Application of ASIC3 inhibitor in preparation of tumor treatment medicine

Abstract

The invention discloses an application of an ASIC3 inhibitor in preparation of a tumor treatment drug, and belongs to the technical field of biological medicines. The ASIC3 inhibitor is injected into the tumor to specifically block the tumor promoting effect of the mouse breast cancer sensory nerves, so that the solid tumor progress is inhibited. The application provides favorable support evidence for further development of a strategy of inhibiting tumors through a sensory nerve pathway, particularly in the aspect of safety, side effects caused by traditional tumor denervation are reduced, good enlightenment is provided for new use of old drugs of the marketed nervous system regulating drugs with good tolerance characteristics, and the application has broad application prospects. The method has important value and application potential.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological medicine, and particularly relates to an application of an ASIC3 inhibitor in preparation of a tumor treatment drug. BACKGROUND

[0002] Clinical pathological studies have shown that a variety of solid tumors are characterized by rich innervation. Although surgical operation or drugs can achieve the purpose of denervation and thus inhibit tumor progression, each method has certain defects. Peripheral nerves contain different types of nerve fibers, and surgical denervation causes the fracture of all nerve fibers in the specific region, resulting in the influence of other nerve fibers. On the other hand, blocking nerves through pharmacological approach causes systemic reactions of multiple organs and influences non-intended targets. Therefore, it is difficult to realize the resolution of the function of specific nerve fibers in the tumor microenvironment using these methods, and the observed results may not only be due to the action of target neurons, but also be interfered by other types of innervation. In summary, the conclusion drawn from the research based on surgical or drug denervation may not be accurate enough.

[0003] Sensory nerves in the tumor microenvironment promote angiogenesis, tumor proliferation and metastasis by releasing neuropeptides (such as CGRP and substance P). For example, CGRP promotes tumor immune escape by inhibiting natural killer (NK) cell function. The FDA-approved migraine treatment drug rimegepant (CGRP receptor antagonist) can block this pathway in a pancreatic cancer model, significantly reducing tumor burden, prolonging survival and relieving tumor pain (Wang, K., et al., Nociceptor neurons promote PDAC progression and cancer pain by interaction with cancer-associated fibroblasts and suppression of natural killer cells. Cell Res, 2025.). Acidification (pH 6.5-6.9) of the tumor microenvironment is one of the key factors for activating sensory nerves. Acid-sensing ion channel 3 (ASIC3) is highly expressed in sensory nerve terminals, responds to acidification and mediates calcium influx, triggering the release of various neuropeptides, but there is no research to explore whether local application of ASIC3 inhibitors can block the activation of sensory nerves in the tumor microenvironment and thus inhibit tumor progression. SUMMARY

[0004] The purpose of the present application is to provide the application of ASIC3 inhibitor in preparation of tumor treatment drug.

[0005] This invention uses mouse breast cancer as a model and employs the ASIC3 inhibitor A-317567 via intratumoral injection. The results showed that A-317567 significantly inhibited the progression of mouse breast cancer. First, the mouse breast cancer cell line E0771 was used to construct an orthotopic mouse breast cancer model via injection into the mammary fat pad. After 21 days of growth, tumors were harvested. Following tumor harvesting, immunofluorescence staining was used to detect the neural innervation type of the mouse breast cancer. The results showed that the dominant neural innervation type was TRPV1. + / CGRP + The tumor was located in a sensory nerve and highly expressed ASIC3. A-317567 was then injected intratumorally. Samples were taken 14 days after the tumor was established (day 7 of the model) to assess the tumor-suppressive effect (tumor volume and weight) and safety (behavioral testing of mice from the same batch before sampling). Results showed that local injection of the ASIC3 inhibitor reduced tumor volume by 30% ± 15.2% (P<0.05), and no abnormalities were found in behavioral testing (such as hyperalgesia or motor dysfunction).

[0006] A-317567 is a potent ASIC3 inhibitor with antidepressant and analgesic effects. This invention utilizes an intratumoral injection of an ASIC3 inhibitor to specifically block the pro-tumor effects of sensory nerves in mouse breast cancer, thereby inhibiting solid tumor progression. This application provides strong supporting evidence for the further development of strategies to inhibit tumors through sensory nerve pathways, particularly in terms of safety, reducing the side effects caused by traditional tumor denervation. It also offers valuable insights into the repurposing of well-tolerated, marketed neuromodulatory drugs, demonstrating significant value and application potential. Attached Figure Description

[0007] Figure 1 Results of nerve innervation type and ASIC3 expression in mouse breast cancer neurons.

