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1. WO2020118136 - SELECTIVELY CLEAVABLE THERAPEUTIC NANOPARTICLES

Note: Text based on automatic Optical Character Recognition processes. Please use the PDF version for legal matters

[ EN ]

CLAIMS

What is claimed is:

1. A therapeutic nanoparticle comprising stimuli-cleavable crosslinks, and at least one physiologically active agent, wherein the nanoparticle has a particle size of between about 100-1,000 nm, and wherein the nanoparticle undergoes stimulus-directed degradation.

2. The nanoparticle of claim 1, wherein the nanoparticle has a particle size from about 50-900 nm, from about 100-800 nm, from about 100-700 nm, from about 100-600 nm, from about 100-500 nm, from about 100-400 nm, from about 100-300 nm, from about 100-200 nm, from about 200-900 nm, from about 200-800 nm, from about 200- 700 nm, from about 200-600 nm, from about 200-500 nm, from about 200-400 nm, from about 200-300 nm, from about 300-900 nm, from about 300-800 nm, from about 300-700 nm, from about 300-600 nm, from about 300-500 nm, or from about 300-400 nm.

3. The nanoparticle of claim 1, wherein the at least one physiologically active agent comprises one or more therapeutic agents or diagnostic agents.

4. The nanoparticle of claim 3, wherein the at least one physiologically active agent comprises at least one therapeutic protein, small molecule therapeutic agent, peptide, aptamer, or nucleic acid.

5. The nanoparticle of claim 4, wherein the at least one physiologically active agent comprises a protein having a molecular weight between about 10,000 Da and 100,000 kDa, between about 100,000 Da and 100,000 kDa, between about 500,000 Da and 100,000 kDa, between about 1-100,000 kDa, between about 1-75,000 kDa, between about 1-50,000 kDa, between about 1-25,000 kDa, between about 1-10,000 kDa, between about 1-5,000 kDa, between about 1-2,500 kDa, between about 1-1,000 kDa, between about 1-500 kDa, between about 10-500 kDa, between about 20-500 kDa, between about 20-400 kDa, between about 20-300 kDa, between about 20-200 kDa, between about 20-150 kDa, between about 20-100 kDa, between about 20-75 kDa, between about 20-50 kDa, between about 50-500 kDa, between about 50-400 kDa,

between about 50-300 kDa, between about 50-200 kDa, between about 50-150 kDa, between about 50-100 kDa, between about 50-75 kDa, between about 100-400 kDa, between about 100-300 kDa, or between about 100-200 kDa.

6. The nanoparticle of claim 1, wherein the at least one physiologically active agent comprises a PEGylated protein.

7. The nanoparticle of claim 1, wherein the at least one physiologically active agent comprises at least one antibody.

8. The nanoparticle of claim 1, wherein the at least one physiologically active agent comprises at least one monoclonal antibody.

9. The nanoparticle of claim 1, wherein the physiologically active agent is crosslinked.

10. The nanoparticle of claim 1, wherein the physiologically active agent is dispersed in a matrix.

11. The nanoparticle of claim 1 , wherein the physiologically active agent is dispersed in a matrix comprising one or more polymers or lipids.

12. The nanoparticle of claim 1, wherein the physiologically active agent is dispersed in a crosslinked polymer matrix, wherein the crosslinked polymer is selected from the group consisting of polyphosphazenes, polycyanoacrylates, polyesters,

polyhydroxyalkanoates, polyanhydrides, polydixanones, polyorthoesters,

polyesteramides, polyamido amides, polythioesters, collagen, fibrin, fibrinogen, gelatin, polysaccharides, and combinations thereof.

13. The nanoparticle of claim 1, wherein the crosslinked polymer matrix comprises a polyester selected from the group consisting of poly(lactic acid), poly(glycolic acid), poly(caprolactone), poly(propylene fumarate), copolymers thereof, and combinations thereof.

14. The nanoparticle of claim 1, wherein the crosslinked polymer comprises a

polysaccharide selected from the group consisting of chitosans, celluloses, modified celluloses, alginates, pectins, pullulans, hyaluronic acids, starches, amyloses, dextrans

15. The nanoparticle of claim 1, wherein the physiologically active agent is loaded into smaller nanoparticles having a particle size no greater than 50 nm, said smaller nanoparticles dispersed in a matrix.

16. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks are degraded upon exposure to oxidant, reductant, acid, irradiation, ultrasound, heat or magnetic exposure.

17. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks are degraded upon exposure to oxidant.

18. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks are degraded upon exposure to reductant.

19. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks are degraded upon exposure to acid.

20. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks are degraded upon exposure to heat.

21. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks are degraded by infrared radiation, microwave irradiation, or UV irradiations.

22. The nanoparticle of claim 1 , wherein stimuli-cleavable crosslinks are degraded by reactive oxygen species.

23. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise

functional groups selected from disulfide bonds, trisulfide bonds, diselenide bonds, thioacetals, acetals, oxalates, imines, and peptide bonds.

24. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks comprise

disulfide bonds.

25. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise

trisulfide bonds.

26. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks comprise

diselenide bonds.

27. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise

thioacetals.

28. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise

acetals.

29. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise

oxalates.

30. The nanoparticle of claim 1, wherein the stimuli-cleavable crosslinks comprise imines.

31. The nanoparticle of claim 1 , wherein the stimuli-cleavable crosslinks comprise

peptide bonds.

32. The nanoparticle of any preceding claim, wherein the nanoparticle has a particle size from about 50-900 nm, from about 100-800 nm, from about 100-700 nm, from about 100-600 nm, from about 100-500 nm, from about 100-400 nm, from about 100-300 nm, from about 100-200 nm, from about 200-900 nm, from about 200-800 nm, from about 200-700 nm, from about 200-600 nm, from about 200-500 nm, from about 200- 400 nm, from about 200-300 nm, from about 300-900 nm, from about 300-800 nm, from about 300-700 nm, from about 300-600 nm, from about 300-500 nm, or from about 300-400 nm.

33. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises one or more therapeutic agents or diagnostic agents.

34. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises at least one therapeutic protein, small molecule therapeutic agent, peptide, aptamer, or nucleic acid.

35. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises a protein having a molecular weight between about 10,000 Da and 100,000 kDa, between about 100,000 Da and 100,000 kDa, between about 500,000 Da and 100,000 kDa, between about 1-100,000 kDa, between about 1-75,000 kDa, between about 1-50,000 kDa, between about 1-25,000 kDa, between about 1- 10,000 kDa, between about 1-5,000 kDa, between about 1-2,500 kDa, between about 1-1,000 kDa, between about 1-500 kDa, between about 10-500 kDa, between about 20-500 kDa, between about 20-400 kDa, between about 20-300 kDa, between about 20-200 kDa, between about 20-150 kDa, between about 20-100 kDa, between about 20-75 kDa, between about 20-50 kDa, between about 50-500 kDa, between about 50- 400 kDa, between about 50-300 kDa, between about 50-200 kDa, between about 50- 150 kDa, between about 50-100 kDa, between about 50-75 kDa, between about 100- 400 kDa, between about 100-300 kDa, or between about 100-200 kDa.

36. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises a PEGylated protein.

37. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises at least one antibody.

38. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises at least one monoclonal antibody.

39. The nanoparticle of any preceding claim, wherein the at least one physiologically active agent comprises a monoclonal antibody selected from abagovomab, abciximab, abituzumab, abrezekimab, abrilumab, actoxumab, adalimumab, adecatumumab, aducanumab, afasevikumab, afelimomab, alacizumab pegol, alemtuzumab, alirocumab, altumomab pentetate, amatuximab, anatumomab mafenatox,

andecaliximab, anetumab ravtansine, anifrolumab, anrukinzumab, apolizumab, aprutumab ixadotin, arcitumomab, ascrinvacumab, aselizumab, atezolizumab, atidortoxumab, atinumab, atorolimumab, avelumab, azintuxizumab vedotin, bapineuzumab, basiliximab, bavituximab, BCD- 100, bectumomab, begelomab, belantamab mafodotin, belimumab, bemarituzumab, benralizuma, fasenramab, berlimatoxumab, bermekimab, bersanlimab, bertilimumab, besilesomab,

