MACROMOLÉCULAS HIPER-RAMIFICADAS TIPO DENDRÍMERO À BASE DE POLI(ETILENO GLICOL), MÉTODOS DE PREPARAÇÃO E USO DAS MESMAS

Abstract

In certain embodiments, the present invention relates to dendrimer-like hyperbranched macromolecules for several uses such as medical or biopharmaceutical applications, or non- medical or industrial uses, such as antibody purification, cosmetic applications, catalytic applications, applications in electronics, agriculture, food, filtration and further applications. In embodiments, the present invention relates to a hyperbranched macromolecule for drug delivery, comprising polyethylene glycol (PEG) units and at least one active agent conjugated to the hyperbranched macromolecule. Further, methods for synthesizing, purifying and characterizing such hyperbranched macromolecules and methods of treatment of a medical condition such as treatment of an ocular disease are provided.

Description

[0001] In certain embodiments, the present invention relates to hyper-branched dendrimer-like macromolecules for various uses, such as medical or biopharmaceutical applications, or non-medical or industrial uses, such as antibody purification, cosmetic applications, catalytic applications, applications in electronics, agriculture, food, filtration, and other applications. In medical applications, the hyper-branched molecules are conjugated with active agents, such as drugs, peptides, or proteins. Furthermore, the present invention relates, in certain embodiments, to a hyper-branched macromolecule for drug delivery, comprising polyethylene glycol (PEG) units and at least one active agent conjugated to the hyper-branched macromolecules. In certain other embodiments, the present invention relates to methods for synthesizing, purifying, and characterizing such hyper-branched dendrimer-like macromolecules.The present invention also relates, in certain embodiments, to methods of treating a medical condition, such as the treatment of an eye disease. BACKGROUND

[0002] Controlled administration and stabilization of therapeutic agents has been a major area of ​​research in recent years. Controlled administration improves therapies, facilitates administration, and leads to greater efficacy, better adherence, fewer side effects, and better overall therapeutic outcomes.

[0003] The eye is a unique organ of perfection and complexity and, in many ways, a microcosm of the body. It offers a great opportunity for nanomedicine, as it is easily accessible, Petition 870250084859, dated 09 / 19 / 2025, page 10 / 290 2 / 208 allowing the direct administration of drugs / genes to maximize the therapeutic effect and minimize side effects. The development of appropriate delivery systems that can sustain and deliver therapeutic agents to target tissues is a fundamental challenge that nanotechnology can address. Current delivery systems for anterior segment ocular disorders, such as punctal plugging, micro- and nanoparticle encapsulation, microneedle systems, iontophoresis, different types of intravitreal implants, etc., represent state-of-the-art tools for sustained and controlled drug delivery in the eye.

[0004] Dendrimers and hyper-branched polymers have attracted the attention of scientists in the field of drug and gene delivery over the past two decades due to their versatility, complexity, and multi-branched properties. Dendrimers are nanostructured, branched, monodisperse, and highly symmetrical, tree-like polymers that possess repeatable building blocks with well-defined size, adaptable structure, and potentially favorable ocular biodistribution. Dendrimers have been widely explored as a novel platform for bioactive drug delivery due to unique biological properties such as high drug loading, lipid bilayer interactions, targeting potential, blood plasma retention time, filtration, intracellular internalization, biodistribution, transfection, good colloidal and biological stability.Several dendrimers have been explored for drug delivery, including polymer-based dendrimers such as polyamidoamine (PAMAM), poly(propylene imine) (PPI), polyester, polyether, poly-L-lysine, triazine, melamine, poly(glycerol-co-succinic acid), poly(glycerol) and poly[2,2-bis(hydroxymethyl)propionic acid], and other types of dendrimers made from peptides, crystal-forming dendrimers. Petition 870250084859, dated 09 / 19 / 2025, page 11 / 290 3 / 208 liquids, carbosilane, etc. (for an overview, see, for example, “Dendrimer as a nanocarrier for drug delivery” Prashant Kesharwani, Keerti Jain, Narendra Kumar Jain, Progress in Polymer Science 39 (2014) 268-307; and “Dendrimer-based drug delivery systems: history, challenges and latest developments” Juan Wang, Boxuan Li, Li Qiu, Xin Qiao and Hu Yang, Journal of Biological Engineering (2022) 16:18).

[0005] Among the applications of dendrimers in drug delivery, those related to the treatment and management of ocular diseases are of particular interest. Ocular drug therapies suffer from some significant disadvantages, including frequent administration, poor penetration, and / or rapid elimination. The use of dendrimers as a strategy to overcome obstacles to the traditional treatment of ocular diseases shows promising progress in this field, and the approach to ocular safety with dendrimers aims to take into account the most advanced science to date. Several ocular applications of dendrimers and dendrimer delivery systems are known, cf. “Dendrimer as a nanocarrier for drug delivery” Prashant Kesharwani, Keerti Jain, Narendra Kumar Jain, Progress in Polymer Science 39 (2014) 268-307.However, most of these applications are still in the early stages of laboratory exploration, and only a few commercial products for treating eye diseases with dendrimer administration are known to date.

[0006] There is therefore a need to provide strategies to optimize drug administration and site-specific targeting. There is also a need to provide sustained-release drug delivery systems that allow for increased half-life of the active agent and improved efficacy and Petition 870250084859, dated 09 / 19 / 2025, page 12 / 290 4 / 208 the bioavailability of the active agents. For the delivery of biomolecules such as peptides and proteins, there is still a need to improve the avidity and extend the half-life of the biomolecules binding to the receptor, providing possibilities for multivalent delivery.

[0007] In addition, dendrimers may have various applications in non-medical fields or industrial uses, such as antibody purification, cosmetic applications, catalysis, electronics, agriculture, food, filtration, energy storage, building materials, and other applications.

[0008] All references cited herein are incorporated by reference in their entirety for all purposes. OBJECTIVES AND SUMMARY OF THE INVENTION

[0009] Therefore, it is an objective of certain embodiments of the present invention to provide a dendrimer platform based on polyethylene glycol building blocks that are highly variable and flexible to be adapted to various uses and applications, and easy to synthesize.

[0010] An additional objective of certain embodiments of the present invention is to provide systems with optimized drug delivery and specific targeting to the site, specifically to treat an ocular condition.

[0011] Another objective of certain embodiments of the present invention is to provide delivery systems for sustained-release drug administration that allow increasing the half-life of the active agent, improving the efficacy, avidity and bioavailability of the active agents.

[0012] Another objective of certain embodiments of the present invention is to provide biodegradable drug delivery systems that can be adjusted for their degradation rate and active agent release rate using a wide range of different groups. Petition 870250084859, dated 09 / 19 / 2025, page 13 / 290 5 / 208 biodegradable molecules, including hydrolyzable groups and linkages, are incorporated into the molecular structure of the drug delivery system. Biodegradable drug delivery systems of certain modalities must be fully reabsorbable and degradable into the initial building blocks, which can be easily eliminated from local tissues and ultimately from the body.

[0013] Another objective of certain embodiments of the present invention is to provide biodegradable drug delivery systems that lead to a greater binding affinity and / or avidity of the active agent, for example, biomolecules such as peptides or proteins, to biological targets.

[0014] A further objective of certain embodiments of the invention, and an aspect, is to provide methods for treating a patient's disease / medical condition.

[0015] An additional objective of certain embodiments of the present invention is to provide dendrimers for applications also in non-medical fields or industrial uses, such as antibody purification, cosmetic applications, catalytic applications, applications in electronics, agriculture, food, filtration, energy storage, building materials, coatings, adhesives, water purification, oil recovery, fragrance release, papermaking, environmental detection and release systems, membranes, textiles, printing inks, surface chemistry applications, thickeners, detergents, rheology modifiers, scaffolding or 3D printing.

[0016] The above objects are solved by the inventions as described in the independent claims. Advantageous modifications are disclosed in the dependent claims.

[0017] Some aspects of the present disclosure are directed to a hyper-branched macromolecule comprising a central unit having at least 3 c-connectivities, a plurality of arms Petition 870250084859, dated 09 / 19 / 2025, page 14 / 290 6 / 208 polymeric arms connected to the central unit in the c-connectivities, at least one of the polymeric arms being connected by a hydrolyzable link to a dendritic constitutional repeat unit (DCRU), the dendritic constitutional repeat unit (DCRU) comprising a branching unit connected to at least two polymeric arms, each comprising a terminal group or being connected to a next dendritic constitutional repeat unit that may again be connected to another dendritic constitutional repeat unit, the polymeric arms of the outermost dendritic constitutional repeat unit each comprising a terminal group; wherein the polymeric arms consist of polyethylene glycol (PEG) units; wherein optionally at least one active agent is conjugated to at least one of the outermost polymeric arms.In some embodiments, at least 10%, preferably about 20 to 100%, of the chemical bonds of the connections can be cleaved by hydrolysis. The bonds cleavable by hydrolysis are preferably ester bonds. In certain embodiments, the ester bonds are introduced using ligands derived from organic diacids.

[0018] In some embodiments, the building blocks or fragments of the hyper-branched macromolecule obtained after cleavage of all hydrolyzable bonds of the connections have an average molecular weight (Mn) of less than 50,000 Daltons. The active agent may be covalently or non-covalently linked to the hyper-branched macromolecule. In certain embodiments, the active agent is covalently conjugated to the hyper-branched macromolecule.

[0019] In certain respects, the hyper-branched macromolecule is a branched macromolecule similar to a G0 generation dendrimer, in which the surface terminal groups of the branched macromolecule are the terminal groups on the arms. Petition 870250084859, dated 09 / 19 / 2025, page 15 / 290 7 / 208 polymeric units connected to the central unit without additional connections to DCRUs. In other respects, the hyper-branched macromolecule is a hyper-branched macromolecule similar to a higher-generation G x dendrimer, with x being an integer from 1 to 10 defining the number of consecutively connected dendritic constitutional repeat units in the hyper-branched macromolecule, the polymeric arms of the outermost dendritic constitutional repeat unit each comprising a terminal group, wherein at least one active agent is conjugated to at least one of the outermost polymeric arms.

[0020] In certain aspects of the present disclosure, the central unit and the branching units of the hyperbranched macromolecule are the same or different and, independently of each other, have a c connectivity of 3 to 10, or 4 to 8, or 4 to 6, or 4. In certain aspects, the central unit and the branching unit are the same or different and are each derived from a polyol with at least 3 hydroxyl groups. In certain aspects, the polyol is selected from the group consisting of glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol, or sorbitol.

[0021] In certain aspects of the present disclosure, the polymeric arms in the hyper-branched macromolecule comprise PEG units with average molecular weights (Mw) in the range of about 1,000 to about 80,000 Daltons, or about 10,000 to about 60,000 Daltons, or about 15,000 to about 50,000 Daltons. In certain aspects, the average molecular weight of the core-linked polymeric arm PEG units is the same as or different from that of the polymeric arms in the dendritic constitutional repeat units. The average molecular weight of the core-linked polymeric arm PEG units may be higher or lower than that of the arms Petition 870250084859, dated 09 / 19 / 2025, p. 16 / 290 8 / 208 polymeric in the dendritic constitutional repeat units. For higher-generation Gx hyper-branched macromolecules, with x being an integer from 2 to 10, the average molecular weight of the PEG units of the polymeric arm may decrease or increase from the innermost polymeric arms to the outermost polymeric arms.

[0022] In certain aspects of the present disclosure, the terminal groups attached to the outermost polymeric arms are grafted onto the ends of the polymeric arms directly or by means of a difunctional linker comprising hydrolyzable linkages, such as a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic, heteroaliphatic or aromatic or heteroaromatic group. In other embodiments, the terminal groups attached to the outermost polymeric arms are functional groups selected from electrophiles, such as activated ester groups, such as succinimidyl esters, succinimidyl carbonates; nitrophenyl carbonates, aldehydes, ketones, acrylates, acrylamides, maleimides, vinyl sulfones, iodoacetamides, alkenes, alkynes, azides, norbornenes, epoxides, mesylates, tosylates, tresylates, cyanurates, orthopyridyl or halogen disulfides;Nucleophiles, such as an amine, such as a primary amine, a hydroxyl group, an alcohol, a thiol, an azide anion, and a carboxyl group; functional groups for click chemistry; functional groups for cycloadditions, such as 1,3-dipolar cycloadditions, [3+2] cycloadditions such as alkene-nitrone cycloadditions or alkyne-nitrone cycloadditions, [4+2] cycloadditions; functional groups for thiol-ene reactions; hetero-Diels-Alder cycloadditions; functional groups for nucleophilic ring openings; functional groups for non-aldol-type carbonyl reactions; functional groups for addition reactions to multiple carbon-carbon bonds, polymerizable vinyl groups, or combinations thereof. Petition 870250084859, dated 09 / 19 / 2025, page 17 / 290 9 / 208

[0023] In certain aspects of the present disclosure, the terminal groups attached to the outermost polymeric arms are ligand-spaced functional groups selected from succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ), and succinimidyl glutaramide (SGA).

[0024] In certain aspects of the present disclosure, the terminal groups attached to the outermost polymeric arms are functional groups selected from an alkyne compound, such as a dibenzocyclo-octyne (DBCO), or a bicyclo[6.1.0]-nonyne (BCN); or a norbornene, or a trans-cyclo-octene (TCO); an azide, a 3,4-dihydroxyphenylacetic acid (DHPA) or a tetrazine (Tz).

[0025] In certain aspects of the present disclosure, the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed by click chemistry.

[0026] In certain aspects of the present disclosure, the connection is formed by click chemistry, wherein the connection is formed by the reaction of a polymer arm functionalized with a tense or terminal alkyne, cycloalkyne or alkene moiety with a polymer arm functionalized with an azide or tetrazine moiety in a SPAAC or IEDDA type click chemistry coupling reaction. In certain aspects, the alkyne moiety is a dibenzocyclooctyne moiety.

[0027] In certain aspects of the present disclosure, the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed between the polymeric arms connected to the central unit and the polymeric arms connected to the branching unit of the dendritic constitutional repeat units and / or between the arms Petition 870250084859, dated 09 / 19 / 2025, page 18 / 290 10 / 208 polymeric arms of a dendritic constitutional repeat unit and the polymeric arms of consecutive dendritic constitutional repeat units.

[0028] In certain aspects of this disclosure, the active agent conjugated to at least one of the terminal groups located on the surface of the hyper-branched macromolecule is selected from the group consisting of therapeutically or diagnostically active agents.

[0029] In certain aspects of this disclosure, the active agent conjugated to at least one of the terminal groups located on the surface of the dendrimer is selected from among steroids; non-steroidal anti-inflammatory drugs (NSAIDs), such as diclofenac, ibuprofen, meclofenamate, mefanamic acid, salsalate, sulindac, tolmetin, ketoprofen, diflunisal, piroxicam, naproxen, etodolac, flurbiprofen, fenoprofen C, indomethacin, celecoxib, ketorolac, nepafenac; intraocular pressure-lowering drugs; antibiotics, such as ciprofloxacin; analgesics, such as bupivacaine; calcium channel blockers, such as nifedipine; Cell cycle inhibitors, such as simvastatin; proteins, such as insulin;hydrophilic small molecule drugs, including carboxylic acid salts and amine salts; hydrophobic small molecule drugs, hydrophilic peptides and protein drugs, such as insulin, single-chain antibody fragments, Fab fragments, IgG antibodies, fusion antibodies, etc.; aptamers; particularly bupivacaine (BPV-HCl or base), ropivacaine (RPV), dexamethasone, travoprost, axitinib, non-steroidal anti-inflammatory drugs (NSAIDs), steroids, antibiotics, analgesics, calcium channel blockers, cell cycle inhibitors, chemotherapeutic agents, antiviral drugs, anesthetics, hormones, anticancer drugs, antineoplastic agents, viruses, gene delivery viruses such as AAVs, protein ligands such as nanobodies, affibodies, ankyrins, DARPins, etc., or any others; Petition 870250084859, dated 09 / 19 / 2025, page 19 / 290 11 / 208 combinations of these.

[0030] In certain aspects of the present disclosure, the active agent covalently or non-covalently conjugated to at least one of the terminal groups located on the surface of the hyperbranched macromolecule is a peptide selected from the group consisting of compstatin, APL-1 and Fc-III-4C, Beovu (Brolucizumab), Zimura (Avacincaptade Pegol), Pegcetacoplan, Abicipar Pegol, Lampalizumab, Fovista, Risuteganib, AXT107, Elamipretide, THR149, ALM201, VGB3 and Largazole.

[0031] In certain aspects of the present disclosure, the active agent is covalently linked to at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or even 100% of the terminal groups located on the surface of the hyper-branched macromolecule.

[0032] In certain forms of disclosure, the dendritic constitutional repeating unit is represented by formula (i): A Í-La4-í-OCH2CH2-j—O----X--O—í-CH2CH2O-4 Í-LbT---B ' 'm' 'n L 'o ' 'p Jy where A is a connection to a polymeric arm that is connected to the central unit, or A is a connection to B of a preceding dendritic constitutional repeating unit represented by Formula (i), La is a ligand, m is 0 or 1, n is an integer from 20 to 2000, o is an integer from 20 to 2000, neo may be different or the same, X is the branching unit, as a polyol-derived unit, Lb is a ligand, p is 0 or 1, B comprises a terminal group located on the surface of the hyper-branched macromolecule or is a connection to A of a consecutive dendritic constitutional repeating unit or a connection to an active agent, La and Lb may be different or the same, m and p may be different or the same, ey is an integer from 2 to 9, where y = c - 1 with c being the connectivity c of the branching unit X;and in which the dendritic constitutional units in hypermolecules Petition 870250084859, dated 09 / 19 / 2025, p. 20 / 290; 12 / 208 branched branches may be the same or different.

[0033] In certain additional aspects, A comprises a functional group formed by click chemistry, such as a triazole or dihydropyrazine and / or the La and / or Lb ligands comprise a diacid group and / or a diamido acid group, a carboxyl group and / or a carboxamide group, such as succinate, glutarate, adipate, azelate or glutaramide. In certain embodiments, the La and / or Lb ligand comprises a structure represented by Formula (ii): 10 where U1 and U2 are independently NH or O and may be the same or different, and where t is an integer from 0 to 10. The La and / or Lb ligand may further comprise a polyethylene glycol unit between the linkage to B and the carboxyl group, carboxamide group or structure of Formula (ii).

[0034] In certain embodiments of the disclosure, the present invention provides a method for fabricating a hyper-branched macromolecule as described herein by divergent synthesis, comprising the steps of (a) providing a core unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the core unit having functional groups suitable for click chemistry at the ends of the polymeric arms; (b) providing precursors of dendritic constitutional repeating units comprising a polymeric arm comprising a functional group suitable for forming a click chemistry connection with the corresponding functional groups of the core-connected polymeric arms (such as an azide, alkyne, alkene or tetrazine) and at least two polymeric arms comprising non-reactive functional groups in click chemistry; (c) forming a click chemistry connection between the core-connected polymeric arms and the arms Petition 870250084859, dated 09 / 19 / 2025, p. 21 / 290 13 / 208 polymeric precursors of the dendritic constitutional repeat unit, (d) optionally convert the functional groups of at least two polymeric arms comprising non-reactive functional groups in click chemistry into functional groups suitable for click chemistry, and (e) conjugate an active agent comprising a functional group to the outermost polymeric arms by reaction with the functional groups of the outermost polymeric arms, thus forming a hyperbranched macromolecule-active agent conjugate. For a higher-generation hyperbranched macromolecule Gx, with x being an integer from 2 to 10, step (d) may be mandatory and consecutive dendritic constitutional repeat unit precursors are connected to the functional groups suitable for click chemistry obtained in step (d) by click chemistry to the hyperbranched macromolecule before conjugation of the active agent in step (f).

[0035] In certain modalities of the method, the precursor of the dendritic constitutional repeating unit in step (c) is represented by Formula (iii): where C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide, or tetrazine), D comprises functional groups unsuitable for click chemistry, such as succinimidyl, and La, m, η, X, o, Lb, pey are as defined above, and wherein the dendritic constitutional repeating units may be the same or different.

[0036] In another embodiment of the method, the invention relates to a method for manufacturing a hyper-branched macromolecule as described herein by convergent synthesis, comprising the steps of (I) providing repeating unit precursors Petition 870250084859, dated 09 / 19 / 2025, page 22 / 290 14 / 208 constitutional dendritic repeating units comprising a polymeric arm comprising a functional group suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine) and at least two polymeric arms comprising non-reactive functional groups in click chemistry; (II) conjugate active agents comprising a functional group to at least one of the at least two polymeric arms comprising non-reactive functional groups in click chemistry of the precursors of constitutional dendritic repeating units; (III) provide a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry (such as an azide, alkyne, alkene or tetrazine) at the ends of the polymeric arms;and (IV) form a click chemistry connection between the polymeric arms connected to the core provided in step III) and the polymeric arm comprising a functional group suitable for forming a click chemistry connection of the dendritic constitutional repeat unit precursors conjugated to the active agent obtained in step II), thus forming a hyper-branched macromolecule-active agent conjugate. In embodiments of this method, the dendritic constitutional repeat unit precursor in step I) is represented by Formula (iii) as described above.;

[0037] For a higher-generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10, the precursors of constitutional dendritic repeat units conjugated to the active agent obtained in step II) can be connected by click chemistry to precursors of reverse constitutional dendritic repeat units comprising a polymeric arm comprising a non-reactive functional group in click chemistry and at least two polymeric arms comprising functional groups suitable for Petition 870250084859, dated 09 / 19 / 2025, page 23 / 290 15 / 208 click chemistry (such as an azide, alkyne, alkene, or tetrazine), in which the non-reactive functional group in the click chemistry of a polymeric arm is subsequently converted into a functional group suitable for click chemistry before connecting to other precursors of reverse constitutional dendritic repeating units or before forming a click chemistry connection with the polymeric arms connected to the core in step IV), thus forming higher-generation hyper-branched macromolecules.In one aspect, precursors of dendritic constitutional repeat units having different active agents conjugated to the polymeric arms can be obtained by performing steps I) and II) for each active agent-conjugated dendritic constitutional repeat unit precursor, and a mixture of the obtained active agent-conjugated dendritic constitutional repeat unit precursors is then used for step IV), thus forming a hyper-branched macromolecule-active agent conjugate having different active agents in different regions of the hyper-branched macromolecule surface.

[0038] In another embodiment, the invention relates to a hyper-branched macromolecule as described herein, for use as a medicament. In another embodiment, the invention relates to a treatment method, wherein the method comprises treating a disease or medical condition in a patient with a hyper-branched macromolecule of embodiments of the invention. In one aspect thereof, the hyper-branched macromolecule is used for an eye treatment, such as the treatment of an eye disease, such as diseases of the fundus of the eye, such as any posterior segment eye disease affecting the vasculature and integrity of the retina, macula or choroid, leading to visual acuity disturbances, vision loss or blindness, particularly posterior segment disease states resulting Petition 870250084859, dated 09 / 19 / 2025, p. 24 / 290 16 / 208 of age, trauma, surgical interventions, such as age-related macular degeneration (AMD), cystoid macular edema (CME), diabetic macular edema (DME), posterior uveitis and diabetic retinopathy.

[0039] In other respects, the hyper-branched macromolecule is used in the treatment of a selected ocular disease from the group consisting of retinal neovascularization, choroidal neovascularization, wet AMD, dry AMD, retinal vein occlusion, diabetic macular edema, retinal degeneration, hyphema, presbyopia, corneal graft rejection, retinoblastoma, melanoma, miosis, mydriasis, glaucoma, conjunctivitis, intraocular infections, choroidal neovascularization (CNV), intraocular tumors, retinal neuroinflammation, inflammation, autoimmune uveitis, uveitis, proliferative vitreoretinopathy and corneal degeneration, acute and chronic macular neuroretinopathy, central serous chorioretinopathy, macular edema, acute multifocal placoid pigment epitheliopathy, Behçet's disease, bird-lead retinochoroidopathy, posterior uveitis, scleritis posterior, serpiginous choroiditis, subretinal fibrosis, uveitis.syndrome, Vogt-Koyanagi-Harada syndrome, retinal artery occlusive disease, central retinal vein occlusion, disseminated intravascular coagulopathy, retinal vein branch occlusion, hypertensive fundus changes, ocular ischemic syndrome, retinal arterial microaneurysms, Coat's disease, parafoveal telangiectasia, hemiretinal vein occlusion, papillophlebitis, carotid artery disease (CAD), ground branch angiitis, sickle cell retinopathy, angioid streaks, familial exudative vitreoretinopathy, Eales disease, proliferative vitreous retinopathy, diabetic retinopathy, tumor-associated retinal disease, congenital retinal pigment epithelium hypertrophy (RPE), posterior uveal melanoma, choroidal hemangioma, choroidal osteoma, choroidal metastasis, combined hamartoma of the retina and retinal pigment epithelium, retinoblastoma, tumors Vasoproliferative disorders of the fundus of the eye, retinal astrocytoma, tumors. Petition 870250084859, dated 09 / 19 / 2025, page 25 / 290 17 / 208 intraocular lymphoid tissue, myopic retinal degeneration, acute retinal pigment epithelium, glaucoma, endophthalmitis, cytomegalovirus retinitis, retinal cancers, retinitis pigmentosa, Leber congenital amaurosis, choroideremia, X-linked retinitis pigmentosa, vitelliform macular dystrophy, X-linked retinoschisis, CNGA3 achromatopsia, CNGB3 achromatopsia, LHON, Stargardt disease, Usher syndrome, Norrie disease, Bardet-Biedl syndrome, and red-green color blindness.

[0040] In certain embodiments, the hyper-branched macromolecule is formulated for direct injection into a patient's treatment site, for example, by parenteral administration, intratumoral injection, injection into the eye, such as intravitreal, intracameral, subconjunctival, retrobulbar, subtenon, subretinal or suprachoroidal injections. The hyper-branched macromolecule can be administered by direct injection, by oral application, incorporated into gels or incorporated into implants. DEFINITIONS

[0041] The terms “hyper-branched macromolecule” or “hyper-branched polymer,” or simply “branched polymer” or “branched macromolecule” are all used interchangeably herein to designate branched macromolecules similar to dendrimers or polymers that have a tree-like structure, such as dendrimers, and the term “dendrimer” is used herein synonymously with them. However, while dendrimers are monodisperse, highly symmetrical molecules of precisely defined composition, the hyper-branched macromolecules of the present invention are polydisperse molecules because they include polyethylene glycol arms or units that have a certain polydispersity, like most synthetic polymeric structures. The polydispersity of PEG chains and precursor molecules, including them, may be small, but Petition 870250084859, dated 09 / 19 / 2025, page 26 / 290 18 / 208 this implies polydispersity also for hyperbranched dendrimer-like macromolecules, as described here.

[0042] Polydispersity is given by the polydispersity index D with D = Mw / Mn, where Mw is the weight-average molar mass and Mn is the number-average molar mass, determined by gel permeation chromatography. For most polyethylene glycol (PEG) materials, the polydispersity index is a parameter provided by the manufacturer in the product specifications as an indicator of the uniformity and quality of the material. The polydispersity of multi-arm PEG precursors can be less than 1.3, less than 1.2, or less than 1.1.

[0043] The term “biodegradable” refers to a material or object (such as the hyper-branched macromolecules according to the present invention) that degrades in vivo, i.e., when placed in the human or animal body, or in vitro when immersed in an aqueous solution under physiological conditions, such as pH 7.2-7.4 at 37 °C. In the context of the present invention, as disclosed in detail below, the hyper-branched macromolecules, once administered or deposited in the human or animal body, biodegrade slowly and are eliminated over time. In certain embodiments, biodegradation occurs at least in part through ester hydrolysis in the aqueous environment of the body. Biodegradation may occur by hydrolysis or enzymatic cleavage of covalent or conjugation bonds, in linker groups and / or within the polymer arms. The hyper-branched macromolecules disintegrate slowly, resulting in elimination via physiological pathways.In certain embodiments, the hyper-branched macromolecules of the present invention are stable against degradation for long periods of time (for example, about 1 month, 3 months, or 6 months). In certain embodiments, the hyper-branched macromolecules only biodegrade for... Petition 870250084859, dated 09 / 19 / 2025, p. 27 / 290 19 / 208 example, until the active agent or at least a larger quantity (for example, at least 50%, at least 75% or at least 90%) of it has been released.

[0044] The terms “precursor” or “component” or “building block” herein refer to molecules or compounds that react with each other and are therefore connected by means of covalent bonds to form a hyper-branched macromolecule.

[0045] The parts of the precursor molecules that are still present in a final hyper-branched dendrimer-like macromolecule are also referred to as “units” or “polymer arms” herein. The “units” or “polymer arms” therefore belong to the main building blocks or constituents of a hyper-branched polymeric macromolecule. For example, a hyper-branched macromolecule suitable for use in the present invention may contain identical or different polyethylene glycol units or arms, in addition to core units and branched units, as disclosed herein.

[0046] The term “central unit,” as used herein, refers to a constitutional unit at the center of a hyperbranched macromolecule, from which emanate the polymeric arms or dendritic constitutional repeat units (DCRUs) or dendrons. The central unit has at least 3 c-connectivities (or valences) to each of which a polymeric arm or a dendritic constitutional repeat unit is connected, i.e., covalently linked. For example, for a branched PEG macromolecule with multiple G0 generation arms, the central unit may be derived from a polyol compound, which is poly(ethoxylated) at each of its hydroxyl groups.

[0047] An exemplary central unit structure with three connectivities with the connectivities c shown as OH is Petition 870250084859, dated 09 / 19 / 2025, page 28 / 290 20 / 208 shown below: HO OH xz I OH

[0048] The terms “branching unit” or “branching point” used herein refer to a constitutional unit within a dendritic constitutional repeat unit with at least 3 c' connectivities (or valences) to each of which a polymeric arm or another dendritic constitutional repeat unit is connected. The branching unit may have the same chemical structure or a different structure from the central unit.

[0049] The term “dendritic constitutional repeat unit” (DCRU), sometimes referred to as “dendron”, as used herein, refers to a constitutional repeat unit of c' > 3 connectivity, including a branching point and polymeric arms emanating from it. It may be connected to a total of c polymeric arms emanating from the central unit and / or other DCRUs consecutively to form a hyper-branched, dendrimer-like structure.

[0050] The term “terminal group,” as used herein, refers to a constitutional unit, for example, a functional group, that is located at one end of a polymeric arm or DCRU. In a hyperbranched macromolecule, terminal groups on the outermost surface of the hyperbranched macromolecule can be used to conjugate or link active agent molecules to the hyperbranched macromolecule. The terminal group may consist of a ligand with hydrolyzable groups connected to a terminal functional group.

[0051] The term “generation”, abbreviated “G”, refers to the set of dendritic constitutional repeating units separated from the free valence of a dendron by the same number of units Petition 870250084859, dated 09 / 19 / 2025, pp. 29 / 290 21 / 208 dendritic constitutional repetitions.

[0052] The term “dendron”, as used herein, refers to a part of the hyper-branched macromolecule with only one free valence, comprising exclusively DCRUs and terminal groups, and in which each path from the free valence to any terminal group comprises the same number of constitutional repeating units.

[0053] The term “conjugate”, as used herein, includes the covalent or non-covalent linkage of an active agent to a hyperbranched macromolecule. Conjugation comprises a non-covalent linkage, such as to a terminal group of a hyperbranched macromolecule with affinity for the active agent molecule, which may also be a means of linking an active agent molecule to the hyperbranched macromolecule.

[0054] The term “release” (and consequently the terms “released”, “releasing”, etc.) as used herein refers to the chemical separation and delivery of active agents from the hyperbranched macromolecules of the present invention to the surrounding environment. The released agents may or may not have molecular fragments of the hyperbranched macromolecule still attached to them. The surrounding environment may be an in vitro or in vivo environment, as described herein. In certain specific embodiments, the surrounding environment is the vitreous humor and / or ocular tissue, such as the retina and choroid. The attachment of the API to the hyperbranched macromolecule may be a covalent bond, in which the API may be separated from the hyperbranched macromolecule by a chemical event, such as the hydrolysis of a bond formed by a ligand group.Furthermore, multiple hydrolyzable linkage chemistries can be employed to release the API at multiple rates from the same hyper-branched macromolecules or blended to achieve a desired release profile. Additionally, the hyper-branched macromolecule can be functionalized with groups. Petition 870250084859, dated 09 / 19 / 2025, page 30 / 290 22 / 208 terminals that bind to an API in a non-covalent manner, releasing the API according to the binding affinity kinetics of the terminal group-API pair of the hyper-branched macromolecule. Several non-covalently linked terminal group-API pairs can be employed to achieve a desired release profile.

[0055] The term “100% release of the active agent” should be interpreted as 95% to 100%. The way this controlled release is achieved is through a series of parameters that are characteristic of the drug delivery system, as disclosed here. Each of these characteristics of the drug delivery system, alone or in combination with the others, can be responsible for controlled release.

[0056] The term “sustained release” for the purposes of the present invention is intended to characterize products such as biodegradable hyper-branched macromolecules, which are formulated to make an active agent available over a long period of time, thus allowing a reduction in dosing frequency compared with an immediate-release dosage form, such as a solution of an active agent that is applied topically to the eye (i.e., eye drops). Other terms that may be used interchangeably with “sustained release” herein are “prolonged release” or “controlled release”. In the sense of the invention, the term “sustained release” includes steady-state release of the active agent, gradual release of the active agent, ascending release of the active agent, as well as any combination thereof, such as a steady-state release of the active agent followed by a gradual release of the active agent.In the meaning of the invention, the term "tapered" or "tapering" refers to a decrease in the release of the active agent over time. Specifically, the term "sustained release" refers to the release of an active agent from... Petition 870250084859, dated 09 / 19 / 2025, page 31 / 290 23 / 208 hyper-branched macromolecules or the drug delivery system, including them in a predetermined manner, and contrasts with immediate release, such as a bolus injection. In certain embodiments, controlled release refers to the amount of active agent released over the total number of days required for the release of 100% of the active agent in an aqueous solution under physiological in vitro conditions, such as pH 7.2-7.4 and 37 °C.

[0057] The term "extended time period," as used in this document, refers to any period of time that would be considered by those skilled in the art to be extended in relation to the treatment of a disease and, in particular, refers to periods such as at least about 1 week, or at least about 1 month or more, such as up to about 12 months, or any intermediate periods, such as about 1 to about 6 months, about 2 to about 4 months, about 2 to about 3 months, or about 3 to about 4 months, or as otherwise disclosed in this document.

[0058] A “zero-order” release or “substantially zero-order” release or “near-zero-order” release is defined as one that exhibits a relatively straight line in a graphical representation of the percentage of active agent released as a function of time. In certain embodiments of the present invention, a substantially zero-order release is defined as the amount of active agent released that is proportional within 20% to the elapsed time.

[0059] The terms API, active (pharmaceutical) ingredient, active (pharmaceutical) agent, active (pharmaceutical) principle, therapeutic (active) agent, active and drug are used interchangeably in this document and refer to the substance used in a finished pharmaceutical product (FPP), as well as the substance used in the preparation of such finished pharmaceutical product, intended Petition 870250084859, dated 09 / 19 / 2025, page 32 / 290 24 / 208 to provide pharmacological activity or otherwise have a direct effect on the diagnosis, cure, mitigation, treatment or prevention of a disease, or have a direct effect on the restoration, correction or modification of a patient's physiological functions.

[0060] The active agent used according to the present invention may be an active agent for the treatment and / or prevention of a disease or disorder, or a diagnostic agent, such as a marker. In one embodiment of the invention, the active agent is a low water-soluble active agent (i.e., with a water solubility less than about 1000 μg / mL or less than about 100 μg / mL). In other embodiments of the invention, the active agent is a highly water-soluble active agent (i.e., with a water solubility greater than about 1000 μg / mL or even greater than 10 mg / mL). This definition is independent of the agent's approval by a governmental agency.

