Application of spermidine and composition thereof in preparation of medicine for preventing or treating radioactive diseases and related complications thereof

Abstract

The invention discloses application of spermidine and a composition thereof in preparation of medicines for preventing or treating radioactive diseases and related complications thereof. According to the application, firstly, an in-vitro experiment shows that spermidine can be used for remarkably relieving proliferation arrest of radiation-induced human colonic dry organs and inhibiting apoptosis of colonic epithelium organs; furthermore, animal in-vivo experiments show that spermidine can repair radioactive injury, including enteritis, intestinal injury, intestinal fibrosis and the like caused by radiation; the application firstly proposes and proves that spermidine can be used as a radiation protective agent and a safe medicine component for treating radioactive injury, and has a good clinical application prospect and application value.

Description

Technical Field

[0001] This application relates to the use of spermidine and its compositions in the preparation of medicaments for the prevention or treatment of radiation-induced diseases and related complications, and belongs to the field of biomedical technology. Background Technology

[0002] Radiotherapy, one of the three traditional methods of cancer treatment, primarily utilizes high-energy ionizing radiation such as X-rays and gamma rays to kill cancer cells. More than 60% of cancer patients require radiotherapy as a primary or adjuvant treatment. Its mechanism of action involves direct ionization and ROS-mediated indirect damage, disrupting the DNA, proteins, and other biomolecules of tumor cells, thereby achieving cell killing and shrinking the tumor. However, this indiscriminate attack characteristic can simultaneously damage adjacent normal tissues, leading to serious radiation-related complications. Radiation damage exhibits a two-stage pathological characteristic: the acute phase is dominated by oxidative stress, senescence, and apoptosis in rapidly proliferating cells (such as epithelial cells), triggering an acute inflammatory response in tissues; the chronic phase, due to abnormal repair, leads to progressive fibrosis, microvascular disease, and multiple organ dysfunction, significantly negatively impacting the patient's treatment progress and prognosis.

[0003] Radiation damage caused by different irradiation sites exhibits organ-specific characteristics: 1) High-dose whole-body irradiation (>60% of body surface area) primarily induces myelosuppression syndrome; 2) Head and neck radiotherapy often leads to skin ulcers, oral mucositis, salivary gland dysfunction, and nerve damage; 3) Chest and abdominal treatment easily causes esophagitis, radiation pneumonitis, and liver damage; 4) Pelvic irradiation is often accompanied by enteritis, cystitis, and reproductive system damage. These complications not only significantly reduce patients' quality of life but also affect the efficacy of radical tumor treatment due to the need for controlled radiation doses.

[0004] Currently, there are no effective interventions for the prevention of radiation injury in clinical practice. While amifostine, the only approved radiation protectant, can relieve radiation-related dry mouth and oral mucositis in the head and neck region, its short half-life (approximately 8 minutes) and narrow therapeutic window (requiring strict administration 30 minutes before radiotherapy) significantly limit its clinical application. Simultaneously, there is a lack of corresponding therapies for the clinical treatment of acute or chronic radiation injury and its complications, and no officially approved effective drugs exist. Developing drugs with both broad-spectrum preventative efficacy and good safety profile has become a key scientific issue in improving the efficacy of tumor radiotherapy. Summary of the Invention

[0005] The purpose of this invention is to address the key technical problem of developing drugs that have both radiation damage prevention and treatment efficacy and good safety, which is crucial for reducing radiotherapy complications and improving the prognosis of tumor radiotherapy. This application aims to provide the use of spermidine and its compositions in the preparation of drugs for the prevention or treatment of radiation diseases and their related complications.

[0006] To achieve the above objectives, this application adopts the following technical solution: This application provides for the use of spermidine and / or its pharmaceutically acceptable salt forms in the preparation of medicaments for the prevention and / or treatment of radiation sickness or its related complications.

[0007] In some embodiments, the radiation sickness or its related complications are injuries or complications induced by exposure to ionizing radiation, the radiation source of which includes one or more of X-ray radiation, gamma-ray radiation, radionuclide radiation, electron radiation, neutron radiation, and proton radiation.