bevacizumab, bezlotoxumab, biciromab, bimagrumab, bimekizumab, birtamimab, bivatuzumab mertansine, bleselumab, blinatumomab, blontuvetmab, blosozumab, bococizumab, brazikumab, brentuximab vedotin, briakinumab, brodalumab, brolucizumab, brontictuzumab, burosumab, crysvitamab, cabiralizumab,

camidanlumab tesirine, camrelizumab, canakinumab, cantuzumab mertansine, cantuzumab ravtansine, caplacizumab, capromab pendetide, carlumab, carotuximab, catumaxomab, CBR96-doxorubicin immunoconjugate, cedelizumab, cemiplimab, cergutuzumab amunaleukin, certolizumab pegol, cetrelimab, cetuximab, cibisatamab, cirmtuzumab, citatuzumab bogatox, cixutumumab, clazakizumab, clenoliximab, clivatuzumab tetraxetan, codrituzumab, cofetuzumab pelidotin, col tux im ah ravtansine, conatumumab, concizumab, cosfroviximab, CR6261, crenezumab, crizanlizumab, crotedumab, cusatuzumab, dacetuzumab, daclizumab, dalotuzumab, dapirolizumab pegol, daratumumab, dectrekumab, demcizumab, denintuzumab mafodotin, denosumab, depatuxizumab mafodotin, derlotuximab biotin, detumomab, dezamizumab, dinutuximab, diridavumab, domagrozumab, dorlimomab aritox, dostarlimab, drozitumab, ds-8201, duligotuzumab, dupilumab, durvalumab, dusigitumab, duvortuxizumab, ecromeximab, eculizumab, edobacomab,

edrecolomab, efalizumab, efungumab, eldelumab, elezanumab, elgemtumab, elotuzumab, elsilimomab, emactuzumab, emapalumab, emibetuzumab, emicizumab, hemlibra, enapotamab vedotin, enavatuzumab, enfortumab vedotin, enlimomab pegol, enoblituzumab, enokizumab, enoticumab, ensituximab, epitumomab cituxetan, epratuzumab, eptinezumab, erenumab, erlizumab, ertumaxomab, etaracizumab, etigilimab, etrolizumab, evinacumab, evolocumab, exbivirumab, fanolesomab, faralimomab, faricimab, farletuzumab, fasinumab, fbta05, felvizumab, fezakinumab, fibatuzumab, ficlatuzumab, figitumumab, firivumab, flanvotumab, fletikumab, flotetuzumab, fontolizumab, foralumab, foravirumab, fremanezumab, fresolimumab, frovocimab, frunevetmab, fulranumab, futuximab, galcanezumab, galiximab, gancotamab, ganitumab, gantenerumab, gatipotuzumab, gavilimomab, gedivumab, gemtuzumab ozogamicin, gevokizumab, gilvetmab, gimsilumab, girentuximab, glembatumumab vedotin , golimumab, gomiliximab, gosuranemab, guselkumab, ianalumab, ibalizumab, IBI308, ibritumomab tiuxetan, icrucumab, idarucizumab, ifabotuzumab, igovomab, iladatuzumab vedotin, IMAB362, imalumab, imaprelimab, imciromab, imgatuzumab, inclacumab, indatuximab ravtansine, indusatumab vedotin, inebilizumab, infliximab, inolimomab, inotuzumab ozogamicin, intetumumab , IOMAB-B, ipilimumab, iratumumab, isatuximab, iscalimab, istiratumab, itolizumab, ixekizumab, keliximab, labetuzumab, lacnotuzumab, ladiratuzumab vedotin, lampalizumab, lanadelumab, landogrozumab, laprituximab emtansine, larcaviximab, lebrikizumab, lemalesomab, lendalizumab, lenvervimab, lenzilumab, lerdelimumab, leronlimab, lesofavumab, letolizumab, lexatumumab, libivirumab, lifastuzumab vedotin, ligelizumab, lilotomab satetraxetan, lintuzumab, lirilumab, lodelcizumab, lokivetmab, loncastuximab tesirine, lorvotuzumab mertansine, losatuxizumab vedotin, lucatumumab, lulizumab pegol, lumiliximab, lumretuzumab, lupartumab amadotin, lutikizumab, mapatumumab, margetuximab, marstacimab, maslimomab, matuzumab, mavrilimumab, mepolizumab, metelimumab, milatuzumab,

