Inhibitors of GSK-3 and crystal structures of GSK-3 protein and protein complexes

A compound selected from the group consisting of:

• 3-amino-4(4-chlorophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 2);
• 3-amino-4,5-bis(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 3);
• 3-amino-4-phenyl-5-(4-fluorophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 4);
• 3-amino-4-(4-fluorophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 5);
• 3-amino-4-(3-fluorophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 6);
• 3-amino-4-phenyl-5-(4-pyridyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 7);
• 3-amino-4-phenyl-5-(3-fluorophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 8);
• N-(4,5-diphenyl-1H-pyrazolo[3,4-c]pyridazin-3-yl)-acetamide (Compound 9);
• N-(4,5-diphenyl-1H-pyrazolo[3,4-c]pyridazin-3-yl)-benzamide (Compound 10);
• 3-amino-4-phenyl-5-(4-methyl-phenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 11);
• 3-amino-4-phenyl-5-(2-methyl-phenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 12);
• 3-amino-4-phenyl-5-(3-methyl-phenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 13);
• 3-amino-4-phenyl-5-(2-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 14);
• 3-amino-4-phenyl-5-(2-fluorophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 15);
• 3-amino-4-phenyl-5-(4-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 16);
• 3-amino-4(2-cyanophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 17);
• 3-amino-4(3-cyanophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 18);
• 3-amino-4(4-cyanophenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 19);
• 3-amino-4-phenyl-5-(2-cyanophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 20);
• 3-amino-4-phenyl-5-(3-cyanophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 21);
• 3-amino-4-phenyl-5-(4-cyanophenyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 22);
• 3-amino-4-phenyl-5-(2-pyridyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 23);
• 3-amino-4-phenyl-5-(3-pyridyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 24);
• N-(4,5-diphenyl-1H-pyrazolo[3,4-c] pyridazin-3-yl)-benzylamine (Compound 25);
• 3-amino-4(2-pyridyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 26);
• 3-amino-4(3-pyridyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 27);
• 3-amino-4(4-pyridyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 28);
• 3-amino-4-phenyl-5-(2-thienyl)-1H-pyrazolo[3,4-c]pyridazine (Compound 29);
• 3-amino-4(2-furanyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazine (Compound 30);
• 3-amino-4-phenyl-5-naphthyl-1H-pyrazolo[3,4-c]pyridazine (Compound 31);
• 4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 32);
• 5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 33);
• 5-phenyl-4-o-tolyl-1H-pyrazolo[3,4-c] pyridazin-3-ylamine (Compound 35);
• 5-phenyl-4-m-tolyl-1H-pyrazolo[3,4-c] pyridazin-3-ylamine (Compound 36);
• 5-phenyl-4-p-tolyl-1H-pyrazolo[3,4-c] pyridazin-3-ylamine (Compound 37);
• 4-(4-nitro-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 38);
• 4-(3-amino-5-phenyl-1H-pyrazolo[3,4-c] pyridazin-4-yl)-benzenesulfonamide (Compound 39);
• 4-(3-fluoro-4-methyl-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 40);
• 4-(3-amino-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)-benzamide (Compound 41);
• 5-(2-amino-phenyl)-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 42);
• 4-(4-ethyl-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 43);
• 4-(2,6-difluoro-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 44);
• 4-(3,4-dimethoxy-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 45);
• 4-(3-fluoro-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 46);
• 4-(3-nitro-phenyl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 47);
• 5-(2-chloro-phenyl)-4-(2,6-difluoro-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 48);
• 5-(2-chloro-phenyl)-4-(3,4-dimethoxy-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 49);
• 5-(2-chloro-phenyl)-4-(3-fluoro-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 50);
• 5-(2-chloro-phenyl)-4-(4-dimethylamino-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 51);
• 4-[3-amino-S-(2-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-4-yl]-benzonitrile (Compound 52);
• 5-(2-chloro-phenyl)-4-pyridin-4-yl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 53);
• 5-(2-chloro-phenyl)-4-(3-nitro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 54);