[0008] In some specific implementations, the radiation sickness or its related complications include radiation damage or related complications caused by radiotherapy for tumors.

[0009] In some implementations, the radiation damage or related complications include: 1) organ-specific damage, inflammation, and chronic tissue fibrosis; 2) skin and mucous membrane reactions; 3) myelosuppression syndrome; 4) systemic reactions; and 5) other local reactions.

[0010] In some implementations, the organ-specific damage, inflammation, and chronic tissue fibrosis include: 1) intestinal damage, radiation enteritis, and intestinal fibrosis, specifically manifested as diarrhea, abdominal pain, hematochezia, and intestinal obstruction; 2) radiation-induced lung damage, radiation-induced pneumonia, and pulmonary fibrosis, specifically manifested as cough, shortness of breath, and fever; 3) radiation-induced esophageal damage, radiation-induced esophagitis, and esophageal fibrosis; 4) radiation-induced bladder damage, radiation-induced reproductive system damage, and related tissue fibrosis; 5) radiation-induced liver damage, radiation-induced kidney damage, and liver / kidney tissue fibrosis. The skin and mucous membrane reactions include: 1) redness, dryness, peeling, and itching of the skin in the irradiated area, and in severe cases, wet peeling and ulceration; 2) oral mucositis, salivary gland dysfunction, dry mouth, ulceration, and painful swallowing caused by head and neck radiotherapy. The bone marrow suppression syndrome includes: a decrease in white blood cells, platelets, and red blood cells, and the resulting decrease in immunity, susceptibility to infection, easy bleeding, and anemia; The systemic reactions mainly include immunosuppression and nerve damage, manifested as fatigue, loss of appetite, nausea and vomiting, headache, and dizziness. Other local reactions include: 1) radiation-induced bone injury; 2) brain radiation-related complications, including headache, nausea, limb weakness, and seizures; and 3) head and neck radiation-related complications, including altered taste and difficulty opening the mouth.

[0011] In some embodiments, the drug comprises a pharmaceutically effective ingredient and pharmaceutically acceptable excipients, wherein the pharmaceutically effective ingredient includes spermidine.

[0012] In some implementations, the active ingredient is spermidine as the sole active ingredient.

[0013] In some embodiments, the active ingredient further comprises a second therapeutic agent and / or adjuvant ingredient other than spermidine.

[0014] The second therapeutic agent includes one or more of the following: antibiotics, antiviral drugs, immunosuppressants, peptide drugs, hormonal drugs, anti-inflammatory drugs, exosomes, and nucleic acid drugs; The auxiliary ingredients include probiotics and / or prebiotics.

[0015] In some embodiments, the pharmaceutically acceptable excipient is one or more of the following: diluent, binder, wetting agent, lubricant, disintegrant, solvent, emulsifier, cosolvent, preservative, pH adjuster, osmotic pressure adjuster, surfactant, coating material, antioxidant, and buffer.

[0016] In some embodiments, the dosage form of the drug is an oral formulation, an injectable formulation, an inhaled formulation, or a topical formulation (including topical solid, semi-solid, or liquid formulations).

[0017] Compared with the prior art, this application has the following beneficial effects: This application demonstrates through in vitro and in vivo experiments that spermidine can repair radiation damage caused by radiotherapy. In specific embodiments, by constructing radiation enteritis models and colorectal cancer-radiotherapy models, it is shown that spermidine can help alleviate colorectal cancer and promote intestinal repair; it can also alleviate fibrosis caused by chronic radiation damage to the intestine, showing the potential for "one drug with multiple uses". Spermidine is an endogenous polyamine in the human body with reliable safety and has been widely used in various health care products; it can be used as a radiation protectant and a safe drug component for treating radiation damage, showing good application prospects and facilitating rapid translation. Attached Figure Description

[0018] Figure 1 This is a diagram showing the reduction of radiation-induced organoid proliferation arrest in the human colonic stem in Example 1 of this invention by spermidine.

