LN-isobutyrylcysteine ​​(L-NIBC)-bound gold cluster Au18(L-NIBC)14, its composition and uses

Abstract

LN-isobutyrylcysteine ​​(L-NIBC)-bound gold cluster Au 18 (L-NIBC) 14 , multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), or glaucoma, and its use for the manufacture of a medicament for the treatment of multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), or glaucoma. Compositions and methods for treating multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), or glaucoma.

Description

[Technical field]

[0001] The present invention relates to a gold cluster (AuC), in particular a LN-isobutyrylcysteine ​​(L-NIBC)-conjugated gold cluster molecule AuC. 18 (L-NIBC) 14 , gold cluster molecule Au 18 (L-NIBC) 14 A composition comprising a gold cluster molecule Au for preparing a medicament for the treatment of various diseases. 18 (L-NIBC) 14 The use of gold cluster molecules Au 18 (L-NIBC) 14 and methods of using the compositions to treat various diseases. [Background technology]

[0002] Gold cluster (AuC) refers to a type of material that contains a ligand-bound gold core, with the diameter of the gold core being less than 3 nm. Because gold clusters exhibit molecular-like properties due to the splitting of energy levels, each gold cluster molecule in a gold cluster can be represented by a molecular formula. However, in a gold cluster, the number of gold atoms in a gold cluster molecule can range from 2 to 1000, and the number of ligands can also range from 1 to 1000. To further complicate the situation, even if gold clusters have the same number of gold atoms, the number of ligands varies depending on the type of ligand and preparation conditions. Therefore, it remains unclear which gold cluster molecules with a specific number of gold atoms and a specific number of ligands may exist in the same ligand-bound gold cluster. Summary of the Invention

[0003] The present invention relates to a method for producing a gold cluster molecule Au bound to L-NIBC. 18 (L-NIBC) 14 and a gold cluster molecule Au bound to L-NIBC for use in treating a disease in a subject. 18 (L-NIBC) 14 , L-NIBC bound gold cluster molecule Au 18 (L-NIBC)14 A composition comprising: 18 (L-NIBC) 14 The present invention provides a method of treating a disease in a subject using the compound.

[0004] In certain embodiments, the present invention provides a LN-isobutyrylcysteine ​​(L-NIBC)-conjugated gold cluster molecule, comprising a gold core and a ligand L-NIBC bound to the gold core, and comprising: Au 18 (L-NIBC) 14 The present invention provides an L-NIBC-conjugated gold cluster molecule having the formula:

[0005] In certain embodiments, the present invention provides a method for the preparation of a gold cluster molecule Au bound to L-NIBC for use in the manufacture of a medicament for the treatment of a disease in a subject. 18 (L-NIBC) 14 In certain embodiments, the disease includes multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), and glaucoma.

[0006] In certain embodiments, the present invention provides a composition for treating a disease in a subject, the composition comprising an L-NIBC-conjugated gold cluster molecule Au. 18 (L-NIBC) 14 In certain embodiments, the composition comprises a gold cluster molecule Au 18 (L-NIBC) 14 When used as a component in a mixture of gold cluster molecules in a composition, the L-NIBC-bound gold cluster molecule Au 18 (L-NIBC) 14 has a chromatographic content of at least 7% in the mixture. In certain embodiments, the diseases include multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), and glaucoma.

[0007] In certain embodiments, the present invention provides a method of treating a disease in a subject, comprising administering to a subject an L-NIBC-conjugated gold cluster molecule, Au 18 (L-NIBC) 14 to a subject, wherein the disease is selected from the group consisting of multiple sclerosis, Alzheimer's disease (AD), cirrhosis, type 2 diabetes, Parkinson's disease (PD), and glaucoma.

[0008] The objects and advantages of the present invention will become apparent from the following detailed description of the preferred embodiment of the invention taken in conjunction with the accompanying drawings.

[0009] Preferred embodiments according to the present invention will now be described with reference to the drawings, in which like reference numerals refer to like elements and in which: [Brief description of the drawings]

[0010] [Figure 1] Size exclusion chromatography (SEC) and mass spectra of S8(Au18(L-NIBC)14). Figure 1A shows the SEC of S8(Au18(L-NIBC)14), Figure 1B shows the ESI mass spectrum (wide range) of S8(Au18(L-NIBC)14), and Figures 1C-1F show the mass spectral isotopic peaks displayed after amplification of mass spectral peaks with various mass-to-charge ratios (M / z), where in Figure 1C, z=-2, in Figure 1D, z=-3, in Figure 1E, z=-4, and in Figure 1F, z=-5. [Diagram 2] FIG. 1 shows a typical UV-Vis absorption spectrum of S8(Au18(L-NIBC)14). [Diagram 3] FIG. 1 shows the infrared absorption spectrum of S8(Au18(L-NIBC)14) (upper spectrum) and the infrared absorption spectrum of the ligand L-NIBC (lower spectrum). [Figure 4] FIG. 1 shows the change in distribution of the S8 drug in the aqueous and oil phases with changes in the pH value of the initial aqueous solution. [Diagram 5]FIG. 1 shows behavioral clinical scores of EAE model drug trials. [Figure 6-1] 6A shows the results of HE staining experiments in EAE mouse models. Figure 6A shows the statistical results of histological scores of exemplary drug-administered groups, where ###: comparison with blank control group (P<0.001), *: comparison with model control group (P<0.05), **: comparison with model control group (P<0.01), and Figures 6B to 6F show exemplary images of the blank control group, model control group, S1 drug administration group, S8 drug administration group, and S9 drug administration group, respectively. [Figure 6-2] This figure continues from Figure 6-1. [Figure 7] FIG. 1 shows behavioral clinical scores for S8 drugs, S9 drugs, and S9+S8 drugs in an EAE model drug study. [Figure 8-1] Figure 8 shows the effect of drug administration of S1, S2, S8, and S9 for 180 consecutive days on the performance of male mice in the Morris water maze test. Figure 8A shows the platform exploration latency during training in the positioning navigation experiment, Figure 8B shows the number of platform crossings, Figure 8C shows the swimming distance in the target quadrant, and Figure 8D shows the time spent in the target quadrant, where #: compared with the normal blank control group (P<0.05), ##: compared with the normal blank control group (P<0.01), *: compared with the model control group (P<0.05), **: compared with the model control group (P<0.01). [Figure 8-2] This figure continues from Figure 8-1. [Figure 9] Figure 1 shows the effect of various gold cluster drugs on the formation of Aβ1-40 plaques in the hippocampus and cortex of APP / PS1 double transgenic AD model mice, where *: comparison with the model control group (P<0.05), **: comparison with the model control group (P<0.01), $: comparison with the S9 drug-administered group (P<0.05). [Figure 10-1]10A shows the effect of drugs on Aβ plaque formation in the brain in a 5xFAD transgenic mouse model. Figure 10A shows the number of Aβ plaques in the cerebral cortex, Figure 10B shows the number of Aβ plaques in the hippocampus region, Figure 10C shows an exemplary image of Aβ plaques in the cerebral cortex of mice in a model control group, Figure 10D shows an exemplary image of Aβ plaques in the cerebral cortex of mice in a S8 drug administration group, Figure 10E shows an exemplary image of Aβ plaques in the hippocampus of mice in a model control group, and Figure 10F shows an exemplary image of Aβ plaques in the hippocampus of mice in a S8 drug administration group, where **: compared with the model control group (P<0.01), $: compared with the S9 drug administration group (P<0.05). [Figure 10-2] This figure continues from Figure 10-1. [Figure 11-1] 11 shows the effect of drugs on the number of Iba1+ positive cells and GFAP+ cells in the brain cortex and hippocampus of 5xFAD transgenic AD model mice. Figure 11A shows the number of Iba1+ positive cells in the brain cortex, Figure 11B shows the number of Iba1+ positive cells in the hippocampus, Figure 11C shows the number of GFAP+ positive cells in the brain cortex, and Figure 11D shows the number of GFAP+ positive cells in the hippocampus, where *: compared with the model control group (P<0.05), **: compared with the model control group (P<0.01), $: compared with S9 (P<0.05). [Figure 11-2] This figure continues from Figure 11-1. [Figure 12-1] 12 shows the effect of drugs on serum indicators in liver cirrhosis model mice modeled with CCl4. Figure 12A shows the serum level of ALT, Figure 12B shows the serum level of AST, Figure 12C shows the serum level of TBIL, Figure 12D shows the serum level of MAO, and Figure 12E shows the serum level of ALB, where #: comparison with blank control group (P<0.05), ##: comparison with blank control group (P<0.01), *: comparison with model control group (P<0.05), **: comparison with model control group (P<0.01), $: comparison with S9 drug administration group (P<0.05). [Figure 12-2] This figure continues from Figure 12-1. [Figure 12-3] This figure continues from Figure 12-2. [Figure 13-1] 13 shows the exemplary HE staining images and pathological scores of liver tissues of the model mice of liver cirrhosis modeled with CCl4 administered with various drugs. Figure 13A shows the blank control group, Figure 13B shows the model control group, Figure 13C shows the S1 drug administration group, Figure 13D shows the S8 drug administration group, Figure 13E shows the S9 drug administration group, and Figure 13F shows the pathological scores, where ###: compared with the blank control group (P<0.001), *: compared with the model control group (P<0.05), **: compared with the model control group (P<0.01), $: compared with the S9 drug administration group (P<0.05). [Figure 13-2] This figure continues from Figure 13-1. [Figure 14] Figure 14 shows the results of fasting blood glucose and glucose tolerance levels from protocol I of simultaneous modeling and drug administration in a mouse diabetes model. Figure 14A shows fasting blood glucose levels, and Figure 14B shows glucose tolerance levels at various time points after intragastric glucose administration, where ##: compared with blank control group (P<0.01), *: compared with model control group (P<0.05), **: compared with model control group (P<0.01). [Figure 15] Figure 15 shows the results of fasting blood glucose and glucose tolerance from protocol II, which administers drugs in various ways after modeling. Figure 15A shows the results of glucose tolerance after acute drug administration, Figure 15B shows the results of fasting blood glucose after one month of continuous drug administration, Figure 15C shows the results of fasting blood glucose after two months of continuous drug administration, and Figure 15D shows the results of glucose tolerance after two months of continuous drug administration, where #: compared with blank control group (P<0.05), ##: compared with blank control group (P<0.01), *: compared with model control group (P<0.05), $: compared with S9 drug administration group (P<0.05). [Figure 16-1]Figure 16 shows the results of insulin immunohistochemistry from protocol I with simultaneous modeling and drug administration. Figure 16A shows an exemplary image of the blank control group, Figure 16B shows an exemplary image of the model control group, Figure 16C shows an exemplary image of the S8 drug administration group, and Figure 16D shows the average optical density, where #: comparison with the blank control group (P<0.05), *: comparison with the model control group (P<0.05), Figure 16E shows an exemplary image of HE staining of the blank control group, Figure 16F shows an exemplary image of HE staining of the model control group, and Figure 16G shows an exemplary image of HE staining of the S2 drug administration group. [Figure 16-2] This figure continues from Figure 16-1. [Figure 17-1] 17 shows the results of the drug effect on the behavior of MPTP injury-induced PD model mice. Figure 17A shows the moving distance, Figure 17B shows the moving speed, Figure 17C shows the swimming distance, Figure 17D shows the swimming time, Figure 17E shows the falling latency, and Figure 17F shows the falling rate, where #: comparison with blank control group (P<0.05), ##: comparison with blank control group (P<0.01), ###: comparison with blank control group (P<0.001), *: comparison with model control group (P<0.05), **: comparison with model control group (P<0.01), ***: comparison with model control group (P<0.001), $: comparison with S9 drug administration group (P<0.05). [Figure 17-2] This figure continues from Figure 17-1. [Figure 17-3] This figure continues from Figure 17-2. [Figure 18] Figure 1 shows the results of WB test of the effect of drugs on TH expression in the striatum of mouse brain in MPTP injury-induced PD model, where ##: comparison with blank control group (P<0.01), *: comparison with model control group (P<0.05), **: comparison with model control group (P<0.01), ***: comparison with model control group (P<0.001), $: comparison with S9 drug administration group (P<0.05). [Figure 19-1]Figure 19 shows the effect of various drugs on rat ERG DA3.0 and DA10.0 in ischemia-reperfusion animal model. Figure 19A shows the exemplary ERG (DA3.0) in the left eye (normal eye) (right panel) and right eye (modeling eye) (left panel) of rats in the model control group on the 7th day (D8) after modeling, Figure 19B and Figure 19C show the results of the b-wave amplitude of the full-field ERG DA3.0 and DA10.0 in the right eye of rats in each group, respectively, and Figure 19D shows the exemplary ERG DA3.0 in the left eye (normal eye) (right panel) and right eye (modeling and drug-administered eye) (left panel) of rats in the S8 drug-administered group on the 7th day (D8) after administration, where *: compared with the model control group (P<0.05), **: compared with the model control group (P<0.01), $: compared with the S9 drug-administered group. [Figure 19-2] This figure continues from Figure 19-1. [Figure 20] 20A shows the pathological changes of the fundus induced by ischemia-reperfusion modeling and drug administration by staining ultrathin sections with toluidine blue. Figure 20A shows the results of pathological scores, Figure 20B shows an exemplary image of the left eye as a blank control, Figure 20C shows an exemplary image of the right eye in the model control group, and Figure 20D shows an exemplary image of the right eye in the S8 drug administration group, where ###: compared with the blank control group, *: compared with the model control group (P<0.05), ***: compared with the model control group (P<0.001), $: compared with the S9 drug administration group. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] The present invention may be understood more readily by reference to the following detailed description of certain specific embodiments of the invention.

