WO2016069760A1

WO2016069760A1 - Method of treating dementia by intranasal administration of vegf gene therapy

Info

Publication number: WO2016069760A1
Application number: PCT/US2015/057841
Authority: WO
Inventors: Steven Yu
Original assignee: Steven Yu
Priority date: 2014-10-31
Filing date: 2015-10-28
Publication date: 2016-05-06

Abstract

Treatment of dementia (e.g. Alzheimers disease) using a polynucleotide agent (e.g. DNA or RNA) that contains a nucleotide sequence that encodes a vascular endothelial growth factor (VEGF) polypeptide. The polynucleotide agent is administered into the patients nose such that it is transported to the brain through or by way of neural pathways associated with the olfactory nerve. The polynucleotide agent may be contained inside a carrier vector, such as a viral vector.

Description

Method of Treating Dementia by Intranasal

Administration of VEGF Gene Therapy

Cross-References

This application claims the benefit of U.S. Provisional Applications No.

62/073,451 filed on 31 October 2014 and No. 62/157,533 filed on 6 May 2015, both of which are incorporated by reference herein in their entirety.

Technical Field

My invention relates to the treatment of dementia, such as Alzheimer's disease

Background

Alzheimer's disease is associated with reduced blood flow to the brain along with a reduction in proangiogenic factors, such as vascular endothelial growth factor (VEGF) and transforming growth factor-βΐ (TGF-βΙ]. This suggests that vascular dysfunction plays an important role in the development of Alzheimer's dementia. Focusing on the role of VEGF, this angiogenic factor not only promotes cerebral neovascularization, it has also been shown to have a direct neuroprotective effect on neural cells.

Because crossing the blood-brain barrier is a major obstacle for delivering any therapeutic agent to the brain, there has been growing interest in the intranasal route and the olfactory nerve as a pathway for direct delivery to the brain. The olfactory epithelium receives an extension of the central nervous system (CNS] via the olfactory nerve. Fibers of the olfactory nerve are unmyelinated axons of olfactory receptor cells that are located in the very top (i.e. superior one-third] of the nasal cavity just under the cribiform plate of the ethmoid bone that separates the nasal and cranial cavities. The dendrites of these sensory neurons extend into the nasal cavity, and the axons collect into nerve bundles that project to the olfactory bulb. See FIG. 1. This understanding of the anatomical relationships has lead to prior work observing that drugs and other therapeutic agents can be delivered to the brain through this olfactory pathway.

Summary

My invention relates to the treatment of dementia, which can result from various diseases that affect the brain, including ischemic brain disease, Alzheimer's disease

(including mild cognitive impairment], Parkinson's disease, multiple sclerosis, etc. The method involves using a polynucleotide agent [e.g. DNA or RNA) that contains a nucleotide sequence that encodes a vascular endothelial growth factor (VEGF) polypeptide. The treatment method comprises administering the polynucleotide agent into a patient's nose and having the polynucleotide agent transported to the brain through or by way of neural pathways associated with the olfactory nerve.

In my invention, the polynucleotide agent is administered intranasally so that it is deposited onto the olfactory mucosa of the nose. From there, the polynucleotide agent is transported along the olfactory nerve to the brain. The polynucleotide agent may be transported along the olfactory nerve by one or more of several possible routes. For example, the polynucleotide agent may be taken up into the olfactory neurons and taken along by intracelluar axonal transport mechanisms. Another possibility is that extracellular or paracellular transport mechanisms may be involved, such as through intercellular clefts in the olfactory neuroepithelium, bulk flow and diffusion within perineuronal channels (e.g. lymphatic) or the perivascular space of a blood vessel running with a neuron or neural pathway.

The target area for applying the polynucleotide agent in the patient's nose can be characterized in various ways. In general, the polynucleotide agent can be applied to the olfactory region of the nasal cavity. The olfactory region has the olfactory epithelium and is generally located in the upper one -third of the nasal cavity.

Via transport along the olfactory neural pathway, the polynucleotide agent reaches the brain (e.g. the olfactory bulb located on the inferior side of the brain). The olfactory bulb has widespread connections with various regions of the brain including the amygdala, orbitofrontal cortex, and hippocampus. As such, the polynucleotide agent may disperse further to other areas of the brain.

