Use of aminoquinolines for the preparation of a medicament for the treatment of hyperparathyroidism

Abstract

This invention relates to the field of pharmaceutical biotechnology, providing the application of aminoquinoline compounds (such as hydroxychloroquine and chloroquine) in the preparation of drugs for treating hyperparathyroidism. These compounds restore the expression of calcium-sensitive receptors (CaSRs) on the cell membrane surface by inhibiting the lysosomal degradation pathway of CaSRs in parathyroid cells. Experiments have confirmed that 4-aminoquinoline compounds, used alone or in combination with calcimimetics (such as cinacalcet), significantly reduce serum PTH and calcium levels in a model of refractory secondary hyperparathyroidism (SHPT) and inhibit glandular hyperplasia. In particular, the combination therapy regimen exhibits a significant synergistic effect, effectively reversing calcimimetics resistance. This invention is the first to discover a novel use for the antimalarial / antirheumatic drug hydroxychloroquine in the treatment of hyperparathyroidism, realizing a new use for an existing drug; and providing a safe and effective new drug treatment strategy for clinically refractory SHPT.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical biotechnology, specifically to the application of aminoquinoline compounds in the preparation of drugs for treating hyperparathyroidism. Background Technology

[0002] Secondary hyperparathyroidism (SHPT) is the most common and serious complication in end-stage renal disease patients undergoing dialysis. It is characterized by nodular hyperplasia of the parathyroid glands, excessive PTH secretion, and calcium and phosphorus metabolism disorders, leading to bone disease, vascular calcification, and a significantly increased risk of cardiovascular death. Downregulation of membrane CaSR expression is a key pathological mechanism in the progression of SHPT and resistance to calcimimetic agents, highlighting the urgent clinical need for treatments that fundamentally restore CaSR function.

[0003] Current clinical practice primarily uses calcimimetic agents (such as cinacalcet) to allosterically activate CaSR to inhibit PTH secretion, supplemented by vitamin D and phosphate binders to control metabolic indicators; for patients who do not respond to drug therapy, only high-risk parathyroidectomy can be performed. Existing treatments all revolve around "activating residual CaSR" and do not intervene in the mechanism of CaSR loss.

[0004] Current treatments cannot block the excessive internalization and lysosomal degradation of CaSR. The continuous loss of membrane receptors leads to a decline in the efficacy of calcimimetic agents or even complete drug resistance. High-dose medications are prone to causing hypocalcemia, gastrointestinal reactions, and rapid rebound after drug withdrawal. At the same time, they cannot effectively inhibit glandular nodular hyperplasia. Ultimately, most patients can only undergo surgery, which has high risks and many complications. Clinically, there is a lack of safe and effective drugs to restore CaSR from the root and reverse drug resistance. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a drug and its application for treating hyperparathyroidism. It solves the problem that current treatments for secondary hyperparathyroidism rely solely on activating CaSR, failing to prevent its excessive internalization and lysosomal degradation, leading to widespread calcimimetic drug resistance. High-dose medications cause significant side effects, rebound effects upon discontinuation, and the inability to inhibit glandular hyperplasia, ultimately requiring high-risk surgery for most patients. Clinically, there is a lack of safe and effective drugs that fundamentally restore CaSR expression and reverse drug resistance.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution: the application of aminoquinoline compounds in the preparation of drugs for treating hyperparathyroidism, wherein the compounds restore the expression level of cell membrane CaSR by inhibiting the endocytosis and lysosomal degradation of calcium-sensitive receptors (CaSR), thereby inhibiting excessive secretion of parathyroid hormone (PTH), reducing blood calcium, and reversing calcimimetic drug resistance.

[0007] Preferably, the drug uses a 4-aminoquinoline compound or a pharmaceutically acceptable salt thereof as its sole active ingredient.

[0008] Preferably, the 4-aminoquinoline compound is at least one of hydroxychloroquine and chloroquine, and pharmaceutically acceptable salts include sulfates and phosphates.