[0012] Throughout this application, publications are referenced, the disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains.

[0013] As used herein, "administration" refers to oral ("po") administration, administration as a suppository, topical contact, intravenous ("iv") administration, intraperitoneal ("ip") administration, intramuscular ("im") administration, intralesional administration, intrahippocampal administration, intraventricular administration, intranasal administration, or subcutaneous ("sc") administration to a subject, or implantation of a sustained release device, such as a mini-osmotic pump or an erodible implant. Administration is by any route, including parenteral and transmucosal (e.g., oral, nasal, vaginal, rectal, or transdermal). Parenteral administration includes, for example, intravenous, intramuscular, intraarteriolar, intradermal, subcutaneous, intraperitoneal, intraventricular, and intracranial. Other modes of delivery include, but are not limited to, the use of liposomal formulations, intravenous infusion, transdermal patches, and the like.

[0014] The terms "systemic administration" and "administered systemically" refer to a method of administering a compound or composition to a mammal such that the compound or composition is delivered via the circulatory system to a site within the body, including the target site of pharmaceutical action. Systemic administration includes, but is not limited to, oral, intranasal, rectal, and parenteral administration (i.e., other than through the digestive tract, such as intramuscular, intravenous, intraarterial, transdermal, and subcutaneous), except that as used herein, systemic administration does not include administration directly to brain regions by means other than via the circulatory system, such as intrathecal injection and intracranial administration.

[0015] As used herein, the terms "treat" and "treatment" refer to delaying the onset, slowing or reversing the progression, or alleviating or preventing either the disease or condition to which the term applies, or one or more symptoms of such disease or condition.

[0016] The terms "patient," "subject," or "individual" refer interchangeably to mammals, such as humans, or non-human mammals, including primates (e.g., macaques, pan troglodytes, pongos), domesticated mammals (e.g., cats, dogs), agricultural mammals (e.g., cows, sheep, pigs, horses), and laboratory mammals or rodents (e.g., rats, mice, rabbits, hamsters, guinea pigs).

[0017] The present invention provides LN-isobutyrylcysteine ​​(L-NIBC)-conjugated gold cluster molecules. The L-NIBC-conjugated gold cluster molecules include a gold core and a ligand, L-NIBC, bound to the gold core. The L-NIBC-conjugated gold cluster molecules are Au 18 (L-NIBC) 14 It has the formula:

[0018] The present invention provides a method for the treatment of a disease in a subject, comprising administering to a subject a L-NIBC-conjugated gold cluster molecule, Au 18 (L-NIBC) 14 The diseases include multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), and glaucoma.

[0019] The present invention provides compositions for use in treating a disease in a subject, including multiple sclerosis, Alzheimer's disease (AD), cirrhosis of the liver, diabetes, Parkinson's disease (PD), and glaucoma.

[0020] The present invention provides methods of treating diseases in a subject by administering a composition to the subject, including multiple sclerosis, Alzheimer's disease (AD), cirrhosis, diabetes, Parkinson's disease (PD), and glaucoma.

[0021] In certain embodiments, the composition comprises an L-NIBC-bound gold cluster molecule Au 18 (L-NIBC) 14 Includes.

[0022] In certain embodiments, the L-NIBC-bound gold cluster molecule Au 18 (L-NIBC) 14 When used as a component in a mixture of gold cluster molecules in a composition, the L-NIBC-bound gold cluster molecules Au in the mixture 18 (L-NIBC) 14 are at least 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109 ...9%, 109%, 109%, 109%, 109 having a chromatographic content of 5%, 56%, 57%, 58%, 59%, 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99%.

[0023] In certain embodiments, the composition further comprises a pharma- ceutically acceptable excipient, hi certain embodiments, the excipient is a phosphate buffer solution or physiological saline.

[0024] The present invention relates to a method for the preparation of a medicament for the treatment of multiple sclerosis, Alzheimer's disease (AD), liver cirrhosis, diabetes, Parkinson's disease (PD), or glaucoma, comprising administering to the patient a gold cluster molecule conjugated to L-NIBC. 18 (L-NIBC) 14 Provide for the use of.

[0025] In certain embodiments, the L-NIBC-bound gold cluster molecule Au 18 (L-NIBC) 14The dosage of is at least 0.001mg / kg / day, 0.005mg / kg / day, 0.01mg / kg / day, 0.05mg / kg / day, 0.1mg / kg / day, 0.5mg / kg / day, 1mg / kg / day, 2mg / kg / day, 3mg / kg / day, 4mg / kg / day, 5mg / kg / day, 6mg / kg / day, 7mg / kg / day, 8mg / kg / day, 9mg / kg / day, 10mg / kg / day, 15 ...10mg / kg / day, 15mg / kg / day, 10mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / kg / day, 15mg / The effective dosage for each disease can be ascertained by routine clinical trials.

[0026] The following examples are provided for the sole purpose of illustrating the principles of the present invention and are not intended to limit the scope of the invention in any way.

[0027] Embodiment Example 1. Sample preparation 1.1. Protocol I Under conditions of ice bath and stirring, 2500 ml of anhydrous chloroauric acid in methanol, with a content of 5 g of chloroauric acid, was added to the reactor, followed by 500 ml of acetic acid. After sufficient cooling, 500 ml of a methanol solution of L-N-isobutyrylcysteine ​​(L-NIBC), with a content of 36 g of L-NIBC, was added. After reacting for 60 minutes, NaBH 4 1500ml of NaBH 4An ethanol solution of 1000 ml of acetone was added while stirring. At this time, the solution quickly turned brownish black and a large amount of gas was generated. After reacting for 30 minutes, 5000 ml of acetone was added to stop the reaction, and the product precipitated. After centrifugation in a centrifuge, the lower precipitate was collected and the upper supernatant was discarded. The precipitate obtained by centrifugation was dissolved in 500 ml of ultrapure water and left at room temperature 25°C and under neutral conditions for 7 days. It was then dialyzed in ultrapure water using a dialysis bag with a retention molecular weight of 500 Daltons, and the water was changed every day. After 7 days, the dialysis bag was untied and the solution in the bag was collected. The collected solution was freeze-dried to obtain a crude gold cluster product.

[0028] Although the gold cluster crude product may contain many gold cluster molecules with various molecular formulas, it is completely unknown what kind of gold cluster molecules are actually contained in the gold cluster crude product. There is no way to know the actual content of specific gold cluster molecules in the gold cluster crude product or their therapeutic effects, and these unknown factors bring great difficulties to the drug discovery potential and new drug application of the gold cluster crude product. Therefore, it is necessary to separate the various components contained in the gold cluster crude product. For this purpose, the above gold cluster crude product was dissolved in water and separated by preparative chromatography, and fractions with different retention times were collected according to the effluent time. Each fraction was dialyzed using a 500 Dalton dialysis bag, and a series of gold cluster samples were obtained after lyophilization. Several samples were selected, and the molecular formulas of the main components in each sample were determined by ultra-high resolution magnetic resonance mass spectrometry (Bruker Daltonics, Solarix 7.0T ESI) combined with other methods. Their molecular formulas are as follows: Sample 1 (S1): Au as the main component 25 (L-NIBC) 18 , Sample 2 (S2): Au as the main component 4 (L-NIBC) 4 , Sample 3 (S3): Au as the main component 7 (L-NIBC) 5 , Sample 4 (S4): Au as the main component 10 (L-NIBC) 6 , Sample 5 (S5): Au as the main component 5 (L-NIBC) 5 , Sample 6 (S6): Au as the main component 11 (L-NIBC) 9 , Sample 7 (S7): Au as the main component 13 (L-NIBC) 9 , Sample 8 (S8): Au as the main component 18 (L-NIBC) 14 , For ease of comparison, crude gold clusters were included in the experiments and designated as follows: Sample 9 (S9): A mixture of three batches of crude gold clusters.

[0029] 1.2. Protocol II Under conditions of ice bath and stirring, 1300 ml of anhydrous chloroauric acid in methanol, with a chloroauric acid content of 5 g, was added to the reactor, followed by 200 ml of acetic acid, 300 ml of L-isobutyrylcysteine ​​(L-NIBC) in methanol, with a L-NIBC content of 12 g, and 1 ml of ultrapure water. The mixture was reacted for 30 minutes with stirring, and then NaBH 4 The content of NaBH in 500 ml is 18 g. 4A methanol solution of 100 ml of acetone was added, at which time the solution quickly turned brownish black and a large amount of gas was generated. After reacting for 30 minutes, 3000 ml of acetone was added to stop the reaction, and the product precipitated. After centrifugation in a centrifuge, the lower precipitate was collected and the upper supernatant was discarded. The precipitate obtained by centrifugation was dissolved in 500 ml of ultrapure water and left to sit at 25°C under acidic conditions for 7 days. After centrifugation, the precipitate was discarded. The supernatant was then dialyzed in weakly acidic water using a dialysis bag with a retention molecular weight of 500 Daltons, and the water was changed every day. After 7 days, the dialysis bag was unwound and the solution in the bag was collected. The collected solution was freeze-dried to obtain a crude gold cluster product. Fractionation of the main components in the sample and determination of the molecular formula were carried out as described in Protocol I of 1.1 above. This result indicated that Au 18 (L-NIBC) 14 was found to be undetectable.

[0030] Embodiment 2.S8 (Au 18 (L-NIBC) 14 ) Characterization Molecular formula Au 18 (L-NIBC) 14 is a typical molecular formula of S8 given by ultra-high resolution mass spectrometry. Due to the presence of carboxyl groups in L-NIBC, in different solutions or solvents, different numbers of hydrogen ions may be replaced by corresponding metal cations or other cations due to the difference in pH value and the presence of metal cations or other cations in the solution or solvent.

[0031] Figure 1 shows the structure of S8(Au 18 (L-NIBC) 14 As shown in FIG. 1A, S8 (Au 18 (L-NIBC) 14 The SEC spectrum of S8(Au) reveals a purity of up to 99.54%. 18 (L-NIBC) 141C-1F show the large-scale ESI mass spectrum of S8 (Au) with various mass-to-charge ratios (M / z), which indicates that the spectrum is very clean, indicating that there are almost no impurities. 18 (L-NIBC) 14 ) mass spectrum isotope peaks are shown.

[0032] Figure 2 shows the S8 (Au 18 (L-NIBC) 14 2 shows a typical UV-Vis absorption spectrum of 1,2-dichloro-1,2-diphenyl-2,3-tetrahydrofuran (1,2-diphenyl-2,4-tetrahydrofuran) (Figure 2). As shown in Figure 2, there is a clear absorption peak in the range of 500 nm to 700 nm.

[0033] Figure 3 shows the S8 (Au 18 (L-NIBC) 14 The infrared absorption spectrum of the ligand L-NIBC (lower spectrum) is shown. -1 It can be seen that the characteristic absorption peak of the nearby thiol group (-SH) of the ligand L-NIBC is no longer visible in S8 (shown in the box), whereas the characteristic absorption peaks of other functional groups are still present, indicating that the ligand L-NIBC in S8 is bound to the gold atom via the thiol group.

