Phenoxazine analogs useful as amyloid aggregation inhibitors and treatment of alzheimer's disease and disorders related to amyloidosis

Disclosed are compounds of the Formula I 1 wherein: R 1 is hydrogen, lower alkyl, or cycloalkyl; R 2 is hydrogen, lower alkyl, lower alkoxy, halogen, hydroxy, aryl, heteroaryl, arylalkyl, heteroarylalkyl, arylalkoxy, heteroarylalkoxy, cyano, carboxy, alkoxycarbonyl, carbamoyl, sulfamoyl, nitro, trifluoromethyl, amino, or mono- or dialkylamino; and R 3 and R 4 independently are hydrogen, lower alkoxy, aryl, heteroaryl, halogen, hydroxy, cyano, carboxy, alkoxycarbonyl, carbamoyl, sulfamoyl, nitro, trifluoromethyl, amino, mono- or dialkylamino, or unsubstituted or substituted lower alkyl or lower alkenyl; or R 3 and R 4 together form an unsubstituted or substituted carbocyclic group. Also provided is a method of inhibiting the aggregation of amyloid proteins using a compound of Formula I and a method of imaging amyloid deposits.

EXAMPLE 1 Synthesis of 3-nitrobenzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (Compound 21) 38 1. 2-&lsqb;(3-Hydroxy(2-naphthyl))amino&rsqb;-3,5-dinitrobenzoic acid 39 A mixture of 3-aminonaphthalen-2-ol (4.65 g, 0.029 mol), 2-chloro-3,5-dinitrobenzoic acid (7.20 g, 0.029 mol), water (30 mL), and 2N sodium acetate (NaOAc) (15 mL) is stirred and heated under reflux for 15 minutes. To the resulting thick paste is added 2N sodium hydroxide (NaOH) (15 mL), and the mixture is stirred and heated for another 15 minutes. The dark purple sodium salt is washed with cold brine. An aqueous solution of the filtrate is acidified with dilute HCl and the free acid precipitates as a dark colored solid. The precipitate is triturated in boiling 10% MeOH/H 2 O, filtered, rinsed with cold 10% MeOH/H 2 O, and dried at room temperature in a vacuum oven overnight to yield an orange solid (9.00 g, 0.024 mol, 84%) as the desired product; mp 154-156° C. Analysis for C 17 H 11 N 3 O 7 .0.75 H 2 O: Calcd: C 53.34; H 3.29; N 10.98. Found: C 53.00; H 2.94; N 10.58. 2. 3-Nitrobenzo&lsqb;b&rsqb;phenoxazinecarboxylic acid A mixture of 2-&lsqb;(3-hydroxy(2-naphthyl))amino&rsqb;-3,5-dinitrobenzoic acid (8.50 g, 0.023 mmol), water (30 mL), and 2N sodium hydroxide (20 mL) is stirred and heated under reflux overnight. The resulting dark purple sodium salt is washed with cold brine and filtered. An aqueous solution of the salt is acidified with dilute HCl and the free acid precipitates as a dark colored solid. The filtrate is acidified with dilute HCl and the free acid precipitates as a dark colored solid. The precipitate is triturated in boiling 10% MeOH/H 2 O, filtered, rinsed with cold 10% MeOH/H 2 O, and dried at room temperature in a vacuum oven overnight to yield a brown solid (0.68 g, 0.002 mol, 7%) as the desired product (compound 21); mp>230° C. Analysis for C 17 H 10 N 2 O 5 .0.29 H 2 O: Calcd: C 62.35, H 3.26, N 8.55. Found: C 61.96, H 2.95, N 8.96. 