[0009] Preferably, the dosage form of the drug is selected from oral solid dosage forms, oral liquid dosage forms, or injections.

[0010] Preferably, the drug further comprises at least one calcimulator, wherein the 4-aminoquinoline compound or a pharmaceutically acceptable salt thereof is used in combination with the calcimulator.

[0011] Preferably, the calcium-simulating agent is selected from at least one of cinacalcet, etalcalcetide, evocratet, upacicalcet, or a pharmaceutically acceptable salt thereof.

[0012] Preferably, the mass ratio of hydroxychloroquine to the calcium aspirin is 1:1 to 20:1.

[0013] Preferably, the pharmaceutically acceptable salt of hydroxychloroquine is selected from hydroxychloroquine sulfate or hydroxychloroquine phosphate.

[0014] Aminoquinoline compounds are used in the preparation of drugs for the treatment of hyperparathyroidism, which include pharmaceutically acceptable excipients and active ingredients. The active ingredients include hydroxychloroquine or pharmaceutically acceptable salts thereof and calcium mimics.

[0015] Preferred for non-surgical conservative treatment of chronic kidney disease patients undergoing dialysis, patients with relapse after SHPT, and elderly or high-risk patients who are not eligible for surgery.

[0016] This invention provides the application of aminoquinoline compounds in the preparation of drugs for treating hyperparathyroidism. It has the following beneficial effects: 1. This invention is the first to reverse calcimimetic drug resistance at its root cause. By inhibiting the degradation of CaSR lysosomes, the number of membrane receptors is restored, making resistant SHPTs sensitive to calcimimetic drugs again, thus breaking through the bottleneck of traditional treatment.

[0017] 2. The present invention produces a significant synergistic effect when used in combination with calcimimetic agents, which can more effectively reduce PTH and blood calcium, reduce the dosage of calcimimetic agents, and reduce the risk of side effects.

[0018] 3. This invention directly inhibits the proliferation of parathyroid cells and glandular hyperplasia, delays or even reverses nodular hyperplasia, and reduces the need for surgery.

[0019] 4. The present invention has a clear target and mechanism, blocks only pathological pathways without affecting normal physiological regulation, and has high safety and is suitable for long-term maintenance.

[0020] 5. This invention enables the repurposing of existing drugs, allowing marketed medications to be rapidly used for refractory SHPT, providing a new non-surgical option for high-risk patients without effective medical interventions. Attached Figure Description

[0021] Figure 1 This invention presents a single-cell transcriptome profile and cellular heterogeneity of parathyroid diseases, wherein: AC, Unified manifold approximation and projection (UMAP) plots of all cells after quality control filtering, colored according to (A) sample origin and (B) main cell type; (C) The violin diagram shows the expression levels of classic marker genes used to identify major cell lineages, including epithelial cells, endothelial cells (ECs), parietal cells, and immune cells; DE, transcriptomic heterogeneity of parathyroid epithelial cells and expression of CaSR in SHPT cells; (D) Re-clustered UMAP visualization of epithelial cells, identifying six distinct subpopulations (Ep #1–#6). (E) The stacked bar chart shows the proportion of the six epithelial cell subpopulations in each sample.

[0022] F, a UMAP visualization of different evolutionary fates of epithelial cells and functional divergences of pathological eosinophils, overlaid with pseudotime values ​​defined by Monocle3 analysis.