[0034] To investigate the hydrophilic / lipophilic properties of S8 drug in aqueous solutions with various pH values, 10 mg of S8 drug was dissolved in 10 ml of aqueous solutions with various pH values ​​(ultrapure water was added with 5 M HCl to adjust its pH value to the range of 1-7, and the specific pH value was measured by a pH meter, and the measured pH value was defined as the pH value of the initial aqueous solution), and then 10 ml of n-octanol was added to the top of the aqueous solution, respectively, and the aqueous solution and n-octanol were thoroughly mixed by shaking, and then allowed to stand until the n-octanol phase and the aqueous phase separated into two layers. The weight-specific concentration of gold (ppm) in the upper n-octanol phase and the lower aqueous phase was measured by a graphite furnace atomic absorption spectrometer, and the lipid-water partition coefficient P was calculated according to the formula (1): logP = log(Co / Cw)(1) where Co and Cw represent the concentrations of the S8 drug in the oil phase and the aqueous phase, respectively.

[0035] As shown in FIG. 4, at high pH values ​​of the initial aqueous solution (e.g., pH>3.4), the S8 drug is mainly dissolved in the aqueous phase, but its content in the n-octanol phase is very low, and its lipid-water partition coefficient is less than -2 (P<0.01). However, when the initial aqueous solution is below pH 3.4, its content in the aqueous phase decreases rapidly with the decrease in pH value, whereas its content in the n-octanol phase increases rapidly. When the pH value of the initial aqueous solution decreases to 3.0, the lipid-water partition coefficient reaches about 1 (P=9.7), and when the pH value of the initial aqueous solution is 2.8, the lipid-water partition coefficient reaches 1.31 (P=20.5). It can be seen that the S8 drug has undergone a significant reversal of hydrophilic / lipophilic properties within a very narrow range of pH values ​​of the initial aqueous solution (as shown in the central area of ​​the dotted line above). These properties are significantly different from those of conventional drugs, which may have great significance in the formulation design of drugs for clinical applications.

[0036] Embodiment 3. Animal experiments on multiple sclerosis (experimental autoimmune encephalomyelitis (EAE) model) 3.1.EAE model experiment C57BL / 6N female mice, 7-9 weeks old. Animals were housed and maintained in accordance with regulations.

[0037] On day 0, mice were randomized by weight and then subcutaneously injected with 200 μl of myelin oligodendrocyte glycoprotein (MOG) peptide in complete Freund's adjuvant (CFA) (Hooke Laboratories) at 100 μl per site in the right and left flanks. Pertussis toxin (PTX) was injected intraperitoneally at 0 and 48 hours after MOG immunization. The above AuC samples were given on days 1 to 28, respectively, according to Table 1. Animals (n=10) were scored daily for clinical signs of EAE.

[0038] Hematoxylin-eosin staining (HE) is one of the most commonly used staining methods for tissues and cells. Hematoxylin staining solution has a high affinity for the nucleus, staining it blue, while eosin staining solution has a high affinity for the cytoplasm, staining it pink or red. Three mice were selected from each group, tissues were frozen in a -80°C freezer, and frozen tissues were removed for tissue sectioning and placed in a freezing slicer (box temperature, -20°C) to equilibrate for 15 min. Tissue sections were 10 μm.

[0039] HE staining was as follows: After behavioral testing, the mice were anesthetized and perfused transcardially, and then the mouse brains were removed and fixed in 4% paraformaldehyde. After gradient dehydration, the frozen coronal sections were attached to adhesive slides. The slides with the brain slices attached were placed in an oven at 37°C for 15 minutes, the slides were stained with hematoxylin staining solution for 8 minutes, excess dye was washed off with tap water for 10 minutes, the slides were rinsed with purified water for a few seconds, the slides were differentiated with differentiation solution for 30 seconds (optional), rinsed with tap water for 10 minutes, then stained with eosin for 1 minute, rinsed with tap water for 10 minutes, 70% ethanol for 10 seconds, 80% ethanol for 10 seconds, 90% ethanol for 10 seconds, 100% ethanol for 10 seconds, xylene for 5 minutes, fresh xylene for 5 minutes, sealed with a neutral gum seal, and microscopic observation revealed blue nuclei and pink cytoplasm.

[0040] [Table 1]

[0041] [Table 2]

[0042] 3.2. Results and Analysis FIG. 5 shows the results of the behavioral clinical score test of EAE model mice in the drug administration groups represented by S1, S2, S8, and the gold cluster crude product S9. As shown in FIG. 5, S1 (Au 25 (L-NIBC) 18 ) drugs and S2(Au 4 (L-NIBC) 4 ) drug did not significantly improve the behavioral clinical scores of EAE model mice. However, S8 (Au 18 (L-NIBC) 14 ) All mice in the drug group were almost the same as normal mice for the first 13 days, and showed a weak increase in behavioral clinical scores from the 14th day, with behavioral clinical scores less than 1 throughout the entire experimental period. Compared with the mice in the model group, a large improvement was observed (from the 9th day, P<0.001), which indicates that the S8 drug had an attractive effect on suppressing the behavioral disorders of mice caused by the MOG drug. Compared with the S8 drug group, only the S9 (gold cluster crude product) drug administration group showed a significant difference from the model control group until the 19th day (P<0.05), with the highest behavioral clinical score exceeding 2 points, while in the follow-up administration, the score improved rapidly and decreased to less than 1 point from the 22nd day, but still showed that the healing effect of the S9 drug was significantly lower than that of the S8 drug. The test results of the other five drugs (S3-S7) were similar to those of S1 and S2, and no obvious pharmacodynamic effects were observed, so they will not be described in detail in this specification.

[0043] Figure 6-1 and Figure 6-2 show the test results of HE staining. In the EAE animal model, a large number of immune cells will invade the bone marrow. The infiltration of immune cells into the bone marrow can be confirmed by HE staining. Figure 6A shows the statistical results of the histological scores of the exemplary drug administration groups. As shown in Figure 6A, the S8 drug can significantly reduce the increase in histological scores caused by EAE modeling (P<0.01, ** ), and administration of S9 (crude gold cluster product) drug can also significantly reduce the score (P<0.05, *), but significantly less effective than S8 drug, and S1 and S2 have no obvious effect. Drugs S3 to S7 are similar to drugs S1 and S2 and have no obvious effect, so they will not be described in detail in this specification. Figures 6B to 6F are exemplary HE staining images of the blank control group, model control group, S1 administration group, S8 administration group, and S9 administration group, respectively. In the model control group, as shown in the box, it can be seen that a large number of immune cells invaded the bone marrow (Figure 6C), whereas such a phenomenon was not observed in the blank control group mice (Figure 6B). Figure 6D is an exemplary HE staining image of the S1 drug administration group, and as shown in the box, the infiltration of a large number of immune cells can be confirmed as in the model control group mice. S8 drug administration (Figure 6E) can significantly reduce the infiltration of immune cells into the bone marrow, and the infiltration of immune cells is almost invisible, which indicates that the effect is excellent. Although the S9 drug administration group has a certain therapeutic effect, a certain degree of immune cell infiltration can still be confirmed in the bone marrow (Figure 6F). Similar to S1, S2–S7 did not significantly suppress the infiltration of immune cells into the bone marrow. These results are consistent with the behavioral results shown in Figure 5.

[0044] The above results indicate that the S8 drug (Au 18 (L-NIBC) 14 It is demonstrated that S8 can significantly improve the intracerebral inflammation and behavioral clinical scores of EAE model mice, and has obvious advantages compared with other drugs in this experiment. The effect of S8 drug is superior to S9 (gold cluster crude product) drug. Therefore, S8 drug has a wide application prospect in multiple sclerosis and other immune-related diseases.

[0045] Gold cluster molecule Au 18 (L-NIBC) 14 showed unique efficacy, so Au in the S9 sample (crude gold cluster product) 18 (L-NIBC) 14 The content of Au was detected by chromatography (detection wavelength 220 nm). 18(L-NIBC) 14 The purity of the chromatographic peak was calculated to be 2.8% by area percentage method. 18 (L-NIBC) 14 It was revealed that the chromatographic content of Au varied depending on the preparation method, preparation conditions, and preparation batch. 18 (L-NIBC) 14 The chromatographic content of was in the range of 1%-4% depending on different preparation batches.

[0046] The fact that the potency of the S9 drug (crude gold cluster product) was significantly lower than that of the S8 drug indicates that the gold cluster molecule Au 18 (L-NIBC) 14 This indicates that the content of S8 drug is one of the important factors affecting the efficacy of the sample. To further investigate this point, various amounts of S8 drug were added to S9 drug (gold cluster crude product) to measure the amount of gold cluster molecule Au 18 (L-NIBC) 14 A series of S9+S8 drugs were obtained so that the chromatographic content of was different in each S9+S8 drug, and then tested using the EAE model. The results showed that the gold cluster molecule Au in the S9+S8 drug 18 (L-NIBC) 14 It was found that such drugs showed great potency when the chromatographic content of Au reached or exceeded 7%. 18 (L-NIBC) 14 These results indicate that L-NIBC-AuC products with a chromatographic content of 7% or more are effective in treating MS.

[0047] FIG. 7 shows the behavioral clinical scores of the EAE model drug test of S8 drug, S9 drug, and S9+S2 mixed drug, in which the gold cluster molecule Au 18 (L-NIBC) 14The chromatographic content of was about 21%. It can be seen that adding a small amount of S8 to the gold cluster crude product S9 significantly increased the therapeutic effect on EAE model mice, which is almost the same as that of S8.

[0048] Embodiment 4. Alzheimer's Disease (AD) Animal Model Experiment 4.1. APP / PS1 double transgenic AD model mouse experiments 4.1.1. Method Male C57BL / 6 germline APP / PS1 transgenic AD model mice were randomly divided into model control group, S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and gold cluster crude product S9 drug administration group. At the same time, age-matched C57BL / 6 wild-type mice were set up as normal control groups. Each group contained 15 mice. Groups S1, S2, S3, S4, S5, S6, S7, S8, and S9 were intraperitoneally injected with saline solutions of the corresponding drugs once a day at a dose of 10 mg per kg of mouse body weight and an injection volume of 100 μl. Mice in the model control group and normal control group were intraperitoneally injected with the same volume of saline.

[0049] Drug administration began when the mice were 3 months old. After 180 days of continuous administration, the cognitive and memory functions of all animals were assayed by the Morris water maze test.

[0050] Positional navigation experiment: The Morris water maze test system consisted of a circular water pool and an automatic video recording and analysis system. The camera on the top of the water pool was connected to a computer. The water maze consisted of a circular water pool with a diameter of 120 cm and a height of 60 cm and a platform with a diameter of 9 cm. The water level was 0.5 cm higher than the platform, and the water temperature was 22±0.5°C. A white pigment was used to dye the water milky white. The positional navigation experiment was used to measure the learning and memory ability of mice in the water maze, which lasted for 5 days. The water maze was divided into four quadrants in four directions: east, west, south, and north. The platform was located in a fixed quadrant. The position of the platform was fixed throughout the experiment. During training, mice were gently placed in the water with their heads facing the wall of the pool near the outer wall from 1 / 2 radian in various quadrants. The experiment was stopped by recording the time it took the mouse to climb onto the hidden platform or when the recording time reached 60 seconds. After climbing onto the platform, the mouse was allowed to remain on the platform for 30 seconds. If the mouse could not find the platform within 60 seconds, the experimenter guided the mouse to climb onto the platform and remain on the platform for 30 seconds. The escape latency of the mouse to explore the platform was recorded by a camera tracking system. After the experiment, all mice were moved to another place and gently dried with a hair dryer. Each mouse was trained four times a day for five consecutive days with a 20-minute interval between training sessions.

[0051] Spatial probe test: After completing the training on the 5th day, on the 6th day, the platform was removed and the mouse was gently placed in the water from the point farthest from the platform toward the wall of the pool. The movement trajectory of the mouse for 60 seconds was recorded by a camera, and the number of times the mouse crossed the platform, the time spent in the target quadrant, and the swimming distance in the target quadrant were analyzed by software.

[0052] The experimental result is x -The mean mean ± SEM was calculated. All data were processed by SPSS software (SPSS 21) and analyzed by post-hoc Dunnett's test. P<0.05 indicates that the difference is statistically significant. Statistical analysis of data not belonging to normal distribution was performed using Kruskal-Wallis H test and Mann-Whitney U test.