[0061] For the purposes of the present invention, an active agent in all its possible forms, including free acid, free base, polymorphs or any pharmaceutically acceptable salts, anhydrates, hydrates, cocrystals or other solvates or derivatives, such as prodrugs or conjugates, may be used. To conjugate to the hyper-branched macromolecule, the active agent may need to be functionalized, unless it already comprises a functional group suitable for conjugation. Whenever in this description or in the claims an active agent is mentioned without further specification, even if not explicitly stated, it also refers to the active agent in the form of any polymorphs, pharmaceutically acceptable salts, anhydrates or solvates (including hydrates) thereof. With respect to the active agent, suitable solid forms include, without limitation, the pure substance form in any physical form known to a person of knowledge. Petition 870250084859, dated 09 / 19 / 2025, page 33 / 290 25 / 208 common in the technique.

[0062] As used herein, the term “therapeutically effective” refers to the amount of active agent required to produce a desired therapeutic result after administration. For example, in the context of the present invention, a desired therapeutic result would be a reduction in symptoms associated with dry eye disease (DED), for example, as measured by in vivo tests known to those skilled in the art, such as an increase in a Schirmer tear test score, a reduction in staining values ​​measured by conjunctival lissamine green staining or corneal staining with fluorescein, a reduction in the severity score of ocular dryness and / or frequency of ocular dryness on a visual analog scale (VAS), a reduction in the Ocular Surface Disease Index and / or the patient's standard assessment of ocular dryness, as well as a reduction in best-corrected visual acuity.In one embodiment, “therapeutically effective” refers to an amount of active agent in a sustained-release intracanalicular insert capable of achieving a tear fluid concentration that is equivalent in terms of therapeutic effect to a cyclosporine concentration of 0.236 pg / mL (which is considered necessary for immunomodulation, Tang-Liu and Acheampong, Clin. Pharmacokinet. 44(3), pp. 247-261)) over a long period of time and, in particular, for substantially the entire remaining period of use of the insert, once said tear fluid concentration is achieved.

[0063] The term “patient” here includes both human and animal patients. Biodegradable drug delivery systems according to the present invention are therefore suitable for human or veterinary medical applications. Generally, an “individual” is an individual (human or animal) to whom a drug delivery system according to the invention is administered. Petition 870250084859, dated 09 / 19 / 2025, page 34 / 290 26 / 208 present invention. A “patient” is an individual who requires treatment due to a specific physiological or pathological condition. A patient does not necessarily have a diagnosis of a specific physiological or pathological condition prior to receiving the drug delivery system.

[0064] The molecular weight of a hyper-branched macromolecule, polymeric precursor, polymeric unit, arm or similar, as used for the purposes of the present invention and as disclosed herein, can be determined by analytical methods known in the art. The molecular weight of polyethylene glycol can, for example, be determined by any method known in the art, including gel electrophoresis, such as SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis), gel permeation chromatography (GPC), including GPC with static light scattering (SLS) or dynamic light scattering (DLS) detectors, liquid chromatography (LC), as well as mass spectrometry, such as matrix-assisted laser desorption / ionization spectrometry (MALDI-TOF) or electrospray ionization mass spectrometry (ESI).The molecular weight of a polymer, including a polyethylene glycol precursor as disclosed herein, is an average molecular weight (based on the molecular weight distribution of the polymer) and can therefore be indicated by several average values, including the weight-average molecular weight (Mw) and the number-average molecular weight (Mn). In the case of multi-arm PEG precursors, as used in some aspects of the present invention, the molecular weight indicated herein is the number-average molecular weight (Mn) determined by gel permeation chromatography using a suitable molecular weight standard, such as a polyethylene glycol or polystyrene standard, according to standard methods known in the art. Typically, the materials, especially multi-arm precursors, are... Petition 870250084859, dated 09 / 19 / 2025, page 35 / 290 27 / 208 acquired with a specific molecular weight and polydispersity, defined by the supplier. Suitable PEG precursors are available, for example, from various suppliers such as Jenkem Technology and Xiamen SinoPeg Biotech Co. Ltd., and others.

[0065] The term “day 1”, as used herein, refers to a point in time that immediately follows “day 0”. Thus, whenever “day 1” is used, it refers to a period of time that has already elapsed of one day or approximately 24 hours after administration of the drug delivery system.

[0066] As used in this document, the term "approximate," in connection with a measured quantity, refers to the normal variations in such measured quantity, as expected by someone skilled in the art performing the measurement and exercising a level of care consistent with the measurement objective and precision of the measuring equipment.

[0067] The term at least about in connection with a measured quantity refers to normal variations in the measured quantity, as expected by someone skilled in the art performing the measurement and exercising a level of care consistent with the measurement objective and precision of the measuring equipment, and any quantities greater than that.

[0068] The term “mean”, as used herein, refers to a central or typical value in a data set (points), which is calculated by dividing the sum of the data (points) in the set by their number (i.e., the average value of a data set).

[0069] As used in this document, the singular forms “um”, “uma”, oe “a” include plural references unless the context clearly dictates otherwise.

[0070] The term and / or as used in a sentence as A and / or B in this document is intended to include both A and B as well as A or B. Petition 870250084859, dated 09 / 19 / 2025, p. 36 / 290 28 / 208

[0071] Open terms such as include, including, contain, containing, and the like mean comprising. These open transition phrases are used to introduce an open list of elements, method steps, or the like that do not exclude additional unrecorded elements or method steps.

[0072] The term “up to”, when used here together with a given value or number, means to include the respective value or number.

[0073] The terms “from A to B”, “from A to B” and “from A to B” are used interchangeably herein and all refer to a range from A to B, including the upper and lower limits A and B.

[0074] Throughout this disclosure, various aspects of this invention are presented in a range format. It should be understood that the range description is simply for convenience and brevity and should not be interpreted as an inflexible limitation on the scope of the invention. Therefore, the description of a range should be considered as having specifically disclosed all possible subranges, as well as individual numerical values ​​within that range. For example, the description of a range as 1 to 6 should be considered as having specifically disclosed subranges, such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5 and 6. This applies regardless of the range width. The recited numerical ranges are inclusive of the numbers that define the range and include every integer within the defined range.

[0075] The abbreviation “PBS” when used here means phosphate-buffered saline.

[0076] The abbreviation “PEG” when used here means polyethylene glycol. BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 schematically illustrates a)-c) different methods of Petition 870250084859, dated 09 / 19 / 2025, page 37 / 290 29 / 208 synthesis of dendrimers or hyper-branched macromolecules. Figure 2 schematically illustrates the generations of a dendrimer or hyper-branched macromolecule. Figure 3 shows a) a schematic image of the formation of a hyper-branched PEG macromolecule via DBCO-azide coupling; and b) schematically illustrates a 3-D model of an 8-armed PEG core with eight branched, 4-armed PEG repeating units conjugated with peptides. Figure 4 illustrates the structure of the peptides compstatin, APL-1 (Mod2) and APL-1 (Mod3). Figure 5 illustrates a purification setup using a) dialysis and b) SEC column filtration from Examples 6 and 7. Figure 6 is a diagram of a UHPLC analysis of dialysis-purified hyper-branched macromolecule-compstatin conjugates from Example 6. Figure 7 is a diagram of a UHPLC analysis of hyper-branched macromolecule-compstatin conjugates purified by SEC column filtration from Example 7. Figure 8 is a UHPLC plot of a) a standard curve of the compstatin sample (12.5, 25, 50, 100, 200 pg / mL) with a calibration plot and b) a 4-arm hyper-branched macromolecule conjugate G0compstatin from Example 1. Figure 9 shows a comparison graph of the substitution rate of different PEG ligands with compstatin and compstatin-lysine based on Example 8. Figure 10 is a graph of the optimization conditions for the conjugation of compstatin from Example 8. Figure 11 illustrates the SPR results from Example 9: C3 binding: a) KD of 4 compstatin samples from different suppliers; b) equilibrium analysis. Petition 870250084859, dated 09 / 19 / 2025, page 38 / 290 30 / 208 Figure 12 illustrates the SPR results from Example 9: C3b binding: a) KD of 4 compstatin samples from different suppliers; b) equilibrium analysis. Figure 13 illustrates the SPR results from Example 9: C3 and C3b binding: a) APL-1 (acetylated amine), b) APL-1 (amine, acetate salt), c) APL-1 (lysine end). Figure 14 illustrates the SPR results of IgG binding: ac) Fc-III 4C ed) Fc-III. Figure 15 illustrates a comparison of SPR of free compstatin and multivalence compstatin. Figure 16 illustrates the SPR results of a) 8α-40k-PEG-[(4α2kPEG-(comp)3]8, b) 4α-40kPEG-SGA-(comp)4, c) 4α-40kPEG-SS(comp)4 and d) SS-comp (hydrolysis) compared to free compstatin. Figure 17 is a plot of the dissociation constant Kd of hyper-branched macromolecule-compstatin conjugates in relation to the corresponding number of peptide substitutions. Figure 18 is an illustration of an alternative route (AP) hemolysis assay. Figure 19. AP hemolysis results of IC50 for hyper-branched macromolecule-compstatin conjugates. Figure 20 is a calibration curve of the hydrodynamic radius Rh versus the half-life T1 / 2 determined in the vitreous humor of the New Zealand white rabbit to predict the sustained release of dendrimer drug conjugates. Figures 21 a) ac) show the degradation effect of the temperature variation from 35 °C to 39 °C at a constant pH of 7.4. Figures 22 a) ac) show the degradation effect of varying the pH from pH 7.0 to pH 8.5 at a constant temperature of 37°C. DETAILED DESCRIPTION OF THE INVENTION

[0077] The present invention is directed, in certain respects, to Petition 870250084859, dated 09 / 19 / 2025, page 39 / 290 31 / 208 Hyper-branched macromolecules (dendrimers) comprising polyethylene glycol polymer units and an active agent covalently linked or conjugated to at least one of the outermost arms of the hyper-branched macromolecule. Conjugation comprises covalent and non-covalent linkage, such as to a terminal group of a peptide hyper-branched macromolecule with affinity for the active agent molecule, and this may also be a means of linking an active agent to the hyper-branched macromolecule.

[0078] Dendrimers are monodisperse macromolecules with multiple reactive end groups on their surface. Dendrimers are often compared to tree-like structures, i.e., a branched molecular architecture that provides a wide variety of possible end groups and extraordinary structural control. Elements are added to a dendrimer structure by a series of chemical reactions and build a branched spheroidal structure from an initial atom or central unit. The central core unit has at least two or at least three reactive functional groups, and the repeating branches are arranged in a series of “radially concentric layers” called “generations.” Hyper-branched macromolecules can have the same molecular architecture as dendrimers without being monodisperse, as they can be constructed using precursors or polydisperse units.

[0079] It has been found that strategies to optimize drug delivery and site-specific targeting using dendrimer-like hyper-branched macromolecules, as described herein, provide several advantages in drug delivery and can utilize dendrimer advantages also for hyper-branched macromolecules that have a dendrimer-like structure without being monodisperse molecules. For example, dendrimer-like hyper-branched macromolecules. Petition 870250084859, dated 09 / 19 / 2025, page 40 / 290 32 / 208 provide several terminal functionalities that can be used to adjust the hydrophobicity / hydrophilicity of the hyperbranched macromolecule used as a carrier for an active agent, or can be used as conjugation precursors for target molecules to improve the interaction between the API and the hyperbranched macromolecule, for example, by multivalent binding to receptors and / or improving the avidity of conjugated biomolecules such as peptides or proteins.

[0080] The multiple number of surface groups in hyper-branched macromolecules can anchor more APIs with desired binding methods and achieve controlled release through different degradation conditions or degradation kinetics. Hyper-branched macromolecule-drug conjugates can increase the stability and solubility of therapeutic agents to be administered, reduce systemic effects, and increase efficacy at the target site compared to free drugs. Furthermore, a hyper-branched macromolecule can have a symmetrical structure that provides numerous intramolecular cavities to capture unbound API molecules. Additionally, the large external hydration radius of specifically PEG-based dendrimer structures extends the in vivo half-life, such as in the glassy body, of dendrimer drug conjugates, which can be used to control and adjust the sustained release of active agents.

[0081] Furthermore, biodegradable synthetic dendrimers offer the advantage of controllably degradable functional groups, such as hydrolyzable or enzymatically cleavable linkages, which after degradation produce lower molecular weight fragments with a smaller hydration radius and a different half-life that determines their elimination from the body. The incorporated degradable groups can be used to adapt and control the release rate of active agents associated with the dendrimer. Petition 870250084859, dated 09 / 19 / 2025, page 41 / 290 33 / 208 Hyper-branched macromolecules

[0082] In certain aspects of the present invention, a hyper-branched macromolecule is provided that is formed from various building blocks of the dendritic structure (not including the active agent), such as the central unit, polymeric arms, branching unit, ligands and / or extenders, and dendritic constitutional repeat units. In one embodiment, a hyper-branched macromolecule is provided comprising a central unit with at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit at the c-connectivities, each polymeric arm comprising a terminal group or being connected to a dendritic constitutional repeat unit comprising a branching unit connected to at least two polymeric arms,each comprising a terminal group or being connected to a next dendritic constitutional repeat unit that may again be connected to other dendritic constitutional repeat units, the polymeric arms of the outermost dendritic constitutional repeat unit each comprising a terminal group; wherein the polymeric arms comprise or consist of linear polyethylene glycol (PEG) units; and wherein at least one active agent is conjugated to at least one of the terminal groups located on the outermost polymeric arms of the hyper-branched macromolecule. The hyper-branched macromolecule includes chemical bonds that can be cleaved by hydrolysis, making the hyper-branched macromolecule biodegradable in aqueous environments.

[0083] In some embodiments, the hyper-branched molecule is formed from building blocks at least partially connected by hydrolyzable bonds or connections located at positions such that complete hydrolysis of all hydrolyzable bonds in the macromolecule produces hydrolysis fragments, each Petition 870250084859, dated 09 / 19 / 2025, page 42 / 290 34 / 208 one with a molecular weight less than 40 kDa. This can be achieved by selecting suitable building blocks or precursors with a molecular weight less than 40 kDa and connecting them via hydrolyzable chemical linkages, typically acid-labile, such as esters or amide linkages, as described in more detail here. For example, the inclusion of diacid linkers to connect constitutional repeating units, each with a molecular weight less than 40 kDa, allows hydrolytic degradation, producing hydrolysis fragments that meet the desired molecular weight limit. If non-hydrolyzable connections are used, for example, linkages formed by some click chemistry reactions such as alkyneazide coupling, they must be located between building blocks that together meet the molecular weight limit for hydrolysis fragments of less than 40 kDa.

[0084] In certain aspects of the invention, the overall molecular size and the number of surface groups of hyperbranched macromolecules gradually increase with the addition of successive layers of monomers, which is called generation. Biodegradable hyperbranched macromolecules can be synthesized by divergent or convergent synthesis, or a combination of both, see FIG. 1. The divergent method involves the addition of monomers or so-called dendritic constitutional repeating units (DCRUs) in repeated sequence and starts from a multivalent core to surface molecules with a continuous increase in the number of branches. The molecular size and the number of surface groups gradually increase with the addition of successive layers of monomers, which is called generations.While the convergent method involves the synthesis of hyper-branched macromolecules from the surface to the core and leads to the formation of wedge-shaped conical units or dendrons, these are joined to a core. Petition 870250084859, dated 09 / 19 / 2025, page 43 / 290 35 / 208 multivalent in the last stage.

[0085] The hyper-branched macromolecules of certain embodiments of the present invention include as main building units a central unit and, optionally, a plurality of branching units that may be derived from polyols, a plurality of polymeric arms comprising polyethylene (PEG) units, optional hydrolyzable linking groups, connecting groups between building units, terminal groups and conjugated active agents, such as, for example, peptides. All these constituent elements or building units are described in more detail below.

[0086] The connections formed between different polymeric arms in the hyper-branched macromolecules may include hydrolyzable linkages by the introduction of suitable linking groups between the PEG arms and the functional groups to connect the different units to form the hyper-branched macromolecule.These ligands that form hydrolyzable bonds facilitate biodegradation in aqueous environments, such as the human or animal body, in vivo. For example, hydrolyzable chemical bonds can be acid-labile, to facilitate cleavage in more acidic environments that can be found, for example, in tumors at the cellular level.

[0087] Hydrolyzable chemical linkages may include selected linkages or bonds from the group consisting of amine, amide, urethane, ester, anhydride, ether, acetal, ketal, nitrile, isonitrile, isothiocyanate, or imine linkages, and combinations thereof. These linkages are normally formed by condensation reactions or click chemistry of suitably functionalized precursors during the synthesis of the hyper-branched macromolecule. In certain preferred embodiments, the hydrolyzable linkages are ester linkages, such as ester linkages formed by the use of diacid ligands, such as succinic acid, glutaric acid, adipic acid, and higher homologs. Petition 870250084859, dated 09 / 19 / 2025, page 44 / 290 36 / 208

[0088] An exemplary structure of a hyperbranched macromolecule of certain embodiments of the invention is shown below:

[0089] In this Formula of a hyper-branched G1 generation macromolecule, X designates a central unit, or a branching unit derived from a polyol, such as glycerol, the central or branching unit being connected to three polyethylene glycol arms, and the branching units being connected to one of their polyethylene glycol arms by means of a linking group Y to the polyethylene glycol arms of the central unit at the center. The linker Y comprises hydrolyzable linkages, such as ester or amide linkages, as defined herein, n designates the number of polyethylene glycol repeating units in the polymeric arm.

[0090] The building blocks from which hyper-branched macromolecules are formed comprise a central unit, polymeric arms, such as arms consisting of polyethylene glycol (PEG), bifunctional linking groups or ligands, bifunctional extenders, dendritic constitutional repeating units comprising a branching unit, functional end groups. In embodiments of the invention, the building blocks have a weight Petition 870250084859, dated 09 / 19 / 2025, page 45 / 290 37 / 208 average molecular weight (Mn) of less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons. Central unit

[0091] The central unit is the center of the hyperbranched macromolecule from which the polymeric arms or dendritic constitutional repeat units (DCRUs) or dendrons emanate. The central unit has at least 3 c-connectivities (or valences) to each of which a polymeric arm or a dendritic constitutional repeat unit is connected, i.e., covalently linked. The polymeric arms can be connected to the central unit by a hydrolyzable bond, preferably by non-hydrolyzable bonds, such as ether bonds.

[0092] In certain embodiments, the central unit has a c connectivity of 3 to 10, or 4 to 8, or 4 to 6, or 4. The central unit may be derived from a molecule or chemical structure with several c functional groups to which polymeric arms are attached. For example, in certain embodiments, the central unit is derived from a polyol with at least 3 hydroxyl groups, or 4, 5, 6, 7, 8, 9, or 10 hydroxyl groups.

[0093] In such embodiments, the polyol may be selected from glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol, or sorbitol. In certain embodiments, the core unit derived from a polyol is ethoxylated at each of its hydroxyl groups to form a multi-arm precursor with the arms being polymeric PEG arms that are terminated with a terminal group or functional group.

[0094] An exemplary central unit structure with three connectivities with the connectivities c shown as OH: Petition 870250084859, dated 09 / 19 / 2025, page 46 / 290 38 / 208 HO OH X OH

[0095] The central unit of a multi-armed precursor that can be used to form the hyper-branched macromolecule of certain embodiments of the present invention is, therefore, a suitable structure to provide the desired number of precursor arms. For example, for 4-armed precursors, the central unit can be a pentaerythritol or ethylenediamine structure, while for 8-armed precursors, the central unit can be a hexaglycerol structure. In certain embodiments of the invention, the central unit is pegylated at its c-connectivities with polyethylene glycol arms, as in the structure shown below. The connectivities at the end are again shown as OH groups, to which a ligand, functional group or other DCRU can be attached.

[0096] Connectivity or terminal groups at the end are again shown as OH groups, where a ligand, functional group, extender or other DCRU can be attached for future generations of hyper-branched macromolecules.

[0097] In embodiments of the invention, a building block is used consisting of a multi-armed PEG precursor, as defined below, derived from polyol core units. Petition 870250084859, dated 09 / 19 / 2025, page 47 / 290 39 / 208 ethoxylated, and these multi-armed PEG precursor building blocks have an average molecular weight (Mn) of less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons. Generations

[0098] As with dendrimers, the sequence of constitutional repeating units in the hyper-branched macromolecules of the invention can be designated by generations. In certain embodiments, the hyper-branched macromolecule can be a G0 hyper-branched macromolecule or a G1 to G10 hyper-branched macromolecule, such as a G1, G2, G3, G4 or G5 hyper-branched macromolecule, generally a Gx hyper-branched macromolecule, with x being an integer from 1 to 10. The abbreviation G refers to the generation, and the number designates the total number of constitutional repeating dendritic units that are consecutively linked to each other in a row.

[0099] As shown in FIG. 2, branching and the number of outermost terminal groups increase from generation to generation.

[0100] In an exemplary embodiment, the hyperbranched macromolecule is a G0 branched macromolecule in which the terminal groups located on the surface of the hyperbranched macromolecule are the terminal groups of the polymeric arms connected to the central unit. The G0 branched macromolecule can also be described as a multi-armed PEG molecule with an active agent covalently linked to at least one of its arms.

[0101] In another exemplary embodiment, the hyperbranched macromolecule is a higher-generation Gx hyperbranched macromolecule, with x being an integer from 1 to 10, defining the number of connected dendritic constitutional repeating units. Petition 870250084859, dated 09 / 19 / 2025, page 48 / 290 40 / 208 consecutively in the hyper-branched macromolecule, the polymeric arms of the outermost dendritic constitutional repeating unit each comprising a terminal group, wherein at least one active agent is conjugated to at least one of the outermost polymeric arms.

[0102] Examples of embodiments of the present invention include hyper-branched macromolecules G1 to G10, such as G1 to G8, G1 to G6, or G1 to G4, such as hyper-branched macromolecules G1, G2, G3 or G4.

[0103] The same or different DCRUs can be used to form different generations in a hyper-branched macromolecule, such as DCRUs with different molecular weights (due to different PEG arm lengths) or different numbers of arms. Furthermore, different generations of DCRUs within a hyper-branched macromolecule can be connected to each other using the same ligand and functional groups or with different ligands and functional groups, for example, to control the degradation rate at different junctions within the hyper-branched macromolecule. Remification units

[0104] Branching units can be selected from the same chemical entities as the main units described above. Like the central unit, a branching unit is a branched chemical structure that includes a branching point and a plurality of connectivities.

[0105] While the central unit is at the center of the hyper-branched macromolecule and occurs only once in the hyper-branched macromolecule, a branching unit occurs within the dendritic constitutional repeat units (DCRU) of the hyper-branched macromolecule. A G0 hyper-branched macromolecule includes a central unit but no branching units. Petition 870250084859, dated 09 / 19 / 2025, page 49 / 290 41 / 208 A higher-generation hyper-branched macromolecule of the Gx generation includes a plurality of branching units. The branched units in a hyper-branched macromolecule may have the same chemical structure as the central unit or may be different. For example, the central unit of a G1 hyper-branched macromolecule may be derived from pentaerythritol with a c = 4 connectivity, and the hyper-branched macromolecule may include 4 DCRUs with branching units, each derived from glycerol with a c' = 3 connectivity, so that the G1 hyper-branched macromolecule overall has 8 terminal (outermost) groups. If the 4 DCRUs also have a branched unit derived from pentaerythritol, like the central unit, the hyper-branched macromolecule will have a total of 12 outermost terminal groups. Polymer arms

[0106] In certain embodiments, the polymeric arms of hyper-branched macromolecules are made of polyethylene glycol (PEG) polymer units. In a G0 generation branched macromolecule, the polymeric arms are connected to the central unit, for example, via ether linkages, and have terminal groups located on the surface of the branched macromolecule. The active agent is covalently linked to at least some of these terminal groups. In a higher generation Gx hyper-branched macromolecule, the polymeric arms additionally occur in consecutively connected dendritic constitutional repeating units.

[0107] Thus, in some embodiments, the polymeric arms comprised in the hyper-branched macromolecule are made of or include at least one polyethylene glycol unit. Polyethylene glycol (PEG, also known as polyethylene oxide) refers to a polymer with a repeating group (CH2CH2OX) with n being by Petition 870250084859, dated 09 / 19 / 2025, page 50 / 290 42 / 208 minus 3.

[0108] A polymeric arm that has a polyethylene glycol therefore has at least three of these repeating groups connected to each other in a linear series. The PEG polymeric arm terminates in a stop group, such as a nucleophile or electrophile, a dibenzocyclo-octyne (or other strained alkyne), a strained alkene, a tetrazine or an azide, and can be used for conjugation with an active agent or for connection with a DCRU precursor to build the next-generation hyper-branched macromolecule.

[0109] Polymer arms may comprise units of PEG with an average molecular weight (Mn) in the range of about 1,000 to about 100,000 Daltons, or about 10,000 to about 60,000 Daltons, or about 15,000 to about 50,000 Daltons.

[0110] In certain embodiments, the average molecular weight (Mn) of the PEG units of the core-attached polymer arm may be the same as or different from that of the polymer arms in the dendritic constitutional repeat units. For example, the average molecular weight of the core-attached polymer arm PEG units may be higher or lower than that of the polymer arms in the dendritic constitutional repeat units. In one aspect of this, for a higher-generation biodegradable hyper-branched macromolecule G x, with x being an integer from 2 to 10, the average molecular weight of the polymer arm PEG units may decrease or increase from the innermost polymer arms to the outermost polymer arms. For example, the polymer arms attached to the core unit may have a large molecular weight, and those of the DCRU may have a lower molecular weight, or vice versa. The molecular weight of the polymer arms may also vary from DCRU generation to generation.As an example, a hyper-branched G2 macromolecule with 24 outermost conjugation sites can be constructed from a single core. Petition 870250084859, dated 09 / 19 / 2025, p. 51 / 290 43 / 208 4-arm 40k PEG connected to four 4-arm 20k PEG DCRUs, which can be further connected to twelve 3-arm 30k PEGs. K in this context refers to kilo Daltons (kDa), so a 4-arm 40k PEG has 4 polymeric PEG arms and a total molecular weight of 40 kDa. Constitutional dendritic repeating units

[0111] A dendritic constitutional repeating unit (DCRU) is a partial structure within the higher-generation hyper-branched macromolecule Gx, as defined herein, having a c' connectivity > 3 and including a branching point and polymeric arms emanating from it. It may be connected to a total of c' polymeric arms emanating from the central unit and / or other DCRUs consecutively to form a tree-like dendrimeric structure.

[0112] In certain modalities, the dendritic constitutional repeating unit in the dendrimer can be represented by the general formula (i):

[0113] Within this Formula (i), A is a connection to a polymeric arm that is connected to the central unit, or A is a connection to B of a preceding dendritic constitutional repeat unit represented by Formula (i), La is a linking group, m is 0 or 1, meaning that the linker may be absent or present, n is an integer from 3 to 2000, or 20 to 2000, o is an integer from 3 to 2000, or 20 to 2000, while neo may be different or the same, X is a branching unit, Lb is a linking group, p is 0 or 1, meaning that the linker may be absent or present, B comprises a terminal group located on the surface of the hyper-branched macromolecule or is a connection to A of a consecutive dendritic constitutional repeat unit or a connection to Petition 870250084859, dated 09 / 19 / 2025, page 52 / 290 44 / 208 an active agent, La and Lb may be different or the same, mep may be different or the same, ey ​​is an integer from 2 to 9, wherein y = c - 1 with c being the c connectivity of the branching unit X. In certain embodiments of the hyper-branched macromolecule, including DCRUs of Formula (i), the connection between A and B may comprise a functional group formed by click chemistry, such as a triazole or dihydropyrazine.

[0114] The branching unit may be derived from a polyol, such as glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol, or sorbitol. In Formula (i) above, the branching unit is ethoxylated at all its c connectivities, therefore it is connected via an ether linkage to a PEG polymer arm connected to A, optionally via the La linkage group, and y PEG polymer arms, each connected to B, optionally via an Lb linkage group. Identical or different DCRUs may be used in a hyperbranched macromolecule, such as DCRUs with different molecular weights (due to different PEG arm lengths) or different numbers of arms. Liaison groups

[0115] The inclusion of hydrolysis-labile linking groups in the dendritic constitutional unit allows for biodegradation of the hyper-branched macromolecule under physiological conditions. The high molecular weight hyper-branched macromolecule conjugate can be degraded into small low molecular weight constitutional units and can be eliminated from the body via usual physiological pathways.

[0116] In certain embodiments of the hyper-branched macromolecule, including DCRUs of Formula (i), the La and / or Lb linking groups comprise a dicarboxyl and / or carboxamide moiety or Petition 870250084859, dated 09 / 19 / 2025, page 53 / 290 45 / 208 combinations of these of varying chain length, and these can be derived from diacid groups, such as succinate, glutarate, adipate, azelate, or a diamido acid group, such as glutaramide. These groups can be connected to the polymeric arms of PEG, A and / or B, via ester or amide linkages that are hydrolyzable under physiological conditions in vivo at different rates, depending on the length of the acid chain. In certain embodiments, the linkers form ester linkages and are derived from diacids.

[0117] In certain embodiments, the La and / or Lb ligand comprises a structure represented by Formula (ii): O1o where U1 and U2 are independently NH or O and may be the same or different, and where t is an integer from 0 to 10. For example, in a succinate ligand, U1 and U2 are both oxygen, and t is 2. For a ligand end group, the ligand of Formula (ii) comprises a terminal functional group at one of its ends. In the above example of a succinate ligand, the reaction with N-hydroxysuccinimidyl results in a succinimidylsuccinate ligand end group that can be used to conjugate amine-functionalized active agents to a hyper-branched macromolecule that has this ligand end group on its outermost polymeric arms. In certain embodiments, the La and / or Lb ligand further comprises a polyethylene glycol unit between the linkage to B and the carboxyl group, carboxamide group, or structure of Formula (ii).

[0118] The ligand of Formula (ii) introduces hydrolyzable linkages into the hyper-branched macromolecule that can be used to adjust the degradation rate of the hyper-branched macromolecule and / or the release rate of conjugated active agents from the hyper-branched macromolecule. For example, the biodegradation / hydrolysis rate of linkages Petition 870250084859, dated 09 / 19 / 2025, page 54 / 290 46 / 208 ester in these ligands decreases from succinate (C4) to azelate (C9). In embodiments of the invention, this can be used to control the degradation rate of the hyper-branched macromolecule and / or the release of active agents conjugated via these ligands to the hyper-branched macromolecule. For example, succinimidyl succinate (SS) groups can degrade on the order of a few days, while succinimidyl glutarate (SG) groups degrade on the order of weeks.

[0119] At the other end of the La and Lb linking groups, a terminal group, such as an ester, may be attached, for example, succinimidyl (NHS) groups formed by esterification of the linking acid group with N-hydroxy succinimide or click chemistry functional groups such as DBCO or an azide, as described in more detail below. Extenders

[0120] In addition to or instead of linkers, bifunctional extender units can be incorporated as additional building blocks to extend polymeric arms in length, for example, between arms connected to core and branch units, to provide greater flexibility and / or provide more hydrolytic cleavage points in the dendrimer. These extenders are typically linear bifunctional polymeric chains, such as linear PEG extenders.

[0121] In certain embodiments, the hyper-branched macromolecule comprises at least one extender unit comprising or consisting of polyethylene glycol (PEG) units, wherein the extender unit is linear, difunctional and connected to the polymeric arm of a dendritic constitutional repeat unit or to the polymeric arm connected to the core unit and to a terminal group or to a polymeric arm of a next dendritic constitutional repeat unit.

[0122] In some modalities, the extension unit comprises Petition 870250084859, dated 09 / 19 / 2025, page 55 / 290 47 / 208 at least one ligand, wherein the ligand may be located at either or both ends of the extender unit and is a difunctional ligand comprising hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic, heteroaliphatic or aromatic group or heteroaromatic.

[0123] The inclusion of extenders can be used to enlarge the hydration radius of the dendrimer and increase the half-life of hyper-branched molecules in vivo. PEG precursors for G0 branched macromolecules and DCRU precursors

[0124] The central element of the hyper-branched macromolecule of certain embodiments of the present invention may comprise one or more multi-armed PEG precursors having 2 to 10 arms, or 4 to 8 arms, or 4, 5, 6, 7 or 8 arms. It should be noted that, as multi-armed precursors have a core, a 2-armed PEG precursor, for example, differs from simple linear PEG by the presence of the core structure. The PEG precursors used in a hyper-branched macromolecule may have a different or equal number of arms. In certain embodiments, the PEG precursors used in the hyper-branched macromolecule of the present invention have 3, 4 and / or 8 arms. In certain embodiments, a combination of 4- and 3-armed PEG precursors or a combination of 4- and 8-armed PEG precursors, and any combinations thereof, is used.For example, an 8-armed central unit can be combined with eight 4-armed DCRU precursors, which can then be connected with 24 3-armed precursors, resulting in a hyper-branched macromolecule with 48 conjugation sites on the outermost arms. In another exemplary embodiment, a unit... Petition 870250084859, dated 09 / 19 / 2025, page 56 / 290 The central 48 / 208 4-arm precursor can be combined with four 3-arm precursors, which can in turn be connected to eight 4-arm precursors, resulting in a hyper-branched macromolecule with 24 conjugation sites on the outermost arms. Multi-arm PEG precursors for G0 branched macromolecules and DCRUs in embodiments of the invention are commercially available, for example, from JenKem Technology USA, SinoPEG or Sigma-Aldrich, optionally including several functional terminal groups for further derivatization.

[0125] In certain embodiments of the present invention, polyethylene glycol units used as a central building block or as DCRU precursors have an average molecular weight in the range of about 1,000 to about 80,000 Daltons, or in a range of about 10,000 to about 60,000 Daltons, or in a range of about 15,000 to about 50,000 Daltons. In some embodiments, the polyethylene glycol units have an average molecular weight in the range of about 10,000 to about 40,000 Daltons, or about 20,000 Daltons. PEG precursors with the same average molecular weight may be used, or PEG precursors with different average molecular weights may be combined with each other. The average molecular weight of the PEG precursors used in the present invention is given as the number-average molecular weight (Mn), which, in certain embodiments, can be determined by gel permeation chromatography against a polystyrene standard according to standardized methods.

[0126] In a 4-arm PEG, each arm may have an average arm length (or molecular weight) of the total molecular weight of the PEG divided by 4. A 4a20kPEG precursor, which is a precursor that can be used in the present invention, has 4 arms with an average molecular weight of about 5,000 Daltons (+ / - 500) each, linked to a central pentaerythritol unit. An 8a20k PEG precursor Petition 870250084859, dated 09 / 19 / 2025, page 57 / 290 49 / 208 PEG, which can be used in addition to the 4a20kPEG precursor in the present invention, therefore has 8 arms, each with an average molecular weight of 2,500 (+ / -250) Daltons, linked to a central unit of tripentaerythritol or hexaglycerol.

[0127] In general, when referring to a polymer precursor with a certain average molecular weight, such as a 15KPEG precursor, the indicated average molecular weight (i.e., an Mn of 15,000 or 20,000, respectively) refers to the polymer unit portion of the precursor before the end groups are added (“20k” here means 20,000 Daltons (+ / - 2,000 Da) and “15k” means 15,000 Daltons (+ / - 1,500 Da) – the same abbreviation is used here for other average molecular weights of PEG or other polymer precursors). In certain embodiments, the Mn of the polymer unit portion of the precursor is determined by gel permeation chromatography against a polystyrene standard according to standardized methods. The degree of substitution with end groups as disclosed herein can be determined by 1H-NMR after functionalization of the end group.

[0128] In certain embodiments, the precursors suitable for use in the formation of DCRUs are generally represented by Formula (iii): (m)cí-LA4-(-OCH2CH2-j—o---χ--Q—Í-cH2CH2O-4 çl-Bi---d ' 'm' 'n L ' '°x zp Jyem where C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide, or tetrazine), D comprises functional groups that are not reactive in click chemistry (such as succinimidyl), and La is a ligand group, m is 0 or 1, meaning the ligand may be absent or present, n is an integer from 3 to 2000 or 20 to 2000, o is an integer from 3 to 2000 or 20 to 2000, while neo may be different or the same, X is a branching unit, Lb is Petition 870250084859, dated 09 / 19 / 2025, p. 58 / 290 50 / 208 a linking group, p is 0 or 1, meaning the linker may be absent or present, La and Le may be different or equal, mep may be different or equal, ey is an integer from 2 to 9, where y = c' - 1 with c' being the c' connectivity of the branching unit X.