 EXAMPLE 2 Synthesis of 3-nitro-8-phenylphenoxazinecarboxylic acid (Compound 2) 1. 2-&lsqb;(2-Hydroxy-5-phenylphenyl)amino&rsqb;-3,5-dinitrobenzoic acid 40 The title compound is prepared from 2-amino-4-phenylphenol (5.59 g, 0.03 mol), 2-chloro-3,5-dinitrobenzoic acid (7.45 g, 0.03 mol), water (30 mL), 2N NaOAc (15 mL), and 2N NaOH (15 mL) using the procedure of Example 1, Step 1 as a red solid (9.37 g, 0.023 mol, 78%); mp 215-217° C. Analysis for C 19 H 13 N 3 O 7 .1.13 H 2 O: Calcd: C 54.90; H3.70,N 10.11. Found: C 54.51, H 3.57, N 9.84. 2. 3-Nitro-8-phenylphenoxazinecarboxylic acid 2-&lsqb;(2-Hydroxy-5-phenylphenyl)amino&rsqb;-3,5-dinitrobenzoic acid (8.82 g, 0.022 mmol), water (50 mL), and 10N NaOH (10 mL) are stirred and heated under reflux overnight. The reaction is worked up as in Example 1, Step 2, to afford the final product (compound 2) as a brown solid (5.75 g, 0.017 mol, 75%); mp>250° C. Analysis for C 19 H 12 N 2 O 5 .0.10 H 2 O: Calcd: C 65.18; H 3.51, N 8.00. Found: C 64.79, H 3.30, N 8.18. 
 EXAMPLE 3 Synthesis of 7-&lsqb;2-(3,4-dichlorophenyl)ethyl&rsqb;-3-nitrophenoxazinecarboxylic acid (Compound 5) 1. Bromo&lsqb;(3,4-dichlorophenyl)methyl&rsqb;triphenylphosphorane 41 A mixture of 4-bromomethyl-1,2-dichlorobenzene (2.40 g, 0.01 mol), and triphenylphosphine (5.24 g, 0.02 mol) in toluene (30 mL) is stirred for 16 hours at room temperature. The solid is filtered, rinsed with toluene, and oven dried to yield a white powder (3.95 g, 0.0078 mol, 78%) as the desired product. 1 H NMR (&dgr; ppm): 7.89-7.61 (m, 15H), 7.50 (d, J&equals;2.3 Hz, 1H), 6.97 (dt, J&equals;8.3 Hz, 2.3 Hz, H), 5.20 (d, J&equals;15.9 Hz, 2H). 2. 5-&lsqb;2-(3,4-Dichlorophenyl)vinyl&rsqb;-2-nitrophenol 42 A solution of bromo&lsqb;(3,4-dichlorophenyl)methyl&rsqb;triphenylphosphorane (11.54 g, 22.98 mmol) in dry THF (250 mL) is cooled to −78° C. Sodium hexamethyldisilazane (NaHDMS) (1.0 M/THF, 50.33 mL, 50.33 mol) is added dropwise to maintain the temperature at −78° C. After stirring for 20 minutes, a solution of 3-hydroxy-4-nitrobenzaldehyde (4.22 g, 25.28 mmol) in THF (50 mL) is added dropwise. The resulting mixture is allowed to warm to room temperature within 3 hours. The mixture is then quenched with saturated ammonium chloride (NH 4 Cl) and extracted with ethyl acetate (EtOAc). The organic layers are then washed with 0.1N HCl solution, H 2 O, and brine, respectively. The solution is dried over sodium sulfate (Na 2 SO 4 ) and concentrated in vacuo to give a light brown oil. Purification by flash chromatography (silica gel, 15% EtOAc/hexane) yields 4.29 g (13.8 mmol, 60%) of the desired product; mp 105-106° C. Analysis for C 14 H 9 N 1 O 3 Cl 2 .0.08 EtOAc: Calcd: C 54.23; H 3.06; N 4.42. Found: C 54.33; H 3.04; N 4.02. 3. 2-Amino-5-&lsqb;2-(3,4-dichlorophenyl)ethyl&rsqb;phenol 43 A sample of 5-&lsqb;2-(3,4-dichlorophenyl)vinyl&rsqb;-2-nitrophenol (4.17 g, 13.45 mmol) in THF (100 mL) is reduced in the presence of Raney Nickel (1 g) at 22° C. to 29° C. (&Dgr;P&equals;4.3 psi). The reaction mixture is filtered, and the filtrate is concentrated in vacuo to give a brown solid, 3.5 g (12.4 mmol, 92%) of the desired product; mp 147-149° C. Analysis for C 14 H 13 N 1 O 1 Cl 2 : Calcd: C 59.59; H 4.64; N 4.96. Found: C 59.68; H 4.60; N 4.71. 4. 7-&lsqb;2-(3,4-Dichlorophenyl)ethyl&rsqb;-3-nitrophenoxazinecarboxylic acid A mixture of 2-amino-5-&lsqb;2-(3,4-dichlorophenyl)ethyl&rsqb;phenol (0.73 g, 2.59 mmol), 2-chloro-3,5-dinitrobenzoic acid (0.68 g, 2.59 mmol), water (3 mL), and 2N sodium acetate (1.3 mL) is stirred and heated under reflux for 5 hours. To the resulting thick paste is added 2N sodium hydroxide (15 mL) and the mixture is stirred and heated for another 3 hours. The reaction mixture is acidified with dilute HCl and the free acid forms as a dark precipitate. The precipitate is filtered off and washed with H 2 O. Purification by flash chromatography (silica gel, toluene:EtOAc:AcOH &lsqb;30:5:1&rsqb;) yields 0.21 g (0.47 mmol, 18%) of the desired product (compound 5); mp>250° C. Analysis for C 21 H 14 N 2 O 5 Cl 2 .0.3 toluene.0.25 H 2 O: Calcd: C 58.12; H 3.57; N, 5.87. Found: C 57.77; H 3.29; N 5.52. 