[0053] After behavioral testing, mice were anesthetized by intraperitoneal injection of 7% chloral hydrate, cardiac perfusion connections were established, and tissues were perfused with 4% chloral hydrate for 7 min after a quick flush using saline. After perfusion, brain tissues were carefully collected, placed in 4% perfusion solution, and stored at room temperature for later use. Immunohistochemical methods were used to detect the expression of Aβ1-40 in the hippocampus and cortex. Perfused tissues were dehydrated, hyalinized, waxed, embedded, and sliced ​​with a paraffin slicer. Gradual dewaxing with xylene and absolute ethanol. After microwave antigen retrieval, sections were incubated with hydrogen oxide, and sections were blocked with serum for 30 min. Primary antibody (mouse anti-Aβ1-40, 1:100) was added at room temperature and incubated overnight (15 h) at 4 °C. Primary antibody was discarded, sections were washed with PBS, and HRP-labeled goat anti-mouse secondary antibody was added and incubated at room temperature for 30 min. After washing the slices with PBS, the developer DAB was added for development, and the slices were counterstained with Harris' hematoxylin, dehydrated, and sealed. Photographs were taken using a fluorescent microscope, and quantitative analysis of the slices was performed using Image J.

[0054] 4.1.2.Results Figures 8-1 and 8-2 show the effects of 180 consecutive days of drug administration of S1, S2, S8, and S9 on the performance of male mice in the Morris water maze test. As shown in Figure 8A, during the training of the position navigation experiment, compared with the normal control group (blank), the platform exploration latency of the model control group (model) mice was significantly longer than that of the normal mice from the second to fifth training days (P<0.05, #, P<0.01, ##). Compared with the model control group, the platform exploration latency of the S1 drug administration group and the gold cluster crude product S9 drug administration group was obviously decreased, showing significant differences on the fourth and fifth days (P<0.05, * ). S8 drug administration caused a more significant decrease in platform exploration latency in the modeled mice, with significant differences observed from day 2 to day 5 compared with the model control group (P<0.05, * ). Compared with the S9 drug-administered group, the platform exploration latency of the S8 drug-administered group was significantly shorter on the third day (P<0.05, $). All these results demonstrated that the effect of the S8 drug was better than that of the S1 drug and the gold cluster crude product S9 drug. The S2 drug-administered group showed no obvious decrease in the platform exploration latency.

[0055] As shown in Figures 8B-8D, the results of the spatial probe test revealed that the mice in the model control group showed a significant decrease in swimming distance (Figure 8C) (P<0.01, ##) and time spent in the target quadrant (Figure 8D) (P<0.05, #) compared with the mice in the normal control group, and showed a shorter platform crossing time without significant differences (P>0.05). Compared with the model control group, the S2 drug could not increase the platform crossing time of the mice (Figure 8B), whereas the platform crossing times in the S1 drug administration group, S2 drug administration group, and S9 drug administration group increased, but none of them showed significant differences (P>0.05). As for the swimming distance in the target quadrant and time spent in the target quadrant (Figures 8C and 8D), the S1 drug, S8 drug, and S9 drug could significantly improve these two values ​​(P<0.05,* ), where the increase in the S8 drug group was greater than that in the S1 and S9 drug groups, but the S2 drug could not significantly increase these two values.

[0056] These data reveal that S1 drug, S8 drug and crude gold cluster S9 drug can all significantly improve the cognitive and memory abilities of AD model mice, but S8 drug showed better therapeutic effects than S1 drug and S9 drug, while S2 drug showed no obvious effect.

[0057] The results of administering S3 to S7 drugs for 180 consecutive days were similar to those of S2 drug. Compared with the model control group, there was no obvious increase in the platform exploration latency or platform crossing time during training in the position navigation experiment, the swimming distance in the target quadrant, and the time spent in the target quadrant in the spatial probe test, indicating that S3 to S7 drugs had no obvious effect on improving the cognitive and memory abilities of AD model mice.

[0058] Immunohistochemistry revealed that S8 gold cluster molecules (Au 18 (L-NIBC) 14 It was further confirmed that S8 drug can significantly reduce the formation of Aβ1-40 fibrillary plaques in the brain of APP / PS1 double transgenic AD model mice. Figure 9 shows the comparison of Aβ1-40 plaque numbers in terms of average optical density values ​​in the hippocampus and cortex between the model control group and the S1 drug administration group, the S2 drug administration group, the S3 drug administration group, or the S9 drug administration group. As shown in Figure 9, S8 drug can significantly reduce the number of Aβ1-40 plaques in the hippocampus and cortex, and the reduction is significant compared with the model control group (P<0.01, ** The crude gold cluster product S9 also showed a relatively good reduction effect, and the number of Aβ1-40 plaques in the hippocampus and cortex was significantly lower than that in the model control group (P<0.05, *), but the reduction was significantly lower than that of the S8 drug administration group (P<0.05, $). The S1 drug administration group reduced the number of Aβ fibrous plaques in the brain, similar to the S9 drug administration group, but the reduction was significantly lower than that of the S8 drug administration group. These results demonstrated that the S8 drug has an excellent effect on reducing fibrous plaques in the AD brain, and is superior to the S1 drug and the gold cluster crude product S9. However, the S2 drug did not show any therapeutic effect. The S3 drugs to S7 drugs do not show any therapeutic effect like the S2 drug, so they will not be described in detail in this specification.

[0059] 4.2. Five-transgenic 5xFAD AD model mouse experiment Traditional mouse models (such as the APP / PS1 double transgenic mouse model mentioned above) are useful for early exploration and research of AD pathogenesis and therapeutic drugs. However, because the pathogenesis of AD is complex, more thorough research requires more suitable animal models. 5xFAD is a relatively new APP / PS1 transgenic AD mouse model, which has five familial gene mutations and is accompanied by various AD-related neurological disorders and memory dysfunction. Compared with the APP / PS1 double transgenic mouse model and other AD animal models, the characteristics of neuropathological lesions are more obvious, so 5xFAD has become increasingly widely recognized.

[0060] 4.2.1. Method The model mice were 5xFAD male mice with C57BL / 6 background. The experimental mice were divided into model control group, S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and gold cluster crude product S9 drug administration group. At the same time, age-matched C57BL / 6 wild-type mice were set up as normal control groups. Each group contained 15 mice. Mice in the drug administration groups were intraperitoneally injected with saline solution of the corresponding drug once a day at a dose of 10 mg per kg mouse body weight and an injection volume of 100 μl. Drug administration began when the mice were 2 months old. Mice in the model control group and normal control group were intraperitoneally injected with the same volume of saline according to the same schedule as in the drug administration groups.

[0061] After 3 months of continuous drug administration, the cognitive and memory functions of all animals were assayed by the Morris water maze test.

[0062] Testing spatial learning ability The Morris water maze system consists of a water maze for mouse swimming and an automatic video recording and analysis system. The camera on top of the water maze was connected to a computer. The water maze consists of a circular water pool with a diameter of 110 cm and a height of 60 cm. During swimming training of the mice, the water in the water pool is darkened by black ink and the water temperature is kept at 22±0.5°C. The water maze is divided into north, south, west, and east quadrants. A platform with a diameter of 9 cm is placed in the fixed quadrant. The platform is 0.5 cm lower than the water surface.

[0063] Mice were trained to learn about the hidden platform in a water maze for 5 days. During training, mice were gently placed in water with their heads facing the pool wall in different quadrants. The time it took the mouse to find and climb onto the hidden platform was recorded, and the mouse was allowed to remain on the platform for 30 seconds after climbing onto it. If the mouse could not find the platform within 60 seconds, the training was stopped and the experimenter guided the mouse to climb onto the platform and remain on the platform for 30 seconds. The mouse's platform exploration latency was recorded by a camera tracking system. After the experiment, all mice were moved to another location and gently dried by a hair dryer. Each mouse was trained 4 times a day for 5 consecutive days with a 20-minute interval between training sessions.

[0064] Spatial memory ability test (spatial probe test) Swimming training was completed after 5 days. Probe test was started on the 6th day. The mouse was gently placed in the water from the farthest point from the platform toward the wall of the pool, and the movement trajectory of the mouse for 60 seconds was recorded by a video camera. The number of times the mouse crossed the platform, the time spent in the target quadrant, and the swimming distance in the target quadrant were analyzed by software.

[0065] The experimental result is x - The mean mean ± SEM was calculated. All data were processed by SPSS software (SPSS 21) and analyzed by post-hoc Dunnett's test. P<0.05 indicates that the difference is statistically significant. Statistical analysis of data not belonging to normal distribution was performed using Kruskal-Wallis H test and Mann-Whitney U test.

[0066] Pathological analysis was performed after behavioral testing. Mice were then cultured with CO 2After sacrifice by IVF, cardiac perfusion connections were established. After a quick flush using saline for 5-10 min, tissues were fixed in 4% PFA overnight. The next day, tissues were dehydrated, hyalinized, waxed, embedded in paraffin, and sliced ​​using a paraffin slicer to form paraffin-embedded blocks. After slicing, slices were dewaxed stepwise with xylene and absolute ethanol. Amyloid plaques, astrocytes, and microglia were then detected by immunohistochemical methods. After antigen retrieval by microwave, sections were incubated with hydrogen oxide, and sections were blocked with serum for 30 min. Primary antibodies (mouse anti-Aβ42, 1:100), GFAP, and Iba1 were added at room temperature. The primary antibodies were discarded, sections were washed with PBS, and HRP-labeled goat anti-mouse secondary antibodies were added and incubated at room temperature for 30 min. After washing the slices with PBS, the developer DAB was added for development, and the slices were counterstained with Harris' hematoxylin, dehydrated, and sealed. Photographs were taken using a fluorescent microscope, and quantitative analysis of the slices was performed using Image J.

[0067] 4.2.2.Results The results of the behavioral study revealed that the platform exploration latency of the mice in the model control group was significantly longer than that of the mice in the normal control group from the second to fifth days of training. Compared with the model control group, the platform exploration latency of the mice in the S8 drug administration group was significantly decreased, and a significant difference was observed compared with the model control group from the third day. The S1 drug administration group and the gold cluster crude product S9 drug administration group showed a decrease in platform exploration latency compared with the model control group, but it was not statistically significant. No obvious decrease was observed in the S2 drug administration group to the S7 drug administration group.

[0068] Similar results were obtained in the spatial probe test. S8 drug was able to significantly improve the platform crossing time, swimming distance in the target quadrant, and time spent in the target quadrant. S1 drug administration group and gold cluster crude product S9 drug administration group showed a slight decrease, but it was not statistically significant. No effect was shown in the S2 drug administration group to the S7 drug administration group.

[0069] Figure 10-1 and Figure 10-2 show the results of Aβ42 immunochemical staining of fibrous plaques in the brain cortex and hippocampus of mice in the model control group and drug-administered group. As shown in Figure 10A, Figure 10B, Figure 10C, and Figure 10E, a large number of Aβ plaques appeared in the above areas in the model control group mice at the age of 5 months. Compared with the model control group, the number of Aβ plaques in the brain cortex and hippocampus of S9 drug-administered mice was reduced to a certain extent, but there was no significant difference (P>0.05). However, the S8 drug reduced the number of Aβ plaques in the brain cortex and hippocampus by more than half (P<0.01), which indicates that the S8 drug has an excellent therapeutic effect, and its effect is much better than that of the gold cluster crude product S9. In addition, a significant difference was also found between the gold cluster crude product S9 drug-administered group and the S8 drug-administered group (P<0.05, $), which further demonstrates the difference in effect. 10D and 10F show exemplary images of immunohistochemical staining of the cortex and hippocampus, respectively. S1 drug showed similar effects to S9 drug, and some reduction was observed compared to the model control group, but the difference was not significant. S2 drug administration group showed no reduction compared to the model control group, indicating no effect. S3 drug administration group to S7 drug administration group had no obvious effect like S2 ​​drug administration group.

[0070] Brain inflammation is another important hypothesis regarding the pathogenesis of AD. Microglia and astrocytes are the main cells that cause neuroinflammation. Iba1 and GFAP are markers for microglia and astrocytes, respectively. Increased expression of these proteins indicates an increase in the number of microglia and astrocytes as well as cell activation. Many studies have revealed that the expression of Iba1 and GFA is significantly increased in the brains of AD mice. Activation of these cells leads to the production of large amounts of cytokines, such as interleukins (IL-1β, IL-6), tumor necrosis factor (TNF), and reactive oxygen species, which may cause neuronal dysfunction and death. Therefore, Iba1-positive (Iba1 + ) cells and GFAP positive (GFAP + ) Assessing the effect of a drug on the number of cells can indicate the effect of the drug on inflammation-related processes in the brain.