[0023] Figure 2 This invention relates to the dynamic activation of the endocytosis-lysosome pathway during epithelial cell differentiation, wherein: A. The line graph shows the trend of UCell scores for the specified pathways in cells arranged in a pseudo-chronological order; B. Expression trends of key genes involved in the endocytosis-lysosome pathway (CLTB, AP2B1, RAB7A, VAMP2) with pseudo-time changes; Figure 3 Representative immunohistochemical (IHC) staining images of CaSR protein expression in normal parathyroid gland and SHPT tissues; Figure 4 Immunofluorescence (IF) staining localization and expression images of CaSR in normal and SHPT-derived primary parathyroid cells in this invention; Figure 5 The image shows the restoration of CaSR membrane localization and function in SHPT cells by inhibiting endocytosis and lysosomal pathways in this invention, wherein: A. Immunofluorescence (IF) staining of CaSR (green) in primary SHPT cells treated with HCQ or CPZ showed restoration of membrane expression; B. Western blot analysis of ERK phosphorylation (pERK) levels in primary SHPT cells that received the specified treatment; Treatment conditions: HCQ, 20 μM HCQ treatment for 24 hours; Cina, 30 nM Cinacalcet treatment for 24 hours; Ca2+, 2 mM calcium carbonate treatment for 2 hours; Quantitative analysis of (C) PTH mRNA levels and (D) secretory PTH concentrations under CD and specified drug treatments; Figure 6 This is a schematic diagram of the experimental design of the PDX model in this invention; Figure 7 The images shown are of HCQ and cinacalcet synergistically alleviating SHPT progression in a PDX mouse model, as described in this invention. Dynamic changes in serum iPTH levels in A.PDX mice during a four-week treatment period; B. Time-course analysis of serum calcium (left) and serum phosphorus (right) levels during treatment; Figure 8 The HCQ method is used to recover the membrane localization image of CaSR in the PDX mouse model of this invention; Figure 9 This image shows the synergistic inhibition of SHPT tissue cell proliferation by HCQ and cinacalcet in this invention. Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] In a first aspect, the present invention provides the use of a 4-aminoquinoline compound or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating hyperparathyroidism.

[0026] In a preferred embodiment of the present invention, the 4-aminoquinoline compound is selected from at least one of hydroxychloroquine, chloroquine and its derivatives.

[0027] In one specific embodiment of the present invention, pharmaceutically acceptable salts include, but are not limited to, sulfates, phosphates, and hydrochlorides; preferably selected from hydroxychloroquine sulfate, hydroxychloroquine phosphate, or chloroquine phosphate. Since hydroxychloroquine and chloroquine have the same pharmacological core structure and lysosomal inhibition mechanism, they have equivalent technical effects in the present invention.

[0028] In a preferred embodiment of the present invention, the hyperparathyroidism is secondary hyperparathyroidism (SHPT).

[0029] More preferably, the secondary hyperparathyroidism is refractory or calcimimetic-resistant secondary hyperparathyroidism. This invention has found that 4-aminoquinoline compounds can effectively reverse the resistance state in patients who have developed resistance to calcimimetic agents due to downregulation of membrane CaSR expression.

[0030] The drug application of this invention includes two regimens: monotherapy and combination therapy. Option 1 (Monotherapy): The drug uses 4-aminoquinoline compounds or their pharmaceutically acceptable salts as the sole active ingredient.

[0031] Experimental results show that hydroxychloroquine alone can significantly increase the CaSR level of parathyroid cell membranes and effectively reduce serum PTH and blood calcium concentrations in animal models, thereby inhibiting glandular hyperplasia.

[0032] Option 2 (Combination Therapy): The drug also contains at least one calcimimetic agent, 4-aminoquinoline compounds or their pharmaceutically acceptable salts, used in combination with the calcimimetic agent.

[0033] "Combined drug use" includes preparing two active ingredients into a compound preparation, as well as administering two active ingredients as independent preparations simultaneously or sequentially.

[0034] In a preferred embodiment of the present invention, the calcium mimic is selected from compounds capable of allosterically activating CaSR, specifically including but not limited to: at least one of cinacalcet, etalcalcetide, evoclcet, upacicalcet, or a pharmaceutically acceptable salt, ester, solvate, or prodrug thereof.

[0035] This invention has found that when 4-aminoquinoline compounds are used in combination with calcium-like agents, the two exhibit a significant synergistic effect at a specific mass ratio.