minretumomab, mirikizumab, mirvetuximab soravtansine, mitumomab,

modotuximab, mogamulizumab, monalizumab, morolimumab, mosunetuzumab, motavizumab, moxetumomab pasudotox, muromonab-CD3, nacolomab tafenatox, namilumab, naptumomab estafenatox, naratuximab emtansine, narnatumab, natalizumab, navicixizumab, navivumab, naxitamab, nebacumab, necitumumab, nemolizumab, NEODOOl, nerelimomab, nesvacumab, netakimab, nimotuzumab, nirsevimab, nivolumab, nofetumomab merpentan, obiltoxaximab, obinutuzumab, ocaratuzumab, ocrelizumab, odulimomab, ofatumumab, olaratumab, oleclumab, olendalizumab, olokizumab, omalizumab, omburtamab, OMS721, onartuzumab, ontuxizumab, onvatilimab, opicinumab, oportuzumab monatox, oregovomab, orticumab, otelixizumab, otilimab, otlertuzumab, oxelumab, ozanezumab, ozoralizumab, pagibaximab, palivizumab, pamrevlumab, panitumumab, pankomab, panobacumab, parsatuzumab, pascolizumab, pasotuxizumab, pateclizumab, patritumab, PDR001, pembrolizumab, pemtumomab, perakizumab, pertuzumab, pexelizumab, pidilizumab, pinatuzumab vedotin, pintumomab, placulumab, plozalizumab, pogalizumab, polatuzumab vedotin, ponezumab, porgaviximab, prasinezumab, prezalizumab, priliximab, pritoxaximab, pritumumab, PRO 140, quilizumab, racotumomab, radretumab, rafivirumab, ralpancizumab, ramucirumab, ranevetmab, ranibizumab lucentis, ravagalimab, ravulizumab, raxibacumab, refanezumab, regavirumab, relatlimab, remtolumab, reslizumab, rilotumumab, rinucumab, risankizumab, rituximab, rivabazumab pegol, rmabrabishield,

robatumumab, roledumab, romilkimab, romosozumab, rontalizumab,

rosmantuzumab, rovalpituzumab tesirine, rovelizumab, rozanolixizumab, ruplizumab, SA237, sacituzumab govitecan, samalizumab, samrotamab vedotin, sarilumab, satralizumab, satumomab pendetide, secukinumab, selicrelumab, seribantumab, setoxaximab, setrusumab, sevirumab, SGN-CD19a, SHP647, sibrotuzumab, sifalimumab, siltuximab, simtuzumab, siplizumab sirtratumab vedotin, sirukumab, sofituzumab vedotin, solanezumab, solitomab, sonepcizumab, sontuzumab, spartalizumab, stamulumab , sulesomab, suptavumab, sutimlimab, suvizumab, suvratoxumab, tabalumab, tacatuzumab tetraxetan, tadocizumab, talacotuzumab, talizumab, tamtuvetmab, tanezumab, taplitumomab paptox, tarextumab, tavolimab, tefibazumab, telimomab aritox, telisotuzumab vedotin, tenatumomab, teneliximab, teplizumab, tepoditamab, teprotumumab, tesidolumab, tetulomab, tezepelumab, TGN1412, tibulizumab, tigatuzumab, tildrakizumab, timigutuzumab, timolumab, tiragotumab, tislelizumab, tisotumab vedotin, TNX-650, tocilizumab,

tomuzotuximab, toralizumab, tosatoxumab, tositumomab, tovetumab, tralokinumab, trastuzumab, trastuzumab emtansine, TRBS07, tregalizumab, tremelimumab, trevogrumab, tucotuzumab celmoleukin, tuvirumab, ublituximab, ulocuplumab, urelumab, urtoxazumab, ustekinumab, utomilumab, vadastuximab talirine, vanalimabmab, vandortuzumab vedotin, vantictumab, vanucizumab, vapaliximab, varisacumab, varlilumab, vatelizumab, vedolizumab, entyvio, veltuzumab, vepalimomab, vesencumab, visilizumab, vobarilizumab, volociximab,

vonlerolizumab, vopratelimab, vorsetuzumab mafodotin, votumumab, vunakizumab, xentuzumab, XMAB-5574, zalutumumab, zanolimumab, zatuximab, zenocutuzumab, ziralimumab, zolbetuximab, zolimomab aritox, ponezumab, and combinations thereof.

