve chemotherapy sensitivity, existing technologies have explored a variety of strategies, such as combination therapy, targeting DNA repair mechanisms, and changing the tumor microenvironment. However, these strategies also have disadvantages: combination therapy may increase drug toxicity and side effects, affecting patients' quality of life. Drugs targeting DNA repair mechanisms may increase genomic instability due to compensatory activation of other repair pathways, thereby inducing secondary drug-resistant mutations. Methods that change the tumor microenvironment may cause damage to normal cells, leading to side effects. Therefore, there is an urgent need to provide a new strategy to improve chemotherapy sensitivity. Summary of the Invention

[0005] The purpose of the present invention is to provide an application of SOX4 in the preparation of a preparation for improving the sensitivity of cervical cancer to chemotherapy drugs, thereby solving the problem of cervical cancer cells' resistance to platinum compounds in the prior art.

[0006] The technical solution adopted in the present invention is: The present invention provides an application of SOX4 in preparing a preparation for improving the sensitivity of cervical cancer to chemotherapy drugs. The amino acid sequence of the SOX4 is shown in SEQ ID NO.1.

[0007] Preferably, the chemotherapy drug is a platinum chemotherapy drug.

[0008] Preferably, the platinum chemotherapy drug includes at least one of carboplatin and oxaliplatin.

[0009] Preferably, the nucleic acid molecule is shRNA targeting SOX4 or siRNA targeting SOX4.

[0010] Preferably, the nucleic acid molecule is delivered to the cervical cancerous tissue via an interfering vector.

[0011] Preferably, the interference vector is selected from any one of a lentiviral vector, an adeno-associated viral vector or a plasmid vector.

[0012] Preferably, the preparation further comprises pharmaceutically acceptable excipients.

[0013] Preferably, the pharmaceutically acceptable excipient is one or more of a diluent, a disintegrant, a precipitation inhibitor, a glidant, a binder, a dispersant, a suspending agent, an isotonic agent, a thickener, an emulsifier, a preservative and a stabilizer.

[0014] Preferably, the diluent includes any one of starch, lactose, sucrose and mannitol.

[0015] Preferably, the disintegrant includes any one of starch, microcrystalline cellulose and low-substituted hydroxypropyl cellulose.

[0016] Preferably, the precipitation inhibitor comprises any one of sodium lauryl sulfate, Tween-80, polyvinyl pyrrolidone and hydroxypropyl methylcellulose.

[0017] Preferably, the glidant includes any one of cationic polyacrylamide, polydiallyldimethylammonium chloride and cationic starch.

[0018] Preferably, the binder comprises any one of starch slurry, hydroxypropyl methylcellulose and povidone.

[0019] Preferably, the dispersant includes any one of sodium lauryl sulfate, polyvinyl pyrrolidone and sodium carboxymethyl cellulose.

[0020] Preferably, the suspending agent comprises any one of gum arabic, gum tragacanth, sodium carboxymethylcellulose and hydroxypropyl methylcellulose.

[0021] Preferably, the isotonic agent includes any one of sodium chloride, glucose and mannitol.

[0022] Preferably, the thickener includes any one of gum arabic, xanthan gum and sodium carboxymethyl cellulose.

[0023] Preferably, the emulsifier includes any one of sodium lauryl sulfate, benzalkonium chloride and sorbitan fatty acid.

[0024] Preferably, the preservative includes any one of benzoic acid, sorbic acid, methyl paraben and benzalkonium bromide.

[0025] Preferably, the stabilizer includes any one of sodium sulfite, sodium bisulfite, tocopherol and disodium edetate.

[0026] Preferably, the acceptable dosage form of the preparation includes one of tablets, capsules, granules, injections, pills, powders, pastes and oral liquids.