[0129] In certain embodiments, the useful PEG precursor for forming the DCRUs of the hyper-branched macromolecule is a dicarboxylic acid ester-terminated NHS multiarm PEG precursor derived from commercially available multiarm PEG compounds, such as Formula (iv), an example of a 4-arm structure derived from pentaerythritol.

[0130] A useful PEG precursor for forming a DCRU of certain modalities can be represented by the following Formula (v): where n is determined by the molecular weight of the respective PEG arm, m is an integer from 0 to 10 and, specifically, is 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, ex is the number of arms (and thus can, for example, be 2, 4, 8, etc., see above). Where m is 1, each arm is terminated with a succinimidylsuccinate (SS) end group, where m is 2, each arm is terminated with a succinimidylglutarate (SG) group, where m is 3, each arm is terminated with a succinimidyladipate (SAP) group, and where m is 6, each arm is terminated with a succinimidylazele (SAZ) group. With these specific electrophilic terminal groups, multi-arm PEG units can be abbreviated in the form of, for example, 4a20kPEG-SAP, referring to Petition 870250084859, dated 09 / 19 / 2025, page 59 / 290 51 / 208 is a 4-armed PEG with a terminal succinimidyl adipate group and a molecular weight of 20,000 Da. In the formula above, R is an appropriate core unit structure to provide the desired number of arms. For 4-armed PEG units and precursors, as shown in the formula above, R can be a pentaerythritol structure, while for 8-armed PEG units and precursors, R can be a hexaglycerol structure.

[0131] In certain applications, the PEG precursor used is 4a20kPEG-SG or 4a20kPEG-SAP.

[0132] Instead of electrophilic end groups, precursors with nucleophilic end groups can also be used. In certain embodiments, nucleophilic end groups for use as PEG precursors of hyper-branched macromolecules are amine end groups (denoted as “NH2”). Thiol end groups (-SH) or other nucleophilic end groups are also possible.

[0133] In certain embodiments, 4 PEG arms with an average molecular weight of about 20,000 Daltons and 4 PEG arms with an average molecular weight of about 40,000 Daltons can be used to form the hyper-branched macromolecules according to the present invention. Functional groups for connecting building blocks of hyper-branched macromolecules.

[0134] To synthesize the hyper-branched macromolecule, the polymeric arms or precursors have pairs of functional groups that react with each other, i.e., a first functional group in a first polymeric arm or precursor is able to react with a second functional group in a second polymeric arm or precursor in a different DCRU precursor.

[0135] In one embodiment, an early precursor of multiple arms, including the central unit and the PEG arms connected to it, Petition 870250084859, dated 09 / 19 / 2025, page 60 / 290 52 / 208 comprises first functional groups, and a second multi-arm DCRU precursor comprises a second functional group capable of reacting with the first functional groups, while all other terminal groups of this second DCRU precursor do not react with the first functional group, the functional groups being located at the ends of the arms of the precursor or DCRU. The first and second functional groups may be grafted directly onto the ends of the arms, or via a ligand, preferably a hydrolyzable ligand as defined elsewhere herein. The functional groups are capable of reacting with each other and forming a covalent bond, for example, in click chemistry reactions or electrophile-nucleophile reactions, or are configured to participate in other chemical crosslinking reactions, as described below.

[0136] In certain embodiments of the invention, the first functional group and the second functional group are selected from among an electrophile and a nucleophile, functional groups for click chemistry, functional groups for cycloadditions, particularly 1,3-dipolar cycloadditions, hetero-Diels-Alder cycloadditions, functional groups for nucleophilic ring openings, functional groups for non-aldol-type carbonyl reactions, functional groups for addition reactions to carbon-carbon multiple bonds, polymerizable vinyl groups, or combinations thereof. Those skilled in the art will know that certain pairs of functional groups can be classified into more than one of these groups. For example, in click chemistry, an azide reacting with dibenzocyclooctyne can also be viewed as an electrophile-nucleophile reaction pair.

[0137] In certain embodiments of the hyperbranched macromolecules of the present invention, the connections between different parts of the hyperbranched macromolecule, such as the polymeric arms connected to the central unit and the DCRUs, are formed by reactions Petition 870250084859, dated 09 / 19 / 2025, page 61 / 290 53 / 208 of click chemistry, such as strain-promoted alkyne-azide cycloaddition (SPAAC), also termed Cu-free click reaction, or inverse electron demand Diels-Alder bonding type click chemistry coupling reactions (IEDDA). An overview of these reaction types is provided in HC Kolb; MG Finn; KB Sharpless (2001).“Click Chemistry: Diverse Chemical Function from a Few Good Reactions”, Angewandte Chemie International Edition, 40 (11): 2004-2021), incorporated herein by reference.

[0138] Other click chemistry reactions suitable for connecting constitutional units of hyper-branched macromolecules of certain embodiments include aldehyde / ketone condensation, cyanobenzothiazole condensation; strain-promoted and oxidation-controlled cyclo-octyno-1,2-quinone (SPOCQ) cycloaddition; 1,3-dipolar cycloadditions, [3+2] cycloadditions, such as alkene-nitrone cycloadditions or alkene-nitrone cycloadditions, [4+2] cycloadditions; and hetero-Diels-Alder reactions.

[0139] SPAAC requires an alkyne with a ring structure, such as dibenzylcyclooctyne (DBCO) and bicyclo[6.1.0]nonyne (BCN), to react with an aliphatic azide. This forced chemistry causes the reaction to occur efficiently, without the need for a copper catalyst required in copper(I)-catalyzed azide-alkyne click reactions (CuAAC). Similarly, IEDDA requires the reaction of norbornene and tetrazine without the need for a catalyst. Therefore, the advantage of SPAAC and IEDDA over CuAAC and electrophilic-nucleophilic reactions, such as NHS-NH2, is that there is no need for a catalyst and no byproduct after the reaction is complete.

[0140] SPAAC and IEDDA coupling reactions are bioorthogonal reactions with selective and quantitative yields under moderate conditions that can occur even within living systems without interfering with native biochemical processes. These reactions Petition 870250084859, dated 09 / 19 / 2025, page 62 / 290 54 / 208 click chemistries utilize a pair of reagents, for example, cyclooctines and azides, which react exclusively and efficiently with each other, remaining inert to the natural functional groups:

[0141] Scheme A: with R1 and R2 being any residues that are the same or different.

[0142] This reaction is suitable for forming hyperbranched macromolecules of embodiments of the invention from correspondingly functionalized precursors and DCRUs, as described herein. Among the considerable number of known cyclooctines, dibenzocyclooctine (DBCO) compounds comprise a class of reagents that possess reasonably fast kinetics and good stability in aqueous buffers. Within the physiological temperature and pH ranges, the DBCO group will not react with amines or hydroxyls that are naturally present in many biomolecules or present as different functional groups in parts of the hyperbranched macromolecule. Furthermore, the reaction of the DBCO group with the azide group is significantly fast and high-yielding.

[0143] The advantages of DBCO-based SPAAC include, for example, its biocompatibility, as it does not require cytotoxic copper catalysts that can remain in undesirable traces in hyper-branched macromolecules. Another advantage is the use of mild reaction conditions: the binding of DCRUs or the conjugation of active agents is possible in aqueous buffered media or common organic solvents under physiological conditions. Furthermore, the DBCO and azide moieties are stable in the long term and have Petition 870250084859, dated 09 / 19 / 2025, page 63 / 290 55 / 208 high selectivity and specificity, since the azide groups react only with DBCO in the presence of amine, hydroxyl, thiol, and acid groups, as well as other protein functional groups. Furthermore, the reactions lead to the formation of a stable triazole with quantitative yield and a high reaction rate, leaving no byproducts. Similar advantages are provided by IEDDA coupling reactions and other types of catalyst-free click chemical reactions mentioned previously.

[0144] In exemplary embodiments, hyperbranched macromolecules including hydrolyzable ligands derived from diacids can be formed using the following precursors for click chemistry: 4-arm PEG-DBCO and any D 4-arms-PEG-(DBCO)3(Azide)1 or Petition 870250084859, dated 09 / 19 / 2025, page 64 / 290 56 / 208 h2n—ç o- N5-^L°ch2ch^nh ywH ^^CH-CHj+C *· ' n y-Í0CH2Cl-;4- wC h2n^ í \ .O—hCHICHO-K 1l4och2ch2+of $ ' 'n n---CH O-^íL λ ϊ_ ϊ_ / 4och2ch2+o V rjn2 n 4-ai 4-arms PEG-Amine(3)Azide(1) ;ou vx 2 f \ *o—FCHjCHjoV r ' * n J a-í-CH^k^ay. n ít ' n ° ' Ι·.(<:Η<:Η;θΚ Ns 4-armed-PEG-Azide . and any / \ / \ XO-fCI-UCI-LO-K +0CH2CH2+0 [ ' 'n / L· _í 2l J °H / CHí-K|X+z+o\ch2c fy Krt N f— NH ff 1 \ / 4-armed-PEG-Amina(3)DBCO(1) O o—\ / \ .oI-ch^choA7 MCCHtC Hj+or 5 ' ” fn '-----' -LL· / 3 > Ol-CH-.CHj^u-o'ch 'j^ GH 4 / f) r í r / -H 4-arms-PEG-NHS(3)DBCO(1) oz OU c ; Petition 870250084859, of 09 / 19 / 2025, p. 65 / 290 57 / 208 4-arm-PEG-Amine and some Petition 870250084859, of 09 / 19 / 2025, p. 66 / 290 58 / 208 where t is m, enem are defined as for formula (v) here before.

[0145] Instead of 4-armed PEG, other core / branch unit connectivities may also be used, as described here. Furthermore, in other embodiments, the above precursors may include hydrolyzable linkages, including carboxamide linkages instead of ester linkages, or ester and amide linkages, as in the SGA linkage units described later in this document.

[0146] Using click chemistry functional groups, in certain embodiments, connections in the hyper-branched macromolecule can be selectively formed using DCRU precursors, such as those above, which include a functional group for click chemistry bond formation, while the other terminal functional groups of the DCRU remain non-reactive and can be used subsequently for other follow-up reactions, such as growth or conjugation of the hyper-branched macromolecule. In other embodiments, the Petition 870250084859, dated 09 / 19 / 2025, page 67 / 290 59 / 208 connections in the hyper-branched macromolecule can be selectively formed using electrophilic-nucleophilic-precursors or other non-reactive functional groups in click chemistry, while the other terminal functional groups of the DCRU include a functional group for click chemistry bond formation and remain non-reactive, and can be used later for other follow-up reactions, such as hyper-branched macromolecule growth or conjugation with click chemistry reactions.

[0147] Functional group pairs for click chemistry can be functional groups selected for cycloadditions, particularly 1,3-dipolar cycloadditions, [3+2] cycloadditions such as alkene-nitrone cycloadditions or alkyne-nitrone cycloadditions, [4+2] cycloadditions, hetero-Diels-Alder cycloadditions; functional groups for thiolene reactions; functional groups for nucleophilic ring openings; functional groups for non-aldol type carbonyl reactions; functional groups for addition reactions to carbon-carbon multiple bonds; functional groups for Michael type additions.

[0148] For example, the first functional group is an alkyne compound, such as a dibenzocyclooctyne (DBCO), or a bicyclo[6.1.0]nonyne (BCN); or a norbornene, or a trans-cyclooctene (TCO); and the second functional group is an azide, a 3,4-dihydroxyphenylacetic acid (DHPA) or a tetrazine (Tz). In these embodiments, the functional groups DBCO, BCN, norbornene, TCO, azide, DHPA and Tz can be grafted onto the ends of the multi-arm precursor by means of a hydrolyzable ligand, such as an acid group, a diacid group, an amide group, a functionalized aliphatic, heteroaliphatic or aromatic or heteroaromatic group, or can be directly connected to the PEG.

[0149] In another embodiment, the first and second functional groups are selected for a [3+2] cycloaddition reaction. Petition 870250084859, dated 09 / 19 / 2025, p. 68 / 290 60 / 208 as alkene-nitrone cycloadditions or alkyne-nitrone cycloadditions. In another embodiment, the first and second functional groups are selected for a [4+2] cycloaddition reaction, particularly a hetero Diels-Alder reaction, wherein the first functional group is an aldehyde or imine compound, and the second functional group is a 1,3-diene compound, an unsaturated carbonyl compound, or a nitroso-alkene compound. In another embodiment, the first and second functional groups are selected for nucleophilic ring openings, wherein the first functional group is selected from an epoxide, tyrane, aziridine, or lactam, and the second functional group is a nucleophile, as mentioned above.In another embodiment, the first and second functional groups are selected for non-aldol type carbonyl reactions, wherein the first functional group is an aldehyde or ketone compound, and the second functional group is a primary amine, a hydrazide, acyl hydrazide, or amino-oxy compound, to form an imine, amide, isourea, hydrazone, acyl hydrazone, or oxime linkage. Conjugation of active agents

[0150] The linking or conjugation of the active agent to the outermost polymeric arms of the hyper-branched macromolecule can also be done by click chemistry, as described above for connecting building blocks of the hyper-branched macromolecule, or by electrophile-nucleophile reactions and other types of coupling reactions, as mentioned here.

[0151] Thus, in one embodiment, the first functional group on the outermost polymeric arms of the hyper-branched macromolecule may be a nucleophile and the second functional group on the active agent may be an electrophile, or vice versa, and the reaction between the first functional group and the second functional group is an electrophile-nucleophile reaction that forms a covalent bond. Petition 870250084859, dated 09 / 19 / 2025, page 69 / 290 61 / 208

[0152] Nucleophiles may be selected from among an amine, such as a primary amine, a hydroxyl group, a thiol group, a carboxyl group, or a hydrazide group. In certain embodiments, one of the functional groups comprises a nucleophile, such as a primary amine.

[0153] Electrophiles that can be used in embodiments of the present invention can be selected from activated ester groups, such as succinimidyl esters, succinimidyl carbonates; nitrophenyl carbonates, aldehydes, ketones, acrylates, acrylamides, maleimides, vinyl sulfones, iodoacetamides, alkenes, norbornenes, epoxides, mesylates, tosylates, tresylates, cyanurates, orthopyridyl disulfides or halogens. These electrophiles comprise functional groups that participate in the electrophile-nucleophile reaction and preferably additionally include reactive groups that form PEG ligands that include hydrolyzable groups or linkages, such as glutarate. For example, in one embodiment of the invention, a succinimidyl ester may comprise a reactive group, such as succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ), or succinimidyl glutaramide.Such electrophilic-nucleophilic reactions for connecting multi-armed PEG precursors are described, for example, in US 2002 / 0042473A1, which is incorporated by reference.

[0154] The active agent may be suitably derivatized with functional groups, as mentioned above, unless it already has a functional group suitable for connecting with the hyper-branched macromolecule. For example, peptides possessing primary amino groups may be conjugated via an electrophilic-nucleophilic reaction to a hyper-branched macromolecule possessing an activated ester group on its surface.

[0155] In certain embodiments, the active agent, particularly peptides, can be conjugated by means of click chemical reactions. Petition 870250084859, dated 09 / 19 / 2025, page 70 / 290 62 / 208 to the hyper-branched macromolecule. In some embodiments, the active agent or peptide having a terminal primary amino group is first reacted with DBCO-NHS or azide-NHS compounds to produce an active agent or peptide functionalized with DBCO or an azide group suitable to react with its equivalent functional group at the terminal ends of the hyper-branched macromolecule, producing conjugates with high reproducibility.

[0156] Suitable reagents for chemical functionalization of active agents or peptides with a terminal primary amino group are, for example, / V-hydroxysuccinimidyl ester of azidoacetic acid (NHSazide), / V-hydroxysuccinimidyl ester of azidobutyric acid or other azidoacid-NHS esters, and / V-hydroxysuccinimidyl ester of dibenzocyclooctyne (DBCO-NHS) of variable acid chain length. Both azide-NHS esters and DBCO-NHS esters can be used with acids of different chain lengths (as discussed as ligands herein earlier) to vary the rate of biodegradation and release of the active agent from hyper-branched macromolecules. These reagents for click chemistry are commercially available, for example, from SigmaAldrich or Thermo Fisher Scientific and other suppliers.

[0157] In certain embodiments, the active agent or peptide with thiol group functionality for conjugation, such as a cysteine ​​thiol group, can be conjugated to the hyper-branched macromolecule via maleimide-thiol click chemical reactions according to the following reaction scheme:

[0158] Scheme B: the with R1 being the terminal end of the hyperbranched macromolecule and R2 being a peptide or an active agent. The thiol reaction... (Petition 870250084859, 19 / 09 / 2025, page 71 / 290) 63 / 208 maleimide is a Michael addition reaction of thiols that produces thio-succinimide linkages. The reaction is fast and chemoselective for thiols at pH 6.5 to pH 7.5.

[0159] For example, maleimide-functionalized terminal ends of the hyperbranched macromolecule can be used to conjugate peptides or active agents via maleimide-thiol reactions. In another embodiment, DBCO- or azide-functionalized terminal ends of the hyperbranched macromolecule can be provided with a maleimide terminal functionalization by reaction with click chemistry ligands having azide or DBCO functionality and a maleimide group at the other end, which is then used for conjugation with thiol groups in a peptide or active agent. Suitable DBCO-maleimide or azidamaleimide ligands can optionally be extended with PEG moieties and are commercially available from Sigma-Aldrich, TCI, Thermo Fisher, etc.

[0160] Examples are compounds such as DBCO-maleimide, DBCO-PEG3-maleimide, DBCO-PEG4-maleimide, azido-PEG3-maleimide, with the following exemplary structures:

[0161] In certain embodiments, the active agent is linked to at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or even 100% of the outermost polymeric arms. The average substitution rate of the active agent conjugated to the surface terminal groups of the hyper-branched macromolecule can be determined by UHPLC, as described in more detail here. Active agents:

[0162] The active agent in biodegradable microparticles of Petition 870250084859, dated 09 / 19 / 2025, page 72 / 290 64 / 208 embodiments of the invention may be a therapeutically active agent or a diagnostically active agent, or combinations thereof. It may be a single active agent or a plurality of active agents.

[0163] In some embodiments, the hyperbranched macromolecule comprises two or more different active agents on different dendrons or regions on the surface of the hyperbranched macromolecule. Two or more active agents may be attached, each with the same or different hydrolyzable groups to control the release of the active agents at different rates. In addition, the active agents may be attached to the dendrimer with or without hydrolyzable linkages or arms / extenders, or combinations thereof, to control the release of the active agents at different rates.

[0164] In certain embodiments, the active agent conjugated to at least one of the outermost polymeric arms of the hyper-branched macromolecule is a peptide selected from the group consisting of Compstatin, APL-1, Fc-III-4C, Beovu (Brolucizumab), Zimura (Avacincaptade Pegol), Pegcetacoplan, Abicipar Pegol, Lampalizumab, Fovista, Risuteganib, AXT107, Elamipretide, THR149, ALM201, VGB3 and Largazole.

[0165] Therapeutically active agents may be steroids; nonsteroidal anti-inflammatory drugs (NSAIDs), such as diclofenac, ibuprofen, meclofenamate, mefanamic acid, salsalate, sulindac, tolmetin, ketoprofen, diflunisal, piroxicam, naproxen, etodolac, flurbiprofen, fenoprofen C, indomethacin, celecoxib, ketorolac, nepafenac; intraocular pressure-lowering drugs; antibiotics, such as ciprofloxacin; analgesics, such as bupivacaine; calcium channel blockers, such as nifedipine; cell cycle inhibitors, such as simvastatin; proteins, such as insulin; hydrophilic small molecule drugs, including carboxylic acid salts and amine salts; hydrophobic small molecule drugs, peptides Petition 870250084859, dated 09 / 19 / 2025, page 73 / 290 65 / 208 hydrophilic and protein drugs, such as insulin, single-chain antibody fragments, Fab fragments, IgG antibodies, fusion antibodies, etc.; aptamers; particularly Bupivacaine (BPV-HCl or base), Ropivacaine (RPV), Dexamethasone, Travoprost, Axitinib, non-steroidal anti-inflammatory drugs (NSAIDs), steroids, antibiotics, analgesics, calcium channel blockers, cell cycle inhibitors, chemotherapeutic agents, antiviral drugs, anesthetics, hormones, anticancer drugs, antineoplastic agents, viruses, gene delivery viruses such as AAVs, protein ligands such as nanobodies, affibodies, ankyrins, DARPins, etc., or any combination thereof.

[0166] In some embodiments, the steroids may be corticosteroids which may include hydrocortisone, loteprednol, cortisol, cortisone, prednisolone, methylprednisolone, dexamethasone, betamethasone, triamcinolone, aldosterone or fludrocortisone.

[0167] In some embodiments, NSAIDs may comprise diclofenac (e.g., diclofenac sodium), flurbiprofen (e.g., flurbiprofen sodium), ketorolac (e.g., ketorolac tromethamine), bromfenac, or nepafenac.

[0168] In some embodiments, IOP-lowering agents and / or glaucoma medications may comprise prostaglandin analogues (e.g., bimatoprost, latanoprost, travoprost or latanoprostene bunod), rho kinase inhibitors (e.g., netarsudil), adrenergic agonists (epinephrine or dipivefrine), beta-adrenergic antagonists, also known as beta-blockers (e.g., timolol, levobunolol, metipranolol, carteolol or betaxolol), alpha2-adrenergic agonists (e.g., apraclonidine, brimonidine or brimonidine tartrate), carbonic anhydrase inhibitors (e.g., brinzolamide, dichlorphenamide, methazolamide, acetazolamide, or dorzolamide), Petition 870250084859, dated 09 / 19 / 2025, page 74 / 290 66 / 208 pilocarpine, ecothiophate, demercarium, physostigmine and / or isofluorophate.

[0169] In some embodiments, the anti-infective may comprise antibiotics including ciprofloxacin, tobramycin, erythromycin, ofloxacin, gentamicin, fluoroquinolone antibiotics, moxifloxacin and / or gatifloxacin; antivirals including ganciclovir, idoxuridine, vidarabine and / or trifluridine; and / or antifungals including amphotericin B, natamycin, voriconazole, fluconazole, miconazole, clotrimazole, ketoconazole, posaconazole, echinocandin, caspofungin and / or micafungin.

[0170] In some embodiments, antimetabolites may comprise methotrexate, mycophenolate, or azathioprine. In some embodiments, antifibrotic agents may comprise mitomycin C or 5-fluorouracil.

[0171] In some embodiments, angiogenesis inhibitors may comprise anti-VEGF agents (e.g., aflibercept, ranibizumab, bevacizumab), PDGF-β inhibitors (e.g., Fovista®), complement antagonists (e.g., eculizumab), tyrosine kinase inhibitors (e.g., sunitinib, axitinib), and / or integrin antagonists (e.g., natalizumab and vedolizumab).

[0172] In some embodiments, nanobodies may be conjugated to hyper-branched macromolecules. Nanobodies are described, for example, in Yang et al. (2020), Nanobodies: Next Generation of Cancer Diagnostics and Therapeutics, Front. Oncol. 10:1182, which is incorporated herein by reference in its entirety. Nanobodies can be selected from 68GaNOTA-Anti-HER2VHH1, 68GaNOTA-Anti-HER2-VHH1, 99mTc-NM-O2, 131I-SGMIB-AntiHER2-VHH1, 68GaNOTA-Anti-MMR-VHH2, 99mTc-Anti-PD-L1, L-DOS47 + Doxorubicin, L-DOS47 + Cisplatin / Vinorelbine, KNO35 + Trastuzumab / Docetaxel, KN035, KN044, TC-210 T cells, CD19 / CD20 bispecific CAR T cells, BCMA CAR T cells or nanobodies Petition 870250084859, dated 09 / 19 / 2025, page 75 / 290 67 / 208 TAS266.

[0173] In some embodiments, non-immunoglobulin affinity proteins, such as affibodies, can be conjugated to hyper-branched macromolecules. Affibody molecules are described, for example, in Stâhl et al., Affibody Molecules in Biotechnological and Medical Applications, Trends in Biotechnology 2017, 35 (8) p.691-712, which is incorporated here by reference in its entirety.

[0174] In some embodiments, binding proteins, such as ankyrins and DARPins, can be conjugated to hyper-branched macromolecules. Ankyrins and DARPins are described, for example, in a review by Caputi et al., Current Opinion in Pharmacology 2020, 51:93-101, which is incorporated here by reference in its entirety. Ankyrins and DARPins can be selected from MP0250, a trispecific DARPin drug candidate that can bind to VEGFA and hepatocyte growth factor (HGF), as well as an MP0250 molecule that binds to two molecules of human serum albumin (HSA); Abicipar pegol (MP0112 or AGN-150998); Brolucizumab, Ranibizumab, or Aflibercept.

[0175] In some embodiments, cytoprotective agents may include ebselen, sulforaphane, oltipraz, or dimethyl fumarate. In some embodiments, neuroprotective agents may include ursodiol, memantine, or acetylcysteine. In some embodiments, anesthetic agents may include lidocaine, proparacaine, or bupivacaine.

[0176] In some embodiments, the active agent may be dexamethasone, ketorolac, diclofenac, vancomycin, moxifloxacin, gatifloxicin, besifloxacin, travoprost, 5-fluorouracil, methotrexate, mitomycin C, prednisolone, bevacizumab (Avastin®), ranibizumab (Lucentis®), sunitinib, pegaptanib (Macugen®), timolol, latanoprost, Petition 870250084859, dated 09 / 19 / 2025, page 76 / 290 68 / 208 brimonidine, nepafenac, bromfenac, triamcinolone, difluprednate, fluocinolide, aflibercept, or combinations thereof. In some embodiments, the agent may be dexamethasone, ketorolac, diclofenac, moxifloxacin, travoprost, 5-fluorouracil, or methotrexate.

[0177] In alternative embodiments, the active agents that can be used with the dendrimers and methods of the present invention include, but are not limited to, immunosuppressants, complement inhibitors (e.g., C5 inhibitors such as eculizumab or avacincaptade pegol), steroids, anti-inflammatory agents such as steroidal and non-steroidal anti-inflammatory drugs (e.g., COX1 or COX2 inhibitors), antivirals, antibiotics, antiglaucoma agents, anti-VEGF agents, analgesics, tyrosine kinase inhibitors, integrin inhibitors, IL-6 blockers, reactive aldehyde species (RASP) inhibitors, nitric oxide donor PgAs, antihistamines, mast cell stabilizers, rho kinase inhibitors, plasma kallikrein inhibitors, BCL-2 blockers, semaphorin antagonists, HtRA1 blockers, IGF1R inhibitors, combination agents of VEGF (multispecific anti-angiogenic agents) and combinations thereof.

[0178] Immunosuppressants include, but are not limited to, cyclosporine, mTOR inhibitors (e.g., rapamycin, tacrolimus, temsirolimus, sirolimus, everolimus, KU-0063794, WYE-354, AZD8055, metformin or Torin-2), cyclophosphamide, atoposide, thiotepa, methotrexate, azathioprine, mercaptopurine, interferons, infliximab, etanercept, mycophenolate mofetil, 15-deoxyspergualin, thalidomide, glatiramer, leflunomide, vincristine, cytarabine, pharmaceutically acceptable salts thereof and combinations thereof.

[0179] Nonsteroidal anti-inflammatory compounds include cyclooxygenase (COX) enzyme inhibitors, such as cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) isoenzymes. The general classes of Petition 870250084859, dated 09 / 19 / 2025, page 77 / 290 69 / 208 Nonsteroidal anti-inflammatory compounds include salicylates, propionic acid derivatives, acetic acid derivatives, enolic acid derivatives, and anthranilic acid derivatives. Examples of nonsteroidal anti-inflammatory compounds include acetylsalicylic acid, diflunisal, salsalate, ibuprofen, dex-ibuprofen, naproxen, fenoprofen, ketoprofen, dex-ketoprofen, flurbiprofen, oxaprozin, loxoprofen, indomethacin, tolmetin, sulindac, etodolac, ketorolac, diclofenac, aceclofenac, nabumetone, piroxicam, tenoxicam, loroxicam, phenylbutazone, mefenamic acid, meclofenamic acid, flufenamic acid, tolfenamic acid, celecoxib, pharmaceutically acceptable salts thereof, and combinations thereof.

[0180] Anti-inflammatory agents that can be used with the dendrimers and methods of the present invention may include agents that target inflammatory cytokines, such as TNFα, IL-1, IL-4, IL-5 or IL-17, or CD20. Such agents may include etanercept, infliximab, adalimumab, daclizumab, rituximab, tocilizumab, certolizumab pegol, golimumab, pharmaceutically acceptable salts thereof and combinations thereof.

[0181] Analgesics that can be used with the dendrimers and methods of the present invention include acetaminophen, acetaminosalol, aminochlortenoxazine, 2-amino-4-picoline acetylsalicylic acid, acetylsalicylic acid, anileridine, benoxaprofen, benzylmorphine, 5-bromosalicylic acid acetate, bucetine, buprenorphine, butorphanol, capsaicin, cinchofen, ciramadol, clometacin, clonixin, codeine, desomorphine, dezocine, dihydrocodeine, dihydromorphine, dimefeptanol, dipyrocethyl, eptazocine, etoxazene, ethylmorphine, eugenol, floctafenine, fosfosal, glafenine, hydrocodone, hydromorphone, hydroxypethidine, ibufenac, p-lactofenetide, levorphanol, meptazinol, metazocine, metopon, morphine, nalbuphine, nicomorphine, norlevorphanol, normorphine, Petition 870250084859, dated 09 / 19 / 2025, page 78 / 290 70 / 208 oxycodone, oxymorphone, pentazocine, phenazocine, phenocol, phenoperidine, phenylbutazone, phenylsalicylate, phenylramidol, salicin, salicylamide, thiorphan, tramadol, diacerein, actarit, pharmaceutically acceptable salts thereof and combinations thereof.

[0182] Antibiotics that can be used with the dendrimers and methods of the present invention include aminoglycosides, penicillins, cephalosporins, fluoroquinolones, macrolides, and combinations thereof. Aminoglycosides may include tobramycin, kanamycin A, amikacin, dibekacin, gentamicin, sisomicin, netilmicin, neomycin B, neomycin C, neomycin E, streptomycin, paromomycin, pharmaceutically acceptable salts thereof, and combinations thereof. Penicillins may include amoxicillin, ampicillin, bacampicillin, carbenicillin, cloxacillin, dicloxacillin, flucloxacillin, mezlocillin, nafcillin, oxacillin, penicillin G, penicillin V, piperacillin, pivampicillin, pivmecillinam, ticarcillin, pharmaceutically acceptable salts thereof, and combinations thereof.Cephalosporins may include cefacetrile, cefadroxil, cephalexin, cephaloglycin, cephalonium, cephaloridine, cephalothin, cefapirin, cefatrizin, cefazaflur, cefazedone, cefazolin, cefradine, cefroxadine, ceftezole, cefaclor, cefamandole, cefmetazole, cefonicid, cefotetan, cefoxitin, cefprozil, cefuroxime, cefuzonam, cefcapene, cefdaloxime, cefdinir, cefditoren, cefetamet, cefixime, cefmenoxime, cefodizime, cefotaxime, cefpimizole, cefpodoxime, cefteram, ceftibuten, ceftiofur, ceftiolene, ceftizoxime, ceftriaxone, cefoperazone, ceftazidime, cefclidine, cefepime, Cefluprenam, cefoselis, cefozoprano, cefpirome, cefquinoma, ceftobiprole, ceftaroline, cefaclomezine, cefaloram, cefaparole, cefcanel, cefedrolor, cefempidone, cefetrazole, cefivitrile, cefmatileno, cefmepidium, cefovecin, cefoxazole, cefrotila, cefsumida, cefuracetime, ceftioxide, pharmaceutically acceptable salts thereof and combinations thereof. Fluoroquinolones may include. Petition 870250084859, dated 09 / 19 / 2025, page 79 / 290 71 / 208 ciprofloxacin, levofloxacin, gatifloxacin, moxifloxacin, ofloxacin, norfloxacin, pharmaceutically acceptable salts thereof and combinations thereof. Macrolides may include azithromycin, erythromycin, clarithromycin, dirithromycin, oxithromycin, telithromycin, pharmaceutically acceptable salts thereof and combinations thereof.

[0183] Antivirals that can be used with the dendrimers and methods of the present invention include nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, fusion inhibitors, integrase inhibitors, nucleoside analogs, protease inhibitors, and reverse transcriptase inhibitors. Examples of antiviral agents include, but are not limited to, abacavir, acyclovir, adefovir, amantadine, amprenavir, ampligen, arbidol, atazanavir, boceprevir, cidofovir, darunavir, delavirdine, didanosine, docosanol, edoxuridine, efavirenz, emtricitabine, enfuvirtide, entecavir, famciclovir, fomivirsen, fosamprenavir, foscarnet, fosfonet, ganciclovir, ibacitabine, immunovir, idoxuridine, imiquimod, indinavir, inosine, type III interferon, type II interferon, type I interferon, lamivudine, lopinavir, loviride, maraviroc, moroxidine, methizazone, nelfinavir, nevirapine, nexavir, oseltamivir, peginterferon alfa-2a, penciclovir,peramivir, pleconaril, podophyllotoxin, raltegravir, ribavirin, rimantadine, ritonavir, saquinavir pyramidal, stavudine, tenofovir, tenofovir disoproxil, tipranavir, trifluridine, trizivir, tromantadine, truvada, valacyclovir, valgancyclovir, vicriviroc, vidarabine, viramidine, zalcitabine, zanamivir, zidovudine, pharmaceutically acceptable salts thereof and combinations thereof.

[0184] Steroidal anti-inflammatory agents that can be used with the dendrimers and methods of the present invention include dexamethasone, budensonide, triamcinolone, hydrocortisone, fluocinolone, loteprednol, prednisolone, mometasone, fluticasone, Petition 870250084859, dated 09 / 19 / 2025, page 80 / 290 72 / 208 rimexolone, fluorometholone, beclomethasone, flunisolide, pharmaceutically acceptable salts thereof and combinations thereof.

[0185] Antiglaucoma agents that can be used with the dendrimers and methods of the present invention include beta-blockers, such as atenolol, propranolol, metipranolol, betaxolol, carteolol, levobetaxolol, levobunolol, timolol, pharmaceutically acceptable salts thereof and combinations thereof; adrenergic agonists or sympathomimetic agents, such as epinephrine, dipivefrine, clonidine, aparclonidine, brimonidine, pharmaceutically acceptable salts thereof and combinations thereof; parasympathomimetic agonists or cholinergic agonists, such as pilocarpine, carbachol, iodophospholine, physostigmine, pharmaceutically acceptable salts thereof and combinations thereof; carbonic anhydrase inhibitors, including topical or systemic agents, such as acetazolamide, brinzolamide, dorzolamide; methazolamide, ethoxzolamide, dichlorphenamide, pharmaceutically acceptable salts thereof and combinations thereof;Mydriatic-cycloplegic agents, such as atropine, cyclopentolate, succinylcholine, homatropine, phenylephrine, scopolamine, tropicamide, pharmaceutically acceptable salts thereof and combinations thereof; prostaglandins, such as prostaglandin F2 alpha, antiprostaglandins, prostaglandin precursors or prostaglandin analogues, such as bimatoprost, latanoprost, travoprost, unoprostone, tafluprost, pharmaceutically acceptable salts thereof and combinations thereof.