[0071] Figures 11-1 and 11-2 show Iba1 expression in the cortex and hippocampus of the 5xFAD transgenic AD model mouse brain. + Positive cells and GFAP + The effect of drugs on the number of Iba1 cells is shown in Figure 11A and Figure 11B. The effect of drugs on the number of Iba1 cells in the cortex and hippocampus of mice in the model control group and each drug administration group is shown in Figure 11A and Figure 11B. + The results of immunohistochemical staining of the cells are shown in Figures 11C and 11D. + The results are from immunohistochemical staining of cells. S1 drug inhibited Iba1 in the brain cortex and hippocampus. + Cells and GFAP + It can be seen that there was no significant effect on the number of cells, which indicates that the S1 drug does not have a significant inhibitory effect on brain inflammation. S2-S7 have no obvious inhibitory effect like S1. The gold cluster crude product S9 inhibited Iba1 in the cortical and hippocampal regions of the brain. + Positive cells and GFAP +Although the number of Iba1 cells was reduced, there was no significant difference compared to the model control group (P>0.05). However, the S8 drug showed a significant inhibitory effect on brain inflammation. As shown in Figure 11A, compared to the model control group, the administration of S8 drug significantly reduced the number of Iba1 cells in the cortex. + The number of cells was reduced by approximately 40%, showing a significant difference (P<0.01, ** As shown in Figure 11B, S8 drug administration significantly increased Iba1 expression in the hippocampus compared to the model control group. + The number of cells was significantly reduced by 27% (P<0.05, * As shown in Fig. 11C and Fig. 11D, GFAP expression in the cortex and hippocampus was significantly increased compared to the model control group. + The number of cells was 24% (P<0.05, * ) and 15% (P<0.05, * These results demonstrate that S8 drugs have a potent inhibitory effect on AD-related brain inflammation.

[0072] The above results show that S8 drug shows excellent therapeutic effect in the quintuple transgenic 5xFAD model mouse test, and can play a positive role in the treatment of AD from at least three aspects: improving cognitive behavior and memory behavior, inhibiting protein fibrillation and brain inflammation. Combined with the test results of the APP / PS1 double transgenic mouse model, it can be seen that S1 drug has a certain therapeutic effect, and S9 drug is superior to S1 drug in terms of overall effect, but cannot be compared with S8 drug. S2 drugs to S7 drugs could not show obvious therapeutic effect.

[0073] These results reveal that the molecular structure of various gold cluster molecules containing the same ligand has a significant effect on the application effect of gold cluster molecules in the prevention and treatment of Alzheimer's disease. 18 (L-NIBC) 14(S8 drug) has significant advantages over the other seven gold cluster molecules and the gold cluster crude product S9, and is more suitable for the development of drugs for the prevention and treatment of Alzheimer's disease.

[0074] Further research revealed that when S8 drug was added to the crude gold cluster product S9 drug to obtain a mixed drug, the gold cluster molecule Au 18 (L-NIBC) 14 When the content reaches 7%, it is obvious that the therapeutic effect of the mixed drug is significantly increased.

[0075] Embodiment 5. Experiments with a mouse model of liver cirrhosis 5.1. Method Modeling of liver cirrhosis in mice SPF male C57BL / 6N mice, aged 6-8 weeks and weighing 16-20 g, were randomly divided into 11 groups (n=10) based on their body weight, namely, blank control group, model group, S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and S9 drug administration group. Except for the blank control group, the mouse cirrhosis model in the other groups was treated with carbon tetrachloride (CCl 4 The modeling protocol was as follows: (1) Each mouse was given 10% CCl 4 (diluted in olive oil) was intraperitoneally injected twice a week at 7 μl per gram of body weight for a total of 8 weeks, and the blank control group was intraperitoneally injected with the same amount of olive oil solvent. (2) From the 6th week, two mice were selected every week and sacrificed 48 hours after the last injection. The appearance of the liver was observed. When the appearance was consistent with the characteristics of liver cirrhosis (8th week), the liver tissue was fixed in formalin. HE staining and Masson staining were used to evaluate the model of liver cirrhosis.

[0076] Drug administration If the modeling was successful, the mice in the S1 drug administration group to the S9 drug administration group were given the corresponding drugs by intraperitoneal injection at 10 mg / kg, and the mice in the blank control group and the model group were given physiological saline intraperitoneally at 10 ml / kg. The administration was performed once a day for 20 consecutive days.

[0077] Biochemical assays After drug administration was completed, blood was collected from the orbit of the mice, and serum was obtained for biochemical testing of albumin (ALB), total bilirubin (TBil), alanine aminotransferase (ALT), aspartate aminotransferase (AST), and monoamine oxidase (MAO) using a Zhongsheng Beikong kit and a biochemical analyzer (Siemens). The detection methods were performed in strict accordance with the kit instructions.

[0078] Pathological examination HE staining After euthanasia, the liver tissue samples of the mice were fixed in 4% paraformaldehyde fixative for more than 48 hours. After fixation, the liver samples were dehydrated through an alcohol gradient and processed with xylene and ethanol. The liver tissue was then immersed and embedded in wax. After trimming, mounting, and repairing the embedded material, the liver tissue was sliced ​​with a paraffin microtome, and the slices had a thickness of 4 μm. The main process of HE staining is as follows: After baking in an oven at 65 °C, the slices were processed with xylene and dehydrated with gradient ethanol. The slices were sequentially stained with hematoxylin, blue enhancement solution, and 0.5% eosin, then processed with gradient ethanol and xylene, and sealed with neutral gum. Fibrosis of the liver tissue was observed under a microscope.

[0079] Masson staining After baking, the mouse liver tissue slices were dewaxed and dehydrated. After chromium treatment, the nuclei were stained with Lugo's hematoxylin staining solution. After washing with water, the slices were stained with Masson's Ponceau Red acid fuchsin, the slices were immersed in 2% glacial acetic acid solution and differentiated with 1% phosphomolybdic acid solution. After direct staining with aniline blue solution or light green solution, the slices were immersed in 0.2% glacial acetic acid solution for a while, then cleared with 95% alcohol, absolute alcohol, and xylene, and then sealed with neutral gum. The liver tissues were observed under a microscope.

[0080] 5.2.Results The liver of mice in the model group was divided into round or oval masses of various sizes by proliferating fibrous septa. As shown in Figures 12-1 to 12-3, serum ALT index, serum TBil index, serum AST index, and serum MAO index were significantly increased compared with those of the blank control group (P<0.01, ##), and serum ALB was significantly decreased compared with those of the blank control group (P<0.05, #), indicating that this experimental modeling was successful.

[0081] Figures 12-1 to 12-3 show the effects of drugs on serum levels of ALT (Figure 12A), AST (Figure 12B), TBil (Figure 12C), MAO (Figure 12D), and ALB (Figure 12E) in liver cirrhosis model mice. The results show that the gold cluster crude product S9 drug can significantly improve serum levels of ALT, AST, and TBil in modeled mice (P<0.05, * ), and that the S8 drug can significantly improve all five indexes in the serum of modeled mice (P<0.05, * , P < 0.01, **) is clear. Compared with the gold cluster crude product S9 drug administration group, the S8 drug administration group showed a more significant improvement in the levels of all five indicators, and significant differences were observed in the levels of ALT, AST, and TBIL (P<0.05, $), which indicates that the effect of the S8 drug on improving serum indicators in liver cirrhosis model mice is significantly better than that of the gold cluster crude product S9 drug. The S1 drug had a similar effect to the S9 drug and had an improving effect, but the improvement range was much lower than that of the S8 drug. The S2 drug did not show obvious improvement of the five indicators in the blood of the model mice. The S3 drug to S7 drug do not have obvious improving effects like the S2 drug, so they will not be described in detail in this specification. The above results make it clear that the S8 drug has a superior therapeutic effect on liver cirrhosis compared to other drugs.

[0082] Figure 13-1 and Figure 13-2 show the results of Masson staining experiments. Figure 13A, Figure 13B, Figure 13C, Figure 13D, and Figure 13E show exemplary images of the blank control group, the model control group, the S1 drug administration group, the S8 drug administration group, and the S9 drug administration group, respectively. As shown in Figure 13A, the normal liver tissue from the mice of the blank control group had a clear structure, intact hepatic lobule, well-arranged hepatic cords with a radial arrangement centered on the central vein, normal hepatic cell nuclei, and only a small amount of fibrous tissue in the catchment area. As shown in Figure 13B, in the liver tissue of the model control group, the hepatocytes were pathological, balloon-like structures appeared, the hepatic lobule was almost lost, pseudolobules were abundantly formed, and numerous proliferated protofibrils were present in the liver tissue, forming round or oval fibrous septa.

[0083] As shown in Figure 13D, compared with the model control group, the liver tissues of the S2 drug-treated mice showed extremely significant recovery from liver injury, exhibiting clear structure, intact hepatic lobules, well-arranged hepatic cell cords, without observable fibroplasia and pseudolobules, and showing no obvious difference from normal liver tissue, indicating extremely significant pharmacodynamics.

[0084] As shown in FIG. 13E, the gold cluster crude product S9 drug administration group also showed a relatively significant effect, but compared with the blank control group, there is a certain amount of fibrous proliferation and pseudolobular structure. Combined with the results of the above serum indicators, it is clear that the overall therapeutic effect of S8 drug is obviously better than that of S9 drug. As shown in FIG. 13C, S1 drug is similar to S9 drug, and although S1 drug has a certain therapeutic effect, it is still inferior to S8 drug. S2 drug to S7 drug could not show therapeutic effect. Here, the hepatocytes were pathological, the hepatic lobule was almost disappeared, the pseudolobule was abundantly formed, and there were many proliferated protofibrils in the liver tissue, forming round or oval fibrous septa. No significant difference was found compared with the model control group.

[0085] Figure 13F shows the effect of S1 drug, S2 drug, S8 drug, and crude gold cluster S9 drug on the histopathological score of liver cirrhosis. It can be seen that the histopathological score of the model control group increased from 0.5 to 3.35 ± 0.42, showing a highly significant difference compared with the blank control group (P < 0.001, ###). The score of the S8 drug administration group significantly decreased to 1.33 ± 0.25 compared with the model control group (P < 0.01, ** The gold cluster crude product S9 drug administration group showed a significant decrease compared to the model control group (P<0.05, *), the reduction was 49% lower than that of the S8 drug group, and there was a significant difference between them (P<0.05, $). These results demonstrated that the S8 drug has an excellent protective effect against liver damage caused by cirrhosis, and the therapeutic effect is significantly better than that of the gold cluster crude product S9 drug. The S1 drug has a therapeutic effect similar to that of the S9 drug, but not as good as that of the S8 drug. The S2 drug showed no obvious effect. The results of the S3 drug administration group to the S7 drug administration group were similar to those of the S2 drug administration group and did not have obvious efficacy, so they will not be described in detail in this specification. The results of HE staining are consistent with those of Masson staining, and will not be described in detail in this specification.

[0086] From the above results, it can be seen that the pathological test results are consistent with the biochemical test results, which collectively confirms that the gold cluster molecule Au 18 (L-NIBC) 14 It is clear that (S8 drug) has a significant therapeutic effect on liver cirrhosis, which is significantly better than the therapeutic effects of the gold cluster crude product S9 drug and other gold cluster drugs. 18 (L-NIBC) 14 (S8 drugs) are more suitable for the development of drugs for the treatment (prevention and cure) of liver cirrhosis.

[0087] Further research revealed that when S8 drug was added to the crude gold cluster product S9 drug to obtain a mixed drug, the Au in the mixed drug was 18 (L-NIBC) 14 When the content reaches 7%, it is obvious that the therapeutic effect of the mixed drug is significantly increased.

[0088] Embodiment 6. Diabetic animal model experiment 6.1. Method 6.1.1. Diabetes Modeling, Drug Administration, and Blood Glucose Testing This study employed two protocols: one in which drugs were administered during modeling, and another in which drugs were administered after successful modeling. These protocols were as follows:

[0089] Protocol I. Simultaneous Modeling and Drug Administration SPF grade C57BL / 6 male mice were randomly divided into blank control group, model control group, S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and S9 drug administration group according to fasting blood glucose level (n=12 per group). A mouse model of diabetes was established by feeding a high-fat diet and a small dose of streptozotocin, and drug administration was started on the first day of modeling. First, the mice in each group were fed a laboratory standard maintenance chow for 1 week, and after 1 week, the other groups except the blank control group were fed a high-fat diet for 3 weeks. After 3 weeks, the mice in the model control group and the S1 drug administration group to S9 drug administration group were intraperitoneally injected with streptozotocin (100 mg / kg) for 5 consecutive days, which caused further damage to the pancreas and obtained a model of type 2 diabetes. The blank control group was injected with the same amount of saline as the control. From the first day of feeding the maintenance feed, the S1 drug administration group to the S9 drug administration group were intraperitoneally administered with a dose of 10 mg per kg of body weight once a day, and the blank control group and the model control group were intraperitoneally injected with the same amount of saline. This administration method was continued for 35 days until the end of the experiment. After the modeling and drug administration tests, all mice were fasted for 5 hours to detect the starting blood glucose (fasting blood glucose), and then glucose (1.5 mg per g of body weight) was intragastrically administered, after which the blood glucose levels were detected at 0.5 hours, 1 hour, 1.5 hours, and 2 hours, respectively.

