Polyethylene Glycol-Based Dendrons

US 20120183578A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2012/0183578 A1 (43) Pub. Date: Sinko et al. (54) (75) POLYETHYLENE GLYCOL-BASED DENDRONS Inventors: Related U.S. Application Data (60) Patrick J. Sinko, Annandale, NJ NJ (US); Stanley Stein, East Brunswick, NJ (US) Int. Cl. A61K 39/00 A61K 47/06 A61P 35/00 A61P 31/12 A61P 37/02 C12Q 1/02 (2006.01) RUTGERS, THE STATE C07K 4/00 (2006.01) Singh, Highland Park, NJ (US); Xiaoping Zhang, Edison, NJ (US); (51) Matthew S. Palombo, Marmora, (73) Assignee: UNIVERSITY OF NEW JERSEY, New Brunswick, NJ (US) (21) Appl. No.: (22) Filed: Provisional application No. 61/413,352, ?led on Nov. 12, 2010. Publication Classi?cation (US); Jieming Gao, New Brunswick, NJ (US); Yashveer Jul. 19, 2012 (52) (57) (2006.01) (2006.01) (2006.01) (2006.01) (2006.01) U.S. Cl. ....... .. 424/278.1; 435/29; 514/773; 530/323 ABSTRACT 13/296,213 The instant invention relates to polyethylene glycol-based dendrons, otherwise known as PEGtide dendrons, composi Nov. 14, 2011 tions thereof and methods of use. Patent Application Publication Jul. 19, 2012 Sheet 1 0f 17 US 2012/0183578 A1 ?ianim Lysina QMM‘VM: Patent Application Publication Jul. 19, 2012 Sheet 2 0f 17 Mm US 2012/0183578 A1 Patent Application Publication Jul. 19, 2012 Sheet 3 0f 17 US 2012/0183578 A1 iiiiigggsésw Z3. \“M iiiifiifi Patent Application Publication Jul. 19, 2012 Sheet 4 0f 17 :1 k' ‘w :1 a ii; M5 w <"I {E 3:3 I a» US 2012/0183578 A1 3 Patent Application Publication Jul. 19, 2012 Sheet 5 0f 17 §1 m as. m: 1% Jim-mans: a»; US 2012/0183578 A1 Patent Application Publication Jul. 19, 2012 Sheet 6 0f 17 US 2012/0183578 A1 w? 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US 2012/0183578 A1 MW Jul. 19, 2012 US 2012/0183578 A1 POLYETHYLENE GLYCOL-BASED DENDRONS CROSS-REFERENCE TO RELATED APPLICATIONS appropriate locations for drug or gene material loading and transportation. Generally, payloads are held inside the den drimer Cavities via covalent attachment, hydrophobic inter action, hydrogen bonds, or charge interaction. The degrad able or cleavable bonds like ester, imine, acetal and ketal are §119(e) to US. Provisional Patent Application Ser. No. 61/413,352, ?led on Nov. 12, 2010, the disclosure ofWhich is more preferred in covalent bonding since they function as the triggers of environmental response release. [0006] The ?rst dendrimer-like structure Was reported by incorporated herein by reference in its entirety. Vogtle and coWorkers in 1978 (Egon Buhleier, W. W., Syn [0001] This application claims priority under 35 USC GOVERNMENT SUPPORT [0002] This invention Was made With government support under A1051214 aWarded by the National Institutes of Health. The federal government has certain rights in this invention. thesis, 155-158, (1978)), Who synthesiZed polypropylen imine (PPI). The term “dendrimer” Was suggested by Tomalia et al. in 1985 Who along With NeWkome et al. (NeWkome, G. R., Journal Of Organic Chemistry 50, 2003-2004 (1985)) synthesiZed dendrimers of higher generation With Well-de ?ned structures (Tomalia, D. A., Polymer Journal 17, 117 132, (1985)). The dendrimers are prepared using either diver FIELD OF THE INVENTION gent (NeWkome, G. R., Journal Of Organic Chemistry 50, [0003] The present invention relates to the synthesis of novel polyethylene glycol-based (PEGTide) dendrons, a car rier for the in vivo delivery of a therapeutic agent, composi 2003-2004 (1985)) or convergent (HaWker, C. 1., Journal Of The American Chemical Society 112, 7638-7647, (1990)) tions thereof and methods of use. In particular, this invention is directed to the PEGtide dendron composed from polyeth ylene glycol (PEG) and amino acids, as a carrier for the in vivo delivery of a therapeutic agent. The invention comprises a dendron structure, a tree like structure, similar in structure to a dendrimer, consisting of peptide subunits interspersed With short monodisperse poly(ethylene glycol)/PEG subunits. The PEG subunits endoW dendrons With favorable pharmaceuti cal (e.g., long circulation time, reduced toxicity, increased Water solubility etc.) and pharmacological (e.g., biocompat ibility, reduced immunogenicity, biodistribution etc.) proper ties, generally knoWn to be associated With PEG. PEGtide dendrons are useful for various applications including drug delivery and diagnostic purposes because they can be modi ?ed With therapeutic, targeting, biologic, and diagnostic agents. [0004] The agent-loading mechanisms include physical encapsulation, charge interaction, covalent binding or their combinations. Payload release is performed through diffu sion, carrier degradation, conjugated bond cleavage or their combinations. Compared to traditional commercial dendrim ers, like polyamidoamine (PAMAM) or polypropyleneimine (PEI), PEGTide dendron oWns improved biocompatibility, higher loading capability and Wider application ?elds. strategies. In a divergent approach, pioneered by Vogtle (Egon Buhleier, W. W., Synthesis, 155-158, (1978)), Donald A. Tomalia, and NeWkome (NeWkome, G. R., Journal Of Organic Chemistry 50, 2003-2004 (1985)), dendrimer syn thesis proceeds outWards from multifunctional core to sur face, Whereas in a convergent approach, pioneered by Frechet and coWorkers (HaWker, C. 1., Journal Of the American Chemical Society 112, 7638-7647, (1990)), dendrimer syn thesis proceeds inward, from surface to core. Some of the most common dendrimers are polyamidoamine (PAMAM); poly(L-lysine) (PLL); polyamide; polyester (PGLSA-OH); polypropylenimine (PEI); and poly(2,2-bis(hydroxymethyl) propionic acid (bis-MPA). [0007] Although dendrimers have found Wide application in drug and gene delivery, and diagnostics, their use is restricted due to reticuloendothelial system (RES) uptake, immunogenicity, hemolytic toxicity, cytotoxicity, hydropho bicity. [0008] Statistics on dendrimer PEGylation shoWs that most of the studies have focused on dendrimer surface modi?ca tion. The toxicity from dendrimer branch/core structure is still a potential threat for safe use of dendrimers. In addition, since the siZes of currently developed dendrimers are less than 100 nanometers, these are prone to excretion by kidney. BACKGROUND OF THE INVENTION [0005] Dendrimers (Greek: dendri: “tree-like” and meros: “part of’) are monodisperse macromolecules With Well-de ?ned branched architecture and symmetrical morphology (Bosman, A. W., Chemical Reviews 99, 1665-1688, (1999)). Dendrimers are comprised of a series of branches extending outWard from an inner core. These branches are arranged in layers, called generations, and represent the repeating units (monomer) of a dendrimer. A typical dendrimer molecule contains an inner core, layers of repeating units, and multiple SUMMARY OF THE INVENTION [0009] The present invention relates to the synthesis of novel polyethylene glycol-based (PEGTide) dendrons, a car rier for the in vivo delivery of a therapeutic agent, and its application in drug delivery, diagnostic applications, vaccines and related methods to treat a condition, and generate an immune response. [0010] Developing novel dendrons With larger siZe (100 to terminal functional groups. The active moieties are either 200 nm) is needed in order to meet the advanced requirements encapsulated into the core/cavities or grafted onto the surface of drug delivery applications in the pharmaceutical ?eld, including better biocompatibility, higher siZe, and tunable of dendrimers. The dendrimers are distinct from other nano carriers in that they possess a tunable structure, empty intramolecular cavity, and multifunctional surface (Duncan, R., Advanced Drug Delivery Reviews 57, 2215-2237, interior void siZe/ structure, design. The development of a PEGtide dendron is disclosed in this patent. [0011] An embodiment of the present invention is a carrier (2005)). The existence of interior void spaces inside the den for the in vivo delivery of a therapeutic agent represented by drimer, particularly the high generation dendrimers, becomes Formula 1: Jul. 19, 2012 US 2012/0183578 A1 (I) R— PEG-La-AA-Lb-PEG-R LC Lo [0012] wherein R is: R1—PEG-Ld-AA1-Le-PEG-R1 L/ | [0024] In certain embodiments, R2 is T1 or T2. [0025] In certain embodiments, AA, AA], AA2, and AA3 [0013] R1 is: are each