[0186] Anti-VEGF agents that can be used with the dendrimers and methods of the present invention include bevacizumab, pegaptanib, ranibizumab, brolucizumab, conbercept, aflibercept, pharmaceutically acceptable salts thereof and combinations thereof. Petition 870250084859, dated 09 / 19 / 2025, page 81 / 290 73 / 208

[0187] Tyrosine kinase inhibitors that can be used with the dendrimers and methods of the present invention include deucravacitinib, axitinib, avapritinib, capmatinib, pegimatinib, ripretinib, selpercatinib, selumetinib, tucatinib, entrectinib, erdaftinib, fedratinib, pexidartinib, upadacatinib, zanubrutinib, baricitinib, binimetinib, dacomitinib, fostamatinib, gilteritinib, larotrectinib, lorlatinib, acalabrutinib, brigatinib, midostaurin, neratinib, alectinib, cobimetinib, lenvatinib, osimertinib, ceritinib, nintedanib, afatinib, ibrutinib, trametinib, bosutinib, cabozantinib, ponatinib, regorafenib, tofacitinib, crizotinib, ruxolitinib, vandetanib, Pazopanib, lapatinib, nilotinib, dasatinib, sunitinib (vorolanib), sorafenib, erlotinib, gefitinib, imatinib, afatinib, bosutinib, cabozantinib, cediranib, ceritinib, crizotinib, dabrafenib, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lestaurtinib,nilotinib, palbociclib, pazopanib, ponatinib, regorafenib, ruxolitinib, semananib, sirolimus, sorafenib, temsirolimus, tofacitinib, trametinib, vandetanib, and vemurafenib. In another embodiment, the tyrosine kinase inhibitor is a tyrosine kinase inhibitor of the Src family, such as, but not limited to, A419259, AP23451, AP23464, AP23485, AP23588, AZD0424, AZM475271, BMS354825, CGP77675, CU201, ENMD 2076, KB SRC 4, KX2361, KX2-391, MLR 1023, MNS, PCI-32765, PD166285, PD180970, PKC412, PKI166, PP1, PP2, SRN 004, SU6656, TC-S7003, TG100435, TG100948, TX-1123, VAL 201, WH-4-023, XL 228, altenusin, bosutinib, damnacanthal, dasatinib, herbimycin A, indirubin, neratinib, lavendustin A, pelitinib, piceatanol, saracatinib, SrcI1, foretinib, motesanib, tivozanib, LY2457546, MGCD-265, MGCD510, tivantinib, AMG458, JNJ-3887, EMD1214063, BMS794833, PHI1665752, SGX-523, INCB280, pharmaceutically acceptable salts thereof and combinations thereof. Petition 870250084859, dated 09 / 19 / 2025, page 82 / 290 74 / 208

[0188] Complement pathway modulators that can be used with the dendrimers and methods of the present invention include those that target, for example, C1 / C1Q, C3, C3 Convertase, C5, C5 convertase, C5a, C5aR, C6, C7, C8, C9, CD59, Factor B, Factor D, Factor H, Factor P, or a combination thereof. Specific agents may include cinryze, berinert, ruconest, sutimlimab, pegcetacoplan (GA), eculiziumab, ravuilizumab, avacopan, pozelimab, nomacopan, zilucopan, vilobelimab, crovalimab, avacincapted pegol), cemdisiran, BDB-001, tesidolumab, avdoralimab, MOR210, ALXN1720, danicopan, vemircopan, ACH-5228, ACH-5548, BCX-9330, AMY-101, ANX005, ANX007, narsoplimab, iptacopan, CLG561, GT103, ARGX117, ALXN1820, NGM621, lampalizumab, NGM621, IONIS-FB-Lrx, GEM103, CLG561, pharmaceutically acceptable salts thereof and combinations thereof.

[0189] Integrin inhibitors that can be used with the dendrimers and methods of the present invention include lifitegrast, vedolizumab, natalizumab, efalizumab, tirofiban, eptifibatide, abciximab, IDL-2965, PLN-74809, PLN-1474, PN-943, 7HP349, MORF-057, OS2966, OTT166, AXT-107, JSM-6427, Risuteganib, THR687 (D / ced), pharmaceutically acceptable salts thereof and combinations thereof.

[0190] Antihistamines that can be used with the dendrimers and methods of the present invention include loradatine, hydroxyzine, diphenhydramine, chlorpheniramine, brompheniramine, cyproheptadine, terfenadine, clemastine, triprolidine, carbinoxamine, diphenylpyralin, phenindamine, azatadine, tripelennamine, dexchlorpheniramine, dexbrompheniramine, methyllazine and trimprazine, doxylamine, pheniramine, pyrilamine, quiorciclizine, tonzylamine, pharmaceutically acceptable salts thereof and combinations thereof. Petition 870250084859, dated 09 / 19 / 2025, page 83 / 290 75 / 208

[0191] IL-6 inhibitors that can be used with the dendrimers and methods of the present invention include sarilumab, tocilizumab, RG6179, pharmaceutically acceptable salts thereof and combinations thereof.

[0192] HtrA1 inhibitors that can be used with the dendrimers and methods of the present invention include IC-500, FHTR2163, RG6147, pharmaceutically acceptable salts thereof and combinations thereof.

[0193] RASP inhibitors that can be used with the dendrimers and methods of the present invention include reproxalap and its pharmaceutically acceptable salts.

[0194] Rho kinase inhibitors that can be used with the dendrimers and methods of the present invention include netardusil, ripasudil, HA-1077, Y-27632, H-1152P, INS-115644, Y-39983, SB772077BS, LX71D1, AR-12286, AMA-0076, AR-13533, pharmaceutically acceptable salts thereof and combinations thereof.

[0195] Plasma kallikrein inhibitors that can be used with the dendrimers and methods of the present invention include ecallantide, lanadelumab, berotralstat, ATN-249, KVD900, KVD824, THR-149, pharmaceutically acceptable salts thereof, and combinations thereof.

[0196] PgAs nitric oxide donors that can be used with the dendrimers and methods of the present invention include Latanoprostene Bunod, NCX470, NCX125, pharmaceutically acceptable salts thereof and combinations thereof.

[0197] Mast cell stabilizers that can be used with the dendrimers and methods of the present invention include lodoxamide, nedocromil, pemirolast, cromoglycate (e.g., sodium cromoglycate), pharmaceutically acceptable salts thereof, and combinations thereof. Petition 870250084859, dated 09 / 19 / 2025, p. 84 / 290 76 / 208 of the same.

[0198] IGF-1R inhibitors that can be used with the dendrimers and methods of the present invention include teprotutumab, VRDN-001, VRDN-002, VRDN-003, ganitumab, figitumumab, MEDI573, cixutumumab, dalotuzumab, robatumumab, AVE1642, BIIB022, xentuzumab, istiratumab, linsitinib, picropodophilin, BMS754807, BMS-536924, BMS-554417, GSK1838705A, GSK1904529A, NVP-AEW541, NVP-ADW742, GTx-134, AG1024, KW-2450, PL-2258, NVP-AEW541, NSM-18, AZD3463, AZD9362, B1I885578, B1893923, TT-100, XL-228, A-928605, pharmaceutically acceptable salts thereof and combinations thereof.

[0199] TRPV1 antagonists that can be used with the dendrimers and methods of the present invention include asivatrep, V116517, azabicyclic compounds, heterocyclic compounds and fused amides, as described, for example, in US Patent Application No. 2004 / 0157849, US Patent Application No. 2004 / 0209884, US Patent Application No. 2005 / 0113576, International Patent Application No. WO 05 / 016890, US Patent Application No. 2004 / 0254188, US Patent Application No. 2005 / 0043351, International Patent Application No. WO 05 / 040121, US Patent Application No. 2005 / 0085512 and Gomtsyan et al., 2005, J. Med. Chem. 48:744-752; fused pyridine derivatives, as described, for example, in US Patent Application No. 2004 / 0138454; pyridylpiperazinyl ureas, as described, for example, in Swanson et al., 2005, J. Med. Chem. 48:1857-1872 and US Patent Application No. 2005 / 0049241, as well as AMG8163 (Bannon et al., 2005, 11th World Congress on Pain) and BCTC (Sun et al., 2003, Chem. Lett.13:3611-3616); 2-(piperazine-1-yl)-1H-Benzimidazole; pyridazinylpiperazines; urea derivatives, as described, for example, in US Patent Application No. 2005 / 0107388, US Patent Application No. 2005 / 0187291 and US Patent Application No. Petition 870250084859, dated 09 / 19 / 2025, page 85 / 290 77 / 208 2005 / 0154230, as well as A-425619 (El Kouhen et al., 2005, J. Pharmacol. Exp. Ther. 314:400-409); cinnamides, including SB-366791 (Gunthorpe et al., 2004, Neuropharmacology 46:133-149) and AMG 9810 (Gawa et al., 2005, J. Pharmacol. Exp. Ther. 313:474-484).

[0200] In some embodiments, the TRPV1 antagonists useful in the methods and compositions disclosed herein include, for example, TRPV-1 antagonists including capsazepine, (E)-3-(4-t-butylphenyl)-N(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acrylamide (commercially available, for example, as AMG9810 from Tocris Bioscience, Bristol, UK) and 4-butyl tertiary cyclohexane (commercially available as SYMSITIVE 1609 from Symrise GmbH of Holzminden, Germany), as well as TRPV1 antagonists as disclosed in U.S. Patents Nos. 8,815,930, 6,933,311, 7,767,705 and in the publications of U.S. Patent Application No. 2010 / 0249203 and 2011 / 0104301, International Application WO / 2008 / 013861.

[0201] In some embodiments, the TRPV1 antagonists useful in the methods, compositions and devices disclosed herein include AMG-517 and AMG-628 (Amgen Inc., Thousand Oaks, California). TRPV1 antagonists useful in the present application are also described, for example, in International Patent Application No. WO 2006065484; International Patent Application No. WO 2003070247; US Patent Application No. US 2005080095; and International Patent Application No. WO 2005007642. Additional TRPV1 antagonists useful in the methods, compositions and devices disclosed herein include TRPV1 antagonists: ABT-102, AMG8562, AMG9810, BCTC, SB366791, JNJ17203212, I-TTX, JYL-1421, A-425619, N-[4-[6[4(Trifluoromethyl)phenyl)pyrimidin-4-yloxy]benzothiazol-2-yl]acetamide (also known as AL-49975 or AMG-517), (R)—N-(4-(6-(4-(1(4-fluorophenyl)ethyl)piperazin-1-yl)pyrimidin-4-yloxy)benzo[d]thiazol-2-yl)acetamide (AL-49976, also known as AMG-628), salts Petition 870250084859, dated 09 / 19 / 2025, page 86 / 290 78 / 208 pharmaceutically acceptable versions thereof and combinations thereof.

[0202] Other TRPV1 antagonists useful in the methods, compositions and devices disclosed herein are those that have low inhibitory activity against CYP3A4, such as, for example, 1-(2-(3,3-dimethylbutyl)-4-(trifluoromethyl)benzyl)-3-(1-methyl-1H-yn-dazol-4-yl)urea; methyl 2,2-dimethyl-4-(2-((3-(1-methyl-1H-indazol-4-yl)ureido)methyl)-5(trifluoromethyl)phenyl)butanoate; 1-(2-(4-hydroxy-3,3-dimethylbutyl)-4(trifluoromethyl)benzyl)-3-(1-methyl-1H-indazol-4-yl)urea; 2,2dimethyl-4-(2-((3-(1-methyl-1H-indazol-4-yl)ureido)methyl)-5-trifluoromethyl)phenyl)butanoic acid; 1-[4-chloro-3-(3,3-dimethylbutyl)benzyl]-3-(1-methyl1H-indazol-4-yl)urea-; 1-(2-isobutyl-4-(trifluoromethyl)benzyl)-3-(1-methyl-1H-indazol-4-yl)urea; 1-(2-isopropyl-4-(trifluoromethyl)benzyl)-3-(1-methyl-1H-indazol-4-yl)urea; 1-(4-chloro-3-isopropylbenzyl)-3-(1-methyl-1Hindazol-4-yl)urea, pharmaceutically acceptable salts thereof and combinations thereof.

[0203] TrkA antagonists that can be used with the dendrimers and methods of the present invention include VM902A, Larotrectinib, Entrectinib, Selitrectinib (LOXO-195, BAY 2731954), repotrectinib (TPX-0005), pharmaceutically acceptable salts thereof and combinations thereof.

[0204] For the purposes of the present invention, an active agent includes all its possible forms, including free acid, free base, polymorphs, pharmaceutically acceptable salts, anhydrites, hydrates, other solvates, stereoisomers, crystalline forms, cocrystals, prodrugs, conjugates (e.g., pegylated compounds), complexes and mixtures thereof.

[0205] Diagnostically active agents may be, for example, imaging agents, markers or visualization agents. Generally, diagnostic agents may be substances used Petition 870250084859, dated 09 / 19 / 2025, page 87 / 290 79 / 208 to examine the body in order to detect impairment of its normal functions. In some cases, diagnostic agents may be agents with a functional purpose, such as for use in detecting ocular deformities, diseases, and pathophysiological aspects. For example, the diagnostic agent may be an important and effective diagnostic aid, such as a dye (e.g., fluorescein dye, indocyanine green, trypan blue, a dark suppression agent such as a cyanine dye, an azo dye, an acridine, a fluorene, an oxazine, a phenanthridine, a naphthalimide, a rhodamine, a benzopyrone, a perylene, a benzanthrone, a parabenzoxanthrone), to aid in visualizing ocular tissues. The diagnostic agent may comprise paramagnetic molecules, fluorescent compounds, magnetic molecules, radionuclides, X-ray imaging agents, and / or contrast media.In some modalities, a diagnostic agent may include radiopharmaceuticals, contrast agents for use in imaging techniques, allergen extracts, activated charcoal, different test strips (e.g., cholesterol, ethanol, and glucose), pregnancy tests, urea-13C breath tests, and various markers / stains. In some modalities, the marking portion is a fluorescent dye or a dark suppressant, selected from the group consisting of a coumarin, a cyanine dye, an azo dye, an acridine, a fluorene, an oxazine, a phenanthridine, a naphthalimide, a rhodamine, a benzopyrone, a perylene, a benzanthrone, and a benzoxanthrone.In non-limiting embodiments, the fluorescent dye is or is the residue of a compound selected from the group consisting of coumarin, fluorescein, cyanine 3 (Cy3), cyanine 5 (Cy5), cyanine 7 (Cy7), Alexa dyes, bodipy derivatives, (E)-2-(4(phenyldiazenyl)phenoxy)acetic acid, 3-(3',3'-dimethyl-6-nitrospiro[chromeno-2,2-indolin]-1'-yl)propanoate (spiropyran), 3,5-dihydroxybenzoate and acid. Petition 870250084859, dated 09 / 19 / 2025, page 88 / 290 80 / 208 (E)-2-(4-(phenyldiazenyl)phenoxy)acetic acid, or combinations thereof.

[0206] In certain embodiments of the invention, the active agent may be further dispersed, incorporated or encapsulated in the voids of the hyper-branched macromolecule. In certain embodiments, the active agent may be in the form of particles. Synthesis

[0207] Various methods for manufacturing hyperbranched macromolecules are known to the skilled craftsman, and these methods can be primarily applied and suitably adapted in embodiments of the present invention.

[0208] In certain embodiments of the invention, synthetic methods have been developed to produce dendrimers, including divergent and convergent synthesis, which are the two most common general methods used by chemists. These methods can, in principle, also be employed to synthesize the hyper-branched macromolecules of certain embodiments of the invention. The divergent method involves the addition of monomers in repeated sequence and begins with a multivalent core up to surface molecules with a continuously increasing number of branches. The molecular size and the number of surface groups gradually increase with the addition of successive layers of monomers, which is called generations (as shown in FIG. 2). While the convergent method involves the synthesis of hyper-branched macromolecules from the surface to the core and leads to the formation of wedge-shaped conical units or dendrons, these are joined to a multivalent core in the last step.Furthermore, the combined divergent / convergent method can also be employed in embodiments of the invention. For example, in one embodiment of a combined convergent / divergent synthesis, as shown in FIG. 1, first-generation DCRUs are connected to a central unit, and second- and level DCRUs. Petition 870250084859, dated 09 / 19 / 2025, page 89 / 290 The higher 81 / 208 molecules are first connected to each other before being connected to the first-generation DRCUs. Any variation of combined convergent and divergent synthesis steps can be used in embodiments of the invention, as desired for the specific hyper-branched macromolecule structure sought.

[0209] In one embodiment of the invention, a method is provided for divergently synthesizing the hyper-branched macromolecule which includes the following steps: (a) Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry at the terminations of the polymeric arms; (b) Providing dendritic constitutional repeat unit precursors comprising a polymeric arm comprising a functional group suitable for forming a click chemistry connection with the corresponding functional groups of the core-connected polymeric arms (such as an azide, alkyne, alkene, or tetrazine), and at least two polymeric arms comprising non-reactive functional groups in click chemistry, (c) Establishing a click chemistry connection between the core-connected polymeric arms and the polymeric arms of the dendritic constitutional repeat unit precursors, (d) Optionally converting the functional groups of the at least two polymeric arms comprising non-reactive functional groups in click chemistry into functional groups suitable for click chemistry, and (e) Conjugating an active agent comprising a functional group to the outermost polymeric arms via reaction with Petition 870250084859, dated 09 / 19 / 2025, pp. 90 / 290 82 / 208 the functional groups of the outermost polymeric arms, thus forming a hyper-branched macromolecule-active agent conjugate.

[0210] In certain embodiments for higher generation hyperbranched macromolecule Gx, with x being an integer from 2 to 10, step (d) is mandatory and precursors of consecutive dendritic constitutional repeating units are connected to functional groups suitable for click chemistry obtained in step (d) by click chemistry to the hyper-branched macromolecule before conjugation of the active agent in step (f).

[0211] For example, step d) can be performed by converting PEG arms with NHS terminal groups SS (succinimidyl succinate), SG (succinimidyl glutarate), SAP (succinimidyl adipate) or SAZ (succinimidyl azelate) into DS (dibenzocyclooctynoamide succinate), DG (dibenzocyclooctynoamide glutarate), DAP (dibenzocyclooctynoamide adipate) or DAZ (dibenzocyclooctynoamide azelate) groups by reacting the NHS group with a DBCO-amine click chemical ligand, such as:

[0212] Similarly, a conversion of PEG-NHS terminals into a PEG arm terminated with azide groups can be done by reacting the NHS group with an azido-amine click chemical ligand, such as azido-PEG2-NH2 or similar. This azido-amine click chemical ligand is commercially available from various suppliers and has a structure as shown below: h2n1 r n3n, with n defining the number of PEG repeating units.

[0213] In the synthesis method, the precursor of the repeating unit Petition 870250084859, dated 09 / 19 / 2025, page 91 / 290 83 / 208 constitutional dendritic in step (c) can be represented by formula (iii): where C comprises a functional group suitable for click chemistry, such as an alkyne, alkene, azide, or tetrazine; D comprises functional groups that are not reactive in click chemistry, such as succinimidyl or a primary amine; La is a linking group; m is 0 or 1, meaning the linker may be absent or present; n is an integer from 3 to 2000 or 20 to 2000; o is an integer from 3 to 2000 or 20 to 2000, while neo may be different or the same; X is a branching unit; Lb is a linking group; p is 0 or 1, meaning the linker may be absent or present; B comprises a terminal group located on the surface of the hyper-branched macromolecule or comprises a linkage connected to A of a consecutive dendritic constitutional repeating unit or to an active agent; La and Lb may be different or the same; mep may be different or the same; and y is an integer from 2 to 9, where y = c' - 1 with c' being the connectivity c' of the branching unit X,and in which the DCRU precursors used to synthesize a hyper-branched macromolecule may be the same or different.

[0214] Exemplary precursors with 4 arms are 4aPEG-NHS(3)Azide(1) or PEG-NHS(3)DBCO(1) compounds with 4 arms and similar structures, with or without hydrolyzable linking groups, such as those connected by ester linkages, amide linkages or a combination of both, see for example the structures below. Petition 870250084859, dated 09 / 19 / 2025, page 92 / 290 84 / 208 or 4-arm PEG-Amine 4-arm PEG-NHS(1)DBCO(3) OR Petition 870250084859, dated 09 / 19 / 2025, page 93 / 290 85 / 208 4-armed PEG-Amine .---N3 λ_fCHzCHsOln__s 4-armed PEG-NHS(1)Azide(3) or the following exemplary precursor pairs: 4-arm PEG-DBCO and any Petition 870250084859, dated 09 / 19 / 2025, pp. 94 / 290 86 / 208 Petition 870250084859, dated 09 / 19 / 2025, pp. 95 / 290 87 / 208 or or and any Petition 870250084859, dated 09 / 19 / 2025, pp. 96 / 290 88 / 208 4-arms-PEG-NHS(1)Azide(3) CH2CH2O· where t is m, and enem are defined as for formula (v) here before.

[0215] Referring to FIG. 3, exemplary synthesis schemes are shown for a 4-unit 4-arm PEG peptide-conjugated hyper-branched macromolecule (FIG. 3 a)) and an 8-4-arm PEG hyper-branched macromolecule according to certain embodiments. In a generic embodiment, a multi-arm PEG with terminal functional groups for click chemistry, such as DBCO or azide, can be used as the core of a hyper-branched macromolecule. Then, another branched PEG with functional groups will react with the central PEG via click chemistry.The branched PEG will contain two types of functional groups; one group, such as azide or DBCO, can couple to the core PEG for hyper-branched macromolecule growth in a click chemical reaction, while the remainder of the branched PEG (i.e., the DCRU) will be inert to the core PEG and can be used for the next generation of hyper-branched macromolecule growth or used as a precursor for terminal bioconjugation (FIG. 3a). Based on this generic method, multiple generations of hyper-branched macromolecules can be synthesized to achieve different numbers of terminal functional groups. FIG. 3b) shows a 3D structure of a hyper-branched macromolecule starting with an 8-armed PEG core and coupled with eight 4-armed PEG branches to achieve 24 terminal groups on the surface and finally conjugated with up to 24. Petition 870250084859, dated 09 / 19 / 2025, page 97 / 290 89 / 208 peptides.

[0216] As shown in the Examples, two cyclic peptides as C3 binding inhibitors, compstatin and APL-1, and an immunoglobulin G (IgG) binding peptide ligand, Fc-III-4C, can be used exemplarily as APIs that are conjugated with a hyper-branched PEG macromolecule. The primary amine groups in the peptides can be used as nucleophiles to react with electrophilic NHS groups in the outermost polymeric arms of the hyper-branched macromolecule.

[0217] Alternatively, by converting the functional groups of the outermost polymeric arms comprising non-reactive functional groups in click chemistry into functional groups suitable for click chemistry and functionalizing the active agent, such as a peptide with a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine), conjugation of the active agent can also be done in a click chemistry reaction on the outermost polymeric arms of the hyper-branched macromolecule.

[0218] Suitable ester groups in the outermost polymeric PEG arms of the hyper-branched macromolecule of certain embodiments, such as succinic (S-), glutaric (G-), adipic (AP-), and azelaic (AZ-), are hydrolyzable under physiological conditions and are degraded under controlled pH conditions to release the peptides in vivo. The controlled release and binding affinity of the peptide moieties can be characterized by ultra-high performance liquid chromatography (UHPLC), sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and surface plasmon resonance (SPR), etc., as described herein.

[0219] In an alternative embodiment, a convergent synthesis for the hyper-branched macromolecules of the present invention is provided, comprising the following steps: Petition 870250084859, dated 09 / 19 / 2025, pp. 98 / 290 90 / 208 I) Supply of precursors for dendritic constitutional repeat units comprising - a polymeric arm comprising a functional group suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine), and - at least two polymeric arms comprising non-reactive functional groups in click chemistry, II) Conjugate active agents comprising a functional group to at least one of the at least two polymeric arms comprising non-reactive functional groups in the click chemistry of the dendritic constitutional repeat unit precursors, III) Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry (such as an azide, alkyne, alkene, or tetrazine) at the ends of the polymeric arms, and IV) Forming a click chemistry connection between the polymeric arms connected to the core provided in step III) and the polymeric arm comprising a functional group suitable for forming a click chemistry connection of the dendritic constitutional repeat unit precursors conjugated to the active agent obtained in step II), thus forming a biodegradable hyper-branched macromolecule-active agent conjugate.

[0220] In this method, the precursor of the dendritic constitutional repeating unit in step I) is represented by Formula (iii) as described above.

[0221] In certain embodiments for biodegradable hyperbranched macromolecules G x of higher generation, with x being an integer from 2 to 10, the precursors of constitutional dendritic repeat units conjugated to the active agent obtained in step II) are Petition 870250084859, dated 09 / 19 / 2025, page 99 / 290 91 / 208 connected by click chemistry to reverse constitutional dendritic repeat unit precursors comprising a polymeric arm comprising a non-reactive functional group in click chemistry and at least two polymeric arms comprising functional groups suitable for click chemistry (such as an azide, alkyne, alkene or tetrazine), wherein the non-reactive functional group in click chemistry of one polymeric arm is subsequently converted into a functional group suitable for click chemistry before connecting to other reverse constitutional dendritic repeat unit precursors or before forming a click chemistry connection with the polymeric arms connected to the core in step IV), thus forming higher generation biodegradable hyper-branched macromolecules.

[0222] In contrast to the divergent synthesis method, the convergent method also allows the synthesis of hyperbranched macromolecules with two or more different active agents in different dendrons or regions on the surface of the hyperbranched macromolecule. This allows the grouping of more than one active agent on the surface of the hyperbranched macromolecule. In certain embodiments of the convergent method of the invention, precursors of dendritic constitutional repeating units with different active agents conjugated to the polymeric arms can be obtained by performing steps I) and II) for each DCRU precursor conjugated to the active agent, and a mixture of the DCRU precursors conjugated to the active agent obtained is used for step IV), thus forming a biodegradable hyperbranched macromolecule-active agent conjugate with different active agents in different regions of the surface of the hyperbranched macromolecule.These clustered hyperbranched macromolecules can, for example, be used for combination therapies involving the administration of more than one active agent. Petition 870250084859, dated 09 / 19 / 2025, pp. 100 / 290 92 / 208

[0223] In other embodiments of the divergent and convergent synthesis methods discussed above, the methods can also be carried out with inversely exchanged functional groups, i.e., using other reactions and functional groups to form connections within the hyper-branched macromolecule and click chemistry functional groups for terminal conjugation. In such “inverse” embodiments of the described synthesis methods, the connections in the hyper-branched macromolecule can be selectively formed using electrophilic nucleophilic precursors or other non-reactive functional groups with click chemistry functional groups, while all other terminal functional groups of the DCRU that do not participate in the connection with the core or previous DCRU include a functional group for click chemistry bond formation and remain non-reactive in the connection formation reaction.These terminal click chemistry functional groups can be used subsequently for other follow-up reactions, such as the growth of hyper-branched conjugation macromolecules with click chemistry reactions.

[0224] Thus, in another embodiment of the invention, a method is provided for divergently synthesizing the hyperbranched macromolecule which includes the following steps: (a) Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having non-reactive functional groups in click chemistry at the terminations of the polymeric arms; (b) Supply of dendritic constitutional repeat unit precursors comprising a polymeric arm comprising a functional group suitable for forming a connection in a reaction other than click chemistry with the corresponding functional groups of the polymeric arms connected to the core (as an electrophile or Petition 870250084859, dated 09 / 19 / 2025, pp. 101 / 290 93 / 208 nucleophile, for example, amine, NHS), and at least two polymeric arms comprising functional groups suitable for click chemistry, (c) Forming a connection between the polymeric arms connected to the nucleus and the polymeric arms of the dendritic constitutional repeating unit precursors, reacting the non-reactive functional groups in click chemistry, (d) Optionally converting the functional groups of the at least two polymeric arms comprising functional groups suitable for click chemistry into non-reactive functional groups in click chemistry, and (e) Conjugation of an active agent comprising a functional group to the outermost polymeric arms by reaction with the functional groups of the outermost polymeric arms, thereby forming a hyper-branched macromolecule-active agent conjugate.

[0225] In certain embodiments for higher generation hyperbranched macromolecule Gx, with x being an integer from 2 to 10, step (d) is mandatory and consecutive precursors of dendritic constitutional repeating units are connected to non-reactive functional groups in click chemistry obtained in step (d) to the hyperbranched macromolecule before conjugation of the active agent in step (f).

[0226] In the synthesis method, the precursor of the dendritic constitutional repeating unit in step (c) can be represented by formula (iii) as described above.

[0227] In a further alternative embodiment of the convergent method, a convergent synthesis for the hyper-branched macromolecules of the present invention is provided, comprising the following steps: I. Supply of repeat unit precursors Petition 870250084859, dated 09 / 19 / 2025, page 102 / 290 94 / 208 constitutional dendritic comprising - a polymeric arm comprising a non-reactive functional group in click chemistry, and - at least two polymeric arms comprising functional groups suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine), II. Conjugate active agents comprising a functional group to at least one of the at least two polymeric arms comprising functional groups suitable for click chemistry of the dendritic constitutional repeat unit precursors, III. Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having non-reactive functional groups in the click chemistry at the terminations of the polymeric arms, and IV. Forming a connection between the polymeric arms connected to the core provided in step III) and the polymeric arm comprising a non-reactive functional group in the click chemistry of the dendritic constitutional repeat unit precursors conjugated to the active agent obtained in step II), thus forming a biodegradable hyper-branched macromolecule-active agent conjugate.

[0228] Exemplary reaction conditions for the formation of dendrimers involve the reaction of core precursors and DCRUs at relatively mild temperatures, such as 10 to 50 °C, or 30 to 45 °C in suitable solvents, such as DMF, for several hours, such as overnight, or even 24 or even 48 hours. Purification and Characterization

[0229] The purification of the reaction mixtures of hyper-branched macromolecules obtained in the synthesis methods described here can be done, for example, by filtration, dialysis, SEC column filtration, Petition 870250084859, dated 09 / 19 / 2025, page 103 / 290 95 / 208 centrifugation or UHPLC.

[0230] In an exemplary embodiment, the synthesis reaction mixtures of hyper-branched molecules, optionally conjugated with active agents such as peptides, are diluted, filtered, for example, through 0.45 µm meshes, then purified by centrifugal ultrafiltration, for example, using a 100 kDa membrane, and can then be lyophilized after the addition of sugar buffers to produce the final product. For administration in therapeutic methods, the lyophilized product can be reconstituted by the addition of solvent, optionally including more sugar buffer.

[0231] Sugar buffers may be added as needed to improve the solubility and stability of the dendrimer peptide or protein conjugates, for example, by preventing peptide precipitation before or after lyophilization. Even with non-peptidic dendrimer conjugates, the addition of sugar buffers improves stability and solubility, as the PEG-based dendrimers of embodiments of the invention exhibit behavior similar to that of synthetic proteins. An exemplary sugar buffer formulation for use with embodiments of the invention may include a sugar solution, such as trehalose, mono- and diphosphates in water at a suitable concentration, such as 3% (or 30 mg / mL) and a pH of about 6.4.

[0232] Dialysis is a common purification method based on the separation of molecules in solution by the difference in their diffusion rates through a semipermeable membrane, such as a dialysis tube. To purify reaction mixtures of hyperbranched macromolecules, which may be a solution containing molecules of different sizes, such as free peptide (MW, for example, of about 1.5 kDa), free PEG / DCRU precursors (MW, for example, of about 10-40 kDa), small conjugates of hyperbranched macromolecules. Petition 870250084859, 19 / 09 / 2025, p. 104 / 290 96 / 208 branched molecules, such as G0 (MW, for example, of about 20-50 kDa) and large conjugates, such as conjugates of higher-generation hyperbranched G x macromolecules (e.g., of about 50 kDa and above), the solution can be loaded into a dialysis tube with a specific pore size membrane defining the cutoff and soaked in a large amount of solvent. Molecules smaller than the pore size will be eluted from the tube into the solvent, while molecules larger than the pore size will remain inside the tube. Dialysis tubes are commercially available, for example, from Spectra / Por® Float-A-Lyzer G2 Dialysis Devices, Spectrum® Laboratories, with various different molecular weight cutoffs as desired for the specific separation task. FIG. 5a) shows a corresponding experimental setup for dialysis purification.

[0233] By selecting suitable molecular weight dialysis tubes and membranes, if necessary, in a series of separation steps with different dialysis tubes, it is possible to remove most impurities, such as excess peptides and unreacted precursors, small-sized intermediates, from the product.

[0234] Another purification method that can be applied to certain aspects of the present invention is the use of size exclusion chromatography (SEC), such as an SEC column. For example, Zeba™ Spin Desalting columns (from ThermoFisher Scientific) designed for protein purification to remove salts and small impurities can be used for the purification of conjugates of hyper-branched macromolecules. Columns with different pore sizes can be used, for example, 7kDa and 40kDa. The purification mechanism is based on size exclusion chromatography, in which small particles will be trapped in the pore of the stationary phase material, and large particles, such as conjugates of hyper-branched macromolecules of certain Petition 870250084859, dated 09 / 19 / 2025, page 105 / 290 97 / 208 embodiments of the invention will be eluted by the column and collected in purified form. FIG. 5b) shows a corresponding experimental setup for purification by column filtration SEC.

[0235] The resulting purified products can be characterized by ultra-high performance liquid chromatography (UHPLC), which is an efficient technique that offers more sensitive analyses with good chromatographic separation and resolution of the analytes. It offers benefits including rapid analysis, high-resolution separations, reduced solvent and sample usage, improved sensitivity and precision, etc. Based on calibration curves with free active agent, free precursor units, and a comparison of the solutions before and after the purification treatment, the desired amounts of product in the purified solution can be determined by peak area integration.

[0236] With dialysis purification, product solutions containing more than 99% of hyperbranched macromolecule-peptide conjugate can be obtained (see Example 6), as determined by UHPLC based on peak area integration. With SEC column purification, product solutions containing more than 98% of hyperbranched macromolecule-peptide conjugate can be obtained (see Example 7), as determined by UHPLC based on peak area integration. Both purification methods described show very efficient purification capabilities.

[0237] To determine the molecular weight of conjugates of hyper-branched macromolecules of certain embodiments of the present invention, SDS-PAGE can be used. SDS-PAGE is an analytical technique for separating materials based on their molecular weight. When samples are separated by electrophoresis under an electrical potential through a gel matrix, smaller compounds migrate faster due to the lower resistance of the gel matrix, while molecules Petition 870250084859, dated 09 / 19 / 2025, page 106 / 290 Larger molecules (98 / 208) migrate more slowly. Sodium dodecyl sulfate (SDS) is a surfactant that can exfoliate large molecules, such as proteins, and eliminates the influence of their structure and charge to separate compounds based solely on their molecular size.

[0238] In embodiments of the invention, the hyperbranched molecules are lyophilized to provide a storage-stable formulation that can be reconstituted with suitable solvents before therapeutic use. Multivalent Receiver Connection

[0239] For biological efficacy, it is desirable that the biomolecules conjugated to the hyper-branched macromolecules of the invention exhibit the same or similar affinity to a receptor. Furthermore, efficacy can be improved if the half-life of the receptor-binding biomolecules is extended by multivalent binding. For example, in antigen binding, affinity is defined as the strength required for an interaction between an antigen-binding site on an antibody and an antigen epitope. Avidity is the total strength required for the interaction between a multivalent antibody and multiple antigenic epitopes. This definition can also be applied to other biomolecules that bind to specific targets or receptor sites. Multivalent binding therefore results in improved avidity. The concept of multivalence and the resulting concept of avidity, and a model for quantifying avidity, were described by Kitov et al., “On the Nature of the Multivalency Effect: A Thermodynamic Model”, JACS 2003, 125, 16271-16284, which is incorporated herein by reference in its entirety.

[0240] Kitov describes the interaction of Shiga-like toxins with a series of multivalent oligosaccharide ligands conjugated to dendrimers based on PANAM dendrimer structures with variable multivalence. Among other things, Kitov discovered that, even Petition 870250084859, dated 09 / 19 / 2025, page 107 / 290 99 / 208 that extra branches of the multivalent ligand dendrimers do not interact with the receptor in a common sense, they increase the probability of interaction with the receptors. Furthermore, Kitov concludes that “in a situation where it is necessary to inhibit all binding sites to achieve a desired effect, the portion of uninhibited binding sites can be precisely controlled by choosing the appropriate number of branches for the assembly of a multivalent inhibitor”. Thus, for multivalent inhibitor systems as an example, extra branches with more conjugated inhibitor molecules can ensure a greater degree of inhibition, even though the individual inhibitors are unable to interact specifically with the receptor, resulting in extended half-life, improved efficacy and avidity, even for possibilities of multivalent binding.

[0241] Khalili et al., “Fab-PEG-Fab as a Potential Antibody Mimetic”, Bioconjugate Chem. 2013, 24, 1870-1882, which is incorporated herein by reference in its entirety, exemplifies a bivalent PEG conjugated to protein-binding ligands, exemplifying an improved avidity of the bivalence.

[0242] Certain embodiments of the invention make use of the concepts described, as shown below with reference to Examples 8, 9 and Figs. 18 and 19.