[0090] Protocol II. Drug administration after successful modeling A high-fat diet-induced diabetic mouse model was used to study the effect of drugs on lowering blood glucose levels. SPF grade B6 mice were randomly divided into blank control group (normally fed), model control group, S1 drug-administered group, S2 drug-administered group, S3 drug-administered group, S4 drug-administered group, S5 drug-administered group, S6 drug-administered group, S7 drug-administered group, S8 drug-administered group, and S9 drug-administered group. The model control group and the nine drug-administered groups were fed a high-fat diet at the age of 2 months. The mice in the blank control group were fed standard laboratory chow. The feeding was continued for 3 months. At the age of 5 months, the blood glucose of the mice was measured to determine whether the mice fed the high-fat diet had elevated blood glucose and impaired glucose metabolism, thereby confirming the success of the modeling. There were two types of drug administration. One was acute administration, in which the mice in the nine drug-administered groups were given a single intraperitoneal injection (5 mg / kg), and the corresponding fasting blood glucose and glucose tolerance were measured 30 minutes later. The other was long-term administration, in which mice in the nine drug administration groups were intraperitoneally injected (5 mg / kg / day) for two months. The blank control group and the model control group were intraperitoneally injected with the same amount of saline. One or two months after drug administration, all mice were fasted overnight for 16 hours to detect fasting blood glucose, and then glucose (1.5 mg / g body weight) was intragastrically administered, after which blood glucose levels were detected at 15 minutes, 30 minutes, 45 minutes, 60 minutes, and 120 minutes, respectively.

[0091] 6.1.2.HE staining After the completion of the above test, the animals in each group were euthanized, and the pancreases of the animals were promptly collected and fixed in 4% paraformaldehyde. The fixed pancreatic tissues of the animals were then placed in an embedding box and washed with running water for 30 minutes to remove the fixative solution. The tissues were dehydrated through an alcohol gradient (50%, 70%, 80%, 95%, absolute ethanol) and treated with an environmental transparent agent. The transparent tissues were sequentially immersed in pure paraffin (I, II, III) for 1 hour each time to embed the tissues and completely remove the remaining transparent agent. The embedded pancreatic tissues were sliced ​​into 3 μm sections with a paraffin slicer in succession, and the slices were baked at 60°C for 1 hour for later use. The slices were dewaxed by placing them in xylene for 15 min, which was repeated three times, and then the slices were dewaxed by immersing them in gradient ethanol (100%, 85%, and 75%) for 5 min each and rinsing with running tap water for 5 min. The slices were stained in hematoxylin dye solution for 5 min, after which the excess dye solution was washed off with tap water, the color was separated using 0.7% hydrochloric acid ethanol for 10 s, and the slices were washed with tap water until the nuclei and nuclear chromatin were clear. The slices were dehydrated in 70% and 90% ethanol for 10 min, stained with 0.5% eosin solution for 5 min, and the excess dye was washed off with tap water. The stained slices were dehydrated in 70%, 80%, 90%, and 100% ethanol for 10 s, hyalinized with a clearing agent, and dried in a ventilated place. After dripping an appropriate amount of neutral gum, the slices were sealed with a cover glass. The pathological sections were observed under a light microscope and photographed.

[0092] 6.1.3. Insulin immunohistochemistry After completion of Protocol I, 3 μm pancreatic slices were sequentially obtained. After dewaxing the 3 μm pancreatic slices, they were frozen in 3% H at room temperature. 2 O 2The slices were incubated with 100 ml of deionized water for 5-10 min to block endogenous peroxides, and washed with phosphate buffered saline (PBS) for 5 min three times. The slices were placed in antigen repair solution and antigen repaired in a microwave oven. After natural cooling, the slices were washed with PBS for 5 min three times. 5% BSA blocking solution was added dropwise, and the slices were incubated at 37 °C for 30 min. The blocking solution was discarded, insulin immunohistochemistry primary antibody incubation solution (antibody ratio: 1:200) was added, incubated at 37 °C for 1-2 h, and rinsed with PBS for 5 min three times to remove excess primary antibody. Horseradish peroxidase-labeled secondary antibody incubation solution was added dropwise, and the slices were incubated at 37 °C for 30 min. The slices were washed with PBS for 5 min three times. The slices were stained with DAB and observed under a microscope to control the color development time, and the staining was terminated by washing with tap water. Nuclei were counterstained with hematoxylin, rinsed with tap water, gradient dehydrated in various concentrations of alcohol (75%, 85%, and 100%), cleared, and sealed with neutral gum. Pancreatic tissues were observed under a microscope, and insulin expression was detected by immunohistochemistry and photographed. Three fields (400x magnification) containing islets were randomly selected from each slice. The average optical density of each field was quantitatively measured by using Image Pro Plus version 6.0 software, and the average value was taken as the average optical density of the slice.

[0093] 6.2.Results Figure 14A shows the test results of fasting blood glucose from protocol I with simultaneous modeling and drug administration. As shown in Figure 14A, the fasting blood glucose of the model control group was 20.3 ± 4.1 mM, which was significantly higher than the fasting blood glucose of normal mice in the blank control group (11.5 ± 2.1 mM, P < 0.01, ##). The fasting blood glucose of the S8 drug administration group was 13.1 ± 2.3 mM, which was significantly lower than the fasting blood glucose of the model control group (P < 0.05, * The fasting blood glucose level in the S9 drug administration group was reduced to 15.1±2.9 mM, which was significantly different from that in the model control group (P<0.05, *). The fasting blood glucose of the S1 drug administration group was reduced to 16.7±1.7mM, which was not significantly different from the fasting blood glucose of the model control group (P>0.05). The fasting blood glucose of the S2 administration group to the S7 administration group was at the same level as the model control group, and had no obvious effect.

[0094] FIG. 14B shows the test results of glucose tolerance level from Protocol I with simultaneous modeling and drug administration, where 0 h corresponds to fasting blood glucose in FIG. 14A. The blood glucose levels at each time point in the model control group were significantly higher than those in the blank control group (P<0.01, ##), indicating that the modeling was successful. The fasting blood glucose levels and blood glucose levels at various time points after glucose feeding in the S1 drug administration group were lower than those in the model control group, where the fasting blood glucose (0 h), 0.5 h, and 2 h data (P<0.05, * ) is significantly different, which indicates that S1 drug has a certain hypoglycemic effect. The S9 drug administration group is similar to the S1 drug administration group, but the fasting blood glucose (0 hours), 0.5 hours, 1.5 hours, and 2 hours data are significantly different from the data of the model control group (P<0.05, * The blood glucose levels of the S8 drug administration group at 0, 0.5, 1, 1.5, and 2 hours were significantly lower than those of the model control group (0 hour, P<0.05, * , remaining time points, P < 0.01; ** ), the extent of blood glucose reduction was much greater than that of S1 drug and gold cluster crude product S9 drug (in the blood glucose results from 0.5 hours to 2.0 hours, the extent of blood glucose reduction was almost more than twice as much). Administration of S2 drug did not significantly reduce blood glucose at any time point. The results of S3 drug to S7 drug were similar to those of S2 drug, and no obvious effects were observed, so they will not be described in detail in this specification.

[0095] FIG. 15 shows the results of fasting blood glucose and glucose tolerance from Protocol II, which administers drugs in various ways after modeling. FIG. 15A shows the results of glucose tolerance after acute drug administration (for ease of comparison, only data from S2, S8, and S9 are shown), where 0 min corresponds to fasting blood glucose. As shown in FIG. 15A, both the fasting blood glucose and blood glucose data at other time points in the model control group are significantly higher than those in the blank control group (P<0.05, #), which indicates the success of modeling. Among them, the fasting blood glucose at 0 hours in the model control group was 10.8±2.3 mM, while the fasting blood glucose at 0 hours in the blank control group was 6.7±1.1 mM. Due to the acute drug administration in each drug administration group, their fasting blood glucose was not significantly different from that of the model control group. In the gold cluster crude product S9 drug administration group, the blood glucose level at each time point after glucose administration was lower than that in the model control group, and the result at 15 minutes was significantly different (P<0.05, * ). The S1 drug administration group was similar to the S9 drug administration group, and will not be described in detail in this specification. For the S8 drug, the reduction in blood glucose at each time point relative to the model control group was greater than that of the S9 drug administration group (the reduction was 39% to 2.2 times higher at 15 to 60 minutes), and significant differences were observed compared to the model control group at 15, 30, 45, and 60 minutes (P<0.05, * The S2 drug administration group showed the same levels as the model control group at all time points. The S3 drug administration groups to the S7 drug administration groups were similar to the S2 drug administration group and showed the same levels as the model control group at each time point.

[0096] Figure 15B and Figure 15C are the results of fasting blood glucose from protocol II after one or two consecutive months of drug administration, respectively (for ease of comparison, only data from S2, S8, and S9 are shown). Fasting blood glucose in the model control group was 12.2 ± 0.7 mM and 10.4 ± 0.6 mM, respectively, which was significantly increased compared to the data from the blank control group (5.8 ± 0.5 mM and 5.5 ± 0.3 mM) (P < 0.01, ##). The values ​​in the S9 drug administration group were decreased, but the difference was not significant compared to the model control group (P > 0.05). The decrease in the S8 drug administration group was larger, with the values ​​significantly decreased to 9.8 ± 0.3 mM and 8.3 ± 0.4 mM, respectively (the decrease was 35% and 3.2 times higher than that of the S9 drug, respectively), which was significant compared to the model control group (P < 0.05, * The S2 drug group showed the same level as the model control group, and no obvious decrease was observed. The S3 to S7 drugs showed the same level as the S1 drug, and no significant decrease was observed, so they will not be described in detail in this specification.

[0097] FIG. 15D shows the results of glucose tolerance after two consecutive months of drug administration of S8. As shown in FIG. 15D, the blood glucose levels of the S8 drug administration group at 0, 15, 30, 60, and 120 minutes were significantly lower than those of the model control group (P<0.05, * ), thereby verifying the excellent hypoglycemic effect of S8 drugs.

[0098] The above results mutually support and collectively show that S8 drug exhibits excellent hypoglycemic effect, and its effect is significantly better than that of S1 drug and crude gold cluster product S9 drug. S2 to S7 drugs did not have obvious hypoglycemic effect.

[0099] Figure 16-1 and Figure 16-2 show the HE staining and insulin immunohistochemistry results from protocol I with simultaneous modeling and drug administration. Figure 16A, Figure 16B, and Figure 16C are exemplary immunohistochemistry images of the blank control group, the model control group, and the S8 drug administration group, respectively. The immunohistochemistry results showed that mice in each group showed yellow to brown insulin positive signals (note: the brown was darker in the gray image and the yellow was lighter in the gray image). The main expression location was pancreatic islet cells. Insulin staining in the blank control group showed strong brown positive signal (this color was very dark in the gray image, as shown by the area surrounded by dotted line in Figure 16A), while insulin staining signal in the model control group was significantly weakened, the color was significantly lighter, sparse light brown to yellow (this color was lighter in the gray image, as shown by the area surrounded by dotted line in Figure 16B), while mice in the S8 drug administration group were similar to normal mice in the blank control group, showing strong brown positive signal (this color was very dark in the gray image, as shown by the area surrounded by dotted line in Figure 16C). Figure 16D shows the statistical results of the average optical density values ​​of the blank control group, the model control group, and the various drug administration groups. As shown in FIG. 16D, the expression of insulin in the model control group was significantly lower than that in the blank control group (P<0.05, #), and the expression of insulin in the S1 drug administration group and the S9 drug administration group was higher than that in the model control group (more pronounced in the S9 drug administration group), but the difference was not significant (P>0.05), indicating that S1 and S9 have some protective effect on pancreatic islets. Compared with the model control group, the S8 drug administration group showed significantly higher insulin expression (P<0.05, *), which indicates that the S8 drug has a very good effect on protecting islets from damage or repairing islets from damage. Since the S2 drug administration group had the same level of insulin expression as the model control group, this indicates that the S2 drug did not have the function of protecting or repairing islets. Since the S3 drug to S7 drug were similar to the S2 drug, they will not be described in detail in this specification. The test results of drug administration after modeling in Protocol II were similar to the results here. The S8 drug also showed a significant effect on protecting islets from damage or repairing islets from damage (compared to the model control group, P<0.01). S9 also has a certain effect that is significantly different from the effect of the model control group (P<0.05), but S9 is much less effective than the S8 drug.