[0036] Specifically, the preferred mass ratio of hydroxychloroquine to calcimimetic agents is 1:1 to 20:1; more preferably 2:1 to 16:1; and even more preferably 4:1 to 10:1. Within this ratio range, hydroxychloroquine significantly amplifies the pharmacological effect of calcimimetic agents activating receptors ("throttling") by increasing the number of membrane receptors ("open source"), thereby achieving a therapeutic effect of "1+1>2", enabling the achievement of ideal PTH control targets at lower doses of calcimimetic agents.

[0037] In one embodiment of the present invention, the dosage form of the drug includes oral formulations and non-oral formulations.

[0038] Specifically, the dosage form is selected from oral solid dosage forms (such as tablets, capsules, granules), oral liquid dosage forms (such as dry suspensions, oral liquids), or injections (such as injections, lyophilized powder injections).

[0039] When the calcium-simulating agent is cinacalcet or evokalcet, it is preferred to prepare it as an oral solid dosage form; When the calcium mimic is etakatide or upacicalcet, it is preferred to prepare it as an injection.

[0040] In a second aspect, the present invention provides a pharmaceutical composition for treating hyperparathyroidism.

[0041] The pharmaceutical composition contains pharmaceutically acceptable excipients and active ingredients; the active ingredients include hydroxychloroquine or its pharmaceutically acceptable salts and calcimidines (such as cinacalcet, itcatide, evokalcet, etc.).

[0042] The preferred types, proportions, and dosage forms of the components in the pharmaceutical composition are consistent with those described in the above applications.

[0043] The following is a description with reference to specific embodiments: Please see the appendix Figure 1 - Appendix Figure 5 To elucidate the microenvironmental characteristics of hyperparathyroidism (HPT) driven by different pathogenic mechanisms, we constructed a high-resolution single-cell transcriptome atlas from 14 human parathyroid specimens, including 1 case of parathyroid carcinoma (PC), 2 cases of parathyroid adenoma (PA), and 4 cases of secondary hyperparathyroidism (SHPT). After rigorous quality control, standardization, and dimensionality reduction, the final dataset contained 178,459 single cells (…). Figure 1 A). The main categories are distinguished by annotations for epithelial cells, endothelial cells, parietal cells, and immune cells, etc. Figure 1 B, C). Epithelial cells from PC, PA, and SHPT were divided into six distinct subclusters (B, C). Figure 1 D, E). To reveal the disease dynamics of HPT progression, we constructed single-cell trajectories of epithelial cells (D, E). Figure 1 F). UCell scores for pathways such as "clathrin-dependent endocytosis" and "lysosomal tissue regulation," as well as related differentially expressed genes (CLTB, RAB7A, AP2B1, VAMP2), gradually increased along the pseudo-time trajectory. Figure 2 The above data suggest that during the progression of PA and SHPT diseases, clathrin-mediated endocytosis gradually increases, which may promote disease progression by internalizing surface CaSR.

[0044] Please see the appendix Figure 6 - Appendix Figure 9 To validate this in vivo, we subcutaneously implanted fresh parathyroid tissue fragments from five different SHPT donors into immunodeficient nude mice. Mice with successful modeling were randomly assigned to four groups: a vehicle control group, a cinacalcet group, an HCQ group, or a combination therapy group, thus establishing a patient-derived xenograft (PDX) model. Subsequently, these recipient mice received a 4-week treatment regimen, and serum biochemical analysis and tissue collection were performed to evaluate the therapeutic effects of HCQ monotherapy or in combination with cinacalcet. Figure 6 Longitudinal monitoring of serum whole-segment human parathyroid hormone (iPTH) levels showed that iPTH levels remained elevated in the solvent control mice after transplantation. Notably, some PDX grafts exhibited resistance to cinacalcet monotherapy, maintaining high levels of iPTH. Figure 7 A). In contrast, iPTH levels in both the HCQ monotherapy group and the combination therapy group began to progressively decrease after 2–3 weeks of intervention, reaching their lowest values ​​at the study endpoint. Statistical analysis confirmed that iPTH levels in the combination therapy group were significantly lower than those in the control group and the cinacalcet monotherapy group. Simultaneously, both HCQ monotherapy and the combination regimen significantly reduced serum calcium levels, while no significant differences in serum phosphorus levels were observed among the treatment groups. Figure 7 B); Subsequently, we performed immunohistochemical (IHC) staining of the collected PDX grafts for CaSR and the proliferation marker Ki67. We found that, compared with the control group and the cinacalcet group: Membrane-localized CaSR expression was significantly restored in both the HCQ monotherapy group and the combination therapy group. Figure 8Representative IHC staining images of CaSR in experimental animal graft tissues. Scale bar: 50 μm. Quantitative analysis of the mean optical density of membrane CaSR. Cina, cinacalcet; Combo. Data are presented as mean ± standard deviation (SD). *, P < 0.05; **, P < 0.01.