[0019] Figure 2 This is a diagram showing how spermidine reduces radiation-induced apoptosis in human colonic epithelial organoids in Example 1 of the present invention.

[0020] Figure 3 This is a diagram illustrating the effect of spermidine on the proliferation of human colonic organoids in Example 1 of the present invention.

[0021] Figure 4 This is a diagram showing the reduction of radiation-induced proliferation arrest of different human-derived colonic stem organoids by spermidine in Example 1 of the present invention.

[0022] Figure 5This is a drug safety evaluation diagram of spermidine in Example 2 of the present invention.

[0023] Figure 6 This is a diagram (A) showing how spermidine alleviates radiation-induced weight loss in mice and an endoscopic image of intestinal damage (B) in Example 3 of the present invention.

[0024] Figure 7 This is a diagram showing the results of spermidine alleviating radiation-induced intestinal damage in mice in Example 3 of the present invention; wherein, A is a picture of the intestinal length measurement of mice in each group on days 4, 8 and 16 after radiation; B is the quantitative statistical result of A; C is a picture of H&E staining of colon tissue sections of mice in each group.

[0025] Figure 8 This is a diagram showing the results of spermidine synergistic radiotherapy for anti-cancer effects and promotion of intestinal repair in a mouse spontaneous tumorigenesis model of intestinal cancer in Example 4 of the present invention; wherein, A is colonoscopy images of mice in each group; B is intestinal length measurement images of mice in each group; C is the quantitative statistical results of B; and D is the statistical count of the number of intestinal tumors in mice in each group.

[0026] Figure 9 This is a diagram showing the results of spermidine synergistic radiotherapy for anti-cancer effects and promotion of intestinal repair in a mouse orthotopic colorectal cancer tumor model in Example 5 of the present invention; wherein, A is colonoscopy images of mice in each group; B is intestinal length measurement images of mice in each group; C is the quantitative statistical results of B; and D is the statistical count of the number of intestinal tumors in mice in each group.

[0027] Figure 10 This is a diagram showing how spermidine alleviates fibrosis caused by chronic radiation damage to the intestines in Example 6 of the present invention. Detailed Implementation

[0028] To make the technical solution of this application clearer and easier to understand, preferred embodiments are described in detail below with reference to the accompanying drawings.

[0029] Unless otherwise specified, the experimental methods used in the following examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0030] Spermidine (SPD) is N-(3-aminopropyl)-1,4-butanediamine, and its chemical formula is as follows:

[0031] Example 1: In vitro experiment on spermidine repairing radiation-induced colonic organoid activity 1.1 Experimental Methods To clarify the effects of spermidine on radiation-induced colonic organoids, intestinal stem organoids or intestinal epithelial organoids were first digested into single cells and evenly seeded into 96-well plates. These were divided into a non-irradiated group, a 6 Gy irradiated control group, and a 6 Gy spermidine-irradiated treatment group, with auxiliary wells in each group. Two hours before 6 Gy irradiation, 20 μM spermidine was added to each well. Subsequently, on days 2 and 4 of gamma ray irradiation, the morphology and viability of the organoids were monitored using a high-content imaging system, and the total ATP production of the organoids was measured using a 3D cell viability assay kit. The organoids were first incubated at 37°C in the dark for 20 min, and the chemiluminescence value of each well was measured using a microplate reader. The level of chemiluminescence reflects the cell viability of the organoids. To clarify the effect of spermidine on the proliferation of colonic stem organoids under non-radiation conditions, intestinal stem organoids were digested into single cells and evenly seeded into 96-well plates, divided into 0, 20 μM, 6 μM, 2 μM, 0.6 μM, and 0.2 μM groups, with subwells for each group, and spermidine of the corresponding concentration was added. Two days after the addition of spermidine, the morphology and viability of the organoids were monitored using a high-content imaging system, and the total ATP production of the organoids was detected using a 3D cell viability assay kit. To clarify the effect of spermidine on organoids from different human sources, organoids from two other human sources were digested into single cells and evenly seeded into 96-well plates, divided into four groups (non-irradiated group, non-irradiated spermidine group, 6 Gy irradiated control group, and 6 Gy irradiated spermidine treatment group), with subwells for each group. Two hours before 6 Gy irradiation, 20 μM spermidine was added to each well. Subsequently, on the second and fourth days of gamma-ray radiation, the morphology and viability of organoids were monitored using a high-content imager, and the total ATP production of organoids was detected using a 3D cell viability assay kit.