[0100] Meanwhile, the HE staining results reveal that compared with the blank control group (Fig. 16E), the pancreas of the model control group (Fig. 16F) showed obvious histological changes, mainly manifested as a decrease in the number of islets, a severe decrease in islet volume, a significant decrease in the number of islet cells, and mild to moderate islet inflammation. In the S8 drug administration group (Fig. 16G), the tissue structure of the islets was significantly improved compared with the model control group, and no abnormal changes or inflammatory cell infiltration in the islets was found compared with the blank control group, which also indicates the excellent protective effect of the S8 drug on the islets. The islet tissue structure of the S9 drug administration group was also significantly improved compared with the model control group, but the effect was much less obvious than that of the S8 drug administration group, more specifically, the islet volume was reduced to a certain extent, the number of islet cells was also reduced, and a small amount of inflammatory cell infiltration also appeared in the islets. Although the S1 drug is less effective than the S9 drug, it shows a relatively good effect compared with the model control group. The S2 drug administration group had the same level as the model control group, which indicates that the S2 drug did not have the function of protecting or repairing the pancreatic islets. Drugs S3 to S7 are similar to drug S2 and do not show obvious efficacy, so detailed explanations are not provided in this specification.

[0101] These results reveal that the S8 drug can not only significantly improve glycemic index, but also protect pancreatic islets from damage or repair the damage of pancreatic islets, and its effect is much better than that of the gold cluster crude product S9 drug and other drugs.

[0102] The above results reveal that the molecular structures of various gold cluster molecules containing the same ligand have a significant impact on the prevention and treatment of type 2 diabetes. 18 (L-NIBC) 14 (S8 drug) shows excellent effects on the prevention and treatment of type 2 diabetes, and has a very significant advantage in the prevention and treatment of type 2 diabetes over the gold cluster crude product and the other seven gold cluster molecules.

[0103] Meanwhile, islet injury is one of the typical characteristics of type I diabetes. The excellent role of S8 drug in islet protection and islet repair has great benefits and prospects for wide application in the prevention and treatment of type I diabetes.

[0104] Further research revealed that when S8 drug was added to the crude gold cluster product S9 drug to obtain a mixed drug, the Au in the mixed drug was 18 (L-NIBC) 14 When the content reaches 7%, it is obvious that the therapeutic effect of the mixed drug is significantly increased.

[0105] Embodiment 7. Parkinson's disease (PD) animal model experiment 7.1. Method The MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) injury model is a widely used animal model in PD research. This animal model was used in this study. The experimental animals were SPF male C57BL / 6 mice. The mice were randomly divided into blank control group, model control group, S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and S9 drug administration group, each of which contained 12 mice. The mice in the model control group were intraperitoneally injected with MPTP (30 mg / kg / day) for 7 days to establish the MPTP-induced PD mouse model, and the blank control group was intraperitoneally injected with the same volume of saline. Each drug group was first intraperitoneally injected with MPTP (30 mg / kg / day), followed 0.5 hours later by intraperitoneal injection of the corresponding drug (10 mg / kg / day) once a day for 7 consecutive days.

[0106] The effects of drugs on the behavior of the model animals were analyzed by open field, rotarod, and swimming tests. The specific methods are as follows:

[0107] Open field: For the experiment, animals are transferred from the feeding cage to the spontaneous activity detector. After the animals are adapted to the new environment for 5 minutes, their spontaneous activity and changes are recorded within 5 minutes, and then the strength of their activity ability is measured by the moving distance and moving speed of the animals within 5 minutes.

[0108] Rotarod test: animals need to balance and move continuously on a rotating rod. This is a widely used experiment to check motor coordination. For the experiment, animals are placed on the rotating rod to prevent slipping. The diameter of the roller is 6 cm, and the rotation speed is 20 rpm / min. After 5 acclimation trials, animals are tested for 15 min each time, and the average value is taken for 3 consecutive trials. When the animal slips off, the lower sensing platform stops accordingly, and the latency to fall off the rotating rod is recorded. In each trial, the latency to fall off the rod and the number of animals to fall off the rod are recorded.

[0109] Swimming test: The mice to be tested are placed in a Morris tank with a water depth of 60 cm and a water temperature of 22° C. The swimming distance and swimming time of the animals within 10 minutes are recorded to measure the strength of their activity ability.

[0110] After behavioral testing, proteins in the striatum were detected by Western blot (WB). The necessary parts of the brain were removed from ice and dissolved in RIPA lysis buffer. The homogenates were centrifuged at 12000g for 30 min at 4°C to extract proteins and prepare samples. SDS-polyacrylamide gel electrophoresis was performed at 55V-60V for 4.5 h, and proteins were transferred to a membrane by the semi-dry method at 60mA for about 1.5 h. The membrane was blocked with 5% skim milk powder at room temperature for 1 h, and rabbit anti-tyrosine hydroxylase (TH) diluted in TBST (dilution ratio 1:300) was added and incubated overnight at 4°C. The antibody was collected and the membrane was washed with TBST three times for 10 min each time. IRDye R 680RD goat anti-rabbit diluted in TBST (dilution ratio 1:3000) was added and washed with TBST three times for 10 min each time. The protein signals were scanned by a two-color infrared laser imaging system.

[0111] All data were processed using Prism 5.0 software, and data were analyzed using t-tests or one-way ANOVA x - Values ​​are expressed ±SD, and P<0.05 indicates that the differences are statistically significant.

[0112] 7.2.Results 17-1 to 17-3 show the results of the behavioral tests.

[0113] Mice in the model control group showed tremors, decreased movement, hunched posture, splayed hind legs, unsteady walking, raised tail, and ruffled hair 3 to 5 minutes after MPTP administration, and one or two mice showed epileptic seizures. The above symptoms gradually alleviated 30 to 60 minutes later, and basically returned to normal after 24 hours. However, although the acute response decreased with increasing administration frequency, the decreased movement, unsteady walking, and sluggish response became increasingly prominent after 24 hours.

[0114] As shown in Figures 17A and 17B, after MPTP was injected continuously for 7 days, the spontaneous movement distance of the mice in the model control group was decreased by about 32% (P<0.01, ##) and the movement speed was decreased by about 29% (P<0.01, ##) compared with the blank control group, indicating the symptoms of motor retardation. The spontaneous movement distance and movement speed of the S8 drug-administered group were significantly higher than those of the model control group (P<0.01, ##). ** ), and almost recovered to the level of the blank control group. The spontaneous movement distance and movement speed of the S9 drug-administered group were also significantly higher than those of the model control group (P<0.05, * ), but the increase was significantly smaller than that of the S8 drug-administered group (the increase in spontaneous movement distance and movement speed in the S8 drug-administered group was 51% and 139% higher, respectively, than that of the S9 drug-administered group, P<0.05). The S1 drug-administered group was similar to the S9 drug-administered group and showed a significant increase compared to the increase in the model control group (P<0.05), but the increase was significantly smaller than that of the S8 drug-administered group. The S2 to S7 drug-administered groups did not show significant differences in spontaneous movement distance and movement speed compared to the model control group (P>0.05).

[0115] In the swimming ability test of mice, the longer the swimming time and the longer the swimming distance, the better the motor coordination of the limbs. As shown in Figures 17C and 17D, compared with the blank control group, the swimming distance of the mice in the model control group for 10 minutes was reduced by about 27% and the swimming time was reduced by about 18% (P<0.01, ##). Compared with the model control group, the swimming distance and swimming time of the S8 drug administration group were significantly increased (P<0.01, ** ), where the swimming distance increased by 33% and the swimming time increased by 21%. Compared with the model control group, the swimming distance and swimming time of the gold cluster crude product S9 drug-administered group also increased significantly (21% and 11%, respectively), but the increase was significantly smaller than that of the S8 drug-administered group (comparison of S9 drug-administered group and S8 drug-administered group, P<0.05, $). The S1 drug-administered group was similar to the S9 drug-administered group and also showed a significant increase compared with the model control group, but the increase was significantly smaller than that of the S8 drug-administered group. The values ​​of the remaining S2 drugs to S7 drugs were similar to those of the model control group and were not significantly different from the model control group (P>0.05).

[0116] As shown in Figures 17E and 17F, in the rotarod behavioral test, the mice in the blank control group showed a fall latency of 13.1 ± 2.1 minutes and a fall rate of 31 ± 1.6%. The fall latency of the mice in the model control group significantly decreased to 5.6 ± 1.8 minutes, and the fall rate significantly increased to 78.1 ± 5.0%, indicating that MPTP modeling reduced motor coordination ability, made grasping unstable, and made them more likely to fall. Compared with the model control group, the fall latency of the S8 drug-administered group was significantly increased (P < 0.01, ** ), and the fall rate was significantly reduced (P<0.0001, *** These results demonstrated that S8 has the ability to significantly improve MPTP-induced motor coordination dysfunction. In the group administered the gold cluster crude product S9, the fall latency was prolonged (P<0.05, * ), and the fall rate was also significantly reduced (P<0.01, **), but the increase and decrease were smaller than those of the S8 drug-administered group (there was a significant difference in the fall rate when comparing S8 and S9, P<0.05, $). The S1 drug-administered group was similar to the S9 drug-administered group, and also showed a significant improvement in fall latency and fall rate compared to the model control group (P<0.05, * ), but the improvement was obviously smaller than that of the S8 drug group. The other drug groups were at the same or similar levels as the model control group in terms of their fall latency and fall rate, and no significant difference was found (P>0.05), indicating that no significant efficacy was found.

[0117] Western blot (WB) was used to further detect the expression of tyrosine hydroxylase (TH) protein in the brain striatum. TH is a key enzyme in the dopamine (DA) biosynthesis pathway. TH immunoblotting can show the expression level of TH in tissues, thus revealing the changes in DA neurons. Figure 18 shows the results of the WB test. As shown in Figure 18, the TH expression level in the model control group was reduced to 47.5 ± 3.7% of that in the blank control group by MPTP treatment. The TH expression level in the modeled mice was restored to 71.1 ± 2.3% of that in the blank control group by treatment with the gold cluster crude product S9, showing a significant difference compared with the model control group (P < 0.01, ** ), which indicates that the gold cluster crude product S9 has a relatively good protective effect against the specific loss of DA neurons. By treatment with S8 drug, the TH expression level in the modeled mice was restored to 91.8±3.3% of the TH expression level in the blank control group, showing a highly significant difference compared with the model control group (P<0.001, ***), and also showed a significant difference compared with the S9 drug-treated group (P<0.05, $), which confirmed that the S8 drug had a superior protective effect against the specific loss of DA neurons, and its protective effect was superior to that of the crude gold cluster S9. Treatment with the S1 drug also restored the TH expression level in the modeled mice to 60.7±4.2% (P<0.05, * ), but the extent of recovery was obviously smaller than that of the S9 group and even smaller than that of the S8 drug group. The TH expression levels in the S2 drug group to the S7 drug group were at the same level as that of the model control group, which indicates that the S2 drug to the S7 drug do not have a significant neuroprotective effect.

[0118] The above results indicate that the molecular structures of various gold cluster molecules containing the same ligand have a significant effect on their prevention and treatment of PD. 18 (L-NIBC) 14 (S8 drug) showed excellent therapeutic effects in the treatment of AD. Gold cluster molecule Au 25 (L-NIBC) 18 Although the S1 drug and the crude gold cluster S9 drug also have obvious therapeutic effects, the effects from either behavioral tests or TH WB were not as clear. 18 (L-NIBC) 14 The other six gold cluster molecules had no obvious therapeutic effect. This is because the gold cluster molecule Au 18 (L-NIBC) 14 It shows that it has great advantages in the prevention and treatment of PD and is more suitable for producing drugs for the prevention and treatment of PD.

[0119] Further research revealed that when S8 drug was added to the crude gold cluster product S9 drug to obtain a mixed drug, the Au in the mixed drug was 18 (L-NIBC) 14When the content reaches 7%, it is obvious that the therapeutic effect of the mixed drug is significantly increased.