[0027] Compared with the prior art, the present invention has the following beneficial effects: The present invention provides a use of SOX4 in the preparation of a preparation for increasing the sensitivity of cervical cancer to chemotherapy drugs, wherein the amino acid sequence of SOX4 is shown in SEQ ID NO. 1. The present invention constructs a stable transfected cell line with SOX4 knockdown in Hela cells, and then adds carboplatin and oxaliplatin to the SOX4 knockdown Hela cells, respectively. It is found that the IC50 values ​​of the cells for carboplatin and oxaliplatin are significantly reduced, indicating that SOX4 knockdown enhances the sensitivity of Hela cells to carboplatin and oxaliplatin. The present invention enhances the sensitivity of Hela cells to chemotherapy drugs such as carboplatin and oxaliplatin by reducing the expression of SOX4, thereby effectively improving the therapeutic effects of chemotherapy drugs such as carboplatin and oxaliplatin. The results of the present invention confirm that inhibiting SOX4 helps restore the chemosensitivity of cervical cancer cells to drugs such as carboplatin and oxaliplatin, which provides a new therapeutic strategy for treating drug-resistant cervical cancer. Furthermore, SOX4 can also be used as a biomarker to predict chemotherapy response, providing a basis for personalized treatment. The research results of the present invention have important potential clinical application significance. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 To successfully construct SOX4 overexpression and interference Hela cell lines, A: After Hela cells were transfected, the transfection efficiency was detected by qRT-PCR; B: After Hela cells were transfected, the transfection efficiency was detected by Western Blot; C: The statistical results corresponding to B.

[0029] Figure 2 Effect of SOX4 on carboplatin sensitivity in cervical cancer cells. A: cell viability; B: IC50 value.

[0030] Figure 3 Effect of SOX4 on oxaliplatin sensitivity in cervical cancer cells. A: cell viability; B: IC50 value. DETAILED DESCRIPTION

[0031] The present invention will be further described below by way of specific examples, but the scope of the present invention is not limited thereto. The details and forms of the technical solution of the present invention may be modified or replaced without departing from the spirit and scope of the present invention, but such modifications or replacements fall within the scope of protection of the present invention.

[0032] The inventive concept of the present invention is as follows: SOX4, short for sex-dertermining region of Y chromosome related high-mobility group box 4, is a member of the SOX transcription factor family, group C. An analysis of the transcriptional profiles of over 3,700 human cancers found that SOX4 was the only gene upregulated among 64 genes commonly found in cancers, suggesting a key role in various tumor types. An analysis of 1,348 cancer patients found that overexpression of SOX4 was associated with poor prognosis in several cancers, including colon and gastric cancer.

[0033] SOX4 is a key transcription factor that plays an important role in cell proliferation, differentiation, migration, and apoptosis. In recent years, the function of SOX4 in tumors has garnered extensive attention. Numerous studies have demonstrated that SOX4 is highly expressed in various cancers, including breast, lung, and prostate cancers, and is closely associated with epithelial-mesenchymal transition (EMT), tumor invasion, and metastasis. SOX4 can promote tumor cell invasiveness and immune evasion by regulating signaling pathways such as Wnt / β-catenin, PI3K / AKT, and TGF-β. In cervical cancer, SOX4 forms a regulatory axis with various noncoding RNAs, such as lncRNAs and circRNAs, regulating the expression of key microRNAs, thereby influencing signaling pathways such as Wnt / β-catenin and promoting tumor proliferation, invasion, and metastasis. Furthermore, SOX4 can inhibit malignant progression in some tumors. Furthermore, elevated SOX4 expression is often associated with poor prognosis, making it a key research topic in tumor diagnosis and treatment. While the role of SOX4 in EMT and tumor metastasis has been extensively studied, its role in tumor metabolic reprogramming and chemotherapy resistance remains unclear. The results of this study suggest that SOX4 may play a role in cervical cancer cell resistance by regulating the expression of metabolism-related genes, affecting metabolic processes such as glycolysis. Therefore, this study focuses on the potential role of SOX4 in chemotherapy resistance, hoping to provide new targets for anti-resistance therapies by revealing its molecular mechanisms.

[0034] The SOX family of transcription factors plays an important role in various cellular processes, including development, differentiation, and tumorigenesis. Among them, SOX4 has attracted much attention due to its dual role in the cellular context: it can both promote tumorigenesis and suppress tumors.

[0035] This study aims to explore the role of SOX4 in inducing chemotherapy resistance in cervical cancer cells. By demonstrating that SOX4 is a multidrug resistance gene, we hope to identify new therapeutic targets for overcoming drug resistance by inhibiting SOX4, thereby improving the treatment of cervical cancer.