 EXAMPLE 4 Synthesis of 7-&lsqb;3 -(3,4-Dichlorophenyl)-3-oxoprop-1-enyl&rsqb;-3-nitrophenoxazinecarboxylic acid (Compound 6) 1. 1-(3,4-Dichlorophenyl)-3-hydroxy-3-(3-hydroxy-4-nitrophenyl)-propan-1-one 44 Sodium hydroxide (4.88 g, 0.122 mmol) is dissolved in water (80 mL) and 95% EtOH (80 mL) and cooled to 0° C. with an ice-H 2 O bath. 1-(3,4-Dichlorophenyl)ethanone (23.10 g, 0.122 mol) is added in one portion. After the addition, the mixture is warmed to 15° C. 3-hydroxy-4-nitrobenzaldehyde (20.42 g, 0.122 mol) is then added with rigorous stirring. After stirring for 5 minutes, the reaction mixture is diluted with 95% EtOH (300 mL). The resulting tan mixture is stirred at room temperature overnight. The reaction mixture is acidified with 3N HCl and stirred for 30 minutes. The resulting yellow solid is filtered off, washed with H 2 O, washed with 5% MeOH/CH 2 Cl 2 , and oven dried (40° C.) to yield a yellow solid (25.00 g, 0.074 mol, 61%) of the title compound; mp 163-166° C. Analysis for C 15 H 9 Cl 2 NO 5 : Calcd: C 50.59; H 3.11; N 3.93. Found: C 50.61; H 2.81; N 3.81. 2. 2-Amino-5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;phenol 45 The title compound is prepared from 1-(3,4-dichlorophenyl)-3-hydroxy-3-(3-hydroxy-4-nitrophenyl)propan-1-one (6.5 g, 18.25 mmol) in the presence of Raney Nickel (4.0 g) in THF (100 mL) at 25° C. (&Dgr;P&equals;6.3 psi) using the procedure described in Example 5, Step 3. The reaction mixture is concentrated in vacuo and dried in an oven vacuum (30° C.) to give a brown solid (5.9 g, 18.08 mmol, 99%) of the desired product. MS: 326.0 (M &plus; ). 1 H NMR (&dgr; ppm): 9.87 (s, 1H), 8.08 (d, J&equals;2 Hz, 1H), 7.87 (dd, J&equals;8.3 Hz, 1.7 Hz, 1H), 7.76 (d, J&equals;8.3 Hz, 1H), 6.67 (s, 1H), 6.53 (d, J&equals;8.1 Hz, 1H), 6.48 (d, J&equals;8.0 Hz, 1H), 5.08 (d, J&equals;4.2 Hz, 1H), 4.82 (ddd, J&equals;4.2, 4.2, 8.4 Hz, 1H), 4.38 (s, 2H), 3.32 (dd, J&equals;8.8, 16.6 Hz, 1H), 3.02 (dd, J&equals;4.4, 15.4 Hz, 1H). 3. 7-&lsqb;3-(3,4-Dichlorophenyl)-3-oxoprop-1-enyl&rsqb;-3-nitrophenoxazinecarboxylic acid The title compound is prepared from 2-amino-5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;phenol (2.00 g, 6.13 mmol), 2-chloro-3,5-dinitrobenzoic acid (1.51 g, 6.13 mmol), water (5 mL), 2N sodium acetate (3 mL), and 2N sodium hydroxide (3 mL) using the procedure described in Example 5, Step 4. This procedure yields a dark solid (0.5 g, 1.06 mmol, 17%) as the desired product (compound 6); mp>200° C. Analysis for C 22 H 12 N 2 O 6 Cl 2 .0.05 H 2 O: Calcd: C 55.96; H 2.58; N 5.93. Found: C 55.57; H 2.85; N 6.07. 