• 5-(2-chloro-phenyl)-4-pyridin-3-yl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 55);
• 4-(2,6-difluoro-phenyl)-5-(3-methoxy-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compoud 56);
• 4-(3,4-dimethoxy-phenyl)-5-(3-methoxy-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 57);
• 4-(3-fluoro-phenyl)-5-(3-methoxy-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 58);
• 4-(4-dimethylamino-phenyl)-5-(3-methoxy-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 59);
• 4-[3-amino-5-(3-methoxy-phenyl)-1H-pyrazolo[3,4-c]pyridazin-4-yl]-benzonitrile (Compound 60);
• 5-(3-methoxy-phenyl)-4-pyridin-4-yl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 61);
• 5-(3-methoxy-phenyl)-4-(3-nitro-phenyl) -1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 62);
• 5-(3-methoxy-phenyl)-4-pyridin-3-yl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 63);
• 4-(2,6-difluoro-phenyl)-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 64);
• 4-(3,4-dimethoxy-phenyl)-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 65);
• 4-(3-fluoro-phenyl)-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 66);
• 4-(4-dimethylamino-phenyl)-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 67);
• 4-(3-amino-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-4-yl)-benzonitrile (Compound 68);
• 4-pyridin-4-yl-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 69);
• 4-(3-nitro-phenyl)-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 70);
• 4-pyridin-3-yl-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 71);
• 3-(3-amino-5-benzo[1,3]dioxol-5-yl-1H-pyrazolo[3,4-c]pyridazin-4-yl)-benzonitrile (Compound 72);
• 3-[3-amino-S-(3-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-4-yl]-benzonitrile (Compound 73);
• 4-[3-amino-4-(3-cyano-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzamide (Compound 74);
• 3-[3-amino-5-(3-nitro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-4-yl]-benzonitrile (Compound 75);
• 3-[3-amino-4-(3-cyano-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzamide (Compound 76);
• 3-(3-amino-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-4-yl)-benzonitrile (Compound 77);
• 3-[3-amino-5-(3-phenoxy-phenyl)-1H-pyrazolo[3,4-c]pyridazin-4-yl]-benzonitrile (Compound 78);
• 4-[3-amino-4-(3-methoxy-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzonitrile (Compound 79);
• 5-benzo[1,3]dioxol-5-yl-4-(4-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 80);
• 4-(4-chloro-phenyl)-5-m-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 81);
• 4-[3-amino-4-(4-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzamide (Compound 82);
• 4-(4-chloro-phenyl)-5-(3-nitro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 83);
• 3-[3-amino-4-(4-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzamide (Compound 84);
• 4-(4-chloro-phenyl)-5-(3-phenoxy-phenyl)-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 85);
• 4-[3-amino-4-(4-chloro-phenyl)-1H-pyrazolo[3,4-c]pyridazin-5-yl]-benzonitrile (Compound 86);
• 5-benzo[1,3]dioxol-5-yl-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 87);
• 5-(3-chloro-phenyl)-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 88);
• 4-(3-amino-4-o-tolyl-1H-pyrazolo[3,4-c] pyridazin-5-yl)-benzamide (Compound 89);
• 5-(3-nitro-phenyl)-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 90);
• 3-(3-amino-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)-benzamide (Compound 91);
• 4-o-tolyl-5-p-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 92);
• 5-(3-phenoxy-phenyl)-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 93);
• 4-(3-amino-4-o-tolyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)-benzonitrile (Compound 94);
• 5-benzo[1,3]dioxol-5-yl-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 95);
• 5-(3-chloro-phenyl)-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 96);
• 5-(3-nitro-phenyl)-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 97);
• 3-(3-amino-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-5-yl)-benzamide (Compound 98);
• 5-(4-tert-butyl-phenyl)-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 99);
• 5-(3-phenoxy-phenyl)-4-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 100);
• 4-cyclohexyl-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 101);
• and
• 4-(2-Methylsulfanyl-pyrimidin-4-yl)-5-phenyl-1H-pyrazolo[3,4-c]pyridazin-3-ylamine (Compound 102).