[0036] In order to make those skilled in the art better understand the technical solution of the present invention and be able to implement it, the present invention is further described below in conjunction with specific examples. In the description of the present invention, if not otherwise specified, the reagents used are all commercially available and the methods used are all conventional techniques in the art.

[0037] The amino acid sequence of SOX4 is shown in SEQ ID NO.1.

[0038] SEQ ID NO.1: MVQQTNNAENTEALLAGESSDSGAGLELGIASSPTPGSTASTGGKADDPSWCKTPSGHIKRPMNAFMVWSQIERRKIMEQSPDMHNAEISKRLGKRWKLLKDSDKIPFIREAERLRLK HMADYPDYKYRPRKKVKSGNANSSSSAAASSKPGEKGDKVGGSGGGGHGGGGGGGSSNAGGGGGGASGGGANSKPAQKKSCGSKVAGGAGGGVSKPHAKLILAGGGGGGKAAAAAAASF AAEQAGAAALLPLGAAADHHSLYKARTPSASASASSAASASAALAAPGKHLAEKKVKRVYLFGGLGTSSSPVGGVGAGADPSDPLGLYEEEGAGCSPDAPSLSGRSSAASSPAAGRSPA DHRGYASLRAASPAPSSAPSHASSSASSHSSSSSSSGSSSSDDEFEDDLLDLNPSSNFESMSLGSFSSSSALDRDLDFNFEPGSGSHFEFPDYCTPEVSEMISGDWLESSISNLVFTY.

[0039] In order to study the role of SOX4 in cervical cancer, the present invention established SOX4 overexpression and knockdown stable cervical cancer cell lines. Figure 1 .

[0040] (1) A stable transfected cell line overexpressing the SOX4 gene was constructed in HeLa cells as follows: When HeLa cells grew to 80% confluence, the SOX4 expression vector containing the SOX4 open reading frame and the negative control vector pEnter were transfected into HeLa cells using Lipofectamin 2000 transfection reagent. Cells were collected 48 hours after transfection, diluted and plated into 96-well plates to generate single-cell clones. After 2 weeks, the cells were digested and continued to be expanded and cultured. The mRNA and protein expression levels of SOX4 were then detected by qRT-PCR and Western Blot to confirm the overexpression effect.

[0041] (2) Construction of a SOX4 knockdown stably transfected cell line in HeLa cells as follows: When Hela cells grew to 80% confluence, the interference vector Lv-shRNA-GP / SOX4 containing the target sequence of the SOX4 gene and the negative mammalian control vector Lv-shRNA-GP / NMC containing a sequence that does not target any known gene were transfected into Hela cells using Lipofectamin 2000 transfection reagent. Cells were collected 48 hours after transfection, diluted and plated into 96-well plates to generate single-cell clones. After 2 weeks, the cells were digested and continued to be expanded and cultured. The mRNA and protein expression levels of SOX4 were then detected by qRT-PCR and Western Blot to confirm the interference effect.

[0042] In the following figures, Hela / SOX4 is the Hela cell group with overexpression of SOX4; Hela / pEnter is the negative control cell group for Hela / SOX4; Hela / shSOX4 is the Hela cell group with knockdown of SOX4, i.e., the SOX4 interference group; and Hela / NMC is the negative control cell group for Hela / shSOX4.

[0043] like Figure 1 As shown, qRT-PCR results showed that the mRNA expression in the SOX4-overexpressing Hela cells increased approximately 22.2-fold compared to the Hela / pEnter control group, while the mRNA expression in the SOX4-knockout group decreased approximately 62.9% compared to the Hela / NMC control group. Western blot analysis further verified the changes in SOX4 protein expression.

[0044] Example 1 The application of SOX4 in the preparation of a preparation for increasing the sensitivity of cervical cancer to chemotherapy drugs is as follows: 1. SOX4 increases the sensitivity of tumor cells to carboplatin.

[0045] Four HeLa cell lines modified with SOX4 were digested and counted, then plated at 4,000 cells per well in a 96-well plate. Different concentrations of carboplatin were added to the culture medium containing the cells according to a designed concentration gradient, with five replicates per well. After 48 hours of carboplatin treatment, CCK8 was added. After 1 hour of CCK8 treatment, the absorbance of each well was measured to calculate cell viability. GraphPad Prism 8 software was then used to calculate the IC50 value of carboplatin for HeLa cells overexpressing SOX4 and knocking down SOX4.