 EXAMPLE 5 Synthesis of 7-&lsqb;3-(3,4-Dichlorophenyl)propyl&rsqb;-3-nitrophenoxazine carboxylic acid (Compound 7) 1. 5-&lsqb;3-(3,4-Dichlorophenyl)-3-hydroxyprop-1-enyl&rsqb;-2-nitrophenol 46 A solution of 1-(3,4-dichlorophenyl)-3-hydroxy-3-(3-hydroxy-4-nitrophenyl)propan-1-one (15.0 g, 42.12 mmol), 15% of NaOH (100 mL), and EtOH (200 mL) is heated to reflux for 30 minutes and then cooled to room temperature and allowed to stir overnight. The reaction mixture is acidified with 10% H 2 SO 4 and stirred for 30 minutes. The precipitate is filtered off, washed with H 2 O, then 95% EtOH, and oven dried (40° C.) to yield a red solid (13.7 g, 40.51 mmol, 96%) of the title compound; mp 178-180° C. Analysis for C 15 H 9 Cl 2 NO 4 .0.4 H 2 O: Calcd: C 52.17; H 2.86; N 4.06. Found: C 51.78; H 2.62; N 3.68. 2. 5-&lsqb;3-(3,4-Dichlorophenyl)propyl&rsqb;-2-nitrophenol and 5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;-2-nitrophenol 47 A solution of 5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxyprop-1-enyl&rsqb;-2-nitrophenol (5.97 g, 17.65 mmol), Et 3 SiH (8.2 g, 70.62 mmol) in TFA (70 mL) is stirred at room temperature for 10 days. The reaction mixture is concentrated to dryness, and the resulting residue is dissolved in H 2 O (100 mL) and extracted with EtOAc. The organic layer is dried (MgSO 4 ) and concentrated to give a brown oil. Purification by flash chromatography (silica gel, 1% MeOH/CH 2 Cl 2 ) affords 3.8 g (11.65 mmol, 66%) of the desired product: 5-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;-2-nitrophenol. MS: 321.9 (M&plus;) and 2.0 g (5.8 mmol, 33%) of 5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;-2-nitrophenol; mp 90-93° C. Analysis for C 15 H 15 N 1 O 4 Cl 2 : Calcd: C 58.34; H 4.39; N 4.07. Found: C 52.38; H 3.97; N 3.91. 3. 2-Amino-5-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;phenol 48 The title compound is prepared from 5-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;-2-nitrophenol (7.0 g, 21.46 mmol) and Raney Nickel (3.0 g) in THF (100 mL) at 24° C. to 28° C. (&Dgr;P&equals;38.1 psi) using the procedure described in Example 5, Step 3. Purification by flash chromatography (silica gel, 20%, 50%, 80% EtOAc/Hexane) affords a white solid (1.16 g, 3.92 mmol, 18%) of the desired product; mp 79-82° C. HRMS analysis for C 15 H 15 N 1 O 1 Cl 2 : Calcd: 296.0609. Found: 296.0621 (&Dgr;&equals;4.05 ppm). 4. 7-&lsqb;3-(3,4-Dichlorophenyl)propyl&rsqb;-3-nitrophenoxazine carboxylic acid The title compound is prepared from 2-amino-5-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;phenol (1.11 g, 3.75 mmol), 2-chloro-3,5-dinitrobenzoic acid (0.93 g, 3.75 mmol), water (3 mL), 2N sodium acetate (2 mL), and 2N sodium hydroxide (2 mL) using the procedure described in Example 5, Step 4. This procedure yields a dark solid (1.50 g, 3.27 mmol, 87%) as the desired product (compound 7); mp 215-217° C. Analysis for C 22 H 16 N 2 O 5 Cl 2 .0.45 H 2 O.0.05 C 7 H 3 N 2 O 6 Cl (2-chloro-3,5-dinitrobenzoic acid): Calcd: C 55.96; H 3.58; N 6.13. Found: C 55.60; H 3.20; N 5.88. 