[0243] The embodiments of the invention relate to methods of treating a disease with antibodies bound to dendrimers, as described herein. For example, the dendrimers of embodiments of the invention can be used to improve the pharmacokinetics of antibodies administered as a delivery target bound or conjugated to hyperbranched molecules, as described herein. Suitable delivery targets are, for example, selected from anti-VEGF, aflibercept, faricimab, bevacizumab, anti-TNF-α, infliximab, etanercept, adalimumab, anti-IL-6R, sarilumab, anti-IL Petition 870250084859, dated 09 / 19 / 2025, page 108 / 290 100 / 208 6, siltuximab, anti-C5, ravuilizumab, eculizumab, anti-CD20, ocrelizumab, rituximab, anti-IGF-1R, or teprotumumab. These antibodies, bound and released by dendrimers, bind non-covalently to the antibody drugs and chaperone them, prolonging the half-life in the blood after intravenous administration or after injection into the vitreous humor (IVT). The high molecular weight of the dendrimer-antibody conjugate prevents the elimination of the bound antibody from the blood through the kidneys and delays diffusion to the therapeutic target site, such as the vitreous humor.

[0244] Antibodies remain functional while bound to dendrimers of embodiments of the invention. The gradual release of the dendrimer allows unrestricted delivery of antibodies to the target tissue. The dendrimer can be engineered to degrade into lower molecular weights, such as fragments with less than 50,000 kDa, as described herein, for eventual clearance via the kidney.

[0245] The nanoscale size of the dendrimer-antibody conjugates of the invention further enables passive targeting of leaky vessels, for example, tumors or choroidal neovascularization (CNV) through the enhanced permeability and retention (RPE) effect. RPE can enable subcutaneous (SC) or intravenous (IV) administration routes, reducing undesirable side effects. This could allow SC or IV administration to CNV areas in the eye.

[0246] Exemplary diseases that can be treated with antibody-dendrimer conjugates in certain modalities include wet AMD, cancer (e.g., with anti-VEGF dendrimer conjugates, IVT or SC); RA, PsA, COPD (e.g., with anti-TNF-α dendrimer conjugates, IV or SC administration); PNH, aHUS, MG, glomerular disease, GA (e.g., with dendrimer conjugates containing anti-C5, ravuilizumab or eculizumab, IV, IVT or SC administration). Petition 870250084859, dated 09 / 19 / 2025, page 109 / 290 101 / 208 SC); RA (e.g., with dendrimer conjugates containing rifuximab, IV administration); or TED (e.g., with dendrimer conjugates containing anti-IGF-1R, IV or SC administration).

[0247] Other embodiments of the invention relate to methods of treating a disease with peptides bound to dendrimers, as described herein. For example, the dendrimers of embodiments of the invention can be used to improve the pharmacokinetics of peptides distributed as an administration target bound or conjugated to hyper-branched molecules, as described herein. Suitable delivery targets are, for example, selected from anti-C3, C3B, Syfovre, GLP-1RA, liraglutide, Victosa, Saxenda, Semaglutide, Ozempic, Rybelsus, Wegovy, Exenatide, hormone therapy, HGH (somatotrypneumonia), insulin, estrogen, etc.

[0248] Compared to larger proteins, peptides have the advantage of lower immunogenicity and better stability. However, peptides have the disadvantage of rapid elimination and may have low solubility, which can limit their usefulness in therapy. In embodiments of the invention, conjugation to dendrimers can be a successful strategy for administering peptides in therapeutic treatments, for example, Syfovre dendrimer conjugates, to increase solubility and prolong half-life. Syfovre also benefits from divalent conjugation for improved binding avidity. Conjugation with a dendrimer, as in embodiments of the invention, can go beyond simple conjugation with PEG to provide higher molecular weight, longer half-life, and higher valence – greater avidity. The gradual biodegradability of the dendrimer into smaller fragments allows for the elimination of these high molecular weight molecules and prevents accumulation in the body.

[0249] Exemplary diseases that can be treated with dendrimer peptide conjugates of certain modalities include Petition 870250084859, dated 09 / 19 / 2025, page 110 / 290 102 / 208 GA, PNH (e.g., with anti-C3, C3B, IVT, IV, or SC dendrimer conjugates); TD2, obesity (e.g., with GLP-1RA dendrimer conjugates); hormone deficiency syndromes (e.g., with hormone dendrimer conjugates, inhalation, IV, or SC).

[0250] Other embodiments of the invention relate to methods of treating a disease with aptamers linked to dendrimers, as described herein. For example, the dendrimers of embodiments of the invention can be used to improve the pharmacokinetics of aptamers distributed as a delivery target linked or conjugated to hyper-branched molecules, as described herein. Suitable delivery targets are, for example, selected from anti-C5, Izervay, anti-VEGF165, Macugen, Anti-CXCL12 / SDF-1 or NOXA12.

[0251] Aptamers are similar to peptides in terms of low immunogenicity. However, in vivo stability has been a problem, which can be solved by conjugation to dendrimers, as described here. Aptamers also exhibit good water solubility. Conjugation with PEG has been a successful strategy for aptamers, for example, Macugen and Izervay, to prolong the half-life. Conjugation with a dendrimer, as described here, can go beyond simple PEG conjugation to provide higher molecular weight, longer half-life, and higher valence – greater avidity. The gradual biodegradability of the dendrimer into smaller fragments allows for the elimination of these high molecular weight molecules and prevents accumulation in the body.

[0252] Exemplary diseases that can be treated with dendrimer-aptamer conjugates of certain modalities include wet AMD (e.g., with anti-VEGF165 dendrimer conjugates); PNH, aHUS, MG, glomerular disease, GA (e.g., with conjugates Petition 870250084859, dated 09 / 19 / 2025, page 111 / 290 103 / 208 anti-C5 or Izervay dendrimer); CLL, pancreatic cancer (e.g., with anti-CXCL12 / SDF-1 dendrimer conjugates).

[0253] To determine the biological effectiveness of hyper-branched macromolecule conjugates of certain embodiments of the invention, binding assay tests can be performed to analyze the binding affinity of peptides conjugated to hyper-branched macromolecules.

[0254] Complement activation is essential for the development of normal inflammatory responses against foreign pathogens; however, its inappropriate activation has been a cause of tissue injury in many disease states. The C3 component of complement is a common denominator in the activation of the classical, alternative, and lectin pathways of complement activation. Uncontrolled complement activation can lead to a wide range of debilitating or life-threatening disorders.

[0255] Compstatin, a 13-mer peptide (le-Cys-Val-Val-Gln-AspTrp-Gly-His-His-Arg-Cys-Thr-NH2) cyclized via a disulfide bridge, is a novel and promising inhibitor of complement system activation and was initially isolated from a library of random peptides displayed in phages and screened against C3b.

[0256] APL-1 (le-Cys-Val-MeTrp-Gln-Asp-Trp-Gly-Ala-His-Arg-CysThr-NH2) has a structure similar to that of compstatin, with 2 different amino acids in the sequence. As reported in the literature, the dissociation constant Kd of APL-1 to C3 is 10 nM, while the Kd of compstatin is 13 μM, which is a difference of about hundreds of times in the binding affinity of C3. The structure of the compstatin and APL-1 peptide sequences is shown below:

[0257] Scheme C: Petition 870250084859, dated 09 / 19 / 2025, page 112 / 290 104 / 208 APL-1 Ac-lle-Cys-Val-MeTrp-G|n-Asp-Trp-G|yA|a-His-Arg-Cys-Thr-NH2I I Compstatin Ac-lle-Cys-Val-Val-Gln-Asp-Trp-Gly-His-His-Arg-Cys-Thr-NH2

[0258] Fc-III-4C is an immunoglobulin G (IgG)-binding peptide ligand composed of 15 residues, in which the 4 cysteine ​​residues form 2 disulfide bonds to generate a double cyclic structure. The proposed structure of the Fc-III-4C double cyclic peptide is shown below:

[0260] The binding affinity of the Fc-lll-4C peptide to human IgG was determined to be 2.45 nM (Kd), which is higher than that of IgG with Protein A / G (Pro-A / G). It is important to note that the Fell I-4C peptide showed high affinity for several IgGs from different species, therefore it was also reported as being used as a peptide-based antibody affinity marker.

[0261] The three peptides mentioned above can be used to analyze the binding affinity of hyper-branched macromolecule conjugates of these peptides using a surface plasmon resonance (SPR) configuration from Mosaic Biosciences, Inc., USA. The surface plasmon resonance (SPR) binding analysis methodology can be used to study molecular interactions. SPR is an optical technique for detecting the interaction of Petition 870250084859, dated 09 / 19 / 2025, page 113 / 290 105 / 208 two different molecules, in which one is mobile and the other is fixed to a thin film. In such an analysis, the C3 target will be fixed to the surface of a thin film or chip, and a hyper-branched macromolecule-peptide conjugate solution will be passed along it. A difference in signal is monitored when the hyper-branched macromolecule-peptide conjugate associates / dissociates at the C3 target.

[0262] These assays demonstrate that the binding of C3 and C3b of free compstatin, APL-1 and Fc-III 4C corresponds well to the values ​​reported in the literature for the dissociation constant KD, therefore the chosen assay is a reliable tool.

[0263] The same assay can be applied to test the binding affinity of conjugated peptides of hyperbranched macromolecules, in which C3 is immobilized on the assay chip and hyperbranched macromolecule-peptide conjugated solutions are flown over at a variety of concentrations. If multivalent binding of several conjugated peptides to a hyperbranched macromolecule occurs, the hyperbranched macromolecule should dissociate very slowly, while the free peptide is expected to dissociate rapidly. This measurement allows comparison of dissociation rates of hyperbranched macromolecules from different generations.

[0264] This theoretical prediction was confirmed by SPR measurement. A comparison of the binding affinity of hyper-branched macromolecule conjugates of compstatin, APL-1, and Fc-III 4C from certain embodiments of the invention in the same assay shows a rapid association rate. During the dissociation time, the response of free compstatin drops rapidly, the rate being the same as the association rate, as might be expected. On the other hand, hyper-branched macromolecule-peptide conjugates exhibit a much faster dissociation rate. Petition 870250084859, dated 09 / 19 / 2025, page 114 / 290 106 / 208 slower. Without wanting to get stuck on one theory, it is believed that this is caused by the multiple interaction of several conjugated peptides in the hyper-branched macromolecule, for example, multivalent linkage. The phase associated with the slower dissociation indicates a cooperative binding of the multivalent conjugated hyper-branched macromolecule peptides to the C3 surface.

[0265] Through similar experiments, it can be demonstrated that the peptide, after being hydrolyzed from the hyperbranched macromolecule during biodegradation, has approximately the same binding affinity as the free peptide. The hydrolyzed peptide contains an acidic ester linkage from degradation (a part of the ligand in the hyperbranched macromolecule), and it can be shown that this has no effect on its C3 linkage according to the similar SPR signal. It is therefore believed that the ester linkage does not alter the bioactivity of the peptide.

[0266] Furthermore, it can be observed in a quantitative KD analysis that the KD of conjugates of hyper-branched macromolecules shows a decrease when there is more substituted peptide in the hyper-branched macromolecule.

[0267] With reference to FIGS. 18 and 19, an IC50 (half the maximum inhibitory concentration) was measured by an alternative pathway hemolysis assay (AP) with four different hyperbranched macromolecules conjugated with compstatin of the invention, as described in Example 10. It can be observed that the hyperbranched macromolecule G1 PEG compstatin showed improved IC50s compared to free compstatin, suggesting an improvement in potency through avidity. Without wanting to get stuck on one theory, it is believed that multiple binding or multivalent binding events possible with higher-generation hyperbranched macromolecular peptide conjugates are contributing to receptor inhibition and Petition 870250084859, dated 09 / 19 / 2025, page 115 / 290 107 / 208 increasing efficiency. Furthermore, conjugates of hyper-branched macromolecules with longer polymeric arms conjugated to the peptide appear to have improved IC50s, likely due to greater flexibility in interacting with receptors than shorter polymeric arms which may have steric repulsion problems. These results were confirmed with a classical hemolysis assay (CP), showing that higher valence compstatin conjugates of certain embodiments of the invention are more effective in inhibiting CP hemolysis compared to free compstatin alone. Additionally, as shown in Example 10, dendrimer-bound agents, such as the APL-1 used in this example, can maintain their stability and activity in vivo for extended periods of time. Release Kinetics

[0268] In certain embodiments, the hyperbranched macromolecules of the invention can be used for sustained-release drug delivery. In general, conjugation of a therapeutically active agent to a hyperbranched macromolecule can extend the in vivo half-life of the agent. The structure of the hyperbranched macromolecule can be adapted to modify the release of an active agent conjugated into the hyperbranched macromolecule by various measures, to provide a drug delivery system based on a hyperbranched macromolecule. For example, the appropriate adaptation or selection of the precursor components and DCRUs that form the hyperbranched macromolecule, such as the length and molecular weight of the polymeric arms, the type of ligands used, and the connections formed between the parts of the hyperbranched macromolecule used for conjugation, etc., influence the release of the active agent.

[0269] In addition, the release of active agents with multiple Petition 870250084859, dated 09 / 19 / 2025, page 116 / 290 The 108 / 208 binding sites on the dendrimer can be delayed by multiple binding of the active agent to the terminal functional groups of the dendrimer, either intramolecularly and / or intermolecularly, i.e., connecting two or more dendrimers through an active agent with multiple bindings. For example, multiple binding to dendrimers can be used to increase the half-life of active agents, since the release of the active agent from the dendrimer requires that more than one conjugation bond be cleaved for complete release of the active agent.

[0270] Dendrimers for drug delivery can be viewed as large carrier or transport vehicles. Due to their highly symmetrical and regular spheroidal structure, they can be used to extend the hydration or hydrodynamic radius Rh of the active agents bound to them. A large hydrodynamic radius of specifically PEG-based dendrimer structures can be used to further extend the in vivo half-life, such as in the vitreous body, of dendrimer drug conjugates, which can be used to control and adjust the sustained release of active agents. As can be estimated by the Stokes-Einstein equation, kBT 6πηKHa, the diffusion rate D of a spherical particle is approximately inversely proportional to the hydrodynamic radius Rh of the particle, given that the temperature T and viscosity η are relatively constant in physiological fluids in vivo.Thus, the larger the radius of the dendrimer-drug conjugate, the slower the diffusion rate and the longer the half-life T1 / 2 of the active agent in vivo.

[0271] Consequently, embodiments of the invention make use of the large size of dendrimers to delay the release of active agents in vivo, appropriately adjusting the overall size of the dendrimer-drug conjugate. As the hydrodynamic radius Rh can be Petition 870250084859, dated 09 / 19 / 2025, page 117 / 290 109 / 208 easily determined, for example, by size exclusion chromatography (SEC), it is possible to predictably adjust the release rate or half-life of an active agent bound to the dendrimer from calibration information from SEC measurements. Example 11 below shows illustratively how to correlate dendrimer size with release rate.

[0272] Thus, biodegradable synthetic dendrimers of embodiments of the invention offer the advantage of incorporated controlled degradable functional groups which, after degradation, produce smaller fragments with a gradually smaller hydrodynamic radius Rh and different half-lives that determine their mobility and / or elimination from the body. As an example, a generation 1 (G1) dendrimer constructed from a 4-arm 40kDa PEG core and 4-arm 20 kDa dendrons conjugated with 12 peptides or proteins of 1.7 kDa each (e.g., 4a40k-PEG(SGA)-[4a20k-PEG(SG)-(FcIII-4C)3]4) has a molecular weight of about 145 kDa. Cleavage of one, two, three, or all four dendrons will gradually reduce the molecular weight to produce fragments of approximately 115 kDa, 90 kDa, 65 kDa, and finally, will leave the core of 40 kDa and 4 dendrons of approximately 25 kDa each, with each fragment having a different hydrodynamic radius and diffusion rate.If linear PEG extenders of 20 kDa are constructed between the arms of the central and branched unit in the same macromolecule, the molecular weight cascade includes a dendrimer of approximately 22 kDa and degradation fragments of approximately 175 kDa, 130 kDa, 85 kDa, 45 kDa, 40 kDa, 25 kDa, and 20 kDa, with the dendrimer having a different and prolonged release rate, as well as a wider distribution of fragment half-lives.

[0273] The multiple half-life aspect of degradable dendrimers in embodiments of the invention is based on the initial Rh of the dendrimer itself (G1, G2, etc.), followed by another half-life based Petition 870250084859, dated 09 / 19 / 2025, page 118 / 290 110 / 208 in degradable fragments (dendrons or dendron-like structures or dendron-like structures with linear PEG extensions) formed by the cleavage of hydrolyzable bonds within the hyper-branched macromolecule structure. These multiple species, which degrade successively, all with different Rh values ​​as they degrade and separate from the initial dendrimeric structure, exhibit a variety of substructures with different hydrodynamic radii, producing different clearance rates and half-lives. The Rh of these different species is correlated to the different building blocks of different molecular weights, number of arms, linear PEG extensions, and / or ligands.In constructing hyper-branched dendrimer macromolecules from building blocks of different molecular weights and numbers of arms, using linear PEG extenders and difunctional linkers of different lengths between hydrolyzable bonds, the dendrimers of embodiments of the invention can be designed for each individual active agent and / or therapeutic purpose / or form of administration to degrade and clear fragments at multiple rates and half-lives.

[0274] Specifically, incorporated degradable linkage groups (such as diacid-derived esters of succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ) based ligands, such as succinate diesters (S), glutarate diesters (G), adipate diesters (AP), or azelate diesters (AZ), etc.) can be used to tailor and control the release rate of the active agent conjugated or associated with the dendrimer. For example, in one embodiment, the dendron building blocks can be made with different degradable linkages (such as SS, SG, SAP, SAZ, etc.), producing a homogeneous dendrimer with all dendrons degrading at the same rate if the ester linkage is the same. Petition 870250084859, dated 09 / 19 / 2025, p. 119 / 290 111 / 208

[0275] In another embodiment, the dendron building blocks can be made with different degradable linkages (such as S, G, AP, AZ, etc.), producing a heterogeneous dendrimer with dendrons degrading at different rates if the ester linkages are different. For example, in one embodiment, a dendrimer can be made with S, G, AP, or AZ dendrons or a mixture of such dendrons, clicked into the central structure via click chemistry links.

[0276] In other embodiments, a mixture of several homogeneous dendrimers can be combined to adapt a specific release profile and adjust the half-life clearance rate by dry or wet mixing.

[0277] Embodiments of the invention based on PEG, due to their large hydrodynamic radius with low solids content, can also be described as nanodroplets. With weight or solids content similar to that of the agent or peptide itself, dendrimers have a much larger hydrodynamic radius (see Figure 20 and Example 11) and therefore a slower diffusion rate and a longer half-life T1 / 2 of the conjugated active agent in vivo.

[0278] In certain embodiments, the ligand of Formula (ii) used in the structure of the hyper-branched macromolecule introduces hydrolyzable linkages into the hyper-branched macromolecule that can be used to modify the degradation rate of the hyper-branched macromolecule and / or the release rate of conjugated active agents from the hyper-branched macromolecule. For example, the biodegradation / hydrolysis rate of ester linkages in these ligands increases from succinate (C4) to azelate (C9). The shorter the length of the diacid linkage chain, the more rapidly the ester linkages it forms are hydrolyzed. Thus, the hydrolysis rate decreases from SS>SG>SAP>SAZ>SGA ester linkages. In embodiments of the invention, this can be used to control the degradation rate of Petition 870250084859, dated 09 / 19 / 2025, pp. 120 / 290 112 / 208 hyper-branched macromolecule and / or the release of active agents conjugated through these ligands to the hyper-branched macromolecule. For example, esters formed from succinimidyl succinate (SS) groups can degrade on the order of a few days, while esters from succinimidyl glutarate (SG) groups degrade on the order of weeks. Different ligands can be used within the hyper-branched macromolecule to control the degradation rates between the junctions of different generations of DCRUs in the hyper-branched macromolecule and to conjugate the active agents to control the release of the active agent from the hyper-branched macromolecule.

[0279] For connections formed by click chemical reactions, an extension of the spacer structure, for example, an alkylene chain or a pegylation, between the DBCO / Azide functional group and the functional group with which it binds to a polymeric arm can be used in disclosure embodiments to retard the hydrolysis of neighboring ester linkages as well. The greater the distance between the DBCO / Azide functional group and the next hydrolyzable ester group, the slower the ester hydrolysis occurs. Different ligands within the hyper-branched macromolecule and at the conjugation site of the active agent can be used to allow control of degradation. By using short-chain linking groups, such as succinates, for DCRU connections and long-chain linkers, such as SAZ, at the conjugation sites, it is possible, during degradation, to first break the ester groups within the hyper-branched macromolecule and only then break the DCRU-active agent conjugation.Conversely, the first cleavage of conjugation sites for release of the active agent with short-chain ligands can be purposefully engineered. This can be used to control the half-life of an active agent and modify the release kinetics.

[0280] In addition, depending on the chain length of Petition 870250084859, dated 09 / 19 / 2025, pp. 121 / 290 113 / 208 diacid ligands, the hydrolysis of ester linkages will depend on the pH and / or temperature of the environment. This can be used in certain modalities to control the release of the active agent, for example, for targeted release in certain tumor cells that have a higher pH than the surrounding cells.

[0281] In certain embodiments, the sustained-release hyper-branched drug macromolecule of the present invention is formulated to make an active agent available for a long period of time, thus allowing a reduction in dosing frequency compared to an immediate-release dosage form, such as a solution of an active agent that is applied topically to the eye (i.e., eye drops). In certain embodiments, the release of the active agent comprises constant release of the active agent, gradual release of the active agent, as well as any combination thereof, such as a constant release of the active agent followed by a gradual release of the active agent. The “sustained release” can be measured in vitro in an aqueous solution under physiological conditions such as pH 7, 27, 4 and 37 °C and is considered to be the same or substantially the same when the hyper-branched macromolecule is administered in vivo to an individual.

[0282] In several embodiments of the present invention, the release of the active agent follows zero-order release kinetics or substantially zero-order release kinetics, preferably without an “explosion” of active agent at the beginning of the period.

[0283] The embodiments of the present invention can provide the release of a therapeutically effective amount of the active agent for a period of time, such as up to 1 year, up to 9 months, up to 6 months, up to 3 months, up to 1 month, or up to about 25 days after administration. Other embodiments of the present invention can provide a Petition 870250084859, dated 09 / 19 / 2025, pp. 122 / 290 114 / 208 release of a therapeutically effective amount of the active agent for up to about 14 days, or up to about 21 days after administration, or a release of a therapeutically effective amount of the active agent for a period of about 6 hours or more after administration, or for a period of about 12 hours, or 24 hours or more, or about 48 hours or more, or about 72 hours or more, or about 7 days or more, or about 10 days or more after administration. The present invention contemplates all the above lower and upper time periods in any combination of ranges.

[0284] Some aspects of this disclosure are directed to a pharmaceutically acceptable hyper-branched macromolecule for controlled release of an active agent conjugated to a hyper-branched macromolecule, wherein controlled release is characterized by: the amount of active agent released on day 1 is 0 to 50% of the total amount of active agent, the amount of active agent released per day from day 2 to the last day of release is 0 to 50% of the total amount of active agent, and / or the number of days required for release of 100% of the total amount of active agent is at least 2 days.

[0285] In one embodiment, controlled release of the active agent is characterized by the amount of active agent released on day 1 being 0 to 50% of the total amount of active agent, the amount of active agent released per day from day 2 to the last day of release being 0 to 50% of the total amount of active agent, and / or the number of days required for the release of 100% of the total amount of active agent being at least 2 days. In another embodiment, the amount of active agent released on day 1 is 0 to 25%, 0 to 20%, 0 to 10%, 0 to 5%, or approximately 0% of the total amount of active agent, the amount of active agent released per day from day 2 to the last day of release is 0 to 50% of the total amount of active agent, and / or the number of days Petition 870250084859, dated 09 / 19 / 2025, pp. 123 / 290 The time required for the release of 100% of the total quantity of the active agent, 115 / 208, is at least 3 days, but not longer than 30 days, 25 days, or 16 days. Sustained-release drug delivery system

[0286] The hyper-branched macromolecules of certain embodiments of the invention can be used for drug delivery to a patient and, for example, for ophthalmic drug delivery, since they offer a number of advantages as a delivery system. The hyper-branched macromolecules can be used for drug delivery, gene delivery, antioxidant delivery, peptide delivery, biomedical imaging and genetic testing in ophthalmology.

[0287] Hyper-branched macromolecules are capable of transporting into and out of cells. Different ocular delivery routes can be used for administering drugs with hyper-branched macromolecules, and their adjustable properties, such as water solubility, permeability, bioavailability, and biocompatibility, can be widely varied depending on the specific needs of different medical applications.

[0288] In certain embodiments, a biodegradable sustained-release drug delivery system comprising the hyper-branched macromolecules as described herein is provided. In certain embodiments of the invention, the hyper-branched macromolecules or the drug delivery system comprising them may be formulated for direct or indirect administration by various routes, such as oral, parenteral, or by operative insertion or injection.

[0289] To formulate the drug delivery system, the Petition 870250084859, dated 09 / 19 / 2025, pp. 124 / 290 116 / 208 hyper-branched macromolecules can be incorporated into a suitable vehicle, such as a solvent or solvent mixture, or they can be incorporated into a hydrogel or organogel.

[0290] In certain modalities, hyperbranched macromolecules are formulated for direct injection into a patient's treatment site, for example, by parenteral administration or intratumoral injection, injection into the eye, such as intravitreal, intracameral, subconjunctival, retrobulbar, subtenon, subretinal and suprachoroidal injections.It can be formulated for injection into the anterior chamber, vitreous, episcleral, posterior subtenon space (inferior fornix), subconjunctival, intracameral, peribulbar, retrobulbar, subtenon, retinal, subretinal, intracanalicular, intravitreal, intrascleral, choroidal, suprachoroidal, a retina, subretinal or lens, a corneal or conjunctival surface, points (canaliculus, superior / inferior canaliculus), ocular fornix, superior / inferior ocular fornix, subtenon space, choroid, suprachoroidal, Tenon, cornea, cancerous tissue, organ, prostate, breast, joint space, subdural, dental, subcutaneous, carpal tunnel, perivascular, surgically created space or lesion, empty space, and potential space.

[0291] In embodiments of the invention, the drug delivery system is used to produce or form a medical implant, in which hyper-branched macromolecules are incorporated or dispersed in a hydrogel or organogel matrix. Treatment methods

[0292] According to certain embodiments of the invention, the hyper-branched macromolecules or the biodegradable drug delivery system comprising the hyper-branched macromolecules are configured for use as a medicament, such as for use in the treatment of a disease or medical condition of a patient. Petition 870250084859, dated 09 / 19 / 2025, pp. 125 / 290 117 / 208

[0293] In one embodiment, the method for treating a patient's disease or medical condition comprises administering hyper-branched macromolecules to the patient to release the active agent over a long period of time.

[0294] A treatment method of one embodiment of the invention comprises an ocular treatment. In this treatment, the hyper-branched macromolecule is used to release the active agent over a long period of time in the eye. In one embodiment thereof, the disease or medical condition to be treated is an ocular disease, such as fundus diseases, such as any posterior segment ocular disease affecting the vasculature and integrity of the retina, macula or choroid, leading to visual acuity disturbances, vision loss or blindness, particularly posterior segment disease states resulting from age, trauma, surgical interventions, such as age-related macular degeneration (AMD), cystoid macular edema (CME), diabetic macular edema (DME), posterior uveitis and diabetic retinopathy.

[0295] The treatment method may also involve treatment of glaucoma, ocular hypertension, hyphema, presbyopia, cataract, retinal vein occlusion, inflammation, miosis, mydriasis, conjunctivitis, intraocular infections, choroidal neovascularization (CNV), intraocular tumors, and retinal neuroinflammation.

[0296] Eye disease may also include retinal neovascularization, choroidal neovascularization, wet AMD, dry AMD, retinal vein occlusion, diabetic macular edema, retinal degeneration, corneal graft rejection, retinoblastoma, melanoma, glaucoma, autoimmune uveitis, uveitis, proliferative vitreoretinopathy and corneal degeneration, acute and chronic macular neuroretinopathy, central serous chorioretinopathy, macular edema, acute multifocal placoid pigment epitheliopathy, Behçet's disease, retinochoroidopathy of Petition 870250084859, dated 09 / 19 / 2025, pp. 126 / 290 118 / 208 birdshot, posterior uveitis, posterior scleritis, serpiginous choroiditis, subretinal fibrosis, uveitis syndrome, Vogt-Koyanagi-Harada syndrome, retinal artery occlusive disease, central retinal vein occlusion, disseminated intravascular coagulopathy, branch retinal vein occlusion, hypertensive fundus changes, ocular ischemic syndrome, retinal arterial microaneurysms, Coat's disease, parafoveal telangiectasia, hemiretinal vein occlusion, papillophlebitis, carotid artery disease (CAD), frozen branch angiitis, sickle cell retinopathy, angioid streaks, familial exudative vitreoretinopathy, Eales disease, proliferative vitreous retinopathy, diabetic retinopathy, tumor-associated retinal disease, congenital retinal pigment epithelium hypertrophy (RPE), uveal melanoma posterior, choroidal hemangioma, choroidal osteoma, choroidal metastasis, combined hamartoma of the retina and retinal pigment epithelium, retinoblastoma,Vasoproliferative tumors of the fundus, retinal astrocytoma, intraocular lymphoid tumors, myopic retinal degeneration, acute retinal pigment epithelium, glaucoma, endophthalmitis, cytomegalovirus retinitis, retinal cancers, retinitis pigmentosa, Leber congenital amaurosis, choroideremia, X-linked retinitis pigmentosa, vitelliform macular dystrophy, X-linked retinoschisis, CNGA3 achromatopsia, CNGB3 achromatopsia, LHON, Stargardt disease, Usher syndrome, Norrie disease, Bardet-Biedl syndrome, and red-green color blindness.

[0297] The methods described in this section may also include the administration of hyper-branched macromolecules in combination with another agent, also referred to as “combination therapy”.

[0298] In one embodiment, combination therapy comprises administering the hyper-branched macromolecules in combination with one or more additional agents on the same day or on different days. In Petition 870250084859, dated 09 / 19 / 2025, pp. 127 / 290 119 / 208 In one embodiment, the additional agent to be administered in a combination therapy may be a liquid formulation of the agent, or it may be contained in an oral dosage form. Thus, the additional agent may be any small molecule, large molecule, a protein, a nanoparticle, or any other of the active agents described herein. In another embodiment, hyper-branched macromolecules having more than one active agent conjugated to them, such as those available from the convergent synthesis described above, may be used for combination therapies involving the administration of more than one active agent. With the hyper-branched macromolecules of certain embodiments, it is possible to conjugate different regions on the surface of the hyper-branched macromolecule with different agents.

[0299] The treatment method involving the administration of hyper-branched macromolecules may include any of the following injections: intravitreal, intracameral, subconjunctival, retrobulbar, subtenon, subretinal, and suprachoroidal. The method of administration may also be topical or oral.

[0300] The active agent or the additional agent to be administered in combination therapy may also be a diagnostic agent. Diagnostic agents have been described above and may be substances used to examine the body in order to detect impairment of its normal functions. In some cases, diagnostic agents may be agents with a functional purpose, such as for use in detecting ocular deformities, diseases, and pathophysiological aspects.

[0301] Examples of medical treatment modalities involving drug-conjugated dendrimers of the invention are summarized in Table A below.

[0302] Table A Treatment, specific use and advantages of dendrimers Petition 870250084859, dated 09 / 19 / 2025, pp. 128 / 290 120 / 208 Drug Delivery: Dendrimers can encapsulate drugs, improving their solubility, stability, and targeted delivery to specific cells or tissues. Drug Delivery Systems: The renal clearance capability, despite their high molecular weight, makes these PEG dendritic polymers excellent candidates for creating drug delivery systems. They can be designed to encapsulate therapeutic agents within their structure or attach them to modifiable end groups, allowing for controlled release. Their biodegradability ensures they can be safely decomposed in the body after fulfilling their drug delivery function. Imaging Agent: Dendrimers can be functionalized with imaging molecules (e.g., fluorescent dyes or MRI contrast agents) for medical diagnostics and imaging applications.Imaging and Diagnostics: The unique structure of these dendritic PEGs can improve the delivery of contrast agents for imaging techniques such as magnetic resonance imaging (MRI) or positron emission tomography (PET). Their biocompatibility and renal clearance capability make them safe carriers that can be functionalized with imaging agents. Nanotechnology: Dendrimers serve as building blocks for nanomaterials and nanodevices, such as sensors, nanoelectronics, and nanocomposites. Gene Therapy: Dendrimers can deliver nucleic acids (e.g., by... Petition 870250084859, dated 09 / 19 / 2025, pp. 129 / 290 121 / 208 For example, DNA or RNA) for gene therapy applications, allowing efficient transfection and modulation of gene expression. Gene Therapy The ability to introduce different terminal groups allows these dendrimers to be adapted for gene delivery applications. They can be used to bind and protect nucleic acids, such as DNA or RNA, facilitating their delivery to cells for gene therapy purposes. The biodegradable nature ensures that the carrier degrades after delivering its genetic payload, reducing potential cytotoxicity. Targeted Cancer Therapy Adjustable diffusivity and the ability to modify surface groups make these polymers ideal for targeted cancer therapies. By conjugating targeting ligands to the surface, these dendritic polymers can be targeted to specific cancer cells, releasing therapeutic agents directly where needed, minimizing side effects on healthy tissue.Biosensors: Dendrimers can be incorporated into biosensors to detect various analytes (e.g., biomolecules, pathogens, or pollutants) with high sensitivity and selectivity. Biomedical Materials: Dendrimers can be used in tissue engineering, wound healing, and regenerative medicine due to their biocompatibility and ability to modify surface properties. Drug Delivery Systems: Dendrimers are used in transdermal drug delivery systems. Petition 870250084859, dated 09 / 19 / 2025, pp. 130 / 290 122 / 208 Increasing solubility for drug administration and offering targeted drug delivery capabilities. Medical Applications: biomedical studies, contrast agents for magnetic resonance imaging, improved stability of formulations, and as nanodrug carriers. Nanoparticle Synthesis: Dendrimers play a role in the synthesis of metallic nanoparticles, such as PAMAM dendrimers used for nanoparticle formation. Therapeutic Vectors: Effective amphiphilic dendrimer vectors share two important elements: hydrophilic chemical entities to bind RNA and stabilizing hydrophobicity of the RNA complex. These two combined characteristics allow the encapsulation of RNA within a stable complex before its release into the cytosol after endocytosis. This hydrophilic / hydrophobic balance allowed by the structural characteristics of amphiphilic dendrimers plays a determining role in the success of RNA delivery.Antimicrobials, antiseptics, diagnostics, tissue engineering. Their ability to form hydrogels can be exploited in the creation of scaffolds for tissue engineering. These structures can aid in cell growth and tissue regeneration, with the added advantage of controlled release of growth factors or other molecules. Petition 870250084859, dated 09 / 19 / 2025, pp. 131 / 290 123 / 208 Bioactive compounds to promote healing. Globular protein mimics. Therapeutic vaccines. Tissue engineering. The ability to form dendrimers through cross-linking makes multi-arm PEG ideal for creating structures that mimic the extracellular matrix, aiding in cell growth and tissue regeneration. Wound healing: These PEG-based hydrogels can be applied as wound dressings that not only provide a moist environment but can also be designed to release therapeutic agents to promote healing. Tissue engineering: Dendrimeric polymers can be designed to mimic the extracellular matrix, aiding in cell attachment and growth. Their multi-arm structure can be exploited to present multiple signals to cells, promoting tissue formation.Hydrolysis degradation allows these structures to gradually break down as new tissues form, making them ideal for regenerative medicine. Antibacterial and antifouling surfaces: By attaching antimicrobial agents to some arms and other functional groups that resist protein adsorption to others, dendrimeric polymers can be used to create surfaces that are both antibacterial and antifouling. This application is particularly useful in medical devices and... Petition 870250084859, dated 09 / 19 / 2025, pp. 132 / 290 124 / 208 Implants, where preventing bacterial colonization is crucial. Wound healing: Dendrimers can be applied as wound dressings that not only provide a moist environment but can also be designed to release therapeutic agents to promote healing. Targeted therapy: By attaching therapeutic molecules to modifiable end groups, dendrimers can be designed to release drugs in a controlled manner. This is particularly useful for cancer therapy, where targeted delivery can reduce side effects and improve efficacy. Targeted drug delivery: Dendrimeric polymers can be designed to carry multiple therapeutic agents simultaneously, each attached to different arms or end groups. This allows for the co-delivery of drugs that may work synergistically together, potentially increasing the effectiveness of treatments for complex diseases such as cancer.The biodegradability of these structures ensures that they can safely decompose after their therapeutic load has been administered. Long-term treatment: For chronic conditions, these polymers can provide sustained drug release, reducing dosing frequency and improving patient adherence. Bioadhesives: creation of bioadhesives that can be used in surgical repairs, such as sealants to prevent bleeding. Petition 870250084859, dated 09 / 19 / 2025, page 133 / 290 125 / 208 Leakage of bodily fluids or to attach medical devices to tissues. Intelligent Delivery Systems Dendrimeric polymers can be engineered to respond to specific stimuli (pH, temperature, enzymes) to release their cargo. This can be particularly useful in agriculture for the controlled release of pesticides or fertilizers, minimizing environmental impact and improving efficiency. Diagnostic Imaging With different terminal groups, dendrimeric polymers can be functionalized to carry imaging agents (such as MRI contrast agents or fluorescent molecules) and target ligands that seek specific disease markers. This dual functionality makes them excellent candidates for use in targeted diagnostic imaging, allowing for early detection of diseases at the molecular level.Self-healing materials: The incorporation of PEG dendritic polymers into composite materials can lead to self-repairing materials. Reversible hydrolyzable bonds may allow the material to repair itself when damaged by artificial enzymes. Industrial and non-medical applications

[0303] In embodiments of the invention, the hyperbranched molecules / dendrimers can also be used for non-medical or industrial applications. In certain embodiments, the dendrimers do not include hydrolyzable linkages. In other embodiments thereof, the dendrimers do not include hydrolyzable linkages as described herein. Petition 870250084859, dated 09 / 19 / 2025, pp. 134 / 290 126 / 208

[0304] Table B below provides an overview of non-medical and industrial applications and exemplary uses of the dendrimers of embodiments of the present invention.