[0046] like Figure 2 As shown in the results, the IC50 of Hela / SOX4 cells to carboplatin was significantly increased, reaching 2.5 times that of the control group. In contrast, the IC50 of Hela / shSOX4 cells to carboplatin was significantly decreased, reaching 32.5% of the control group. These results indicate that overexpression of SOX4 significantly enhances carboplatin resistance in cervical cancer cells, while decreased SOX4 expression can enhance cell sensitivity to carboplatin by inhibiting key resistance mechanisms, thereby effectively improving treatment efficacy. This phenomenon suggests that targeted regulation of SOX4 expression may provide a new therapeutic strategy to overcome carboplatin resistance in cervical cancer.

[0047] 2. SOX4 increases the sensitivity of tumor cells to oxaliplatin.

[0048] Four HeLa cell lines modified with SOX4 were digested and counted, then plated at 4,000 cells per well in a 96-well plate. Different concentrations of oxaliplatin were added to the culture medium containing the cells according to a designed concentration gradient, with five replicates per well. After 24 hours of oxaliplatin exposure, CCK8 was added. After 1 hour of CCK8 exposure, the absorbance of each well was measured to calculate cell viability. GraphPad Prism 8 software was then used to calculate the IC50 of oxaliplatin against HeLa cells overexpressing and knocking down SOX4.

[0049] like Figure 3 As shown, the results showed that the IC50 of Hela / SOX4 cells to oxaliplatin was significantly increased, reaching 1.5 times that of the control group. In contrast, the IC50 of Hela / shSOX4 cells to oxaliplatin was significantly decreased, reaching 48.0% of the control group. These results indicate that overexpression of SOX4 significantly enhances oxaliplatin resistance in cervical cancer cells, while decreased SOX4 expression can enhance cell sensitivity to oxaliplatin by inhibiting key resistance mechanisms, thereby effectively improving treatment efficacy. This phenomenon suggests that targeted regulation of SOX4 expression may provide a new therapeutic strategy to overcome oxaliplatin resistance in cervical cancer.

[0050] Figures 1 to 3*p<0.05, **p<0.01, ***p<0.001. Data are expressed as mean ± SD, n = 3 independent experiments, two-tailed Student's t-test was used, and p<0.05 was considered significant.

[0051] The technical features of the above-mentioned embodiments can be combined arbitrarily. In order to make the description concise, not all possible combinations of the technical features in the above-mentioned embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0052] The above-described embodiments merely illustrate several implementations of the present invention. While the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that a person skilled in the art would be able to make numerous variations and improvements without departing from the spirit of the present invention, and all such variations and improvements fall within the scope of protection of the present invention.

Claims

1. Application of SOX4 in the preparation of a preparation for increasing the sensitivity of cervical cancer to chemotherapy drugs, characterized in that: The amino acid sequence of SOX4 is shown in SEQ ID NO.

1.

2. The use according to claim 1, characterized in that The chemotherapy drug is a platinum chemotherapy drug.

3. The use according to claim 2, characterized in that The platinum chemotherapy drug includes at least one of carboplatin and oxaliplatin.

4. The use according to claim 1, wherein The preparation comprises a nucleic acid molecule that interferes with SOX4 expression, and the nucleic acid molecule is shRNA targeting SOX4 or siRNA targeting SOX4.

5. The use according to claim 4, characterized in that The nucleic acid molecule is delivered to the cervical cancer tissue via an interfering vector.

6. The use according to claim 5, characterized in that The interference vector is selected from any one of a lentiviral vector, an adeno-associated viral vector or a plasmid vector.

7. The use according to claim 1, wherein The preparation also includes pharmaceutically acceptable excipients.

8. The use according to claim 7, characterized in that The pharmaceutically acceptable excipients are one or more of diluents, disintegrants, precipitation inhibitors, glidants, binders, dispersants, suspending agents, isotonic agents, thickeners, emulsifiers, preservatives and stabilizers.

9. The use according to claim 1, wherein The acceptable dosage forms of the preparation include one of tablets, capsules, granules, injections, pills, powders, pastes and oral liquids.

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