A method for inhibiting GSK-3 activity in a patient comprising the step of administering to said patient:

• 228
• or a pharmaceutically acceptable derivative thereof, wherein:
• R1 is selected from H, aliphatic, RC(O)—, RS(O)n—, ROC(O)—, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl;
• wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• R2 and R3 are each independently selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, heteroaralkyl, —N(R)2, —NRCOR, —NRCO2R, —NRSO2R, —S(O)nR, —SO2N(R)2, —SR, —OR, —CF3, halo, —NO2, —CN, —C(O)R, —CO2R, —OC(O)R, —CON(R)2, or —OC(O)N(R)2, wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• or wherein R2 and R3 taken together with the intervening atoms optionally form a five- to nine-membered ring that is fused to the pyridazinyl ring of formula I, said fused ring having 0-2 heteroatoms;
• R is selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl, wherein each member of R except H is optionally substituted;
• and
• n is 1 or 2;
• or
• (b) a pharmaceutical composition comprising said compound and a pharmaceutically acceptable carrier, adjuvant, or vehicle;
• in an amount effective to inhibit GSK-3 activity.

A method for enhancing glycogen synthesis or lowering blood levels of glucose in a patient in need thereof, comprising the step of administering to said patient:

• 229
• or a pharmaceutically acceptable derivative thereof, wherein:
• R1 is selected from H, aliphatic, RC(O)—, RS(O)n—, ROC(O)—, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl;
• wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• R2 and R3 are each independently selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, heteroaralkyl, —N(R)2, —NRCOR, —NRCO2R, —NRSO2R, —S(O)nR, —SO2N(R)2, —SR, —OR, —CF3, halo, —NO2, —CN, —C(O)R, —CO2R, —OC(O)R, —CON(R)2, or —OC(O)N(R)2, wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• or wherein R2 and R3 taken together with the intervening atoms optionally form a five- to nine-membered ring that is fused to the pyridazinyl ring of formula I, said fused ring having 0-2 heteroatoms;
• R is selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl, wherein each member of R except H is optionally substituted;
• and
• n is 1 or 2;
• or
• (b) a pharmaceutical composition comprising said compound and a pharmaceutically acceptable carrier, adjuvant, or vehicle;
• in an amount sufficient to enhance glycogen synthesis or lower blood glucose levels.

A method for inhibiting the production of hyperphosphorylated Tau protein in a patient in need thereof, comprising the step of administering to said patient:

• 230
• or a pharmaceutically acceptable derivative thereof, wherein:
• R1 is selected from H, aliphatic, RC(O)—, RS(O)n—, ROC(O)—, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl;
• wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• R2 and R3 are each independently selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, heteroaralkyl, —N(R)2, —NRCOR, —NRCO2R, —NRSO2R, —S(O)nR, —SO2N(R)2, —SR, —OR, —CF3, halo, —NO2, —CN, —C(O)R, —CO2R, —OC(O)R, —CON(R)2, or —OC(O)N(R)2, wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• or wherein R2 and R3 taken together with the intervening atoms optionally form a five- to nine-membered ring that is fused to the pyridazinyl ring of formula I, said fused ring having 0-2 heteroatoms;
• R is selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl, wherein each member of R except H is optionally substituted;
• and
• n is 1 or 2;
• or
• (b) a pharmaceutical composition comprising said compound and a pharmaceutically acceptable carrier, adjuvant, or vehicle;
• in an amount sufficient to inhibit hyperphosphorylation of Tau protein.