[0305] Table B Application Fields: Specific Use of Dendrimers / Hyper-branched Molecules: Antibody Purification: High avidity can reversibly bind antibodies and allow separation of low molecular weight impurities by size-based techniques or by coupling to a solid substrate. Fragrance Release: Can be formulated as an aqueous application that slowly releases fragrance from the dendrimer by hydrolysis. Hair Products: Hair surface binding agents, dynamic coloring agents (change color over time as different pigments are hydrolyzed). Water Purification: Can bind specific impurities, allowing separation by size of the bound complex. Possibly flocculation and / or filtration.Recyclable. Coupling agents for high-tech adhesives. The structure allows for bispecific bonding. Sunscreens for the skin. Dendrimers may contain portions that absorb titanium dioxide / UV, as well as other portions that can bind to the skin (water resistance), but are released at high pH (soap). Petition 870250084859, dated 09 / 19 / 2025, pp. 135 / 290 127 / 208 Solar antennas, photocatalysis, battery catalysis: dendrimers can be used to stabilize catalytic agents on surfaces / chips / in solution and can still be recovered and separated from the final products. In catalysis, dendrimers have been proposed to fill the gap between homogeneous catalysis and surface catalysis because their sizes allow for the separation of reaction products by modern membrane techniques. Dendrimers with catalytic functionalities can be used in chemical reactions to accelerate or control reactions, improving efficiency and selectivity. The dendrimeric structure, with its large surface area and multiple functional sites, can serve as a support for catalysts. This application can revolutionize several industrial processes, including organic synthesis and the production of renewable energy sources.Magnetic materials Electronic devices Dendrimers can be used in electronic devices, such as organic light-emitting diodes. Petition 870250084859, dated 09 / 19 / 2025, pp. 136 / 290 128 / 208 (OLEDs) and photovoltaic cells, for improved performance and stability. Environmental Remediation Dendrimers can be used for pollutant capture, water purification, and remediation of contaminated sites due to their high surface technique and functionalization capabilities. Environmental Remediation The customized attachment of specific functional groups makes these dendrimers suitable for capturing and removing pollutants from the environment. For example, certain terminal groups can selectively bind to heavy metals or organic pollutants, allowing their efficient removal from water or soil. Water Treatment Flocculation and Coagulation: Water-soluble polymers are used to aggregate suspended solids in water treatment processes, facilitating their removal. They are used in the treatment of municipal and industrial effluents.Water treatment Sludge dewatering: polymers aid in sludge dewatering by binding water molecules, making the sludge easier to filter and dry. Agriculture Soil conditioning: improves soil structure, promoting water retention and aeration Agriculture Water retention agents: used. Petition 870250084859, dated 09 / 19 / 2025, page 137 / 290 129 / 208 In hydrogels to help soils retain water, reducing the need for frequent irrigation. Coatings and adhesives: Dendrimers can improve the properties of coatings and adhesives, providing better adhesion, corrosion resistance, and mechanical strength. Coatings and adhesives: For agrochemical delivery, these systems can be used to release fertilizers, pesticides, or herbicides in a controlled manner, minimizing environmental impact and improving efficiency. Paper manufacturing: Strength additives: used to increase paper strength. Retention aids: Dendrimers improve the retention of fillers and fines, reducing material loss and improving the efficiency of the paper manufacturing process. Light harvesting materials: Photodynamic therapy: Transdermal patches / dressings with skin adhesion and localized administration of antihistamines / antibiotics / sunscreens, etc.Separating Agents Enhanced Oil Recovery (EOR) Water-soluble polymers are injected into oil reservoirs to increase the viscosity of the floodwater, improving its effectiveness. Petition 870250084859, dated 09 / 19 / 2025, pp. 138 / 290 130 / 208 in pushing oil towards production wells. Environmental Detection and Release Systems By incorporating detection mechanisms that respond to environmental stimuli (pH, temperature, enzymes), these dendrimers can be designed to release their cargo only under specific conditions, increasing the precision of treatments or protecting sensitive ingredients. Pollutant Capture and Release Adjustable diffusivity and surface functionality can be exploited in the design of systems for controlled capture and release of pollutants or in the delivery of environmental remediation agents. Their biodegradability ensures minimal environmental impact. Nanofiltration Membranes The size and structure of these dendritic polymers may make them suitable for creating or enhancing nanofiltration membranes, potentially improving water purification technologies by selectively filtering pollutants while allowing water molecules to pass through.Textile sizing agents: applied to yarns to strengthen and protect them during weaving. Textile finishing agents: impart desired properties to fabrics. Petition 870250084859, dated 09 / 19 / 2025, pp. 139 / 290 131 / 208 such as softness, crease resistance, and water repellency. Printing inks / nanotubes Thickening agents Used in cosmetics and personal care products to adjust viscosity and improve texture in cosmetics Thickening Agents: Used in cosmetics and personal care products to adjust viscosity and improve texture in cosmetics In cosmetic formulations, dendrimers can be used to encapsulate active ingredients, such as vitamins or antioxidants, protecting them from degradation and ensuring their controlled release for prolonged benefits. Detoxification materials Food Industry Thickeners, gelling agents, and stabilizers: improve the texture, consistency, and stability of food products.Construction: Concrete additives: improve concrete properties such as workability, strength, and drying time. Paint coatings: Additives for paints and coatings: used to adjust viscosity, improve adhesion, and increase water resistance. Food packaging: The incorporation of these dendrimers into packaging materials may... Petition 870250084859, dated 09 / 19 / 2025, pp. 140 / 290 132 / 208 To allow the controlled release of preservatives or antioxidants to extend the shelf life of food products, without direct addition to the food.

[0306] Other examples of application and use include those of Table C:

[0307] Table C Application Specific Use Surface Chemistry Tribology, Roughness, Contact Angle, Coatings, Lubricants Cosmetics Color, Skin Care Detergents Soaps and Dishwashing Membranes Ion-Specific Membranes Fillers Coatings, Reinforcements Scaffolding / 3D Printing Tissue Engineering, Bone Generation Phase Change Materials Diapers, Mattresses Filtration Size / Specific Separation Wound Healing Antimicrobial Adhesives Surgical Kits, Surfaces Rheology Modifiers Xanthan Gum, Starches Energy Storage Adhesives Urethanes and Epoxies Food Science Flavors, Sugars (Chewing Gum) Emulsions Polymer Synthesis Stimuli-Responsive Light-Sensitive / pH-Sensitive / Pressure-Sensitive EXAMPLES

[0308] The following examples are included to demonstrate certain embodiments of the invention as described in the claims. It should be appreciated by those skilled in the art, however, that the description a Petition 870250084859, dated 09 / 19 / 2025, pp. 141 / 290 133 / 208 below is for illustrative purposes only and should not be considered in any way as a restriction of the invention.

[0309] Materials and abbreviations used in the examples: 4-arm PEG-(NHS)3-(azide)1 difunctional degradable (10kJ, 20kJ and 40kDa) and 4-arm PEG-DBCO (1kJ, 20kJ, 40kDa) were purchased from XIAMEN SINOPEG BIOTECH Co. Ltd. All NHS-terminated PEGs were purchased from JenKem Technology USA.

[0310] Compstatin (ICVVQDWGHHRCT, disulfide bridge: Cys2-Cys12, TFA and acetate salt forms) was acquired from MedChemExpress. APL-1 (ICV{L-1-Me-Trp}QDWGAHRCT, disulfide bridge: Cys2-Cys12, TFA and acetate salt forms) and Fc-lll4C (CDCAWHLGELVWCTC, disulfide bridge: Cys1-Cys15, Cys3-Cys13, TFA and acetate salt forms) were acquired from Alan Scientific. The structure is shown in FIG. 4. Solvents and other reagents, including methanol, acetonitrile, PBS buffer, and triethylamine, were purchased from VWR. Example 1

[0311] Divergent synthesis of branched GO-peptide macromolecule conjugates:

[0312] Scheme 1: G0-peptide

[0313] A quantity of peptide was weighed and dissolved in anhydrous methanol. 4 arms of PEG-SS-NHS (MW=40 kDa) with a molar ratio of 1:6 to peptide was slowly added to the peptide solution (compstatin) with vigorous stirring. A small amount of triethylamine (5-10 pL) was added to the reaction mixture. The reaction was conducted at room temperature for 1-4 hours. The final product was collected and purified via dialysis against a weight-cut tube. Petition 870250084859, dated 09 / 19 / 2025, pp. 142 / 290 134 / 208 molecular weight 10 kDa in methanol for 24 hours. The purified hyperbranched macromolecule G0 peptide conjugate was collected from the dialysis tube and the solvent was removed in a rotary evaporator. The dry powder was stored at -20°C for characterization. The formulation is shown in Table 1:

[0314] Table 1: Synthesis of the G0-comp conjugate Peptide-amine Compstatin ester 4 arms 40kPEG(SS)-NHS End group NH2 NHS # of functionality / molecule 1 4 MW 1600 40000 Theoretical reaction ratio 4 1 Experimental excess reaction ratio 6.25 1 mmol of molecule 0.0125 0.002 mmol of functionality 0.0125 0.008 weight (mg) 20 80 Solvent: anhydrous methanol (mL) 2 2 conc. in reaction mixture (mg / mL) 5 20 EXAMPLE 2

[0315] Divergent synthesis of G1-peptide branched macromolecule conjugates:

[0316] Scheme 2: End functional group conversion Petition 870250084859, dated 09 / 19 / 2025, pp. 143 / 290 135 / 208 DBCO-amine 4-arm PEG-NHS Endfunctional 4-arm PEG-(N) (NHS) 1 3 4-arm PEG-DBCO>j macromolecule formation 4arms PEG-Í 4arms PEG (NHS) 1 = G1 4 Conversion of PEG-DBCO 4-arm end functional group:

[0317] A quantity of PEG-NHS 4-arms (MW=40kDa) was dissolved in anhydrous methanol. DBCO-amine with a molar ratio of 1:1 to PEG-NHS 4-arms was weighed and dissolved in anhydrous acetonitrile. The DBCO-amine solution was added dropwise to the PEG-NHS 4-arms solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product was used in the next step of the synthesis. The formulation is shown in Table 2 Step 1. Formation of a hyper-branched 4-armed PEG macromolecule [4-armed PEG-(NHS)3]4(G1):

[0318] A quantity of 4-armed PEG-DBCO was dissolved in a mixed solvent of anhydrous methanol:acetonitrile = 1:1. 4-armed PEG(N3)i(NHS)3de (MW = 20 kDa) with a molar ratio of 1:1 to 4-armed PEG-DBCO was weighed and dissolved in anhydrous methanol. The 4-armed PEG-(N3)i(NHS)3 solution was slowly added to the 4-armed PEG-DBCO solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product can be further purified by dialysis if necessary. The formulation is shown in Table 2, Step 2. Petition 870250084859, dated 09 / 19 / 2025, pp. 144 / 290 136 / 208 Peptide conjugation of the hyper-branched macromolecule G1-conjugated peptide:

[0319] A quantity of peptide was weighed and dissolved in anhydrous methanol. 4-armed PEG-[4-armed PEG-(NHS)3]4 (G1) with a molar ratio of 1:18 to peptide was slowly added to the peptide solution with vigorous stirring. A small amount of triethylamine (5-10 pL) was added to the reaction mixture. The reaction was conducted at room temperature for 1-4 hours. The final product was collected and purified via dialysis against a 40 kDa molecular weight shear tube in methanol for 24 hours. The purified G1-peptide hyper-branched macromolecule conjugates were collected from the dialysis tube and the solvent was removed in a rotary evaporator. The dry powder was stored at -20°C for characterization. The formulation is shown in Table 2, Step 3.

[0320] Table 2 Step 1: Synthesis of 4-armed 40k PEG-SG-(DBCO)4 amine Ester DBCO-amine 4-armed 40kPEG-SG-NHS End group NH2 NHS # of functionality / molecule 1 4 MW 276 40000 Theoretical reaction ratio 4 1 Experimental excess reaction ratio 6 1 mmol of molecule 0.15 0.025 mmol of functionality 0.15 0.1 weight (mg) 42 1000 Solvent: anhydrous acetonitrile:methanol = 1:1 (mL) 2 10 concentration in reaction mixture (mg / mL) 3.5 83 Petition 870250084859, dated 09 / 19 / 2025, pp. 145 / 290 137 / 208 Step 2: 4 arms 40k PEG-SG-[ 4 arms 20k PEG-SG- (NHS)3]4 = G1 azide DBCO 4 arms 20k PEGSG-(N3)i(NHS)3 4 arms 40k PEG-SG(DBCO)4 End group N3 DBCO # of functionality / molecule 1 4 MW 20000 40000 Theoretical reaction ratio 4 1 Experimental excess reaction ratio 4 1 mmol of molecule 0.0125 0.003125 mmol of functionality 0.0125 0.0125 weight (mg) 250 125 Solvent: anhydrous acetonitrile:methanol = 1:1 (mL) 1 conc. in reaction mixture (mg / mL) 125 62.5 Step 3: Peptide conjugation of 4 arms 40k PEG-SG-[4 arms 20k PEG-SG(Comp)3]4 amine Compstatin ester 4 arms 40k PEG-SG[4 arms 20k PEG-SG(NHS)3]4 = G1 NH2 end group NHS # of functionality / molecule 1 3 MW 1550 120000 theoretical reaction ratio 12 1 experimental excess reaction ratio 24 1 mmol of molecule 0.0129 0.00054 mmol of functionality 0.0129 0.00648 weight (mg) 20 64.5 Solvent: acetonitrile 1 1 Petition 870250084859, dated 09 / 19 / 2025, pp. 146 / 290 138 / 208 anhydrous:methanol=1:1 (mL) conc. in reaction mixture (mg / mL) 10 32 EXAMPLE 3

[0321] Divergent synthesis of hyperbranched G2-peptide macromolecule conjugates:

[0322] Scheme 3: 4-arm PEG-(N) (NHS) I | Macromolecule Formation I G2-peptide conjugate 4-armed PEG-{4-armed PEG-[4-armed PEG (NHS)3]} = G2 4-arm PEG end functional group conversion [4-arm PEG (DBCO)3]4:

[0323] A quantity of 4-armed PEG-[4-armed PEG-(NHS)3]4 (G1) obtained in step 2 of Example 2 was dissolved in anhydrous methanol. DBCO-amine with a molar ratio of 1:1 to 4-armed PEG-[4-armed PEG(NHS)3]4 (G1) was weighed and dissolved in anhydrous acetonitrile. The DBCO-amine solution was added dropwise to the 4-armed PEG[4-armed PEG-(NHS)3]4 (G1) solution with vigorous stirring. The reaction was carried out at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product was used in the next step of the synthesis. The formulation is shown in Table 3 Step 1. Formation of a hyper-branched macromolecule with 4 PEG arms. PEG-1 arms. PEG (NHS)3]3}4

[0324] A quantity of 4-arm PEG-[4-arm PEG (DBCO)3]4 Petition 870250084859, dated 09 / 19 / 2025, pp. 147 / 290 139 / 208 was dissolved in anhydrous mixed solvent methanol:acetonitrile=1:1. 4-arm PEG-(N3)i(NHS)3 (MW=10kDa) with a molar ratio of 1:1 to 4-arm PEG-[4-arm PEG (DBCO)3]4 was weighed and dissolved in anhydrous methanol. The 4-arm PEG-(N3)1(NHS)3 solution was slowly added to the 4-arm PEG-[4-arm PEG (DBCO)3]4 solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product can be further purified by dialysis if necessary. The formulation is shown in Table 3, Step 2. Peptide conjugation of the hyper-branched macromolecule G2-peptide conjugate:

[0325] A quantity of peptide was weighed and dissolved in anhydrous methanol. 4-armed PEG-{4-armed PEG-[4-armed PEG (NHS)3]3}4 (G2) with a molar ratio of 1:18 to peptide was slowly added to the peptide solution with vigorous stirring. A small amount of triethylamine (5-10 pL) was added to the reaction mixture. The reaction was conducted at room temperature for 1-4 hours. The final product was collected and purified via dialysis against a 40 kDa molecular weight shear tube in methanol for 24 hours. The purified hyper-branched macromolecule conjugates of the G2 peptide were collected from the dialysis tube and the solvent was removed in a rotary evaporator. The dry powder was stored at -20°C for characterization. The formulation is shown in Table 3, Step 3.

[0326] Table 3: Step 1: Synthesis of 4-arm 40k PEG-SG-(DBCO)4 amine Ester DBCO-amine 4-arm 40k PEG-SG-[4-arm 20k PEG-SG-(NHS)3]4 = G1 NH2 NHS end group # of functionality / molecule 1 12 Petition 870250084859, dated 09 / 19 / 2025, pp. 148 / 290 140 / 208 MW 276 120000 Theoretical reaction ratio 12 1 Experimental excess reaction ratio 24 1 mmol of molecule 0.024 0.001 mmol of functionality 0.024 0.012 weight (mg) 6.6 120 Solvent: anhydrous acetonitrile:methanol=1:1 (mL) 1 9 conc. in reaction mixture (mg / mL) 0.66 12 Step 2: 4 arms 40k PEG-SG-{4 arms 20k PEG-SG-[4 arms 10k PEG-SG-(NHS)3]3}4 = G2 azide DBCO4a 10k PEGSG(N3)1(NHS)3 4 arms 40k PEG-SG-[4 arms 20k PEG-SG(DBCO)3]4 End group N3 DBCO # of functionality / molecule 1 12 MW 10000 120000 theoretical reaction ratio 12 1 Experimental excess reaction ratio 12 12 mmol of molecule 0.024 0.001 mmol of functionality 0.024 0.012 weight (mg) 240 120 Solvent: anhydrous acetonitrile:methanol=1:1 (mL) 5 5 con. In reaction mixture (mg / mL) 24 12 Petition 870250084859, dated 09 / 19 / 2025, pp. 149 / 290 141 / 208 Step 3: Peptide conjugation of 4 arms 40k PEG-SG-{4 arms 20k PEGSG-[4 arms 10k PEG-SG(Comp)3]3}4 amine Compstatin ester 4 arms 40k PEG-SG- {4 arms 20k PEG-SG- [4 arms 10k PEG-SG- (NHS)3]3}4 = G2 End group NH2 NHS # of functionality / molecule 1 36 MW 1550 240000 theoretical reaction ratio 36 1 Experimental excess reaction ratio 72 1 mmol of molecule 0.006 0.000083 mmol of functionality 0.006 0.003 weight (mg) 10 20 Solvent: anhydrous acetonitrile:methanol=1:1 (mL) 1 1 concentration in reaction mixture (mg / mL) 5 10 EXAMPLE 4 Convergent synthesis of conjugated hyper-branched macromolecules G1-peptide: Diagram 4: 4-arm PEG-(N) (NHS) Conjugates of X Peptide peptide 4 arms PEG-(N3)i(peptide)3 formation of macromolecules 4-arm PEG-DBCO G1-peptide conjugate PEG-(N3)i(peptide)3 4-arm peptide conjugation:

[0327] A quantity of peptide was weighed and dissolved in Petition 870250084859, dated 09 / 19 / 2025, pp. 150 / 290 142 / 208 anhydrous methanol. 4 arms of PEG-(Ns)i(NHS)3 (MW=20kDa) with a molar ratio of 1:4.5 to peptide was slowly added to the peptide solution with vigorous stirring. A small amount of triethylamine (5-10 pL) was added to the reaction mixture. The reaction was conducted at room temperature for 1-4 hours. The solvent was removed in a rotary evaporator. The dry crude product was used in the next step of the synthesis. The formulation is shown in Table 4, Step 1. Formation of a hyper-branched macromolecule conjugate of G1 peptide:

[0328] A quantity of PEG-(N3)1(peptide)3 4-arms was dissolved in anhydrous methanol. PEG-DBCO 4-arms (MW=40kDa) with a molar ratio of 1:1 to PEG-(N3)1(peptide)3 was weighed and dissolved in anhydrous methanol. The PEG-(N3)1(peptide)3 4-arms solution was slowly added to the PEG-DBCO 4-arms solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours. The conjugate was collected and purified via dialysis tubing with a molecular weight cutoff of 40 kDa in methanol for 24 hours. The solvent was removed in a rotary evaporator and the dry powder was stored at -20°C for characterization. The formulation is shown in Table 4, Step 2.

[0329] Table 4: Step 1: Synthesis of 4a 20k PEG-SG-(N3)1(Comp)3 amine Compstatin ester 4a 20k PEG-SG-(N3)1(NHS)3 NH2 end group nhs # of functionality / molecule 1 3 MW 1550 20000 theoretical reaction ratio 3 1 Excess reaction ratio 6 1 Petition 870250084859, dated 09 / 19 / 2025, pp. 151 / 290 143 / 208 Experimental molar mass of molecule 0.012 0.002 mol of functionality 0.012 0.006 weight (mg) 19 40 Solvent: anhydrous methanol (mL) 2 2 conc. in reaction mixture (mg / mL) 5 10 Step 2: 4 arms 40k PEG-SG-[4 arms 20k PEG-SG-(Comp)3]4 azide DBCO 4a 20k PEG-SG(N3)1(Comp)3 4 arms 40k PEGSG-(DBCO)4 End group N3 DBCO # of functionality / molecule 1 4 MW 20000 40000 theoretical reaction ratio 4 1 Experimental excess reaction ratio 4 1 mmol of molecule 0.003 0.00075 mmol of functionality 0.003 0.003 weight (mg) 60 30 Solvent: anhydrous methanol (mL) 2 1 conc. in reaction mixture (mg / mL) 20 10 EXAMPLE 5 Peptide conversion and conjugation of hyperbranched G0 macromolecules: Scheme 5: Petition 870250084859, dated 09 / 19 / 2025, pp. 152 / 290 144 / 208 Route 1:

[0330] Peptide-DBCO Conversion: Step 1. A quantity of peptide (compstatin, MW=1.5kDa) was dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile=1:1). DBCO-NHS was weighed and dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile = 1:1). The peptide solution was added dropwise to the DBCO-NHS solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours. The dry crude product was used in the next step of the synthesis. The formulation is shown in Table 5 Step 1.

[0331] Formation of 8-armed PEG-(peptide)8 dendrimer. Step 2. A quantity of 8-armed 20k PEG-Azide was dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile = 1:1) and then slowly added to the peptide-DBCO solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product can be further purified by dialysis if necessary. The formulation is shown in Table 5 Step 2.

[0332] Table 5: Step 1: Synthesis of DBCO-NHS peptide-azide MW 402 1550 Petition 870250084859, dated 09 / 19 / 2025, pp. 153 / 290 145 / 208 Weight (mg) 20 37.5 mmol 0.05 0.025 -NHS / amine in molecules 1 1 Theoretical reaction ratio 1 1 Experimental excess reaction ratio 2 1 Solvent (acetonitrile:methanol=1:1) 1 mL 1 mL Step 2: Synthesis of 8-arm PEG-(peptide)8 peptideDBCO 8-arm 20k PEG-Azide MW 2k 20k Theoretical reaction ratio 8:1 Experimental excess reaction ratio 16:1 Weight (mg) 57.5 36 mmol 0.02875 0.0018 Solvent (acetonitrile:methanol=1:1) 1 mL 1 mL Route 2:

[0333] Peptide-Azide Conversion: Step 1. A quantity of peptide (compstatin, MW=1.5kDa) was dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile=1:1). Azide-NHS was weighed and dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile = 1:1). The peptide solution was added dropwise to the Azide-NHS solution with vigorous stirring. The reaction was conducted at room temperature for 1-4 hours. The dry crude product was used in the next step of the synthesis. The formulation is shown in Table 6, Step 1.

[0334] Formation of 8-armed PEG-(peptide) dendrimers: Step 2. A quantity of 8 arms 20k PEG-DBCO was dissolved in 1 mL of mixed solvent (anhydrous methanol:acetonitrile = 1:1) and then slowly added to the peptide-azide solution with stirring. Petition 870250084859, dated 09 / 19 / 2025, pp. 154 / 290 146 / 208 vigorous. The reaction was conducted at room temperature for 1-4 hours, and the solvent was removed in a rotary evaporator. The dry crude product can be further purified by dialysis, if necessary. The formulation is shown in Table 6, Step 2.

[0335] Table 6: Step 1: Azide-Azide-NHS Peptide Synthesis MW Peptide 198 1550 Weight (mg) 10 37.5 mmol 0.05 0.025 -NHS / amine in molecules 1 1 Theoretical reaction ratio 1 1 Experimental excess reaction ratio 2 1 Solvent (acetonitrile:methanol=1:1) 1 mL 1 mL Step 2: Synthesis of 8-arm PEG-(peptide)8 peptide-azide 8-arm 20k PEG-DBCO MW 2k 20k Theoretical reaction ratio 8:1 Experimental excess reaction ratio 16:1 Weight (mg) 47.5 30 mmol 0.02375 0.0015 Solvent (acetonitrile:methanol=1:1) 1 mL 1 mL Example 6 Purification of conjugates of macromolecules and hyperbranched peptides by dialysis.

[0336] The products obtained in Examples 1 to 5 were dissolved in methanol to achieve a concentration of 10 mg / mL or higher, and the solution (1-5 mL) was loaded into a dialysis tube (Spectra / Por® Float-A-Lyzer G2 Dialysis Devices, Spectrum® Laboratories) with a Petition 870250084859, dated 09 / 19 / 2025, pp. 155 / 290 147 / 208 specific molecular weight cutoff. The tube was then placed in 500 mL of methanol in a beaker at room temperature for 24 hours to separate the lower molecular weight components that diffused out of the tube. See FIG. 5 a). After treatment, the product was removed from the tube and collected in a glass vial for characterization.

[0337] Table 7: Dialysis tubing selection Molecular weight cut: Purpose: 8-10 kDa Remove free peptide 20-50 kDa Remove free peptide, small PEG 50-100 kDa Remove free peptide, small-large PEG

[0338] The products were then characterized by UHPLC. FIG. This shows a UHPLC analysis of a G1 conjugate 4arm 40k PEG[4arm 20k PEG-SG-(comp)s]4 from Example 2 purified by methanol dialysis with membrane molecular weight shear of 8-10kDa that primarily removes the low molecular weight peptide (MW of approximately 1.5kDa), where the blue line is before purification and the green line is after purification. It can be observed that the peak at a retention time of approximately 4 minutes, attributed to free compstatin, decreased from 45% to less than 1% in the sample based on peak area integration. On the other hand, a starting material in the mixture, 4arm 40k PEG-SG-(DBCO)4 (retention time of approximately 9 minutes, MW of approximately 40kDa), which has a molecular weight greater than 8-10kDa, remained in the tube. Furthermore, the hyperbranched macromolecule conjugate (MW of approximately 120 kDa) remained in the tube, with a peak slightly shifted to the left at approximately 8.5 to 9 minutes.The separation of the hyper-branched macromolecule conjugate from the PEG-DBCO precursor can be done using another dialysis tube with a higher molecular weight cutoff.

[0339] Table 8: Example 2 of the product before and after dialysis purification (data based on UHPLC peak area integration). Petition 870250084859, dated 09 / 19 / 2025, pp. 156 / 290 148 / 208 Sample before purification After purification Free compstatin 45% <1% Conjugated hyperbranched macromolecule-compstatin 55% >99%

[0340] These data show that a very efficient purification with purity greater than 99% of hyper-branched macromolecule-peptide conjugates can be obtained by dialysis. EXAMPLE 7 Purification of hyper-branched macromolecule and peptide conjugates by the SEC column filtration method.

[0341] The SEC column (Zeba™ Spin Desalting columns) was unpacked and rinsed with 1 mL of methanol twice. The products obtained in Examples 1 to 5 were dissolved in methanol to achieve a concentration of 10 mg / mL or higher (1-2 mL) and the solution was loaded into the column, flowing through the column by gravity. See FIG. 5b). The eluate was collected in a glass vial for characterization.

[0342] Table 9: SEC column selection Molecular weight cutoff: Purpose: kDa Free Peptide Remover kDa Free Peptide Remover, small PEG

[0343] FIG. 7 shows a UHPLC analysis of a G0 conjugate. 4arm 40k PEG-SS-compstatin from Example 1 before (black line) and after purification (blue line). Based on peak area integration, the free compstatin content decreased from 38.8% to 1.6%, demonstrating a very efficient purification capability.

[0344] Table 10: Content change before and after filtering the SEC column (data based on UHPLC peak area integration). sample before purification after purification Petition 870250084859, dated 09 / 19 / 2025, pp. 157 / 290 149 / 208 Free icompstatin 38.8% 1.6% Conjugated PEG-compstatin 61.2% 98.4%

[0345] These data show that a very efficient purification with purity greater than 98% of hyper-branched macromolecule-peptide conjugates can be obtained by SEC column filtration. EXAMPLE 8 Purity and replacement rate by UHPLC

[0346] Ultra-high performance liquid chromatography (UHPLC) is an efficient technique that offers more sensitive analyses with good chromatographic separation and resolution of analytes. It provides benefits including rapid analysis, high-resolution separations, reduced solvent and sample usage, improved sensitivity and precision, etc. A Waters XBridge BEH300 C18 column (3.5 µm, 2.1 x 100 mm, PN1860036080) was used to characterize hyperbranched macromolecules and hyperbranched macromolecule-peptide conjugates of Examples 1 to 5, with mobile phases A (0.1% trifluoroacetic acid in water) and B (0.1% trifluoroacetic acid in acetonitrile).

[0347] For UHPLC analysis, the peptide powder was dissolved in PBS: methanol = 9:1 at concentrations of 12.5, 25, 50, 100, and 200 pg / mL was injected using UHPLC. The peak area of ​​each sample was integrated and used as a standard for calculating peptide concentration (as shown in FIGURE 8a). The inset graph is the standard curve of peptide concentration versus peak integration area.

[0348] Dried hyper-branched macromolecule-peptide G0 PEG was dissolved in PBS:methanol = 9:1 at a concentration of 1 mg / mL and analyzed by UHPLC. FIG. 8b shows a typical UHPLC plot of a 4-arm PEG-hyper-branched macromolecule-compstatin G0 conjugate from Example 1, in which the peak with a retention time of 22 minutes is from free compstatin, and the peak centered at 42 minutes is from the 4-arm PEG-hyper-branched macromolecule-compstatin conjugate. Using Petition 870250084859, dated 09 / 19 / 2025, pp. 158 / 290 150 / 208 The standard curve of compstatin, the concentration of each component in the product can be calculated and the peptide substitution can be estimated by the moles of peptide conjugated with hyperbranched macromolecule divided by the moles of PEG, according to the following equation: peptide substitution ________C (peptide conjugate)________ _ molecular weight of the peptide C(total sample) — [C (preset pep.) + C(free pep.)] molecular weight of PEG

[0349] In this equation, C(conjugated peptide) is the concentration of conjugated peptide, C(free peptide) is the concentration of free peptide, C(total sample) is the concentration of total solids in the prepared sample of PEG-hyper-branched macromolecule-peptide conjugate, i.e., including conjugated peptide, free peptide and free (non-conjugated) macromolecule, the equation resulting in moles of conjugated peptide per mole of macromolecule.

[0350] This method provides a practical way to compare and optimize reaction methods. Compstatin peptide samples from different suppliers, Ambeed, Genscript, and MCE, in different salt forms, trifluoroacetate (TFA), acetate, and lysine, were selected to react with the same 4-arm 40k PEG-SGA-NHS conjugation system under the same reaction conditions. Table 11 lists the conjugation results. Overall, the substitution of each peptide in the 4-arm PEG G 0 system is approximately 80%, with a variation of ±10% between them.

[0351] Table 11: Supplier Salt form # of end groups Compstatin conjugate calculated per hyper-branched macromolecule Substitution % Ambeed TFA 4 3.7 92.5 Petition 870250084859, dated 09 / 19 / 2025, pp. 159 / 290 151 / 208 iGenscript acetate 4 2.9 72.5 iGenscript Lys 4 3.3 82.5 MCE free NH2 4 3.2 80.0

[0352] Compstatin and compstatin-lysine replacement in different PEGs, such as 4-arm 40k PEG-SGA and 4-arm 40k PEG-SS, were investigated using the same analytical method. These reactions were performed under the same conditions and with the same molar ratio of each compound, only altering the PEGs. Three replicates of each reaction were applied, and in FIG. 9, these reactions show good reproducibility, with an average substitution of approximately 3 peptides in 4-arm PEGs.

[0353] Other reaction conditions, such as reaction time, solvent and catalyst use, were investigated and the results are shown in FIG. 10. According to the results, it is clear that the reaction in methanol with triethylamine as a catalyst is the best condition to achieve the highest peptide substitution, while the reaction time plays a minor role in altering the number of substitutions.

[0354] Table 12 below lists a series of hyperbranched macromolecules, from G0 to G2, that were synthesized by the convergent or divergent method. The highest molecular weight hyperbranched macromolecule is approximately 240 kDa for a G2 hyperbranched macromolecule, with approximately 36 end functionalities in the structure. In most conjugation results, peptide substitution was above 50%, indicating that these methods have good reproducibility.