[0045] In mice treated with HCQ alone or in combination with cinacalcet, the positivity rate of Ki67 was significantly reduced. Figure 9 Representative IHC staining images of Ki67 in graft tissues of experimental animals. Scale bar: 50 μm. Quantitative analysis of Ki67 positivity rate. Cina, cinacalcet; Combo. Data are presented as mean ± standard deviation (SD). *, P < 0.05; **, P < 0.01; ***, P < 0.001.

[0046] In summary, these findings indicate that HCQ significantly enhances the therapeutic effect of cinacalcet by restoring membrane CaSR expression, thereby inhibiting parathyroid cell proliferation and PTH secretion.

[0047] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. Use of an aminoquinoline compound for the manufacture of a medicament for the treatment of hyperparathyroidism, characterized in that, The compound restores cell membrane CaSR expression levels by inhibiting endocytosis and lysosomal degradation of calcium-sensitive receptors (CaSR), thereby inhibiting excessive secretion of parathyroid hormone (PTH), reducing blood calcium, and reversing calcimimetic drug resistance.

2. The use of the aminoquinoline compound according to claim 1 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The drug uses 4-aminoquinoline compounds or their pharmaceutically acceptable salts as its sole active ingredient.

3. The use of the aminoquinoline compound according to claim 1 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The 4-aminoquinoline compounds are at least one of hydroxychloroquine and chloroquine, and pharmaceutically acceptable salts include sulfates and phosphates.

4. The use of the aminoquinoline compound according to claim 1 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The dosage form of the drug is selected from oral solid dosage forms, oral liquid dosage forms, or injections.

5. The use of the aminoquinoline compound according to claim 4 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The drug also contains at least one calcimulator, wherein the 4-aminoquinoline compound or a pharmaceutically acceptable salt thereof is used in combination with the calcimulator.

6. The use of the aminoquinoline compound according to claim 5 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The calcium-simulating agent is selected from at least one of cinacalcet, etalcalcetide, evocratet, upacicalcet, or a pharmaceutically acceptable salt thereof.

7. The use of the aminoquinoline compound according to claim 6 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The mass ratio of hydroxychloroquine to the calcium apothecium is 1:1 to 20:

1.

8. The use of the aminoquinoline compound according to claim 1 in the preparation of a drug for treating hyperparathyroidism, characterized in that, The pharmaceutically acceptable salt of hydroxychloroquine is selected from hydroxychloroquine sulfate or hydroxychloroquine phosphate.

9. The use of aminoquinoline compounds in the preparation of drugs for treating hyperparathyroidism, as described in any one of claims 1-8, in the preparation of drugs for treating hyperparathyroidism, characterized in that... It contains pharmaceutically acceptable excipients and active ingredients; The active ingredients include hydroxychloroquine or pharmaceutically acceptable salts thereof and calcium mimics.

10. The aminoquinoline compound according to claim 9 in the preparation of a drug for treating hyperparathyroidism, characterized in that, It is used for non-surgical conservative treatment of patients with chronic kidney disease undergoing dialysis, patients with relapse after SHPT, and elderly or high-risk patients who are not eligible for surgery.

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