40. The nanoparticle of any preceding claim, wherein the physiologically active agent is a therapeutic protein selected from Lepirudin, Dornase alfa, Denileukin diftitox, Bivalirudin, Leuprolide, Peginterferon alfa-2a, Alteplase, Interferon alfa-nl,

Darbepoetin alfa, Reteplase, Epoetin alfa, Salmon Calcitonin, Interferon alfa-n3, Pegfilgrastim, Sargramostim, Secretin, Peginterferon alfa-2b, Asparaginase,

Thyrotropin Alfa, Antihemophilic Factor, Anakinra, Gramicidin D, Intravenous Immunoglobulin, Anistreplase, Insulin Regular, Tenecteplase, Menotropins,

Interferon gamma- lb, Interferon Alfa-2a, Recombinant, Coagulation factor Vila, Oprelvekin, Palifermin, Glucagon recombinant, Aldesleukin, Botulinum Toxin Type B, Lutropin alfa, Insulin Lispro, Insulin Glargine, Collagenase, Rasburicase,

Imiglucerase, Alpha- 1 -proteinase inhibitor, Pegaspargase, Interferon beta- la, Pegademase bovine, Human Serum Albumin, Eptifibatide, Serum albumin iodonated, Follitropin beta, Vasopressin, Interferon beta- lb, Hyaluronidase, Insulin, porcine, Digoxin Immune Fab (Ovine), Daptomycin, Pegvisomant, Botulinum Toxin Type A, Pancrelipase, Streptokinase, Alglucerase, Faronidase, Urofollitropin, Serum albumin, Choriogonadotropin alfa, Antithymocyte globulin, Filgrastim, Coagulation factor ix, Becaplermin, Agalsidase beta, Interferon alfa-2b, Oxytocin, Enfuvirtide, Idursulfase, Alglucosidase alfa, Exenatide, Mecasermin, Pramlintide, Galsulfase, Abatacept, Cosyntropin, Corticotropin, Insulin aspart, Insulin detemir, Insulin glulisine, Pegaptanib, Nesiritide, Thymalfasin, Defibrotide, Natural alpha interferon OR multiferon, Glatiramer acetate, Preotact, Teicoplanin, Sulodexide, Teriparatide, Liraglutide, Belatacept, Buserelin, Velaglucerase alfa, Tesamorelin, Taliglucerase alfa, Aflibercept, Asparaginase erwinia chrysanthemi, Ocriplasmin, Glucarpidase, Teduglutide, Insulin, isophane, Epoetin zeta, Fibrinolysin aka plasmin, Follitropin alpha, Romiplostim, Lucinactant, Aliskiren, Ragweed Pollen Extract, Somatotropin Recombinant, Drotrecogin alfa, Alefacept, OspA lipoprotein, Urokinase, Abarelix, Sermorelin, Aprotinin, Albiglutide, Ancestim, Anti thrombin Alfa, Antithrombin III human, Asfotase Alfa, Autologous cultured chondrocytes, Beractant, Cl Esterase Inhibitor (Human), Coagulation Factor XIII A-Subunit (Recombinant), Conestat alfa, Daratumumab, Desirudin, Dulaglutide, Elosulfase alfa, Elotuzumab, Fibrinogen Concentrate (Human), Filgrastim-sndz, Gastric intrinsic factor, Hepatitis B immune globulin, Human calcitonin, Human Clostridium tetani toxoid immune globulin, Human rabies virus immune globulin, Human Rho(D) immune globulin,