 EXAMPLE 6 Synthesis of 7-&lsqb;3 -(3,4-Dichlorophenyl)-3-hydroxypropyl&rsqb;-3-nitrophenoxazine carboxylic acid (Compound 8) 1. 2-Amino-5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;phenol 49 The title compound is prepared from 5-&lsqb;3-(3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;-2-nitrophenol (2.0 g, 5.86 mmol) is Raney Nickel (0.5 g) in THF (50 mL) at 24° C. to 25° C. (&Dgr;P&equals;38.3 psi) using the procedure described in Example 5, Step 3. This procedure yields an orange solid (1.43 g, 4.58 mmol, 79%) of the desired product; mp 121-123° C. Analysis for C 15 H 15 N 1 O 2 Cl 2 : Calcd: C 57.71; H 4,84; N 4.49. Found: C 57.40; H 4.71; N 4.35. 2. 7-&lsqb;3-(3,4-Dichlorophenyl)-3-hydroxypropyl&rsqb;-3-nitrophenoxazine carboxylic acid The title compound is prepared from 2-amino-5-&lsqb;3,4-dichlorophenyl)-3-hydroxypropyl&rsqb;phenol (2.39 g, 7.66 mmol), 2-chloro-3,5-dinitrobenzoic acid (1.89 g, 7.66 mmol), water (6 mL), 2N sodium acetate (4 mL), and 2N sodium hydroxide (4 mL) using the procedure described in Example 5, Step 4. This procedure affords a dark solid (3.40 g, 7.15 mmol, 93%) as the desired product (compound 8); mp 218-220° C. Analysis for C 22 H 16 N 2 O 6 Cl 2 .0.75 H 2 O.0.05 C 7 H 3 N 2 O 6 Cl (2-Chloro-3,5-dinitro-benzoic acid): Calcd: C 53.57; H 3.55; N 5.87. Found: C 53.20; H 3.25; N 5.85. 
 EXAMPLE 7 Synthesis of 3-Amino-7-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;phenoxazine carboxylic acid (Compound 9) The title compound is prepared from 7-&lsqb;3-(3,4-dichlorophenyl)propyl&rsqb;-3-nitrophenoxazinecarboxylic acid (compound 8) (0.86 g, 1.87 mmol) and Raney Nickel (0.2 g) in THF (50 mL) at 24° C. to 26° C. (&Dgr;P&equals;13.3 psi) using the procedure described in Example 5, Step 3. This procedure yields a dark solid (0.13 g, 0.3 mmol, 16%) of the desired product (compound 9); mp 213-216° C. Analysis for C 22 H 18 N 2 O 3 Cl 2 .0.55 H 2 O: Calcd: C 60.16; H 4.38; N 6.38. Found: C 59.77; H 4.20; N 6.03. 
 EXAMPLE 8 The following compounds are prepared essentially according to the procedures described in Examples 1-7 and shown in Schemes 1-4: (a) Phenoxazinecarboxylic acid (compound 1); (b) 3-Nitrophenoxazinecarboxylic acid (compound 3); (c) 3-(Phenylmethoxy)phenoxazinecarboxylic acid (compound 4); (d) 9-Chloro-8-(trifluoromethyl)benzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (compound 10); (e) Benzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (compound 11); (f) 8,9-Dimethylbenzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (compound 12); (g) 8,9-Dihydroxybenzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (compound 13); (h) 8,9-Dichlorobenzo&lsqb;b&rsqb;phenoxazinecarboxylic acid (compound 14); (i) 7-Phenylphenoxazinecarboxylic acid (compound 15); (j) 7-(3,4-Dichlorophenyl)phenoxazinecarboxylic acid (compound 16); (k) 7-Benzylphenoxazinecarboxylic acid (compound 17); (l) 7-&lsqb;(3,4-Dichlorophenyl)methyl&rsqb;phenoxazinecarboxylic acid (compound 18); (m) 7-&lsqb;2-(3,4-Dichlorophenyl)ethyl&rsqb;phenoxazinecarboxylic acid (compound 19); and (n) 8-(3,4-Dichlorophenyl)phenoxazinecarboxylic acid (compound 20). As noted above, the compounds of Formula I are useful because of their ability to inhibit amyloid protein aggregation. The inhibitory activity of the invention compounds has been determined in several biological assays routinely utilized by those skilled in the art to measure such amyloid inhibition. Representative invention compounds have been evaluated in the following amyloid assays. 