[0032] 1.2 Experimental Results By treating human colonic organoids and epithelial organoids with gamma rays combined with spermidine, such as Figure 1 and Figure 2 As shown, spermidine can significantly alleviate radiation-induced proliferative arrest in human colonic stem organoids. Figure 1 ), and inhibit apoptosis of colonic epithelial organoids ( Figure 2 ).like Figure 3 As shown, spermidine did not promote the proliferation of colonic stem organoids under non-radiation conditions. Figure 4 As shown, spermidine has a repair effect on the colonic organoids of different individuals.

[0033] Example 2: Animal experiments to evaluate the safety of spermidine drugs 1.1 Experimental Methods Wild-type SPF-grade male C57 mice aged 8-10 weeks were randomly divided into two groups: 1) control group, given normal drinking water; 2) control group, given 5 mM spermidine in drinking water (free access to water). The experimental endpoint was reached on day 16 after the start of modeling. Wild-type C57 mice were dissected and specimens of the heart, liver, spleen, lungs, and kidneys were collected.

[0034] 1.2 Experimental Results like Figure 5 As shown, the weights of the heart, liver, spleen, lungs, and kidneys were measured, and their safety was tested by H&E staining of the sections. The results showed that spermidine had no significant weight effect or toxicological damage on the organs of mice, and its safety was controllable.

[0035] Example 3: Animal experiment to evaluate the repair effect of spermidine on radiation-induced mouse intestine. 1.1 Experimental Methods Wild-type SPF-grade male C57 mice aged 8-10 weeks were randomly divided into four groups: 1) Control group, given regular drinking water; 2) Control group, given 5 mM spermidine in drinking water (free access to water); 3) Radiation enteritis model group, given regular drinking water; 4) Radiation enteritis model group, given 5 mM spermidine in drinking water starting one day before radiation until the experimental endpoint (free access to water), followed by 16 Gy gamma ray radiation to induce the model. The experimental endpoint was reached on days 4, 8, and 16 after the start of modeling. Detection of enteritis phenotype in radiation-induced enteritis mice: Mouse weight was monitored and recorded daily; on days 4, 8, and 16, small animal endoscopy was used to observe redness, swelling, ulceration, and bleeding of the intestinal mucosa. At the experimental endpoint, mice were anesthetized and euthanized, and intestinal tissue was collected. Wild-type C57 mice were dissected, and colon tissue specimens were collected. Intestinal length was measured, and sections were fixed and embedded in 4% paraformaldehyde. H&E staining was performed on the sections.

[0036] 1.2 Experimental Results like Figure 6 As shown in Figure A, after radiation-induced modeling, mice treated with spermidine showed significant relief from weight loss symptoms. Figure 6 As shown in Figure B, intestinal endoscopy of mice revealed that continuous administration during the recovery period of enteritis significantly improved enteritis in mice. Figure 7 As shown in Figures A and B, on days 4, 8, and 16 post-radiation, the intestinal length of mice in the spermidine-treated group showed significant recovery compared to the irradiated control group. Pathological sections revealed that spermidine significantly alleviated radiation-induced enteritis phenotypes, including intestinal mucosal damage, intestinal muscle layer loosening, and submucosal immune infiltration. Figure 7 As shown in Figure C. Based on the constructed mouse model of acute radiation enteritis, the efficacy of spermidine in vivo treatment was preliminarily confirmed. Speridine significantly promoted the reduction of inflammation and intestinal repair in radiation enteritis.