Hyaluronidase (Human Recombinant), Immune Globulin Human, Turoctocog alfa, Tuberculin Purified Protein Derivative, Simoctocog Alfa, Sebelipase alfa,

Sacrosidase, Prothrombin complex concentrate, Poractant alfa, Pembrolizumab, Peginterferon beta- la, Metreleptin, Methoxy polyethylene glycol-epoetin beta, Insulin Pork, Insulin Degludec, Insulin Beef, Thyroglobulin, Anthrax immune globulin human, Anti-inhibitor coagulant complex, Anti-thymocyte Globulin (Equine), Cl Esterase Inhibitor (Recombinant) , Chorionic Gonadotropin

(Recombinant), Coagulation factor X human, Efmoroctocog alfa, Factor IX Complex (Human), Hepatitis A Vaccine, Human Varicella-Zoster Immune Globulin,

Lenograstim, Pegloticase, Protamine sulfate, Protein S human, Sipuleucel-T, Somatropin recombinant, Susoctocog alfa, Thrombomodulin Alfa, and combinations thereof.

41. The nanoparticle of any preceding claim, wherein the physiologically active agent is crosslinked.

42. The nanoparticle of any preceding claim, wherein the physiologically active agent is dispersed in a matrix.

43. The nanoparticle of any preceding claim, wherein the physiologically active agent is dispersed in a matrix comprising one or more polymers or lipids.

44. The nanoparticle of any preceding claim, wherein the physiologically active agent is dispersed in a crossl inked polymer matrix, wherein the crosslinked polymer is selected from the group consisting of polyphosphazenes, polycyanoacrylates, polyesters, polyhydroxyalkanoates, polyanhydrides, polydixanones, polyorthoesters, polyesteramides, polyamido amides, polythioesters, collagen, fibrin, fibrinogen, gelatin, polysaccharides, and combinations thereof.

45. The nanoparticle of any preceding claim, wherein the crosslinked polymer matrix comprises a polyester selected from the group consisting of poly(lactic acid), poly(glycolic acid), poly(caprolactone), poly(propylene fumarate), copolymers thereof, and combinations thereof.

46. The nanoparticle of any preceding claim, wherein the crosslinked polymer comprises a polysaccharide selected from the group consisting of chitosans, celluloses, modified celluloses, alginates, pectins, pullulans, hyaluronic acids, starches, amyloses, dextrans

47. The nanoparticle of any preceding claim, wherein the physiologically active agent is loaded into smaller nanoparticles having a particle size no greater than 50 nm, said smaller nanoparticles dispersed in a matrix.

48. The nanoparticle of any preceding claim, wherein the stimuli-cleavable crosslinks are degraded upon exposure to oxidant, reductant, acid, irradiation, ultrasound, heat or magnetic exposure.

49. The nanoparticle of any preceding claim, wherein the stimuli-cleavable crosslinks are degraded by infrared radiation, microwave irradiation, or UV irradiations.

50. The nanoparticle of any preceding claim, wherein stimuli-cleavable crosslinks are degraded by reactive oxygen species.

51. The nanoparticle of any preceding claim, wherein the stimuli-cleavable crosslinks comprise functional groups selected from disulfide bonds, trisulfide bonds, diselenide bonds, thioacetals, acetals, oxalates, imines, and peptide bonds.

52. A method of preparing the nanoparticle of any preceding claim, comprising contacting the physiologically active agent with at least one crosslinking agent at a concentration and time sufficient to prepare a crosslinked nanoparticle.

53. The method of any preceding claim, wherein the crosslinking agent has the formula:


wherein L represent a linking group, R1 is in each case independently hydrogen or Ci- ealkyl, or where two R1 groups form a ring; and

E is an electrophilic group.

54. The method of any preceding claim, wherein E comprises an imidoester, an N- hydroxysuccinimide ester, a maleimide, a haloacetyl, or a pyridyl disulfide.

55. The method of any preceding claim, wherein E has the formula:


56. The method of any preceding claim, wherein L is selected from Ci-ioalkyl,

poly(ethylene glycol), or aryl.

57. The method of any preceding claim, wherein the crosslinking agent is provided in an amount from 0.00001-0.1 wt%, relative to the physiologically active agent.