1. BASSR (Beta-Amyloid Self-Seeding Radioassay) An assay for inhibitors of self-seeded amyloid fibril growth Materials: Stock Solutions: Assay Buffer—50 mM sodium phosphate, pH 7.5, 100 mM NaCl, 0.02% NaN 3 , 1 M urea (filter and store at 4° C.) Soluble A&bgr;(1-40) peptide (Bachem, Torrance, Calif.)—2.2 mg/mL in 10 deionized H 2 O (is stored in aliquots at −20° C.; is kept on ice when thawed) will self-seed after 1 week storage. Typically, the solution is stored until no lag phase is seen in the assay. 125 I-labeled A&bgr;(1-40)—150K to 350K cpm/&mgr;L in 100% acetonitrile−0.1% trifluoroacetic acid (TFA)—1% &bgr;-mercaptoethanol (aliquots stored at −20° C). 125 I-labeled A&bgr;(1-40) is made in accordance with the procedure set forth by LeVine H., III, in Neurobiol. Aging, 1995;16:755, which is hereby incorporated by reference, or this reagent may be purchased from Amersham, Arlington Heights, Ill. Final assay conditions: 30 &mgr;M soluble A&bgr;(1-40) in deionized water in assay buffer&plus;20K to 50K cpm 125 I-labeled A&bgr; (1-40) per assay. Compound to be tested is dissolved in dimethylsulfoxide (DMSO), typically 5 to 50 mM stock, such that the final concentration of DMSO is<1% v/v in the assay. Assay: Reaction mixture for 50 assays (on ice) is comprised of 0.1 to 0.2 &mgr;L of 125 I-labeled A 125 I-labeled A&bgr;(1-40)&plus;1 &mgr;L of soluble A&bgr;(1-40)&plus;13.5 &mgr;L assay buffer per assay. The following are the amounts of the components of the reaction mixture sufficient for 50 assay wells. 5-10 &mgr;L 125 I-labeled A&bgr;(1-40) dried down 675 &mgr;L assay buffer 50 &mgr;L soluble A&bgr;(1-40) Assay Method: 1) The reaction mixture of above is prepared by mixing components and storing on ice. b 2 ) 14.5 &mgr;L of the reaction mixture is pipetted into each of 50 wells on a polypropylene U-bottom 96-well microtiter plate on ice (Costar 3794). 3) 1.7 &mgr;L of diluted compound to be tested is added to each well in a column of eight, including solvent control. Serial 3-fold dilutions from 1 mM (100 &mgr;M final) in assay buffer−urea&equals;7 dilutions&plus;zero. Each 96-well plate can therefore accommodate 11 samples&plus;1 Congo Red control (0.039-5 &mgr;M final in 2-fold steps). 4) The plate with aluminum film (Beckman 538619) is sealed and incubated for 10 minutes on ice. 5) The temperature is raised to 37° C. and incubated for 3 to 5 hours (depending on the lot of the peptide). 6) The aluminum film is removed and 200 &mgr;L/well of ice cold assay buffer with urea is added. The radiolabeled fibrils are collected by vacuum filtration through 0.2 &mgr;m pore size GVWP filters in 96-well plates (Millipore MAGV N22, Bedford, Mass.). The radioactivity of the filters is determined by using standard methods well-known to those skilled in the art. 2. BASST (Beta-Amyloid Self-seeding, Thioflavin T) An assay for inhibitors of self-seeded amyloid fibril growth Materials: Stock Solutions: Assay Buffer—50 mM sodium phosphate, pH 7.5, 100 mM NaCl, 0.02% NaN 3 , 1 M urea (filter and store at 4° C.) Soluble A&bgr;(1-40)—2.2 mg/mL in deionized H 2 O (is stored in aliquots at −20° C.; is kept on ice when thawed) will self-seed after 1 week storage. Typically, the solution is stored until no lag phase is seen in the assay. Final assay conditions: 30 &mgr;M soluble A&bgr;(1-40) in deionized water in assay buffer. Compound to be tested is dissolved in DMSO, typically 5 to 50 mM stock, such that the final concentration of DMSO is<1% v/v in the assay. Assay: Reaction mixture for 50 assays (on ice) is comprised of 1 &mgr;L of soluble A&bgr;(1-40)&plus;13.5 &mgr;L assay buffer per assay. The following are the amounts of the components of the reaction mixture that result in each of the 50 assay wells. 