[0037] Example 4: Evaluation of the synergistic effect of spermidine on radiotherapy in cancer prevention and intestinal repair in a mouse model of spontaneous intestinal cancer. 1.1 Experimental Methods First, a spontaneous tumorigenesis model of intestinal cancer was established in mice: C57 / BL mice were intraperitoneally injected with AOM (azoxymethane) 10 mg / kg for one week, then fed 2% DSS (dextran sodium sulfate) for 7 days with water changes for 14 days; after 3 cycles, the mice were randomly divided into 4 groups: a non-radiated normal drinking water group, a non-radiated spermidine 5 mM group, a radiation-induced normal drinking water group, and a radiation-induced spermidine 5 mM group. The mice were given 5 mM spermidine in their drinking water starting one day before radiation until the experimental endpoint (free access to water), followed by 8 Gy gamma ray radiation for modeling, and a second 8 Gy radiation model on day 4. On day 8 after modeling, the intestinal mucosa of the mice with spontaneous tumorigenesis was observed using a small animal endoscope to observe redness, swelling, ulceration, and bleeding. At the experimental endpoint, the mice were anesthetized and euthanized, and intestinal tissue was collected to measure the length and weight of the intestine, as well as the size and number of tumors within the intestine. Sections were fixed and embedded in 4% paraformaldehyde, and H&E staining was used to assess the enteritis phenotype and the repair of intestinal epithelial damage.

[0038] 1.2 Experimental Results We conducted in situ tumor-bearing experiments in mice and performed radiotherapy simulations, during which spermidine intervention was administered or not. Figure 8 As shown in AC, the spermidine administration group significantly reduced the radiation-induced enteritis phenotype, including improvements in intestinal length observed endoscopically in mice. In contrast, under radiation therapy conditions... Figure 8 As shown in Figure D, the growth of tumors and the number of tumors at sample collection were significantly reduced in the spermidine-treated group compared to the control group. Example 5: Evaluation of the synergistic effect of spermidine on radiotherapy in cancer prevention and intestinal repair in a mouse orthotopic colorectal cancer tumor-bearing model.

[0039] 1.1 Experimental Methods First, an in situ tumor-bearing model of intestinal cancer was established in mice: using a microinjection microscope, 1.5 × 10⁻⁶ tumors were injected... 6MC38 cells were seeded into the rectum and anus of C57 mice, with an injection volume of approximately 10 µL each time. After two weeks of rearing, a radiation-induced enteritis model was established. Mice were divided into a radiation-induced group with normal drinking water and a group receiving 5 mM spermidine. One day before radiation, mice were given 5 mM spermidine in their drinking water until the experimental endpoint (free access to water). Then, 8 Gy gamma ray radiation was used to establish the model, and a second 8 Gy radiation model was performed on day 4. On day 8, small animal endoscopy was used to observe the redness, swelling, ulceration, and bleeding of the intestinal mucosa. At the experimental endpoint, mice were anesthetized and euthanized, and intestinal tissue was collected to measure intestinal length and weight, as well as the size and weight of intestinal tumors. Sections were fixed and embedded in 4% paraformaldehyde, and H&E staining was used to assess the enteritis phenotype and the extent of intestinal epithelial damage repair.

[0040] 1.2 Experimental Results We conducted in situ tumor-bearing experiments in mice and performed radiotherapy simulations, during which spermidine intervention was administered or not. Figure 9 As shown in AC, the spermidine administration group significantly reduced the radiation-induced enteritis phenotype, including improvements in intestinal length observed endoscopically in mice. In contrast, under radiation therapy conditions... Figure 9 As shown in Figure D, the growth of tumors and the tumor weight at sample collection were significantly reduced in the spermidine-treated group compared to the control group. Example 6: Animal experiment to evaluate the effect of spermidine in alleviating intestinal fibrosis caused by chronic radiation damage. 1.1 Experimental Methods First, a chronic radiation enteritis model was established in mice: 8-10 week old C57 mice were randomly divided into 3 groups: 1) Control group, given regular drinking water; 2) Chronic radiation enteritis model group, given regular drinking water; 3) Chronic radiation enteritis model group, given 5 mM spermidine in drinking water (free access to water). The experimental endpoint was reached at week 8 after the start of modeling. At week 8, small animal endoscopy was used to observe redness, swelling, ulceration, and bleeding of the mouse intestinal mucosa. At the experimental endpoint, mice were anesthetized and sacrificed, intestinal tissue was collected, and the length and weight of the intestine were measured; sections were fixed and embedded in 4% paraformaldehyde, and the enteritis phenotype and intestinal fibrosis repair were assessed using indicators such as Masson staining.