lysine; La, Lb, LC, Ld, Le, Lf, Lg, Lh, L], Lk, Lm, and R2—PEG-Lg-1|\A2-Lh-PEG-R2 Ln are selected from alanine, arginine or histidine; T l and T2 are ?uorenylmethyloxycarbonyl protected amino groups. [0026] In certain embodiments, AA, AAl, AA2, and AA3 are each lysine; La, Lb, LC, Ld, Le, Lf, Lg, Lh, L], Lk, Lm, and L/ | L” are each alanine; T 1 and T2 are ?uorenylmethyloxycarbo nyl protected amino groups. [0027] In accordance With any of the above embodiments, the invention further comprises at least one therapeutic agent. In a further embodiment, a therapeutic agent may be a phar [0014] R1 is: T1—PEG-Lg-AA2-Lh-PEG-T2 Ln [0015] [0022] In certain embodiments, R may not contain L d or L8. In certain embodiments, R1 may not contain PEG. In certain embodiments, R1 may not contain L8 or Lh. In certain embodiments, R2 may not contain PEG. In certain embodi ments, R2 may not contain Lk or Lm. [0023] In certain embodiments, the T 1 and T2 amino groups are selected from free amino groups, protected amino groups and amino groups modi?ed With therapeutic agent. PEG is linear or branched poly(ethylene glycol); maceutically active, diagnostic, biologic, imaging, targeting agent or an adjuvant. [0028] In accordance With any of the above embodiments, the invention may further comprise at least one therapeutic agent and a pharmaceutically acceptable carrier. In a further embodiment, a therapeutic agent may be an antineopastic or an antiretroviral agent. [0029] In accordance With any of the above embodiments, [0016] AA, AA], AA2, and AA3 are each independently the invention may be a vaccine comprising a composition lysine or ornithine; represented by Formula 1. [0017] La, Lb, LC, Ld, Le, Lf, Lg, Lh, L], Lk, Lm, and Ln are [0030] The present invention further provides a method for treating a condition linked to a hyperproliferative disorder, comprising administering to a patient in need thereof an effective amount of a composition represented by Formula 1 each independently 0-8 amino acids long and selected froma lanine, glycine, val-ine, leucine, isoleucine, statine, phenylg lycine, phenylalanine, cysteine, penicillamine, homocys teine, arginine, histidine, norvaline, norleucine, 2-?uorophenylalanine, 3-?uorophenylalanine, 4-?uorophe nylalanine, [3-cycloheXyl-alanine, 2-aminobutyric acid, 4-aminobutyric acid, 6-aminobutyric acid and ot-amino-bu tyric acid; and [0018] T1 and T2 are each a ?nal PEG terminus indepen dently selected from an amino group, acetyl group, ?uore nylmethyloxycarbonyl group, therapeutic agent, diagnostic agent, biologic agent, targeting agents, or adjuvant. [0019] In certain embodiments, LC may be selected from cysteine, penicillamine, homocysteine and alanine. [0020] In certain embodiments, Formula (I) may not con tain La, Lb, LC, Ld, Le, Lf, Lg, Lh, L], Lk, Lm, or L”. In certain other embodiments, Formula (I) may be represented by: further comprising an antineoplastic agent. [0031] The present invention also provides a method for treating stimulating an immune response, comprising admin istering to a patient in need thereof an effective amount of a vaccine comprising a composition represented by Formula 1 further comprising an antigen. BRIEF DESCRIPTION OF THE DRAWINGS [0032] FIG. 1 depicts the structure of a generation 3.0 (G3. 0) PEGtide dendron. [0033] FIG. 2 is a MALDI-TOF spectrum of a generation 3.0 (G3.0) PEGtide dendron: C-A-K-{A-PEG-K-[A-PEG K-(A-PEG)2l2}2 [0034] FIG. 3 shoWs the results of dynamic light scattering (DLS) analysis of a generation 3.0 (G3.0) PEGtide dendron, Which shoWs that the dendron has a radius of 53.9 nm. R — PEG-AA-PEG-R LC [0035] FIG. 4 depicts the structure of 5-carboxy?uorescein (FAM) and arginine-containing PEGTide dendron G30 [0036] FIG. 5 is a MALDI-TOF spectrum of 5-carboxy ?uorescein (FAM) and arginine-contaning PEGTide dendron G3 .0 With folloWing structure: A-K(FAM)-PEG-K-{A-PEG [0021] In certain embodiments, Formula (I) may not con tain PEG. In certain other embodiments, R may not contain PEG, and Formula (I) may be represented Wherein R is: K-[A-R-R-A-R-A-K-(A-PEG)2]2}2. [0037] FIG. 6 illustrates the structure of DV3-containing tetravalent PEGTide dendron G2.0.