50 &mgr;L soluble A&bgr;(1-40) 675 &mgr;L assay buffer Assay Method: 1) The reaction mix above is prepared by mixing the components and storing on ice. 2) 14.5 &mgr;L of reaction mixture is pipetted into each of 50 wells of a polystyrene U-bottom, 96-well microtiter plate (Corning 25881-96) on ice. 3) 1.7 &mgr;L of diluted compound to be tested is added to each well in a column of eight, including solvent control. Serial 3-fold dilutions from 1 mM (100 &mgr;M final) in assay buffer−urea&equals;7 dilutions&plus;zero. Each 96-well plate can therefore accommodate 11 samples&plus;1 Congo Red control (0.039-5 &mgr;M final in 2-fold steps). 4) The plate with aluminum film is sealed and incubated for 10 minutes on ice. 5) The temperature is raised to 37° C. and incubated for 3 to 5 hours (depending on the lot of the peptide). 6) The aluminum film is removed and 250 &mgr;L/well of 5 &mgr;M thioflavin T (ThT) &lsqb;T-3516, Sigma-Aldrich&rsqb; is added in 50 mM glycine-NaOH, pH 8.5. Fluorescence is read on a plate reader (ex&equals;440 nm/20 nm; em&equals;485 nm/20 nm) within 5 minutes. 3. BAPA (Beta-Amyloid Peptide Aggregation) This assay is used to provide a measure of inhibition by a compound against the aggregation behavior of the beta amyloid peptide. The purpose of this assay is to provide a higher volume method of assaying the amount of beta amyloid aggregation using an endpoint assay based on filtration. In this assay, hexafluoroisopropanol (HFIP) is used to break down the initial amyloid peptide to a monomer state and a concentration of 33 &mgr;M is used, a concentration which is high enough so that aggregation will occur at pH 6.0 in several hours. Method: &bgr;-Amyloid Peptide Aggregation, pH 6.0 (BAPA) To a 96-well plate (Costar 3794) is added 25 &mgr;L 50 mM Phosphate Buffer (pH 6.0), 10 &mgr;L 0.5 mg/mL A&bgr;(1-40) peptide in 20% HFIP&plus;0.1 &mgr;L/assay radioiodinated 125 I A&bgr;(1-40) &lsqb; 125 I A&bgr;(1-40)&rsqb;, and 1 &mgr;L of the compound to be tested, starting at 50 mm with a concentration of DMSO<1%. The reaction is incubated for 2 to 4 hours at room temperature. The reaction is stopped with 200 &mgr;L of 50 mM phosphate buffer, pH 6.0, and filtered through a 0.2 &mgr;m 96-well filter plate (Millipore MAGU N22). The filter plate is washed with 100 &mgr;L of the same phosphate buffer. Aggregation is detected on a Microbeta counter after impregnating the filters with Meltilex (1450-441) and is corrected for background. 4. BATYM ASSAY Methods: Required A&bgr;(1-42) (California Peptide) is dried from its hexafluoroisopropanol (HFIP) stock solution. The A&bgr;(1-42) is dissolved in DMSO and then mixed with phosphate buffered saline (PBS) (pH 7.4). The mixed A&bgr;(1-42) solution is filtered with a GVWP 0.22 &mgr;m syringe filter (Millipore, Bedford, Mass.). The compound to be tested in DMSO (50 times concentrate) is put into each well (0.5 &mgr;L/well) of a 96-well plate. The A&bgr;(1-42) solution is added into each well (25 &mgr;L/well). The plate is centrifuged at 1000 g for 5 minutes and incubated at 37° C. for 1 day (A&bgr;1-42; final concentration 100 &mgr;M). After incubation Thioflavin T (ThT) (30 &mgr;M) solution in glycine-NaOH buffer (pH 8.5, 50 mM) is added into each well (250 &mgr;L/well), fluorescence is measured (ex&equals;440/20 nm, em&equals;485/20 nm) using a fluorescence plate reader. The inhibitory activity is calculated as the reduction of fluorescence with the following formula: Inhibition (%)&equals;&lcub;( F ( A &bgr;)− F ( A &bgr;&plus;compound)&rcub;/&lcub; F ( A &bgr;)− F (solvent−compound)&rcub;×100. The IC 50 s are calculated by a curve-fitting program using the equation given below. The data is obtained from two different experiments in triplicate. F(x)&equals;100−100/&lcub;1&plus;(IC 50 /10 x ) n &rcub;; x&equals;concentration of tested compound (M), IC 50 &equals;(M), n&equals;Hill coefficient. The results of these assays for compounds of the present invention are shown in Table 2. 