[0041] 1.2 Experimental Results Within 8 weeks after the establishment of a chronic radiation enteritis model, spermidine or a solvent control drug was administered to observe the effect of spermidine on intestinal repair. Figure 10 The results shown in AB revealed that the intestinal tract of mice in the spermidine intervention group significantly increased compared to the control group, returning to the level of the non-radioactive control group. Furthermore, as... Figure 10 As shown in Figure C, compared to the control group, the spermidine intervention group showed less intestinal fibrosis. These experiments suggest that spermidine administration can alleviate chronic radiation enteritis in mice.

[0042] The above description is merely a preferred embodiment of this application and is not intended to limit this application in any form or substance. It should be noted that those skilled in the art can make several improvements and additions without departing from this application, and these improvements and additions should also be considered within the scope of protection of this application.

Claims

1. The use of spermidine and / or its pharmaceutically acceptable salt forms in the preparation of medicaments for the prevention and / or treatment of radiation sickness or its related complications.

2. The application according to claim 1, characterized in that, The radiation sickness or its related complications are injuries or complications induced by exposure to ionizing radiation, the radiation source of which includes one or more of X-ray radiation, gamma-ray radiation, radionuclide radiation, electron radiation, neutron radiation, and proton radiation.

3. The application according to claim 1, characterized in that, The radiation-induced damage or related complications include: 1) organ-specific damage, inflammation and chronic tissue fibrosis; 2) skin and mucous membrane reactions; 3) myelosuppression syndrome; 4) systemic reactions; and 5) other local reactions.

4. The application according to claim 3, characterized in that, The organ-specific injuries, inflammations, and chronic tissue fibrosis include: 1) intestinal injury, radiation enteritis, and intestinal fibrosis; 2) radiation lung injury, radiation pneumonia, and pulmonary fibrosis; 3) radiation esophageal injury, radiation esophagitis, and esophageal fibrosis; 4) radiation bladder injury, radiation reproductive system injury, and related tissue fibrosis; and 5) radiation liver injury, radiation kidney injury, and liver / kidney tissue fibrosis. The skin and mucous membrane reactions include: 1) redness, dryness, peeling, itching, pruritus, and ulceration of the skin in the irradiated area; 2) oral mucositis, salivary gland dysfunction, dry mouth, ulceration, and painful swallowing caused by head and neck radiotherapy; The systemic response includes immunosuppression and neurological damage; Other local reactions include: 1) radiation-induced bone injury; 2) brain radiation-related complications, including headache, nausea, limb weakness, and seizures; and 3) head and neck radiation-related complications, including altered taste and difficulty opening the mouth.

5. The application according to claim 1, characterized in that, The drug comprises a pharmaceutically effective ingredient and pharmaceutically acceptable excipients, wherein the pharmaceutically effective ingredient includes spermidine.

6. The application according to claim 5, characterized in that, The active ingredient also includes a second therapeutic agent and / or adjuvant ingredient other than spermidine.

7. The application according to claim 5, characterized in that, The pharmaceutically acceptable excipients are one or more of the following: diluents, binders, wetting agents, lubricants, disintegrants, solvents, emulsifiers, cosolvents, preservatives, pH adjusters, osmotic pressure adjusters, surfactants, coating materials, antioxidants, and buffers.

8. The use according to any one of claims 1 to 7, characterized in that, The dosage form of the drug is an oral preparation, an injectable preparation, an inhaled preparation, or a topical preparation.

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