[0355] Table 12: Hyperbranched macromolecule structure. Synthesis method. MW of macromolecule. Number of end functionalities. Number of substitutions. Substitution (%). Petition 870250084859, dated 09 / 19 / 2025, pp. 160 / 290 152 / 208 4a 40kPEG-[4a 10k PEG-SG- (comp)3]4 convergent 80k 12 10.8 90% 4a 40kPEG-[4a 20k PEG-SG- (comp)3]4 convergent 120k 12 7.0 58% 4a 40k PEG-SS(comp)4 divergent 40k 4 2.9 73% 4a 40k PEGSG-(comp)4 divergent 40k 4 2.6 65% 4a 40kPEG-[4a 10k PEG-SG- (comp)3]4 convergent 80k 12 6 50% 4a 40kPEG-[4a 10k PEG-SG- (comp)3]4 convergent 80k 12 6 50% 4a 40kPEG-[4a 20k PEG-SG-(comp)3]4 convergent 120k 12 5 42% 4a 40kPEG-[4a 20k PEG-SG-(comp)3]4 convergent 120k 12 8 67% 4a 20kPEG-[4a 10k PEG-SG-(comp)3]4 divergent 60k 12 3.5 29% 4a 40kPEG-[4a 10k PEG-SG-(comp)3]4 divergent 80k 12 3.8 32% 4a 20kPEG-[4a 10k PEG-SG-(comp)3]4 convergent 60k 12 6.1 51% 4a 40kPEG-[4a convergent 80k 12 8.8 73% Petition 870250084859, de 19 / 09 / 2025, pág. 161 / 290 153 / 208 10k PEG-SG- (comp)3]4 4a 40k PEG- SGA-(comp)4 divergent 40k 4 3.7 93% 4a 40k PEG- SGA-(comp)4 divergent 40k 4 2.9 73% 4a 40k PEG- SGA-(comp)4 divergent 40k 4 3.3 83% 4a 40k PEG- SGA-(comp)4 divergent 40k 4 3.2 80% 4a 40kPEG-[4a 20k PEG-SG-(comp)3]4 divergent 120k 12 10.8 90% 4a 40kPEG-{[4a 20k PEG-SG- (4a 10k PEG- SG-(comp)3]3}4 divergent 240k 36 29.1 81%

[0356] The same synthesis method was applied using APL-1 and Fc-III 4C for conjugation of hyper-branched macromolecules, and the substitution results are shown in Tables 13 and 14. The substitution of APL-1 and Fc-III 4C is less than that of Compstatin.

[0357] Table 13: Structure of hyperbranched macromolecule synthesis method MW of macromolecule # of functionality of end # of substitution substitution % 4a 40k PEG- SGA-(APL-1)4 divergent 40k 4 1.3 32% 4a 20k PEG-SS(APL-1)4 divergent 20k 4 2.9 72% 4a 40kPEG-[4a 10k PEG-SG- convergent 80k 12 5.1 43% Petition 870250084859, dated 09 / 19 / 2025, pp. 162 / 290 154 / 208 (APL-1)3]4 4a 40kPEG-[4a 20k PEG-SG- (APL-1)3]4 convergent 120k 12 6.4 53% 0358] Table 14: Structure of hyperbranched macromolecule synthesis method MW of macromolecule # of functionality of end # of substitution substitution % 4a 20k PEG-[4a 10k PEG-SG-(Fc-III 4C)3]4 convergent 60k 12 3.3 28% 4a 20k PEG-[4a 20k PEG-SG-(Fc-III 4C)3]4 convergent 100k 12 3.8 32% 8a 20k PEG-[4a 10k PEG-SG-(Fc-III 4C)3]8 convergent 100k 24 3.6 15% 8a 20k PEG-[4a 20k PEG-SG-(Fc-III 4C)3]8 convergent 180k 24 4.5 19% 4a 40k PEG-[4a 10k PEG-SG- (Fc-III 4C)3]4 convergent 80k 12 1.7 14% 4a 40k PEG-[4a 20k PEG-SG- (Fc-III 4C)3]4 convergent 120k 12 3.6 30% 4a-40k PEG- SGA-(Fc-III 4C)4 divergent 40k 4 1.2 30% 4a-40k PEG- SGA-(Fc-III 4C)4 divergent 40k 4 1.1 28% EXAMPLE 9 Petition 870250084859, dated 09 / 19 / 2025, pp. 163 / 290 155 / 208 Bond test

[0359] The surface plasmon resonance (SPR) linkage analysis methodology obtained from Mosaic Biosciences, Inc. was used to study molecular interactions.

[0360] A Biacore 3000 instrument is used to detect signals SPR. Generally, C3 and C3b were immobilized on the sensor chip surface at high density (~20 kRU). Aqueous buffered saline solution at pH 7.4 was passed through the device at a flow rate of 30 pL / min at 25 °C. Hyper-branched macromolecule-peptide conjugates of embodiments of the invention were injected at concentrations ranging from 1 nM to 300 nM (APL-1 derivatives) or 200 nM to 50 μM (Compstatin derivatives). Association was monitored for 4 minutes and dissociation for 10 minutes. Equilibrium analysis was performed for Compstatin analogs, and kinetic analysis with mass transport for APL-1 analogs. Binding affinity of free peptides

[0361] FIGS. 11 and 12 show the binding of C3 and C3b of different types of compstatin, and FIG. 13 shows the binding of C3 and C3b of different types of APL-1. The KD results are summarized in Table 15. From these results it is clear that the free peptides, compstatin and APL-1, show KD values ​​remarkably similar to the reported results. This result is in agreement with previous measurements with APL-1, but has lower affinity than that reported by Apellis for APL-2 (200 pM), possibly due to the avidity effect of the bivalent APL-2.

[0362] Table 15: KD of the affinity of C3 and C3b of compstatin and APL-1 and comparison with reference results. Common Derivative Reagent C3 Affinity, Kd C3b Affinity, Kd Reported C3 Affinity, Kd Petition 870250084859, dated 09 / 19 / 2025, pp. 164 / 290 156 / 208 Compstatin-1 Compstatin * (TFA salt) 13.5 pM 8.8 pM 3-10 pM Compstatin-2 Compstatin** (acetate salt) 6.8 pM 4.8 pM 3-10 pM Compstatin-3 Compstatin** (lysine end) 3.3 pM 2.6 pM 3-10 pM Compstatin-4 Compstatin*** (free amine) 10 pM 6.6 pM 3-10 pM APL-1-1 APL-1** (acetylated amine) 14 nM 15 nM 11 nM (Mosaic) APL-1-2 APL-1 ** (acetate salt) 18 nM 21 nM 11 nM (Mosaic) APL-1-3 APL-1** (lysine end) 19 nM 22 nM 11 nM (Mosaic) *Sample purchased from Ambeed, **Sample purchased from Genscript, ***Sample purchased from MCE.

[0363] The same experiments were also conducted for Fc-lll 4C. For comparison, another peptide, Fc-lll, was also evaluated under the same conditions. Fc-lll has a peptide sequence similar to Fc-lll 4C, but lacks a Cys-Cys bridge. The amino acid sequence structures are as follows:

[0364] Scheme 6: Fc-lll-4C; k„2.45 nM Fc-lll; 16 nM

[0365] This structural difference leads to a significant difference in antibody binding affinity, approximately 8 times more sensitive for Fc Petition 870250084859, dated 09 / 19 / 2025, pp. 165 / 290 157 / 208 III 4C to the antibody (KD is equal to 2.45 nM vs. 16 nM). The SPR results (FIG. 13 and Table 16) also showed that 3 types of Fc-III 4C exhibit much lower KD binding affinity than Fc-III.

[0366] Table 16: KD of Fc-III 4C IgG affinity and comparison with reference results. Common Derivative Reagent IgG Affinity, Kd Reported IgG Affinity, Kd Fc-III-4c-1 Fc-III-4C* (HCl salt) 7.6 nM 2.5 nM Fc-III-4c-2 Fc-III-4C* (acetate salt) 11 nM 2.5 nM Fc-III-4c-3 Fc-III-4C** (salt acetate) 6.7 nM 2.5 nM Fc-III -4 Fc-III** (acetate salt) 62 nM 16 nM *Sample purchased from Genscript, **Sample purchased from Alan Scientific. Binding affinity of conjugated peptides of hyper-branched macromolecules

[0367] The same experiment was applied to test the binding affinity of conjugated peptides of hyper-branched macromolecules, in which C3 was immobilized on the chip and conjugated hyper-branched peptides of macromolecules were flown over at a variety of concentrations.

[0368] FIG. 15 shows a comparison of free compstatin and a multivalence compstatin, 4arm 40k PEG-SGA-(comp)4. When both samples flow over the C3 coated chip, both show a very rapid association rate. After the interaction, buffer solution was passed over the chip to wash the samples associated with it. During this dissociation time, the response of the free compstatin drops very rapidly, the rate is the same as during association. On the other hand, the Petition 870250084859, dated 09 / 19 / 2025, pp. 166 / 290 158 / 208 hyper-branched macromolecule-compstatin conjugates exhibit a much slower dissociation rate, which is caused by the multiple peptide interactions in the hyper-branched macromolecule with the receptors. FIG. 16 ac) shows the SPR results of the comparison of 3 different hyper-branched macromolecule-comp conjugates with free compstatin. The compstatin conjugated to the hyper-branched macromolecule appears to contain a fast and slow dissociation component, where the slower dissociation phase associated with these constructs indicates cooperative binding of the multivalent hyper-branched macromolecule conjugates to the C3 surface.

[0369] The sample in FIG. 16 d) is a compstatin after hydrolysis of 4a 40k PEG-SS-(comp)4. This hydrolyzed peptide contains a succinate ester linkage resulting from the degradation of the conjugation linker group and showed no effects on its C3 linkage, according to the similar SPR signal. This strongly indicates that the ester linkage did not alter the bioactivity of the peptide.

[0370] The same set of high-purity samples was detected again to obtain quantitative KD analysis. The KD results are listed in Table 17 and plotted against the corresponding number of peptide substitutions in each sample. It can be observed that KD shows a decrease when there is more substituted peptide in the hyper-branched macromolecule.

[0371] Table 17: KD of hyperbranched macromolecule-compstatin conjugates. Sample Compstatin Replacement KD [M]* -SS-comp (hydrolysis) 1 1.89E-06 4a-40kPEG-SS-(comp)4 2.2 1.24E-09 4a-40kPEG-SGA-(comp)4 2.4 3.85E-09 4a-40kPEG-SGA-(comp)4 2.5 7.46E-09 Petition 870250084859, dated 09 / 19 / 2025, pp. 167 / 290 159 / 208 4a-40k-PEG-[4a-20kPEG(comp)3]4 7.1 1.09E-09 4a-40k-PEG-[4a-10kPEG(comp)3]4 10.8 1.85E-09 * Kd was calculated using the reported association rate constant for Compstatin (5e5 M-1 s-1) and adjusting the dissociation rate (kd) for multivalent dissociation. Kd is then calculated as kd / ka. Kd for compstatin was measured directly in this experiment. EXAMPLE 10 Alternative pathway hemolysis assay (AP) for IC50 measurement

[0372] The IC50 (half the maximum inhibitory concentration) was measured by an alternative pathway hemolysis assay (AP). Four hyper-branched macromolecules conjugated with compstatin were used in this assay: Samples no. REA638 (4a-40k-PEG-SGA-(comp)n TFA salt) and REA639 (4a-40k-PEG-SGA-(comp)n acetate salt) having 2.6 and 2.5 compstatins per hyper-branched macromolecule, respectively, and samples no. REA640 (4a-40k-PEG-[4a-10kPEG-(comp)n]4) and REA641 (4a-40k-PEG-[4a-20k-PEG-(comp)n]4) having 10.8 and 7.1 compstatins per hyper-branched macromolecule, respectively (as per Tables 10 and 15).

[0373] In a 96-well plate assay experiment (cf. Fig. 18) Inhibitors (50 μL) are diluted in GVBo (GVBo: 0.1% gelatin, 5 mM barbital, 145 mM NaCl, 0.025% NaNS, pH 7.3) and incubated with 1:2 normal human serum:GVBo in a concentration range for 30 minutes at room temperature. Rabbit red blood cells (CompTech) are pelleted at 500 x g for 3 minutes and resuspended at 5.0 x 10⁸ cells / mL in MgEGTA (MgEGTA: 0.1 MMgCl₂, 0.1 M EGTA, pH 7.3). Rabbit RBCs (20 μL) were added and incubated at 37°C for 60 minutes. The reaction was stopped by the addition of 200 μL of GVBE (GVBE: 0.1% gelatin, 5 mM barbital, 145 mM NaCl, 10 mM EDTA, 0.025% NaN3, pH 7.3). Cells pelleted at 500 xg Petition 870250084859, dated 09 / 19 / 2025, pp. 168 / 290 160 / 208 for 5 minutes and the supernatant (150 μL) transferred to a new 96-well plate. Calculate % Hemolysis = (A412 inhibitor / no A412 inhibitor)*100 and fit with the 4PL curve to determine IC50. The results are shown in Table 18.

[0374] Tabela 18: Resultados de IC50 do ensaio de hemolise AP Sample IC50 [μΜ] Compstatin 6.83 REA638 (4a 40k-PEG-SGA-(comp)n (TFA salt)) 11.7 REA639 (4a-40k-PEG-SGA-(comp)n acetate salt) 12.3 REA640 (4a-40k-PEG-[4a-10kPEG-(comp)n]4) 3.91 REA641 (4a-40k-PEG-[4a-20k-PEG-(comp)n]4) 1.35

[0375] From the results (FIG. 19 and Table 18), it can be observed that the 4-arm G0 PEG compstatins (REA638 and REA639 having 2.6 and 2.5 compstatins per hyperbranched macromolecule, respectively) showed worse IC50s per molecule compared to the free compstatin. This is not surprising, as these molecules have approximately 2 compstatins per molecule and therefore probably cannot achieve avidity with such low substitution. On the other hand, the G1 hyperbranched PEG macromolecule compstatins (REA640 and REA641 having 10.8 and 7.1 compstatins per hyperbranched macromolecule, respectively) showed improved IC50s compared to the free compstatin, suggesting an improvement in potency through avidity. Furthermore, the slope of these curves (curve slope) is lower compared to free compstatin and REA 638 and REA639, suggesting multiple binding events that contribute to the inhibition.Interestingly, REA641, which has 7.1 compstatins, performed better than REA640, which has 10.8 compstatins, despite having a higher valence than the latter. It is believed that the 20k PEG dendrons of REA641 provide greater flexibility for interaction with [the underlying components]. Petition 870250084859, dated 09 / 19 / 2025, pp. 169 / 290 161 / 208 C3 than the shorter 10k PEG dendrons of the REA640. Classical Pathway Hemolysis Assay (CP)

[0376] The IC50 (half the maximum inhibitory concentration) was also measured by a classical pathway hemolysis assay (CP), using the same hyper-branched macromolecule conjugates as the AP hemolysis assay above. This assay is similar in principle to the AP hemolysis assay, but uses sensitized sheep red blood cells, since the classical pathway begins with antibody binding to cells.

[0377] Serial dilutions of inhibitors (50 pL) were prepared in veronal gelatin buffer supplemented with Mg2+ and Ca2+GVB++ (GVB++: 0.1% gelatin, 5 mM barbital, 145 mM NaCl, 0.025% NaN3, pH 7.3, containing 0.15 mM calcium chloride and 0.5 mM magnesium chloride) in a 96-well plate. C3-depleted human serum supplemented with 12 nM human C3 was diluted 1:2 in GVB++ (30 pL) and added to each well and incubated for 30 minutes. Sheep erythrocytes sensitized with anti-sheep pAbs at 5.0 x 108 cells / mL in GVB++ (20 pL) were added and incubated at 37°C for 30 minutes. The reaction was stopped by the addition of gelatin veronal buffer with EDTA (GVBE, 200 μL). Cells were pelleted at 500 x g for 5 minutes and the supernatant (150 μL) transferred to a new 96-well plate. Absorbance at 412 nm was measured using a Molecular Devices SpectraMax M5 plate reader, and % hemolysis was calculated by % hemolysis = (A412 inhibitor / A412 without inhibitor)*100.IC50s were determined using a 4PL curve fit in GraphPad Prism. The assay was also performed in the absence of C3 to determine background hemolysis. The results are shown in Table 19.

[0378] Table 19: IC50 Results of Hemolysis Assay AP Sample IC50 [μM] Compstatin 142 Petition 870250084859, dated 09 / 19 / 2025, pp. 170 / 290 162 / 208 REA638 (4a 40k-PEG-SGA-(comp)n (TFA salt)) 56.0 REA639 (4a-40k-PEG-SGA-(comp)n acetate salt) 73.3 REA640 (4a-40k-PEG-[4a-10kPEG-(comp)n]4) 30.8 REA641 (4a-40k-PEG-[4a-20k-PEG-(comp)n]4) 34.6

[0379] Free compstatin had an IC50 = 142 μM. Increasing the valence of the hyperbranched macromolecule to 2.6 and 2.5 for REA638 and REA639 improved the IC50s to 56.0 μM and 73.3 μM, respectively. Further increasing the valence of the hyperbranched macromolecule to 10.8 or 7.1 compstatins (REA640 and REA641) improved the IC50s to 30.8 and 34.6 μM, respectively. Combined, these data suggest that the higher valence compstatins of the invention are more effective in inhibiting CP hemolysis compared to free compstatin alone.

[0380] Similarly to what is described above, the IC50 (half the maximum inhibitory concentration) was also measured by the classical pathway hemolysis assay (CP), using samples of a 12-armed G1 hyperbranched molecule 4a40k-PEG(SGA)-[4a20k PEG(SG)-(APL1)s]4 prepared by convergent synthesis as in Example 4, using APL-1 instead of compstatin. The product has a substitution rate of about 35% of the 12 terminal groups, which was also used in Examples 11 and 12 below. 32 mg x2 (dendrimer equivalent to 20 mg x2) of lyophilized dendrimer were reconstituted in 90 mM aqueous sodium phosphate and 360 mM NaCl, and 50 μL aliquot doses (1.25 mg / eye) were injected into the eyes of the animal (New Zealand white rabbit) for one week. Vitreous humor was collected from 2 animals / time point at 1, 3, 5, and 7 days post-dose and analyzed using the classical pathway hemolysis assay (CP) to determine the IC50 over time.The IC50 value of the dendrimer was determined as a control. The results are summarized in Table 20 below.

[0381] Table 20: Petition 870250084859, dated 09 / 19 / 2025, pp. 171 / 290 163 / 208 Sample Day 1 IC50 [nM] Day 3 IC50 [nM] Day 5 IC50 [nM] Day 7 IC50 [nM] Control dendrimer 54.7 - - - Right eye (OD) 43.2 90.2 54.7 39.4 Left eye (OS) 16.4 - 67.2 50.8

[0382] The results show that APL-1 bound to the dendrimer maintains its stability and activity in vivo for more than 7 days. EXAMPLE 11 Correlation of hydrodynamic radius with in vivo half-life

[0383] The hydrodynamic radius Rh of linear pegylated protein (IgG) of different sizes (IgG 2x40k PEG and 2x20k PEG) and its half-life T1 / 2 in the vitreous humor of the New Zealand white rabbit (NZWVH) was determined and compared to the free protein and several unconjugated active ingredients (APIs) to obtain a calibration curve allowing the estimation of T1 / 2 of APL-1 conjugated to the PEG 12arm120kDa dendrimer based on its Rh determined by SEC. The results are shown in Table 21 below:

[0384] Table 21: API theoretical MW (Mn, kDa) Rh per SEC (nm) NZW T1 / 2 (days) IgG 2x40k PEG IgG 230 10.39 10.2 2x20k PEG IgG 190 9.27 7.3 IgG 150 4.69 3.3 Bevacizumab 150 4.17 3.7 Aflibercept 115 5.4 4.4 12a120k APL-1** ~120+ 9.75 8.7 **12a120k APL-1 is a first-generation dendrimer conjugate of the nominal structure 4a40k-PEG(SGA)-[4a20k-PEG(SG)-(APL-1)3]4 prepared by convergent synthesis as in Example 4, using APL-1 instead of compstatin. The product has a substitution rate of approximately 35% of the end 12 groups. Petition 870250084859, dated 09 / 19 / 2025, pp. 172 / 290 164 / 208

[0385] As can be seen in Table 20 and Figure 20, the hydrodynamic radius Rh determined by SEC allows reliable estimates of the half-life of dendrimer drug conjugates of embodiments of the invention depending on the size of the dendrimer and adjusting its sustained release properties. EXAMPLE 12 Effect of dendrimer degradation on in vitro release kinetics

[0386] A first-generation dendrimer conjugated with APL-1 of the nominal structure 4a-40kPEG-[4a-40kPEG-APL-1]3 prepared by convergent synthesis as in Example 4, using APL-1 instead of compstatin, with a substitution rate of approximately 35% of the 12 terminal groups, was subjected to in vitro degradation tests under various temperature and pH conditions to determine the degradation / hydrolysis behavior. The tests were performed in PBS using HPLC chromatograms to show the disappearance of the dendrimer and the appearance of dendron groups (hydrolysis products) over time.

[0387] Figures 21 a) ac) show the effect of temperature variation from 35 °C to 39 °C at a constant pH of 7.4. Figure 21 a) shows the decrease in dendrimer concentration over time, Figure 21 b) shows the increase in dendron concentration over time, and Figure 21 c) shows the impact of temperature on the % loss rate of dendrimer at pH 7.4 on a logarithmic scale, based on first-order release kinetics, to determine the rate constant K and the half-life T1 / 2 of the active agent release estimated from these degradation rates. The results are in Table 22 below:

[0388] Table 22: Temperature (°C) Tl / 2 K 35 23.59 0.029 Petition 870250084859, dated 09 / 19 / 2025, pp. 173 / 290 165 / 208 37 21.29 0.033 39 21.37 0.032

[0389] It can be observed that the degradation and therefore the release rate of APL-1 from this 1st generation 120kPEG dendrimer are increasing with temperature, with similar trends for all three temperatures in the first 26 days. Overall, the effect of temperature on the release half-life is low.

[0390] Figures 22 a) and c) show the effect of pH value at a constant temperature of 37°C. Figure 22 a) shows the decrease in dendrimer concentration over time, Figure 22 b) shows the increase in dendron concentration over time, and Figure 22 c) shows the impact of pH on the % dendrimer loss rate at pH 7.4 on a logarithmic scale, based on first-order release kinetics, to determine the rate constant K and the estimated release half-life T1 / 2 of the active agent from these degradation rates. The results are in Table 23 below:

[0391] Table 23: PH T1 / 2 K 7 51.43 0.013 7.4 21.36 0.032 7.8 10.28 0.067 8.5 3.58 0.194

[0392] It is observed that the degradation and therefore the release rate of APL-1 from this 1st generation 120kPEG dendrimer increases significantly over time with higher pH. Overall, the effect of pH on the release half-life is significant and follows an exponential model. The higher the pH, the shorter the release half-life. Petition 870250084859, dated 09 / 19 / 2025, pp. 174 / 290 166 / 208