2 TABLE 2 Ex- am- Com- ple pound BASSR BASST BATYM BAPA No. No. (IC 50 &equals; &mgr;M) (IC 50 &equals; &mgr;M) (IC 50 &equals; &mgr;M) (IC 50 &equals; &mgr;M) 1 21 12, 11, 10 1 6.7 67 2 2 100, 60, 70, 1 4.15 5 60 3 5 60, 3, 1 7.89 (P) 0.9 (Q) >100 (ppt) 3.02 (Q) 4 6 21, 22, 25 1 4.61 >100 5 7 30 (ppt), 20, 0.5 1.88 >100 6 6 8 >100, 100, 0.6 4.18 100 100 7 9 60, >100 3 8.75 86 8b 3 8 13, 40, 23, 62, >100 23, 40, 13 (BAPA2) 8c 4 14 8.4, 45, 45, 50, >50 8.44 (BAPA2) ppt &equals; Precipitate and indicates that a precipitate formed at the indicated concentration The above data establishes that representative invention compounds are active in standard assays used to measure inhibition of protein aggregation. The compounds exhibit excellent specificity, for example, as shown in the BASST assay, as well as the BATYM assay. The compounds are thus useful to clinically inhibit amyloid protein aggregation and to image amyloid deposits for diagnostic use. The compounds will be used in the form of pharmaceutical formulations, and the following examples illustrate typical compositions. 
 EXAMPLE 9 3 Tablet Formulation Ingredient Amount Compound of Example 1 50 mg Lactose 80 mg Cornstarch (for mix) 10 mg Cornstarch (for paste) 8 mg Magnesium Stearate (1%) 2 mg 150 mg The compound of Example 1 (Compound 21) is mixed with the lactose and cornstarch (for mix) and blended to uniformity to a powder. The cornstarch (for paste) is suspended in 6 mL of water and heated with stirring to form a paste. The paste is added to the mixed powder, and the mixture is granulated. The wet granules are passed through a No. 8 hard screen and dried at 50° C. The mixture is lubricated with 1% magnesium stearate and compressed into a tablet. The tablets are administered to a patient at the rate of 1 to 4 each day for prevention of amyloid protein aggregation and treatment of Alzheimer's disease. 
 EXAMPLE 10 Parenteral Solution In a solution of 700 mL of propylene glycol and 200 mL of water for injection is added 20.0 g of Compound No. 7 (Example 5). The mixture is stirred and the pH is adjusted to 5.5 with hydrochloric acid. The volume is adjusted to 1000 mL with water for injection. The solution is sterilized, filled into 5.0 mL ampoules, each containing 2.0 mL (40 mg of Compound No. 7), and sealed under nitrogen. The solution is administered by injection to a patient suffering from medullary carcinoma of the thyroid and in need of treatment. 
 EXAMPLE 11 Patch Formulation Ten milligrams of Compound No. 13 (Example 8g) is mixed with 1 mL of propylene glycol and 2 mg of acrylic-based polymer adhesive containing a resinous cross-linking agent. The mixture is applied to an impermeable backing (30 cm 2 ) and applied to the upper back of a patient for sustained release treatment of amyloid polyneuropathy. The invention and the manner and process of making and using it are now described in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, to make and use the same. It is to be understood that the foregoing describes preferred embodiments of the present invention and that modifications may be made therein without departing from the spirit or scope of the present invention as set forth in the claims. To particularly point out and distinctly claim the subject matter regarded as invention, the following claims conclude this specification.