A method for inhibiting the phosphorylation of β-catenin in a patient in need thereof, comprising the step of administering to said patient:

• 231
• or a pharmaceutically acceptable derivative thereof, wherein:
• R1 is selected from H, aliphatic, RC(O)—, RS(O)n—, ROC(O)—, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl;
• wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• R2 and R3 are each independently selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, heteroaralkyl, —N(R)2, —NRCOR, —NRCO2R, —NRSO2R, —S(O)nR, —SO2N(R)2, —SR, —OR, —CF3, halo, —NO2, —CN, —C(O)R, —CO2R, —OC(O)R, —CON(R)2, or —OC(O)N(R)2, wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• or wherein R2 and R3 taken together with the intervening atoms optionally form a five- to nine-membered ring that is fused to the pyridazinyl ring of formula I, said fused ring having 0-2 heteroatoms;
• R is selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl, wherein each member of R except H is optionally substituted;
• and
• n is 1 or 2;
• or
• (b) a pharmaceutical composition comprising said compound and a pharmaceutically acceptable carrier, adjuvant, or vehicle;
• in an amount sufficient to inhibit phosphorylation of β-catenin.

A method for inhibiting GSK-3 activity in a biological sample, comprising the step of contacting the biological sample with:

• 232
• or a pharmaceutically acceptable derivative thereof, wherein:
• R1 is selected from H, aliphatic, RC(O)—, RS(O)n—, ROC(O)—, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl;
• wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• R2 and R3 are each independently selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, heteroaralkyl, —N(R)2, —NRCOR, —NRCO2R, —NRSO2R, —S(O)nR, —SO2N(R)2, —SR, —OR, —CF3, halo, —NO2, —CN, —C(O)R, —CO2R, —OC(O)R, —CON(R)2, or —OC(O)N(R)2, wherein said aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl is optionally substituted;
• or wherein R2 and R3 taken together with the intervening atoms optionally form a five- to nine-membered ring that is fused to the pyridazinyl ring of formula I, said fused ring having 0-2 heteroatoms;
• R is selected from H, aliphatic, carbocyclyl, heterocyclyl, aryl, aralkyl, heteroaryl, or heteroaralkyl, wherein each member of R except H is optionally substituted;
• and
• n is 1 or 2;
• or
• (b) a composition comprising said compound and a carrier, adjuvant, or vehicle;
• in an amount effective to inhibit GSK-3 activity.

A method of obtaining a crystal of a GSK-3β protein complex or GSK-3β homologue protein complex, wherein said protein complex comprises a chemical entity that binds to the substrate-binding groove, comprising the steps of:

• a) producing and purifying GSK-3β protein;
• b) mixing a crystallization solution with the protein complex to produce a crystallizable composition;
• and
• c) subjecting the composition to conditions which promote crystallization.

A computer for determining at least a portion of the structure coordinates corresponding to X-ray diffraction data obtained from a molecule or molecular complex, wherein said computer comprises:

• a) a machine-readable data storage medium comprising a data storage material encoded with machine-readable data, wherein said data comprises at least a portion of the GSK-3β structure coordinates of according to any one of FIGS. 1-7;
• b) a machine-readable data storage medium comprising a data storage material encoded with machine-readable data, wherein said data comprises X-ray diffraction data obtained from said molecule or molecular complex;
• and
• c) instructions for performing a Fourier transform of the machine readable data of (a) and for processing said machine readable data of (b) into structure coordinates.

A method of utilizing molecular replacement to obtain structural information about a molecule or a molecular complex of unknown structure, comprising the steps of:

• a) crystallizing said molecule or molecular complex;
• b) generating an X-ray diffraction pattern from said crystallized molecule or molecular complex;
• and
• c) applying at least a portion of the GSK-3β structure coordinates set forth in any one of FIGS. 1-7 to said X-ray diffraction pattern to generate a three-dimensional electron density map of at least a portion of the molecule or molecular complex whose structure is unknown.