Biomarkers and systems for determining the sensitivity of tumors to radiotherapy

Abstract

This invention discloses biomarkers and systems for determining tumor radiosensitivity. The inventors discovered that high HOXA1 expression is closely associated with radioresistance in head and neck squamous cell carcinoma, low-grade glioma, and nasopharyngeal carcinoma. Patients with high HOXA1 expression have an increased risk of recurrence after radiotherapy and a poorer prognosis. Detecting HOXA1 expression levels can predict tumor radiosensitivity. Inhibiting HOXA1 expression in nasopharyngeal carcinoma cells via interfering RNA can significantly improve the radiosensitivity of these cells. Inhibition of HOXA1 expression can be used for tumor radiosensitization.

Description

Technical Field

[0001] This invention belongs to the field of biology, and specifically relates to biomarkers and systems for determining the sensitivity of tumors to radiotherapy. Background Technology

[0002] Radiation therapy (RT) is an important treatment for malignant tumors. Approximately 50% of patients with malignant tumors require RT. In malignant tumors such as squamous cell carcinoma of the head and neck, lung cancer, and glioma, RT is an important adjuvant therapy for shrinking tumors before surgery and reducing the risk of recurrence after surgery. In some tumors, such as nasopharyngeal carcinoma, RT can also be used as a primary radical treatment due to the difficulty of surgery caused by their special anatomical location. Although RT plays an important role in the treatment of malignant tumors, the radiosensitivity of tumors varies significantly among patients. In some patients, the radioresistance of tumors severely affects the effectiveness of RT, resulting in residual tumors or recurrence after RT, leading to poor prognosis. Therefore, it is of great significance to identify relevant biomarkers to predict tumor radiosensitivity and thereby guide individualized treatment for patients.

[0003] HOXA1 is a member of the homeobox transcription factor gene family. As a transcription factor, it can regulate the expression of related genes by binding to the promoter sequences of downstream genes. Current reports show that HOXA1 is aberrantly expressed in various tumors and may play a role in promoting tumor cell proliferation. However, there are currently no reports on HOXA1's involvement in regulating tumor radiosensitivity. Summary of the Invention

[0004] The purpose of this invention is to overcome at least one deficiency of the prior art and to provide a biomarker and system for determining the sensitivity of tumors to radiotherapy.

[0005] The technical solution adopted in this invention is:

[0006] The first aspect of the present invention provides:

[0007] Application of reagents for quantifying HOXA1 in the preparation of reagents for assessing the sensitivity of tumors to radiotherapy.

[0008] In some applications, the tumor is selected from head and neck squamous cell carcinoma, low-grade glioma, or nasopharyngeal carcinoma.

[0009] In some applications, HOXA1 expression level refers to the expression level of HOXA1 mRNA or protein.

[0010] In some application examples, reagents for quantifying HOXA1 are selected from real-time quantitative qPCR kits for detecting HOXA1 gene mRNA expression or specific HOXA1 antibody and immunohistochemistry kits for detecting HOXA1 gene expression.

[0011] In some applications, the samples tested by reagents for quantifying HOXA1 are tissue samples.

[0012] A second aspect of the present invention provides:

[0013] A system for determining the sensitivity of a tumor to radiotherapy, comprising:

[0014] HOXA1 quantification device: used to quantify the amount or concentration of HOXA1 in a sample;

[0015] Assessment device: Determines the sensitivity of tumors to radiotherapy based on the amount or concentration of HOXA1 in the sample;

[0016] Result output device: Outputs the result given by the judgment device.

[0017] In some system instances, the sample is a tissue sample.

[0018] In some systemic instances, the tumor is selected from head and neck squamous cell carcinoma, low-grade glioma, or nasopharyngeal carcinoma.

[0019] In some system instances, the HOXA1 quantification device is selected from mRNA quantification devices or immunohistochemical detection devices.

[0020] In some systematic examples, the sensitivity criteria are as follows: when HOXA1 protein expression is detected by immunohistochemistry, if the proportion of strongly stained cells is greater than 50%, it is considered that HOXA1 is highly expressed and the sensitivity to tumor radiotherapy is low; conversely, if the proportion of strongly stained cells is greater than 50%, it is considered that HOXA1 is low expressed and the sensitivity to tumor radiotherapy is high.

[0021] A third aspect of the present invention provides:

[0022] Application of reagents that downregulate HOXA1 expression in the preparation of tumor radiotherapy enhancers.

[0023] In some application examples, the reagents used to downregulate HOXA1 expression are selected from HOXA1-specific siRNAs that interfere with HOXA1 expression.

[0024] In some application examples, the nucleotide sequence of the HOXA1-specific siRNA is GUUCCUUUCAGAUGACCUU (SEQ ID NO.: 1).