[0008] Figure 2 The proliferation of mammary cancer in mice after local injection of A-317567. Detailed Implementation

[0009] The preferred embodiments of the present invention will now be described in detail with reference to specific examples. It should be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art can make various modifications and substitutions to the present invention without departing from its spirit and essence.

[0010] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0011] Unless otherwise specified, all materials and reagents used in the following examples are commercially available. Example

[0012] 1. Establish a mouse orthotopic breast cancer xenograft model The experimental method is as follows: (1) Preparation of cell suspension: After digestion and centrifugation, E0771 breast cancer cells were counted to prepare a cell suspension with a concentration of 1×10⁻⁶. 6 Cell suspension of cells / ml.

[0013] (2) Injection of mouse mammary fat pad cancer cells: SPF-grade female C57 / B6 mice, weighing 15-20g, were anesthetized using a gas anesthesia device containing isoflurane and placed on an operating table that could maintain the anesthesia. The abdomen was shaved and the skin was prepared. The surgical site was disinfected with an alcohol swab. 100μl of cell suspension was drawn up using an insulin syringe and injected into the fourth pair of mammary fat pads of the mice. The mice were fed for 14 days after the operation to establish a tumor model.

[0014] 2. Immunofluorescence staining method was used to detect the nerve innervation type and ASIC3 expression in mice with mammary breast cancer. The experimental method is as follows: The established tumor model mice were injected in situ with fluorescent gold (a retrograde nerve tracking agent), perfused, and the dorsal root ganglion tissue was removed, fixed, dehydrated, and sectioned. The expression of sympathetic nerve marker protein tyrosine hydroxylase (TH), parasympathetic nerve marker protein choline acetyltransferase (ChAT), and sensory nerve marker protein TRPV1 was detected by immunofluorescence staining.

[0015] like Figure 1 As shown, the main nerve innervation type of mouse mammary gland is TRPV1. + / CGRP + Sensory nerves, and highly express ASIC3. Example

[0016] 1. Behavioral testing of mice after local injection of A-317567 into the tumor.

[0017] Control group: Starting 7 days after establishing the mouse tumor model, 20 μL of 0.01% DMSO was injected into the tumor once a day for a total of 14 days.

[0018] Group A-317567: 20 μL of 20 μM A-317567 was injected into the tumor once a day for 14 days, starting 7 days after the establishment of the mouse tumor model.

[0019] Mechanical pain threshold: Mice were acclimatized in an elevated metal mesh animal test cage for 2 hours. A von Frey fiber (0.16 g-2.00 g) was used perpendicularly to the test area (the sole of the hind paw) of each animal. The same force was applied each time to bend the fiber for 1-2 seconds, with a 10-second interval between two measurements. If the animal retracted its paw, shook its paw, or licked its paw, it indicated that the animal felt pain and was marked with "×". If the animal's paw did not change, it indicated that the animal did not feel pain and was marked with "o". After 6 measurements, the corresponding mechanical pain threshold was obtained according to the scale. Animals with a pain threshold of 0.98-1.16 g were selected for the experiment.

[0020] 2. The tumor-suppressing effect (tumor volume and weight) was assessed 14 days after local injection of A-317567 into the tumor.

[0021] After perfusing physiological saline into both groups of mice, tumor tissue was removed, weighed using an electronic scale, and its volume was calculated using the water displacement method.

[0022] like Figure 2 As shown, the results indicate that local injection of A-317567 can inhibit the proliferation of breast cancer in mice.

Claims

1. Application of ASIC3 inhibitors in the preparation of tumor therapeutic drugs.

2. The application according to claim 1, characterized in that, The ASIC3 inhibitor is A-317567.

3. The application according to claim 2, characterized in that, The tumor is breast cancer.

4. The application according to claim 3, characterized in that, The ASIC3 inhibitor A-317567 inhibits tumor growth by blocking CGRP signaling in sensory nerves.

5. The application according to claim 4, characterized in that, The ASIC3 inhibitor A-317567 is administered via intratumoral injection.

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