[0393] These release experiments show that hydrolytic degradation of ester bonds between dendritic building blocks or dendrons can be used to alter or retard the release kinetics of active ingredients conjugated with dendrimers. Specific sets of modalities First set of modalities 1. A hyper-branched macromolecule comprising: a central unit with at least 3 connectivity options; a plurality of polymeric arms connected to the central unit in the c-connectivities, each polymeric arm comprising a terminal group or being connected to a dendritic constitutional repeat unit, the dendritic constitutional repeat unit comprising a branching unit connected to at least two polymeric arms, each comprising a terminal group or being connected to a next dendritic constitutional repeat unit that may again be connected to other dendritic constitutional repeat units, the polymeric arms of the outermost dendritic constitutional repeat unit of the hyper-branched macromolecule, each comprising a terminal group; wherein the polymeric arms comprise polyethylene glycol (PEG) units; in which at least one active agent is conjugated to at least one of the outermost polymeric arms; and in which the hyper-branched macromolecule includes chemical bonds that can be cleaved by hydrolysis. 2. The hyper-branched macromolecule according to the aspect 1, being a branched G0 generation macromolecule, in which the terminal groups of the branched macromolecule are the terminal groups Petition 870250084859, dated 09 / 19 / 2025, pp. 175 / 290 167 / 208 of the polymer arms connected to the central unit. 3. The hyper-branched macromolecule according to the aspect 1, being a higher-generation hyper-branched macromolecule G x, with x being an integer from 1 to 10 defining the number of consecutively connected dendritic constitutional repeating units in the hyper-branched macromolecule. 4. A hyper-branched macromolecule according to any of the preceding aspects, in which the central unit and the branching unit are the same or different and, independently of each other, have a c or c' connectivity of 3 to 10, or 4 to 8, or 4 to 6, or 4. 5. A hyper-branched macromolecule according to any of the preceding aspects, wherein the central unit and the branched unit are the same or different and are each derived from a polyol with at least 3 hydroxyl groups. 6. The hyper-branched macromolecule according to the aspect 5, wherein the polyol is selected from the group consisting of glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol or sorbitol. 7. A hyper-branched macromolecule according to any of the preceding aspects, wherein the polymeric arms comprise polyethylene glycol (PEG) units having an average molecular weight (Mn) in the range of about 1,000 to about 100,000 Daltons, or about 10,000 to about 60,000 Daltons, or about 15,000 to about 50,000 Daltons. 8. A hyper-branched macromolecule according to any of the preceding aspects, in which the average molecular weight of the PEG units of the core-linked polymeric arm is the same as or different from that of the polymeric arms in the dendritic constitutional repeat units. 9. A hyper-branched macromolecule according to any Petition 870250084859, dated 09 / 19 / 2025, pp. 176 / 290 168 / 208 one of the previous aspects, in which the average molecular weight of the PEG units of the core-linked polymeric arm is greater or less than that of the polymeric arms in the dendritic constitutional units. 10. A hyper-branched macromolecule according to any of the previous aspects, where for a higher-generation Gx hyper-branched macromolecule, with x being an integer from 2 to 10, the average molecular weight of the PEG units of the polymer arm decreases or increases from the innermost polymer arms to the outermost polymer arms. 11. A hyper-branched macromolecule according to any of the preceding aspects, wherein the terminal groups attached to the outermost polymeric arms are grafted onto the ends of the polymeric arms directly or by means of a suitable difunctional linker comprising hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic, heteroaliphatic or aromatic or heteroaromatic group. 12. A hyper-branched macromolecule according to any of the preceding aspects, wherein the functional groups of the terminal groups and / or terminal linking groups attached to the outermost polymeric arms are functional groups selected from electrophiles, such as activated ester groups, such as succinimidyl esters, succinimidyl carbonates; nitrophenyl carbonates, aldehydes, ketones, acrylates, acrylamides, maleimides, vinyl sulfones, iodoacetamides, alkenes, alkynes, azides, norbornenes, epoxides, mesylates, tosylates, tresylates, cyanurates, orthopyridyl disulfides or halogens; nucleophiles, such as an amine, such as a primary amine, a hydroxyl group, an alcohol, a thiol, an azide and a carboxyl group; functional groups for click chemistry; functional groups for cycloadditions, such as 1,3-dipolar cycloadditions, cycloadditions Petition 870250084859, dated 09 / 19 / 2025, pp. 177 / 290 169 / 208 [3+2] such as alkene-nitrone cycloadditions or alkyne-nitrone cycloadditions, [4+2] cycloadditions; functional groups for thiol-ene reactions; hetero-Diels-Alder cycloadditions; functional groups for nucleophilic ring openings, functional groups for non-aldol type carbonyl reactions; functional groups for addition reactions to carbon-carbon multiple bonds, polymerizable vinyl groups or combinations thereof. 13. A hyper-branched macromolecule according to any of the preceding aspects, wherein the terminal linker groups attached to the outermost polymeric arms are functional groups selected from succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ), and succinimidyl glutaramide (SGA). 14. A hyper-branched macromolecule according to any of aspects 1 to 10, wherein the terminal groups attached to the outermost polymeric arms are functional groups selected from an alkyne compound, such as a dibenzocyclooctyne (DBCO), or a bicyclo[6.1.0]-nonyne (BCN); or a norbornene, or a trans-cyclooctene (TCO); an azide, a 3,4-dihydroxyphenylacetic acid (DHPA) or a tetrazine (Tz). 15. A hyper-branched macromolecule according to any of the preceding aspects, in which the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed by click chemistry. 16. The hyper-branched macromolecule according to its appearance. 15, wherein the connection is formed by the reaction of a polymeric arm functionalized with a tensed or terminal alkyne, cycloalkyne, or alkene moiety with a polymeric arm functionalized with a moiety Petition 870250084859, dated 09 / 19 / 2025, pp. 178 / 290 170 / 208 azide or tetrazine in a SPAAC or IEDDA type click chemical coupling reaction. 17. The hyper-branched macromolecule according to its appearance. 16, where the alkyne portion is a dibenzocyclooctyne portion. 18. A hyper-branched macromolecule according to any of aspects 15 to 17, wherein the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed between the polymeric arms connected to the central unit and the polymeric arms connected to the branching unit of the dendritic constitutional repeat units and / or between the polymeric arms of a dendritic constitutional repeat unit and the polymeric arms of consecutive dendritic constitutional repeat units. 19. A hyper-branched macromolecule according to any of the preceding aspects, in which the active agent conjugated to at least one of the outermost polymeric arms is selected from the group consisting of therapeutically or diagnostically active agents. 20. A hyper-branched macromolecule according to any of the preceding aspects, wherein the active agent conjugated to at least one of the outermost polymeric arms is selected from steroids; non-steroidal anti-inflammatory drugs (NSAIDs), such as diclofenac, ibuprofen, meclofenamate, mefanamic acid, salsalate, sulindac, tolmetin, ketoprofen, diflunisal, piroxicam, naproxen, etodolac, flurbiprofen, fenoprofen C, indomethacin, celecoxib, ketorolac, nepafenac; intraocular pressure-reducing drugs; antibiotics, such as ciprofloxacin; analgesics, such as bupivacaine; calcium channel blockers, such as nifedipine; cell cycle inhibitors, such as simvastatin; proteins, such as insulin; Petition 870250084859, dated 09 / 19 / 2025, pp. 179 / 290 171 / 208 hydrophilic small molecule drugs, including carboxylic acid salts and amine salts; hydrophobic small molecule drugs, hydrophilic peptides and protein drugs, such as insulin, single-chain antibody fragments, Fab fragments, IgG antibodies, fusion antibodies, etc.; aptamers; particularly bupivacaine (BPV-HCI or base), ropivacaine (RPV), dexamethasone, travoprost, axitinib, non-steroidal anti-inflammatory drugs (NSAIDs), steroids, antibiotics, analgesics, calcium channel blockers, cell cycle inhibitors, chemotherapeutic agents, antiviral drugs, anesthetics, hormones, anticancer drugs, antineoplastic agents, viruses, gene delivery viruses, such as AAVs, protein ligands, such as nanobodies, affibodies, ankyrins, DARPins, etc., or any combination thereof. 21. A hyper-branched macromolecule according to any of the preceding aspects, wherein the active agent conjugated to at least one of the outermost polymeric arms is a peptide selected from the group consisting of Compstatin, APL-1 and Fc-III-4C, Beovu (Brolucizumab), Zimura (Avacincaptade Pegol), Pegcetacoplan, Abicipar Pegol, Lampalizumab, Fovista, Risuteganib, AXT107, Elamipretide, THR149, ALM201, VGB3 and Largazole. 22. A hyper-branched macromolecule according to any of the above aspects, in which the active agent is linked to at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or even 100% of the outermost polymeric arms. 23. The hyper-branched macromolecule according to any of the previous aspects, in which a dendritic constitutional repeating unit is represented by Formula (i): A ^LA-j-^OCH2CH2^—o---X--O—^CH2CH2O^ ---B where A is a connection to a polymeric arm that is connected to Petition 870250084859, dated 09 / 19 / 2025, pages 180 / 290 172 / 208 central unit, or A is a connection to B of a preceding dendritic constitutional repeating unit represented by Formula (i), La is a linker, m is any 0 or 1, n is an integer from 20 to 2000, o is an integer from 20 to 2000, neo can be different or equal, X is a branching unit having a c' connectivity, Lb is a linker, p is either 0 or 1, B comprises a terminal group located on the surface of the hyper-branched macromolecule or is a connection with A of a consecutive dendritic constitutional repeat unit or an active agent. La and Lb can be different or the same, mep can be different or the same, ey ​​is an integer from 2 to 9, where y = c' - 1, with c' being the connectivity c' of the branching unit X; and where the dendritic constitutional units in hyper-branched macromolecules can be the same or different. 24. A hyper-branched macromolecule according to aspect 23, in which the connection between A and B comprises a functional group formed by click chemistry, such as a triazole or dihydropyrazine. 25. A hyperbranched macromolecule according to aspects 23 or 24, wherein the La and / or Lb ligand comprises a diacid and / or an acidic diamido group, such as succinate, glutarate, adipate, azelate or glutaramide. 26. The hyper-branched macromolecule according to either of aspects 23 to 25, in which the La and / or Lb ligand comprises a structure represented by Formula (ii): Petition 870250084859, dated 09 / 19 / 2025, pp. 181 / 290 173 / 208 where U1 and U2 are independently NH or O and may be equal or different, and where t is an integer from 0 to 10. 27. The hyper-branched macromolecule according to aspects 25 or 26, wherein the La and / or Lb ligand further comprises a polyethylene glycol unit between the linkage to B and the carboxyl group, carboxamide group or structure of Formula (ii). 28. A method for manufacturing a hyperbranched macromolecule according to any of aspects 1 to 27 by divergent synthesis, comprising the following steps: (a) Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry at the terminations of the polymeric arms; (b) Providing dendritic constitutional repeat unit precursors comprising a polymeric arm comprising a functional group suitable for forming a click chemistry connection with the corresponding functional groups of the core-connected polymeric arms (such as an azide, alkyne, alkene, or tetrazine), and at least two polymeric arms comprising click-chemical non-reactive functional groups, (c) Establishing a click chemistry connection between the core-connected polymeric arms and the polymeric arms of the dendritic constitutional repeat unit precursors, (d) Optionally converting the functional groups of the at least two polymeric arms comprising click-chemical non-reactive functional groups into functional groups suitable for click chemistry, and (e) Conjugation of an active agent comprising a group Petition 870250084859, dated 09 / 19 / 2025, pp. 182 / 290 174 / 208 functional to the outermost polymeric arms through reaction with the functional groups of the outermost polymeric arms, thus forming a hyper-branched macromolecule-active agent conjugate. 29. The method according to aspect 28, wherein for the higher generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10, step (d) is mandatory and consecutive precursors of additional dendritic constitutional repeating units are connected to the functional groups suitable for click chemistry obtained in step (d) by click chemistry to the hyper-branched macromolecule before conjugation of the active agent in step (e). 30. The method according to aspects 28 or 29, in which the precursor of the dendritic constitutional repeating unit in step (c) is represented by Formula (iii): where C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide, or tetrazine), D comprises non-reactive functional groups in click chemistry (such as succinimidyl), and La, m, η, X, o, Lb, pey are as defined in aspects 23 to 27; and wherein the dendritic constitutional units may be the same or different. 31. The method according to aspects 28 to 30, wherein after a penultimate step (d) of converting the functional groups of at least two polymeric arms comprising non-reactive functional groups in click chemistry into functional groups suitable for click chemistry, the active agent in step (e) is first functionalized with a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine) and then conjugated in a click chemistry reaction to the polymeric arms. Petition 870250084859, dated 09 / 19 / 2025, pp. 183 / 290 175 / 208 outer layers of the hyper-branched macromolecule. 32. The method described in aspect 31, in which the active agent functionalized with a functional group suitable for click chemistry is a peptide. 33. A method for manufacturing a hyperbranched macromolecule according to any of aspects 1 to 27 by convergent synthesis, comprising the following steps: I) Supply of precursors for dendritic constitutional repeat units comprising - a polymeric arm comprising a functional group suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine), and - at least two polymeric arms comprising non-reactive functional groups in click chemistry, II) Conjugating active agents comprising a functional group to at least one of the at least two polymeric arms comprising non-reactive functional groups in the click chemistry of the dendritic constitutional repeat unit precursors, III) Providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry (such as an azide, alkyne, alkene or tetrazine) at the ends of the polymeric arms, and IV) Forming a click chemistry connection between the polymeric arms connected to the core provided in step III) and the polymeric arm comprising a functional group suitable for forming a click chemistry connection of the dendritic constitutional repeat unit precursors conjugated to the active agent obtained in step II), thus forming a hyper-branched macromolecule-active agent conjugate. Petition 870250084859, dated 09 / 19 / 2025, pp. 184 / 290 176 / 208 34. The method according to aspect 33, in which a precursor of the dendritic constitutional repeating unit in step i) is represented by Formula (iii): where C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide, or tetrazine), D comprises non-reactive functional groups in click chemistry (such as succinimidyl), and La, Lb, m, η, X, o, pey are as defined in aspects 23-27, wherein the dendritic constitutional units may be the same or different. 35. The method according to aspects 33 or 34, wherein for the higher-generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10, the precursors of conjugated dendritic constitutional repeat units to the active agent obtained in step II) are connected by click chemistry to the precursors of reverse dendritic constitutional repeat units comprising: a polymeric arm comprising a non-reactive functional group in click chemistry, and at least two polymeric arms comprising functional groups suitable for click chemistry (such as an azide, alkyne, alkene, or tetrazine), wherein the non-reactive functional group in click chemistry of the polymeric arm is subsequently converted into a functional group suitable for click chemistry before connecting to other precursors of reverse dendritic constitutional repeating units or before forming a connection by click chemistry with the polymeric arms connected to the core in step IV), thus forming higher-generation hyper-branched macromolecules. 36. The method according to aspects 33 or 34, in which Petition 870250084859, dated 09 / 19 / 2025, pages 185 / 290 177 / 208 precursors of dendritic constitutional repeat units having different active agents conjugated to the polymeric arms are obtained by performing steps I) and II) for each precursor of dendritic constitutional repeat unit conjugated to the active agent, and a mixture of the obtained precursors of dendritic constitutional repeat units conjugated to the active agent is used for step IV), thus forming a hyper-branched macromolecule conjugate with different active agents in different regions of the surface of the hyper-branched macromolecule. 37. The method according to any of aspects 28 to 36, wherein the outermost polymeric arms of the hyperbranched macromolecule have terminal maleimide functional groups, and peptides or active agents are conjugated to them via the maleimidthiol reaction. 38. The method of aspect 37, in which terminal maleimide functional groups are provided by reacting the terminal functional group functionalized with DBCO or azide of the hyperbranched macromolecule with click chemistry ligands having an azide or DBCO functionality connected to a maleimide group, such as DBCOmaleimide, DBCO-PEG3-maleimide, DBCO-PEG4-maleimide or azidoPEG3-maleimide. 39. A hyper-branched macromolecule in accordance with any one of aspects 1 to 27, for use as a medicine. 40. A treatment method, wherein the method comprises treating a disease or medical condition in a patient with a hyper-branched macromolecule in accordance with any of aspects 1 to 27. 41. The hyper-branched macromolecule for use or the treatment method according to aspects 39 or 40, wherein the hyper-branched macromolecule is used for an eye treatment. Petition 870250084859, dated 09 / 19 / 2025, pp. 186 / 290 178 / 208 42. The hyper-branched macromolecule for use or the treatment method according to aspects 39 to 41, wherein the hyper-branched macromolecule is used in the treatment of an eye disease, such as diseases of the fundus of the eye, such as any eye disease of the posterior segment affecting the vasculature and integrity of the retina, macula or choroid, leading to visual acuity disturbances, vision loss or blindness, particularly posterior segment disease states resulting from age, trauma, surgical interventions, such as age-related macular degeneration (AMD), cystoid macular edema (CME), diabetic macular edema (DME), posterior uveitis and diabetic retinopathy. 43. The hyper-branched macromolecule for use or the treatment method according to aspects 39 to 42, wherein the hyper-branched macromolecule is used in the treatment of a selected eye disease from the group consisting of retinal neovascularization, choroidal neovascularization, wet AMD, dry AMD, retinal vein occlusion, diabetic macular edema, retinal degeneration, hyphema, presbyopia, corneal graft rejection, retinoblastoma, melanoma, miosis, mydriasis, glaucoma, conjunctivitis, intraocular infections, choroidal neovascularization (CNV), intraocular tumors, retinal neuroinflammation, inflammation, autoimmune uveitis, uveitis, proliferative vitreoretinopathy and corneal degeneration, acute and chronic macular neuroretinopathy, central serous chorioretinopathy, macular edema, acute multifocal placoid pigment epitheliopathy, disease of Behçet's disease, bird-lead retinochoroidopathy, posterior uveitis, scleritis, serpiginous choroiditis, subretinal fibrosis.uveitis syndrome, Vogt-Koyanagi-Harada syndrome, retinal artery occlusive disease, central retinal vein occlusion, disseminated intravascular coagulopathy, retinal vein branch occlusion, hypertensive fundus changes, ocular ischemic syndrome. Petition 870250084859, dated 09 / 19 / 2025, pp. 187 / 290 179 / 208 retinal arterial microaneurysms, Coats' disease, parafoveal telangiectasia, hemiretinal vein occlusion, papillophlebitis, carotid artery disease (CAD), matte branch angiitis, sickle cell retinopathy, angioid streaks, familial exudative vitreoretinopathy, Eales' disease, proliferative vitreous retinopathy, diabetic retinopathy, tumor-associated retinal disease, congenital retinal pigment epithelium hypertrophy (RPE), posterior uveal melanoma, choroidal hemangioma, choroidal osteoma, choroidal metastasis, combined hamartoma of the retina and retinal pigment epithelium, retinoblastoma, vasoproliferative tumors of the fundus, retinal astrocytoma, intraocular lymphoid tumors, myopic retinal degeneration, acute retinal pigment epithelium, glaucoma, endophthalmitis, retinitis cytomegalovirus, retinal cancers, retinitis pigmentosa, Leber congenital amaurosis, choroideremia, X-linked retinitis pigmentosa, vitelliform macular dystrophy,X-linked retinoschisis, CNGA3 achromatopsia, CNGB3 achromatopsia, LHON, Stargardt disease, Usher syndrome, Norrie disease, Bardet-Biedl syndrome, and red-green color blindness. 44. The hyper-branched macromolecule for use or the method of treatment according to any of aspects 39 to 43 wherein the hyper-branched macromolecule is formulated for direct injection into the treatment site of a patient, for example, by parenteral administration, intratumoral injection, injection into the eye, such as intravitreal, intracameral, subconjunctival, retrobulbar, subtenon, subretinal or suprachoroidal injections. 45. The hyper-branched macromolecule for use or the method of treatment according to any of aspects 39 to 44, wherein the hyper-branched macromolecule is administered by direct injection, by oral application, incorporated into gels or incorporated into implants. 46. ​​The hyper-branched macromolecule for use or the method of Petition 870250084859, dated 09 / 19 / 2025, pp. 188 / 290 180 / 208 treatment according to any of aspects 39 to 46, wherein the hyper-branched macromolecule comprises two or more different active agents in different dendrons or regions on the surface of the hyper-branched macromolecule. 47. The hyper-branched macromolecule for use or the treatment method according to aspect 46, for use in a combination therapy involving the administration of more than one active agent. Second set of modalities 1. A hyper-branched macromolecule, characterized by the fact that it comprises building blocks that include: a central unit with at least 3 connectivity options; a plurality of polymeric arms connected to the central unit in the c-connectivities, at least one of the polymeric arms being connected by a dendritic constitutional repeat unit, the dendritic constitutional repeat unit comprising a branching unit connected to at least two polymeric arms, each comprising a terminal group or being connected by a hydrolyzable link to a next dendritic constitutional repeat unit that may again be connected by a chemical link to other dendritic constitutional repeat units, the polymeric arms of the outermost dendritic constitutional repeat unit of the hyper-branched macromolecule, each comprising a terminal group; wherein the polymeric arms consist of polyethylene glycol (PEG) units. 2. The hyper-branched macromolecule according to the aspect 1, in which at least 10%, preferably about 20 to 100%, of the connections in the macromolecule can be cleaved by hydrolysis. Petition 870250084859, dated 09 / 19 / 2025, pages 189 / 290 181 / 208 3. A hyper-branched macromolecule that meets any of the previous criteria, in which the connections are not hydrolyzable. 4. A hyper-branched macromolecule, according to any of the preceding aspects, is a higher-generation G x hyper-branched macromolecule, with x being an integer from 1 to 10, defining the number of consecutively connected repetitive constitutional dendritic units in the hyper-branched macromolecule. 5. A hyper-branched macromolecule according to any of the preceding aspects, in which the central unit and the branching unit are the same or different and, independently of each other, have a c or c' connectivity of 3 to 10, or 4 to 8, or 4 to 6, or 4. 6. A hyper-branched macromolecule according to any of the above aspects, in which the central unit and the branched unit are the same or different, and are derived from a polyol with at least 3 hydroxyl groups. 7. The hyper-branched macromolecule according to its appearance. 6, wherein the polyol is selected from the group consisting of glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol or sorbitol. 8. A hyper-branched macromolecule according to any of the preceding aspects, wherein the polyethylene glycol (PEG) units of the polymeric arms have an average molecular weight (Mn) in the range of about 1,000 to about 100,000 Daltons, or about 10,000 to about 60,000 Daltons, or about 15,000 to about 50,000 Daltons, or about 10,000 to about 40,000 Daltons. 9. A hyper-branched macromolecule according to any of the preceding aspects, in which the average molecular weight of the PEG units of the core-linked polymeric arm is the same as or different from that of the polymeric arms in the dendritic constitutional repeat units. Petition 870250084859, dated 09 / 19 / 2025, pp. 190 / 290 182 / 208 10. A hyper-branched macromolecule according to any of the previous aspects, in which the average molecular weight of the PEG units of the core-linked polymeric arm is greater than that of the polymeric arms in the dendritic constitutional units. 11. A hyper-branched macromolecule according to any of the previous aspects, in which the average molecular weight of the PEG units of the core-linked polymeric arm is less than that of the polymeric arms in the dendritic constitutional units. 12. A hyper-branched macromolecule according to either of the preceding aspects, wherein for a higher-generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10, the average molecular weight of the PEG units of the polymer arm decreases from the innermost polymer arms to the outermost polymer arms; or wherein the average molecular weight of the PEG units of the polymer arm increases from the innermost polymer arms to the outermost polymer arms. 13. A hyper-branched macromolecule according to any of the preceding aspects, wherein at least one arm connected to the central unit or branching unit is connected to the dendritic constitutional unit by means of a difunctional ligand forming hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic, heteroaliphatic or aromatic or heteroaromatic group. 14. A hyper-branched macromolecule according to any of the preceding aspects, wherein the terminal groups attached to the outermost polymeric arms are grafted onto the ends of the polymeric arms directly, or by means of a difunctional linker comprising or forming hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a group Petition 870250084859, dated 09 / 19 / 2025, pp. 191 / 290 183 / 208 dicarboxamide, a functionalized aliphatic, heteroaliphatic, aromatic, or heteroaromatic group. 15. A hyper-branched macromolecule according to any of the preceding aspects, wherein the functional groups of the terminal groups and / or terminal linking groups attached to the outermost polymeric arms are functional groups selected from electrophiles, such as activated ester groups, such as succinimidyl esters, succinimidyl carbonates; nitrophenyl carbonates, aldehydes, ketones, acrylates, acrylamides, maleimides, vinyl sulfones, iodoacetamides, alkenes, alkynes, azides, norbornenes, epoxides, mesylates, tosylates, tresylates, cyanurates, orthopyridyl disulfides or halogens; nucleophiles, such as an amine, such as a primary amine, a hydroxyl group, an alcohol, a thiol, an azide and a carboxyl group; functional groups for click chemistry; functional groups for cycloadditions, such as 1,3-dipolar cycloadditions, [3+2] cycloadditions such as alkene-nitrone cycloadditions or alkyne-nitrone cycloadditions, [4+2] cycloadditions;Functional groups for thiol-ene reactions; hetero-Diels-Alder cycloadditions; functional groups for nucleophilic ring openings; functional groups for non-aldol-type carbonyl reactions; functional groups for addition reactions to multiple carbon-carbon bonds; polymerizable vinyl groups or combinations thereof. 16. A hyper-branched macromolecule according to any of the preceding aspects, wherein the terminal groups (ligands) attached to the outermost polymeric arms are functional groups selected from succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ), and succinimidyl glutaramide (SGA). 17. A hyper-branched macromolecule according to any of aspects 1 to 14, in which the terminal groups are attached to the arms. Petition 870250084859, dated 09 / 19 / 2025, pages 192 / 290 184 / 208 outermost polymeric functional groups are selected from an alkyne compound, such as a dibenzocyclo-octyne (DBCO), or a bicyclo[6.1.0]-nonyne (BCN); or a norbornene, or a trans-cyclooctene (TCO); an azide, a 3,4-dihydroxyphenylacetic acid (DHPA) or a tetrazine (Tz). 18. A hyper-branched macromolecule according to any of the preceding aspects, wherein the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed by click chemistry, optionally with click chemistry functionalized ligands that include a hydrolyzable linkage. 19. The hyper-branched macromolecule according to the aspect 18, wherein the connection is formed by the reaction of a functionalized polymer arm, optionally via a ligand, with a tense or terminal alkyne, cycloalkyne, or alkene moiety with a polymer arm functionalized with an azide or tetrazine moiety in a SPAAC or IEDDA type click chemistry coupling reaction. 20. The hyper-branched macromolecule according to the aspect 19, where the alkyne portion is a dibenzocyclooctyne portion. 21. A hyper-branched macromolecule according to either aspect 18 or 20, wherein the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed between the polymeric arms connected to the central unit and the polymeric arms connected to the branching unit of the dendritic constitutional repeat units and / or between the polymeric arms of a dendritic constitutional repeat unit and the polymeric arms of dendritic constitutional repeat units. Petition 870250084859, dated 09 / 19 / 2025, pages 193 / 290 185 / 208 consecutive, optionally by means of a dysfunctional linker forming at least one hydrolyzable linkage. 22. A dendritic constitutional repeating unit represented by Formula (i), or the hyper-branched macromolecule according to any of the preceding aspects, including the dendritic constitutional repeating unit represented by Formula (i): wherein A is a connection to a polymeric arm that is connected to the central unit, or A is a connection to B of a preceding dendritic constitutional repeating unit represented by Formula (i), La is a ligand, m is any 0 or 1, n is an integer from 20 to 2000, o is an integer from 20 to 2000, neo may be different or equal, X is a branching unit having a c' connectivity, Lb is a linker, p is either 0 or 1, B comprises a terminal group located on the surface of the hyper-branched macromolecule or is a connection with A of a consecutive dendritic constitutional repeat unit or an active agent. La and Lb can be different or the same, mep can be different or the same, ey ​​is an integer from 2 to 9, where y = c' - 1, with c' being the connectivity c' of the branching unit X; and where the dendritic constitutional units in hyper-branched macromolecules can be the same or different. 23. A hyper-branched macromolecule according to aspect 22, in which the connection between A and B comprises a functional group formed by click chemistry, such as a triazole or dihydropyrazine. Petition 870250084859, dated 09 / 19 / 2025, pp. 194 / 290 186 / 208 24. A hyperbranched macromolecule according to aspects 22 or 23, wherein the La and / or Lb ligand comprises a diacid and / or an acidic diamido group, such as succinate, glutarate, adipate, azelate or glutaramide. 25. The hyper-branched macromolecule according to either of aspects 22 to 24, in which the La and / or Lb ligand comprises a structure represented by Formula (ii): <|Ί)1o where U1 and U2 are independently NH or O and may be equal or different, and where t is an integer from 0 to 10. 26. The hyper-branched macromolecule according to aspects 24 or 25, wherein the La and / or Lb ligand further comprises a polyethylene glycol unit between the linkage to B and the carboxyl group, carboxamide group or structure of Formula (ii). 27. A hyper-branched macromolecule according to any of the preceding aspects, wherein the hyper-branched macromolecule further comprises at least one extender unit comprising polyethylene glycol (PEG) units, wherein the extender unit is linear, difunctional and connected to the polymeric arm of a dendritic constitutional repeat unit or to the polymeric arm connected to the central unit and to a terminal group or to a polymeric arm of a further dendritic constitutional repeat unit. 28. A hyper-branched macromolecule according to aspect 27, wherein the extender unit comprises at least one ligand, wherein the ligand may be located at either or both ends of the extender unit and is a difunctional ligand comprising hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic group, or a heteroaliphatic group. Petition 870250084859, dated 09 / 19 / 2025, pp. 195 / 290 187 / 208 either aromatic or heteroaromatic. 29. A hyper-branched molecule according to any of the preceding aspects, wherein at least one, or all, preferably all, of the building blocks selected from among the central unit, central unit including polymeric arms in the c-connectivities, dendritic constitutional repeat unit, ligands and extenders, between hydrolyzable linkages, have a molecular weight less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons. 30. A hyper-branched molecule according to any of the preceding aspects, in which, after complete hydrolysis of the hydrolyzable bonds, all the fragments formed from the molecule have a molecular weight less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons. 31. A precursor to a dendritic constitutional repeating unit comprising: - a polymeric arm comprising a functional group suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine), and - at least two polymeric arms comprising non-reactive functional groups in click chemistry, wherein the polymeric arms are connected to a branching unit with a c' connectivity, wherein the polymeric parts of the polymeric arms consist of polyethylene glycol (PEG) units. 32. The precursor of aspect 31, in which the compound is represented by Formula (iii): Petition 870250084859, dated 09 / 19 / 2025, pages 196 / 290 188 / 208 where C comprises a functional group suitable for click chemistry (such as an alkyne, alke...

Claims

1. A hyper-branched macromolecule, characterized in that it comprises building blocks comprising: a central unit with at least 3 c-connectivities; a plurality of polymeric arms connected to the central unit at the c-connectivities, at least one of the polymeric arms being connected by a hydrolyzable bond to a dendritic constitutional repeat unit, the dendritic constitutional repeat unit comprising a branching unit connected to at least two polymeric arms, each comprising a terminal group or being connected by a hydrolyzable bond to a next dendritic constitutional repeat unit that may again be connected by a chemical bond to other dendritic constitutional repeat units, the polymeric arms of the outermost dendritic constitutional repeat unit of the hyper-branched macromolecule, each comprising a terminal group;wherein the polymeric arms consist of polyethylene glycol (PEG) units; wherein at least one active agent is conjugated to at least one of the outermost polymeric arms, and wherein the central unit and the branched unit are the same or different and are derived from a polyol with at least 3 hydroxyl groups.

2. Hyper-branched macromolecule, according to claim 1, characterized in that at least 10%, preferably about 20 to 100%, of the connections in the macromolecule can be cleaved by hydrolysis. Petition 870250084859, dated 09 / 19 / 2025, pp. 240 / 290 2 / 15 3. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that each of the building blocks (fragments) of the hyper-branched macromolecule obtained after cleavage of all hydrolyzable bonds of the connections in the macromolecule has an average molecular weight (Mn) of less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons.

4. A hyper-branched macromolecule, according to any of the preceding claims, characterized in that it is a higher-generation G x hyper-branched macromolecule, with x being an integer from 1 to 10, defining the number of consecutively connected repetitive constitutional dendritic units in the hyper-branched macromolecule.

5. Hyper-branched macromolecule, according to claim 1, characterized in that the polyol is selected from the group consisting of glycerol, pentaerythritol, xylitol, dipentaerythritol, tripentaerythritol, hexaglycerol, isomalt, lactitol, maltitol, mannitol or sorbitol.

6. A hyper-branched macromolecule, according to any of the preceding claims, characterized in that the average molecular weight of the PEG units of the core-linked polymeric arm is greater than that of the polymeric arms in the dendritic constitutional units, or in that the average molecular weight of the PEG units of the core-linked polymeric arm is less than that of the polymeric arms in the dendritic constitutional units.

7. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that for a higher generation Gx hyper-branched macromolecule, with Petition 870250084859, dated 09 / 19 / 2025, page 241 / 290 3 / 15 x being an integer from 2 to 10, the average molecular weight of the PEG units of the polymer arm decreases from the innermost polymer arms to the outermost polymer arms; or in that the average molecular weight of the PEG units of the polymer arm increases from the innermost polymer arms to the outermost polymer arms.

8. A hyper-branched macromolecule, according to any of the preceding claims, characterized in that at least one arm connected to the central unit or branching unit is connected to the dendritic constitutional unit by means of a difunctional ligand forming hydrolyzable linkages comprising a carboxyl group, a dicarboxyl group, a carboxamide group, a dicarboxamide group, a functionalized aliphatic, heteroaliphatic or aromatic or heteroaromatic group.

9. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that the terminal linking groups attached to the outermost polymeric arms are functional groups selected from succinimidyl succinate (SS), succinimidyl glutarate (SG), succinimidyl adipate (SAP), succinimidyl azelate (SAZ) and succinimidyl glutaramide (SGA), or in that the terminal groups attached to the outermost polymeric arms are functional groups selected from an alkyne compound, such as a dibenzocyclooctyne (DBCO), or a bicyclo[6.1.0]-nonyne (BCN); or a norbornene, or a trans-cyclooctene (TCO); an azide, a 3,4-dihydroxyphenylacetic acid (DHPA) or a tetrazine (Tz).

10. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that the connection between the polymeric arms connected to the central unit and the first dendritic constitutional repeat unit and / or the connections between consecutive dendritic constitutional repeat units are formed by click chemistry, optionally with click chemistry functionalized ligands that include a hydrolyzable linkage.

11. A hyper-branched macromolecule, according to any of the preceding claims, characterized in that the active agent conjugated to at least one of the outermost polymeric arms is selected from steroids; non-steroidal anti-inflammatory drugs (NSAIDs), such as diclofenac, ibuprofen, meclofenamate, mefanamic acid, salsalate, sulindac, tolmetin, ketoprofen, diflunisal, piroxicam, naproxen, etodolac, flurbiprofen, fenoprofen C, indomethacin, celecoxib, ketorolac, nepafenac; intraocular pressure-reducing drugs; antibiotics, such as ciprofloxacin; analgesics, such as bupivacaine; calcium channel blockers, such as nifedipine; cell cycle inhibitors, such as simvastatin; proteins, such as insulin; hydrophilic small molecule drugs, including carboxylic acid salts and amine salts;hydrophobic small molecule drugs, hydrophilic peptides and protein drugs, such as insulin, single-chain antibody fragments, Fab fragments, IgG antibodies, fusion antibodies, etc.; aptamers;particularly bupivacaine (BPV-HCl or base), ropivacaine (RPV), dexamethasone, travoprost, axitinib, non-steroidal anti-inflammatory drugs (NSAIDs), steroids, antibiotics, analgesics, calcium channel blockers, cell cycle inhibitors, chemotherapeutic agents, antiviral drugs, anesthetics, hormones, anticancer drugs, antineoplastic agents, viruses, gene delivery viruses such as AAVs, protein ligands such as nanobodies, affibodies, ankyrins, DARPins, etc., or any combination thereof, and / or wherein the active agent conjugated to at least one of the outermost polymeric arms is a peptide or aptamer selected from Petition 870250084859, dated 19 / 09 / 2025, p. 243 / 290 5 / 15 group consisting of Compstatin, APL-1 and Fc-lll-4C, Beovu (Brolucizumab), Zimura (Avacincaptad Pegol), Pegcetacoplan, Abicipar Pegol, Lampalizumab, Fovista, Risuteganib, AXT107, Elamipretide, THR149, ALM201, VGB3 and Largazole; 12. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that the active agent is linked to at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or even 100% of the outermost polymeric arms.

13. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that a dendritic constitutional repeat unit is represented by Formula (i): wherein A is a connection to a polymeric arm that is connected to the central unit, or A is a connection to B of a preceding dendritic constitutional repeat unit represented by Formula (i), La is a ligand, m is any 0 or 1, n is an integer from 20 to 2000, o is an integer from 20 to 2000, neo may be different or the same, X is a branching unit having a c' connectivity, Lb is a ligand, p is any 0 or 1, B comprises a terminal group located on the surface of the hyper-branched macromolecule or is a connection to A of a consecutive dendritic constitutional repeat unit or an active agent, La and Lb may be different or the same, Petition 870250084859, of 19 / 09 / 2025, page.244 / 290 6 / 15 mep can be different or the same, ey ​​is an integer from 2 to 9, where y = c' - 1, with c' being the connectivity c' of the branching unit X; and where the dendritic constitutional units in hyper-branched macromolecules can be the same or different.

14. Hyper-branched macromolecule, according to claim 13, characterized in that the connection between A and B comprises a functional group formed by click chemistry, such as a triazole or dihydropyrazine; and / or in that the La and / or Lb ligand comprises a diacid and / or an acidic diamido group, such as succinate, glutarate, adipate, azelate or glutaramide.

15. Hyper-branched macromolecule, according to any of claims 13 to 14, characterized in that the La and / or Lb ligand comprises a structure represented by Formula (ii): 1 o wherein U1 and U2 are independently NH or O and may be the same or different, and wherein t is an integer from 0 to 10.

16. Hyper-branched macromolecule, according to any of the preceding claims, characterized in that the hyper-branched macromolecule further comprises at least one extender unit comprising polyethylene glycol (PEG) units, wherein the extender unit is linear, difunctional and connected to the polymeric arm of a dendritic constitutional repeat unit or to the polymeric arm connected to the central unit and to a terminal group or to a polymeric arm of a next dendritic constitutional repeat unit.

17. Hyper-branched molecule, according to any one of the preceding claims, characterized in that at least one, or all, preferably all, of the building blocks selected from among the central unit, central unit including polymeric arms in the c-connections, dendritic constitutional repeat unit, linkers and extenders, between hydrolyzable linkages, have a molecular weight less than 50,000 Daltons, such as less than 45,000 Daltons, or less than 40,000 Daltons, or less than 35,000 Daltons, or less than 30,000 Daltons.

18. A dendritic constitutional repeating unit precursor, characterized in that it comprises: a polymeric arm comprising a functional group suitable for forming a click chemistry connection (such as an azide, alkyne, alkene or tetrazine), and at least two polymeric arms comprising non-reactive functional groups in click chemistry, wherein the polymeric arms are connected to a branching unit with a c' connectivity, wherein the polymeric parts of the polymeric arms consist of polyethylene glycol (PEG) units; and wherein the compound is represented by Formula (iii): wherein C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine), D comprises non-reactive functional groups in click chemistry (such as succinimidyl), and La, m, η, X, o, Lb, pey are as defined in any one of claims 13 to 15.

19. Precursor of reverse dendritic constitutional repeating unit, characterized by the fact that it comprises: a polymeric arm comprising a functional group. Petition 870250084859, dated 09 / 19 / 2025, p. 246 / 290 8 / 15 non-reactive in click chemistry, and at least two polymeric arms comprising functional groups suitable for click chemistry (such as an azide, alkyne, alkene or tetrazine), and a branching unit having a c' connectivity, wherein the polymeric parts of the polymeric arms consist of polyethylene glycol (PEG) units, and wherein the precursor is represented by Formula (iv): wherein C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine), D comprises non-reactive functional groups in click chemistry (such as succinimidyl), and La, m, η, X, o, Lb, pey are as defined in the preceding claims.

20. Method for fabricating a hyperbranched macromolecule as defined in any one of claims 1 to 19 by divergent synthesis, characterized in that it comprises the following steps: (a) providing a central unit having at least 3 c-connectivities, a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry at the terminations of the polymeric arms; (b) providing dendritic constitutional repeat unit precursors comprising a polymeric arm comprising a functional group suitable for forming a click chemistry connection with the corresponding functional groups of the polymeric arms connected to the core (such as an azide, alkyne, alkene or tetrazine), and at least two polymeric arms comprising functional groups that are not reactive in click chemistry,(c) establishing a connection via chemical click between the polymeric arms connected to the core and the polymeric arms of the precursors of the dendritic constitutional repeating unit, (d) optionally converting the functional groups of at least two polymeric arms comprising non-reactive functional groups in click chemistry into functional groups suitable for click chemistry, and (e) conjugation of an active agent comprising a functional group to the outermost polymeric arms via reaction with the functional groups of the outermost polymeric arms, thereby forming a hyper-branched macromolecule-active agent conjugate, wherein for the higher-generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10,Step (d) is mandatory and consecutive precursors of additional dendritic constitutional repeating units are connected to the functional groups suitable for click chemistry obtained in step (d) by click chemistry to the hyper-branched macromolecule before conjugation of the active agent in step (e).

21. Method according to claim 20, characterized in that the precursor of the dendritic constitutional repeating unit in step (c) is represented by Formula (iii): wherein C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine), D comprises functional groups not reactive in click chemistry (such as succinimidyl), and La, m, η, X, o, Lb, pey are as defined in any of the preceding claims; and wherein Petition 870250084859, dated 09 / 19 / 2025, pp. 248 / 290 10 / 15 the dendritic constitutional units may be the same or different.

22. Method for fabricating a hyperbranched macromolecule as defined in any one of claims 1 to 19 by convergent synthesis, characterized in that it comprises the following steps: V) providing dendritic constitutional repeat unit precursors comprising - a polymeric arm comprising a functional group suitable for forming a connection by click chemistry (such as an azide, alkyne, alkene or tetrazine), and - at least two polymeric arms comprising functional groups that are not reactive in click chemistry, VI) conjugating active agents comprising a functional group to at least one of the at least two polymeric arms comprising functional groups that are not reactive in click chemistry of the dendritic constitutional repeat unit precursors, VII) providing a central unit having at least 3 c-connectivities,a plurality of polymeric arms connected to the central unit having functional groups suitable for click chemistry (such as an azide, alkyne, alkene or tetrazine) at the ends of the polymeric arms, and V III) forming a click chemistry connection between the polymeric arms connected to the core provided in step III) and the polymeric arm comprising a functional group suitable for forming a click chemistry connection of the precursors of conjugated dendritic constitutional repeating units to the active agent obtained in step II), thus forming a hyper-branched active agent macromolecule conjugate.

23. Method according to claim 22, characterized in Petition 870250084859, dated 09 / 19 / 2025, pp. 249 / 290 11 / 15 by the fact that a precursor of the dendritic constitutional repeating unit in step I) is represented by Formula (iii): wherein C comprises a functional group suitable for click chemistry (such as an alkyne, alkene, azide or tetrazine), D comprises functional groups that are not reactive in click chemistry (such as succinimidyl), and La, Lb, m, η, X, o, pey are as defined in any of the preceding claims; and wherein the dendritic constitutional units may be the same or different; and / or wherein for the higher-generation hyper-branched macromolecule Gx, with x being an integer from 2 to 10,The precursors of conjugated dendritic constitutional repeat units to the active agent obtained in step II) are connected by click chemistry to precursors of reverse dendritic constitutional repeat units comprising: a polymeric arm comprising a non-reactive functional group in click chemistry, and at least two polymeric arms comprising functional groups suitable for click chemistry (such as an azide, alkyne, alkene, or tetrazine), wherein the non-reactive functional group in click chemistry of the polymeric arm is subsequently converted into a functional group suitable for click chemistry before connecting to other precursors of reverse dendritic constitutional repeat units or before forming a connection by click chemistry with the polymeric arms connected to the core in step IV), thus forming higher-generation hyper-branched macromolecules.

24. Method according to claims 22 to 23, Petition 870250084859, dated 09 / 19 / 2025, pp. 250 / 290 12 / 15 characterized in that precursors of dendritic constitutional repeat units having different active agents conjugated to the polymeric arms are obtained by performing steps I) and II) for each precursor of dendritic constitutional repeat unit conjugated to the active agent, and a mixture of the precursors of dendritic constitutional repeat units conjugated to the active agent obtained is used for step IV), thus forming a hyper-branched macromolecule-active agent conjugate having different active agents in different regions of the surface of the hyper-branched macromolecule.

25. A method according to any one of claims 20 to 24, characterized in that the outermost polymeric arms of the hyper-branched macromolecule have terminal maleimide functional groups, and peptides or active agents are conjugated to them via the maleimide-thiol reaction, and wherein the terminal maleimide functional groups are provided by the reaction of the DBCO- or azide-functionalized terminal functional group of the hyper-branched macromolecule with click chemistry ligands having an azide or DBCO functionality connected to a maleimide group, such as DBCO-maleimide, DBCO-PEG3-maleimide, DBCO-PEG4-maleimide or azido-PEG3-maleimide.

26. Hyper-branched macromolecule as defined in any one of claims 1 to 19, characterized in that it is for use as a medicament.

27. A treatment method for treating a disease or medical condition in a patient, characterized by the fact that it is with a hyper-branched macromolecule as defined in any one of claims 1 to 19.

28. Hyper-branched macromolecule for use, or treatment method, according to claim 26 or 27, characterized by the fact that the hyper-branched macromolecule is used for an eye treatment.

29. Hyper-branched macromolecule for use, or treatment method, according to any of claims 26 to 28, characterized in that the hyper-branched macromolecule is used in the treatment of an eye disease, such as diseases of the fundus of the eye, such as any eye disease of the posterior segment affecting the vasculature and integrity of the retina, macula or choroid, leading to visual acuity disturbances, vision loss or blindness, particularly posterior segment disease states resulting from age, trauma, surgical interventions, such as age-related macular degeneration (AMD), cystoid macular edema (CME), diabetic macular edema (DME), posterior uveitis and diabetic retinopathy; and / or in which the hyper-branched macromolecule is used in the treatment of a selected eye disease from the group consisting of retinal neovascularization, choroidal neovascularization, wet AMD, dry AMD, retinal vein occlusion, diabetic macular edema, retinal degeneration,Hyphema, presbyopia, corneal graft rejection, retinoblastoma, melanoma, miosis, mydriasis, glaucoma, conjunctivitis, intraocular infections, choroidal neovascularization (CNV), intraocular tumors, retinal neuroinflammation, inflammation, autoimmune uveitis, uveitis, proliferative vitreoretinopathy and corneal degeneration, acute and chronic macular neuroretinopathy, central serous chorioretinopathy, macular edema, acute multifocal placoid pigment epitheliopathy, Behçet's disease, bird-lead retinochoroidopathy, posterior uveitis, scleritis, serpiginous choroiditis, subretinal fibrosis, uveitis syndrome, Vogt-Koyanagi-Harada syndrome, retinal artery occlusive disease, central retinal vein occlusion, disseminated intravascular coagulopathy, retinal vein branch occlusion, hypertensive disorders from the fundus, ocular ischemic syndrome, retinal arterial microaneurysms, Coats' disease, parafoveal telangiectasia, Petition 870250084859, dated 09 / 19 / 2025, pp. 252 / 290 14 / 15occlusion of the hemiretinal vein, papillophlebitis, carotid artery disease (CAD), matte branch angiitis, sickle cell retinopathy, angioid streaks, familial exudative vitreoretinopathy, Eales disease, proliferative vitreous retinopathy, diabetic retinopathy, tumor-associated retinal disease, congenital hypertrophy of the retinal pigment epithelium (RPE), posterior uveal melanoma, choroidal hemangioma, choroidal osteoma, choroidal metastasis, combined hamartoma of the retina and retinal pigment epithelium, retinoblastoma, vasoproliferative tumors of the fundus, retinal astrocytoma, intraocular lymphoid tumors, myopic retinal degeneration, acute retinal pigment epithelium, glaucoma, endophthalmitis, cytomegalovirus retinitis, retinal cancers, retinitis pigmentosa, Leber congenital amaurosis, choroideremia, retinitis X-linked pigmented dystrophy, vitelliform macular dystrophy, X-linked retinoschisis, CNGA3 achromatopsia, CNGB3 achromatopsia, LHON.Stargardt disease, Usher syndrome, Norrie disease, Bardet-Biedl syndrome, and red-green color blindness.

30. Hyper-branched macromolecule for use, or treatment method, according to any one of claims 26 to 29, characterized in that the hyper-branched macromolecule is formulated for direct injection into a patient's treatment site, for example, by parenteral administration, intratumoral injection, injection into the eye, such as intravitreal, intracameral, subconjunctival, retrobulbar, subtenon, subretinal or suprachoroidal injections, and / or wherein the hyper-branched macromolecule is administered by direct injection, by oral application, incorporated into gels or incorporated into implants.

31. Hyper-branched macromolecule for use, or treatment method, according to any of claims 26 to 30, Petition 870250084859, dated 09 / 19 / 2025, pp. 253 / 290 15 / 15 characterized in that the hyper-branched macromolecule comprises two or more different active agents in different dendrons or regions on the surface of the hyper-branched macromolecule.

32. Hyper-branched macromolecule, according to any one of claims 1 to 19, hyper-branched macromolecule for use, or treatment method, according to any one of claims 26 to 31, characterized in that the hyper-branched macromolecule comprises an active agent at different positions or regions on the surface of the hyper-branched macromolecule with different hydrolyzable groups to vary the release of the active agent at different rates, or in that the hyper-branched macromolecule comprises two or more different active agents at different positions or regions on the surface of the hyper-branched macromolecule with different hydrolyzable groups to vary the release of the same or different active agents at different rates.

33. Use of a hyper-branched acromolecule as defined in any one of claims 1 to 17, or of a dendritic constitutional repeat unit precursor as defined in claim 18 or 19, characterized in that it is for the manufacture of a medicament for use in therapy.

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