[0025] The beneficial effects of this invention are:

[0026] The inventors discovered through research that high HOXA1 expression is closely associated with radioresistance in head and neck squamous cell carcinoma, low-grade glioma, and nasopharyngeal carcinoma. Patients with high HOXA1 expression have an increased risk of recurrence after radiotherapy and a poorer prognosis. By detecting HOXA1 expression levels, tumor radiotherapy sensitivity can be predicted.

[0027] The inventors also discovered through research that inhibiting HOXA1 expression in nasopharyngeal carcinoma cells via interfering RNA can significantly improve the radiosensitivity of these cells. Inhibiting HOXA1 expression could be used for tumor radiosensitization. Attached Figure Description

[0028] Figure 1 Data from the TCGA database shows that HOXA1 is upregulated in a variety of malignant tumors, including head and neck squamous cell carcinoma (HNSC) and low-grade glioma (LGG).

[0029] Figure 2 High expression of HOXA1 in patients with head and neck squamous cell carcinoma (A) and low-grade glioma (B) who have received radiotherapy indicates a higher risk of tumor recurrence and a poorer prognosis.

[0030] Figure 3 In nasopharyngeal carcinoma patients undergoing radical radiotherapy, high expression of HOXA1 in the tumor indicates a higher risk of recurrence and a poorer prognosis after radiotherapy.

[0031] Figure 4 Interfering with the expression of HOXA1 in nasopharyngeal carcinoma cells can enhance the radiosensitivity of related cells. Detailed Implementation

[0032] The first aspect of the present invention provides:

[0033] Application of reagents for quantifying HOXA1 in the preparation of reagents for assessing the sensitivity of tumors to radiotherapy.

[0034] In some applications, the tumor is selected from head and neck squamous cell carcinoma, low-grade glioma, or nasopharyngeal carcinoma.

[0035] In some applications, HOXA1 expression level refers to the expression level of HOXA1 mRNA or protein.

[0036] The reagents for quantifying HOXA1 can be existing HOXA1 mRNA or protein quantification reagents, and there are no special requirements for the type of reagent. In some application examples, the reagents for quantifying HOXA1 are selected from real-time quantitative qPCR kits for detecting HOXA1 gene mRNA expression or specific HOXA1 antibody and immunohistochemistry kits for detecting HOXA1 gene expression.

[0037] In some applications, the reagents used to quantify HOXA1 are used to detect tissue samples. Specifically, these tissue samples can be obtained during tumor surgery or biopsy.

[0038] A second aspect of the present invention provides:

[0039] A system for determining the sensitivity of a tumor to radiotherapy, comprising:

[0040] HOXA1 quantification device: used to quantify the amount or concentration of HOXA1 in a sample;

[0041] Assessment device: Determines the sensitivity of tumors to radiotherapy based on the amount or concentration of HOXA1 in the sample;

[0042] Result output device: Outputs the result given by the judgment device.

[0043] In some system instances, the sample is a tissue sample.

[0044] In some systemic instances, the tumor is selected from head and neck squamous cell carcinoma, low-grade glioma, or nasopharyngeal carcinoma.

[0045] In some system instances, the HOXA1 quantification device is selected from mRNA quantification devices or immunohistochemical detection devices.

[0046] The criteria for determining sensitivity can be determined through retrospective studies of existing cases. The criteria may be the same or different for different tumors. In some systematic cases, the criteria for determining sensitivity are: when HOXA1 protein expression is detected by immunohistochemistry, a high proportion of strongly stained cells (greater than 50%) indicates high HOXA1 expression and low sensitivity to radiotherapy; conversely, a low proportion of strongly stained cells indicates low HOXA1 expression and high sensitivity to radiotherapy.

[0047] A third aspect of the present invention provides:

[0048] Application of reagents that downregulate HOXA1 expression in the preparation of tumor radiotherapy enhancers.

[0049] In some application examples, the reagents used to downregulate HOXA1 expression are selected from HOXA1-specific siRNAs that interfere with HOXA1 expression.

[0050] In some application examples, the nucleotide sequence of the HOXA1-specific siRNA is GUUCCUUUCAGAUGACCUU.

[0051] The technical solution of this experiment will be further explained in conjunction with the following experiment:

[0052] Transcriptome gene expression data for over 11,000 cases of cancerous and adjacent normal tissues, including all 33 types of malignant tumors such as head and neck squamous cell carcinoma (HNSC) and low-grade glioma (LGG), were downloaded from The Cancer Genome Atlas (TCGA) database (database updated as of May 3, 2022). The differences in HOXA1 gene expression levels between corresponding cancerous and adjacent normal tissues were then compared using the Mann-Whitney U test. Results are as follows: Figure 1 The results showed that HOXA1 expression levels were significantly elevated in a variety of malignant tumors, including head and neck squamous cell carcinoma (HNSC) and low-grade glioma.