[0120] Embodiment 8. Ischemia-reperfusion glaucoma animal model 8.1. Method One hundred male SD rats were randomly divided into 10 groups according to the ERG b-wave amplitude before modeling. The right eye underwent ocular ischemia-reperfusion modeling, and the left eye was not subjected to modeling treatment (as a blank control), and administration was started on the day of modeling (counted as day 1, D1). Except for the model control group, which was not treated, the rest were all drug-administered groups, including S1 drug administration group, S2 drug administration group, S3 drug administration group, S4 drug administration group, S5 drug administration group, S6 drug administration group, S7 drug administration group, S8 drug administration group, and S9 drug administration group, which were administered by eye drop at a dose of 1 mg per eye, three times a day for 21 consecutive days. The drug administration times on D1 were about 3 hours, 2 hours, and 1 hour before modeling, and the drug administration intervals on the remaining administration days except D1 were 3 hours. During the study, the animals underwent general clinical observation and local ocular observation every day, and their body weights were measured once a week. Animals were examined by full-field electroretinogram (ERG) and optical coherence tomography (OCT) before drug administration and on days 7 (D8), 14 (D15), and 21 (D22) after the first drug administration.

[0121] After the final examination, all animals were euthanized for gross anatomical observation, and both eyeballs were harvested and fixed in Davidson's solution for routine histological processing, including sampling, paraffin embedding, sectioning, and HE staining. After removing the surrounding connective tissue, the optic nerve was quickly placed in pre-cooled PFA fixation solution, quickly repaired, dehydrated, embedded in resin, and ultrathin sections were obtained and stained with toluidine blue. The structure of the eyeball and various parts of the optic nerve were observed under a light microscope, focusing on the morphology of the optic nerve axons, the thickness of each layer of the retina, and the morphological changes of the cells. The sections were examined and evaluated by a pathologist using a light microscope. The lesions were diagnosed and graded using standard terminology. The results of the microscopic examination were graded and scored according to four grading methods: slight (1 point), mild (2 points), moderate (3 points), and severe (4 points), respectively, to facilitate comparison between groups.

[0122] Statistical analysis of data: Data were processed by statistical software SPSS 22.0. First, Levene's test was used to test the homogeneity of data. If the data were homogeneous (P>0.05), one-way ANOVA should be performed, and if the Levene's test analysis shows that the data are non-normally distributed (P≦0.05), Kruskal-Wallis nonparametric test should be performed. If the result of Kruskal-Wallis nonparametric test is significant (P≦0.05), Mann-Whitney U test should be further used to perform pairwise comparison. P<0.05 was considered as statistically significant.

[0123] 8.2.Results Electroretinogram (ERG) is a common method to evaluate optic nerve dysfunction in ischemia-reperfusion glaucoma animals. A complex set of potential changes generated by retinal visual cells under light stimulation can be elicited by electrodes and recorded with an appropriate magnifying device, resulting in an ERG pattern that directly reflects the changes in retinal optic nerve function.

[0124] FIG. 19A shows exemplary ERGs (DA3.0) in the left eye (normal eye) (right panel) and right eye (modeling eye) (left panel) of rats in the model control group on the 7th day (D8) after modeling. As shown in FIG. 19A, there is no significant difference in the amplitude of the a-wave between the left and right eyes, but there is a significant change in the b-wave. As a blank control, the b-wave of the normal left eye was clearly visible with normal amplitude, while the amplitude of the b-wave of the modeling right eye was significantly reduced and the waveform became unclear. The b-wave amplitude of DA10.0 in the right and left eyes was similar to that of DA3.0 and also changed significantly. The above results revealed that the optic nerve function of the modeling eye was severely reduced and the modeling was successful.

[0125] Figures 19B and 19C show the results of full-field ERG DA3.0 and DA10.0 b-wave amplitude in the right eye of rats in each group, respectively. No significant difference was found in the b-wave amplitude in the right eye between rats in each group during modeling and before drug administration. Compared with the model control group at the same time, the b-wave amplitude of DA3.0 in the right eye of rats in the S8 drug administration group (Figure 19B) was significantly higher than that in the model control group at D8, D15, and D22 (D8: P<0.01, ** , D15: P<0.05, * , D22: P<0.05, * ), and the b-wave amplitude of DA10.0 (Fig. 19C) was also significantly higher than that of the model control group (all days, P<0.05, * ).

[0126] FIG. 19D shows exemplary ERG DA3.0 in the left eye (normal eye) (right panel) and right eye (modeling and drug-administered eye) (left panel) of rats in the S8 drug-administered group on the 7th day (D8) after administration. It can be seen that after S8 drug administration, the ERG waveform of the right eye is clear and normal, and there is no significant difference in the shape and amplitude of b-wave between the right eye and the left eye. The b-wave amplitude of ERG DA10.0 in the right eye and the left eye is similar to that of DA3.0, and there is no significant difference between the left eye and the right eye, so this will not be described in detail here. These results reveal that S8 drug can significantly improve the impairment of optic nerve function in a rat model of glaucoma induced by ischemia-reperfusion.

[0127] The amplitude of b-wave DA3.0 in the right eye of rats in the S9 drug-administered group was also higher than that in the right eye of rats in the model control group, showing significant differences at D15 and D22 (P<0.05, * ), but the increase was obviously lower than that of the right eye of rats in the S8 drug-administered group (significant difference was seen at D8, P<0.05, $). The amplitude of b-wave DA10.0 in the right eye of rats in the S9 drug-administered group was also higher than that in the right eye of rats in the model control group, with a significant difference shown at D8 (P<0.05, * These results demonstrated that the gold cluster crude product S9 has the effect of improving the impairment of optic nerve function in a rat model of glaucoma induced by ischemia-reperfusion, but the improvement effect is obviously lower than that of the S8 drug.

[0128] The amplitude of b-wave DA3.0 in the right eye of rats in the S1 drug-administered group was significantly higher than that in the right eye of rats in the model control group at D8 (P<0.05, *), and on D15 and D22, there was a certain degree of increase, but no significant difference. The amplitude of b-wave DA10.0 in the right eye of rats in the S1 drug administration group was significantly higher than that in the right eye of rats in the model control group. These results demonstrate that the S1 drug has the effect of improving the impairment of optic nerve function in the rat model of glaucoma induced by ischemia-reperfusion, but the improvement effect is obviously lower than that of the S8 drug.

[0129] Drugs S2 to S7 did not show any therapeutic effect.

[0130] Optical coherence tomography (OCT) was further used to investigate the thickness change of rat retina. These results revealed that the retinal thickness of the right eye at 750 μm or 1500 μm from the optic disc in the model control group showed a gradual decreasing trend with increasing days. For example, at 1500 μm from the optic disc, the retinal thickness was reduced by about 11% on the 21st day (D22) after modeling compared with before modeling, and the difference was significant (P<0.01). After modeling and drug administration, the thickness of the right eye of rats in the S8 drug administration group was significantly thicker than that of the right eye of rats in the model control group on the corresponding days, and the difference was significant on the 14th day (D15) (P<0.05) and 21st day (D22) (P<0.01). This result also indicates the protective effect of S8 drug on the rat retina.

[0131] FIG. 20 shows the pathological changes of the fundus induced by ischemia-reperfusion modeling and drug administration by staining ultrathin sections with toluidine blue. FIG. 20A shows the results of the pathological scores, and FIG. 20B, FIG. 20C, and FIG. 20D show exemplary images of the left eye as a blank control, the right eye in the model control group, and the right eye in the S8 drug administration group, respectively. Pathological examination showed that the optic nerve structure of the left eye as a blank control was clear, the myelin sheath had rings of various sizes, and no obvious abnormalities were found (FIG. 20B). In the right eye of the model control group, severe optic nerve structure damage and myelin loss were observed after modeling (FIG. 20C). Compared with the right eye of the model control group, the right eye of the rats in the S9 drug administration group showed a better improvement effect, but the optic nerve structure was pathological to a certain extent, the loss of myelin sheath was obvious, and the pathological score was significantly lower than that of the right eye of the model control group (FIG. 20A) (P<0.05, * ). Compared with the right eye of the model control group, the right eye of the S2 drug-administered group showed extremely obvious improvement, the optic nerve structure was as clear as the normal left eye, and only a small amount of slight demyelination was observed (Fig. 20D), and the pathological score was significantly lower than that of the right eye of the model control group (Fig. 20A) (P<0.001, *** ). Compared with the right eye of the rats in the S9 drug group, the pathological score of the right eye of the rats in the S8 drug group was significantly lower (P<0.05, $), which indicates that the therapeutic effect of the S8 drug is much better than that of the gold cluster crude product S9 drug. The S1 drug also showed a certain improving effect similar to the S9 drug, but it was significantly lower than that of the S8 drug, so it is not described in detail in this specification. The S2 drug to S7 drug showed no obvious effect.

[0132] These results reveal that the molecular structures of various gold cluster molecules containing the same ligand have a great influence on their effects in protecting the optic nerve and on the prevention and treatment of glaucoma. 18(L-NIBC) 14 (S8 drug) has an extremely excellent therapeutic effect, and the gold cluster crude product S9 drug and gold cluster molecule Au 25 (L-NIBC) 18 (S1 drug) also has a clear therapeutic effect, but its therapeutic effect is limited by the gold cluster molecule Au 18 (L-NIBC) 14 The other six gold cluster molecules have no obvious therapeutic effect. From the above results, the gold cluster molecule Au 18 (L-NIBC) 14 It can be seen that it has enormous advantages in preventing and curing glaucoma.

[0133] Further research revealed that when S8 drug was added to the crude gold cluster product S9 drug to obtain a mixed drug, the Au in the mixed drug was 18 (L-NIBC) 14 When the content reaches 7%, it is obvious that the therapeutic effect of the mixed drug is significantly increased.

[0134] Although the present invention has been described with reference to specific embodiments, it will be understood that the embodiments are illustrative and do not limit the scope of the invention. Alternative embodiments of the invention will be apparent to those skilled in the art to which the invention pertains. Such alternative embodiments are deemed to be included within the scope of the invention. Accordingly, the scope of the present invention is defined by the appended claims and supported by the above description.

Claims

1. A L-N-isobutyrylcysteine ​​(L-NIBC) conjugated gold cluster molecule, Gold core and a ligand L-NIBC bound to the gold core; Including Au １８ (L-NIBC) １４ The L-NIBC-conjugated gold cluster molecule has the formula:

2. The L-NIBC-conjugated gold cluster molecule Au of claim 1 for use in the manufacture of a medicament for the treatment of a disease in a subject. １８ (L-NIBC) １４ .

3. The use according to claim 2, wherein the disease is multiple sclerosis.

4. The use according to claim 2, wherein the disease is Alzheimer's disease (AD).

5. The use according to claim 2, wherein the disease is cirrhosis of the liver.

6. The use according to claim 2, wherein the disease is diabetes.

7. The use according to claim 2, wherein the disease is Parkinson's disease (PD).

8. The use according to claim 2, wherein the disease is glaucoma.

9. The L-NIBC-bound gold cluster molecule Au １８ (L-NIBC) １４ 13. A composition comprising:

10. The L-NIBC-bound gold cluster molecule Au １８ (L-NIBC) １４ is used as a component in the mixture of gold cluster molecules in the composition, the L-NIBC-bound gold cluster molecule Au １８ (L-NIBC) １４ The composition of claim 9, wherein said mixture has a chromatographic content of at least 7%.

11. The composition according to claim 9 or 10, wherein the disease is multiple sclerosis.

12. The composition of claim 9 or 10, wherein the disease is Alzheimer's disease (AD).

13. The composition according to claim 9 or 10, wherein the disease is cirrhosis of the liver.

14. The composition according to claim 9 or 10, wherein the disease is diabetes.

15. The composition of claim 9 or 10, wherein the disease is Parkinson's disease (PD).

16. The composition according to claim 9 or 10, wherein the disease is glaucoma.

17. 10. A method of treating a disease in a subject comprising administering to the subject a composition comprising the L-NIBC-conjugated gold cluster molecule Au of claim 1. １８ (L-NIBC) １４ A method comprising:

18. The L-NIBC-bound gold cluster molecule Au １８ (L-NIBC) １４ is used as a component in the mixture of gold cluster molecules in the composition, the L-NIBC-bound gold cluster molecule Au １８ (L-NIBC) １４ The method of claim 17, wherein said mixture has a chromatographic content of at least 7%.

19. 19. The method of claim 17 or 18, wherein the disease is multiple sclerosis.

20. 19. The method of claim 17 or 18, wherein the disease is Alzheimer's disease (AD).

21. 19. The method of claim 17 or 18, wherein the disease is cirrhosis of the liver.

22. 19. The method of claim 17 or 18, wherein the disease is diabetes.

23. 19. The method of claim 17 or 18, wherein the disease is Parkinson's disease (PD).

24. 19. The method of claim 17 or 18, wherein the disease is glaucoma.

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