[0053] Further analysis of head and neck squamous cell carcinoma and low-grade glioma data from the TCGA database revealed that all samples with complete clinical treatment and follow-up data were divided into two groups based on whether or not they had received radiotherapy. Then, based on the median overall HOXA1 expression, cases in the radiotherapy-treated group and the non-radiotherapy-treated group were further subdivided into HOXA1 high-expression and low-expression subgroups. Kaplan-Müller survival curve analysis was used to analyze the correlation between HOXA1 expression levels and recurrence risk and prognosis in head and neck squamous cell carcinoma and low-grade glioma patients who received and did not receive radiotherapy. Results are as follows: Figure 2 As shown, in patients with head and neck squamous cell carcinoma and low-grade glioma who received radiotherapy, those with high HOXA1 expression had a significantly higher risk of recurrence and a worse prognosis compared to those with low expression. However, HOXA1 expression levels had no significant predictive value in patients who did not receive radiotherapy.

[0054] In biopsy tumor tissues collected from 50 patients with nasopharyngeal carcinoma who did not recur five years after radical radiotherapy and 20 patients with nasopharyngeal carcinoma that recurred in situ within five years after radiotherapy, the expression of HOXA1 was detected by immunohistochemistry, and the correlation between the expression level of HOXA1 in nasopharyngeal carcinoma tissue and the radiotherapy effect was further analyzed. Results are as follows: Figure 3 As shown, nasopharyngeal carcinoma patients with high HOXA1 expression who received radical radiotherapy had significantly worse prognoses, and high HOXA1 expression also indicated a higher risk of recurrence after radiotherapy.

[0055] In vitro experiments used siRNA to inhibit HOXA1 expression in nasopharyngeal carcinoma cells CNE1 and HNE1. The interference efficiency of siRNA (nucleotide sequence GUUCCUUUCAGAUGACCUU) was verified by real-time quantitative qPCR. After confirming that siRNA effectively interfered with HOXA1 expression in the corresponding cells, a certain number of control and HOXA1-interfering cells were seeded in small culture dishes. After cell adhesion, the cells were irradiated with radiation at 0, 0.5, 1, 2, 4, 6, and 8 Gy, respectively, and cultured for more than 14 days. Colony formation was observed, and the adhesion rate and cell viability fraction were calculated. Radiation survival curves were generated by fitting a linear quadratic curve [S=e^(-αD-βD2)] to the cells, and the differences in radiation survival curves between different treatment groups were compared to determine the effect of HOXA1 interference on cell radiosensitivity. Results are as follows: Figure 4 As shown, after radiation irradiation, the surviving cell clonogenic ability of nasopharyngeal carcinoma cells with HOXA1 expression interference was significantly reduced compared to control cells, the radiation survival curve shifted to the left, and radiosensitivity was enhanced. This result suggests that HOXA1 is involved in inducing radioresistance in tumor cells, and interfering with HOXA1 expression can achieve radiosensitization of nasopharyngeal carcinoma cells.

[0056] The above is a further detailed description of the present invention and should not be considered as a limitation on the specific implementation of the present invention. For those skilled in the art, simple deductions or substitutions without departing from the concept of the present invention are all within the protection scope of the present invention.

Claims

1. Application of reagents for quantifying HOXA1 in the preparation of reagents for assessing the sensitivity of tumors to radiotherapy, wherein the tumor is nasopharyngeal carcinoma and the sample tested by the reagents for quantifying HOXA1 is a tissue sample.

2. The application according to claim 1, characterized in that, The reagents for quantifying HOXA1 are selected from real-time quantitative qPCR kits for detecting the expression level of HOXA1 gene mRNA or specific HOXA1 antibody and immunohistochemistry kits for detecting HOXA1 gene expression.

3. A system for determining the radiotherapy sensitivity of a tumor, comprising: HOXA1 quantification device: used to quantify the amount or concentration of HOXA1 in a sample, wherein the sample is a tissue sample; Judgment device: Based on the amount or concentration of HOXA1 in the sample, the sensitivity of tumor to radiotherapy is determined. The criteria for judging sensitivity are: when HOXA1 protein expression is detected by immunohistochemistry, if the proportion of strongly stained cells is greater than 50%, it is judged that HOXA1 is highly expressed and the sensitivity of tumor to radiotherapy is low; otherwise, it is judged that HOXA1 is low expressed and the sensitivity of tumor to radiotherapy is high. Result output device: Outputs the result given by the judgment device; The tumor is nasopharyngeal carcinoma.

4. The system according to claim 3, characterized in that, The HOXA1 quantification device is selected from mRNA quantification devices or immunohistochemical detection devices.

5. Application of reagents that downregulate HOXA1 expression in the preparation of radiotherapy enhancers for tumors, wherein the tumor is nasopharyngeal carcinoma.

6. The application according to claim 5, characterized in that, The reagents used to downregulate HOXA1 expression were selected from HOXA1-specific siRNAs that interfere with HOXA1 expression.

7. The application according to claim 6, characterized in that, The nucleotide sequence of the HOXA1-specific siRNA is GUUCCUUUCAGAUGACCUU.

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