Brief Description of the Drawings

FIG. 1 depicts a sagittal section of the human nasal cavity, showing the the nasal vestibule (A); atrium (B); respiratory region— inferior turbinate (CI), middle turbinate (C2), superior turbinate (C3); olfactory region (D); and nasopharynx (E) . Taken from M.M. Wen et al., "Olfactory Targeting Through Intranasal Delivery of Biopharmaceutical Drugs to the Brain - Current Development" in Discovery Medicine, 2011 Jun, 11(61):497- 503 Detailed Description

1. VEGF Polypeptide. The polynucleotide agent of my invention contains a nucleotide sequence that encodes a VEGF polypeptide, which can be a naturally occurring VEGF polypeptide or a functionally active polypeptide analogue thereof. In some embodiments, the polynucleotide agent is a DNA agent (deoxyribonucleic acid). In some embodiments, the polynucleotide agent is an RNA agent (ribonucleic acid). The polynucleotide agent may be made by any suitable technique.

1(a). Naturally Occurring VEGF Polypeptide. A "naturally occurring VEGF polypeptide" means a polypeptide from any member of the human vascular endothelial growth factor (VEGF) family, including VEGF-A (conventionally referred to as VEGF), VEGF-B, VEGF-C, VEGF-D, VEGF-E, and VEGF-F. Another member of this family is placenta growth factor (P1GF), which is closely related to VEGF-A. For a naturally occurring VEGF polypeptide, the polynucleotide agent encodes a polypeptide having the same amino acid sequence as a naturally occurring VEGF, i.e. having a native sequence. This definition includes naturally occurring variant forms such as isoforms or alternatively spliced forms. For example, isoforms of VEGF-A include VEGF-Ai65 and VEGF-A121. The native amino acid sequence of the various members of the VEGF family and their natural variants are well known.

1(b). Polypeptide Analogues. A "functionally active polypeptide analogue" of a naturally occurring VEGF polypeptide differs from the naturally occurring VEGF polypeptide, but retains its essential biologic functional properties. Being functionally active means that the polypeptide analogue is capable of binding to and activating a VEGF receptor, such as VEGFR-1 (also known as Flt-1), VEGFR-2 (also known as Flk- 1/KDR), and/or VEGFR-3 (also known as Flt-4). Functional activity can also be indicated by causing proliferation of endothelial cells, presumably through VEGF receptor activation.

Generally, functionally active polypeptide analogues are overall closely similar, and in many regions, identical (i.e. homologous) to the naturally occurring VEGF polypeptide. Analogues of the VEGF polypeptide may be full length or other than full length. The VEGF polypeptide analogue may differ from its native sequence counterpart by any combination of deletion(s), insertion(s), and/or substitution (s) of amino acid residues. Such VEGF polypeptide analogues can be made by a variety of methods well known in the art, including recombinant DNA technologies.

1(c). Amino Acid Sequence Homology. For purposes of defining my invention, in some embodiments, a functionally active polypeptide analogue has at least 85% amino acid sequence identity; in some cases, at least 90% amino acid sequence identity; and in some cases, at least 95% amino acid sequence identity with a naturally occurring VEGF polypeptide (i.e. VEGF-A, VEGF-B, VEGF-C, etc.), or fragment thereof. This identity can be determined by comparing to an amino acid sequence of identical size or by comparing to an aligned sequence as performed by any of the various homology analysis tools known in the art (such as the "Wisconsin Package" of software programs). A VEGF polypeptide analogue can also differ from the naturally occurring counterpart by other modifications, such as glycosylation and other covalently or noncovalently introduced modifications.

1(d). Conserved Amino Acid Residues. All members of the VEGF family (naturally occurring) have been found to have a common VEGF homology domain, which has been shown to be required for biological activity of VEGF. As such, in some embodiments of my invention, the VEGF polypeptide encoded by the polynucleotide agent comprises a VEGF homology domain that has at least 85% amino acid sequence identity with the VEGF homology domain of a naturally occurring VEGF polypeptide (i.e. VEGF-A, VEGF-B, VEGF-C, etc.); in some cases, at least 90% amino acid sequence identity; and in some cases, at least 95% amino acid sequence identity with the VEGF homology domain of a naturally occurring VEGF polypeptide (i.e. VEGF-A, VEGF-B, VEGF-C, etc.).

This VEGF homology domain creates a four-stranded β-sheet within each monomer. VEGF monomers dimerize in an antiparallel way. As such, in some

embodiments of my invention, the VEGF polypeptide encoded by the polynucleotide agent is monomer that can dimerize in an antiparallel way.

The VEGF homology domain contains a cystine knot motif, with eight cysteine residues involved in inter- and intramolecular disulfide bonds. See G Neufeld et al, "Vascular endothelial growth factor (VEGF) and its receptors" in FASEB J, vol 13: 9-22 (1999); N Ortega et al, "Signal relays in the VEGF system" in Front Biosci, vol 4: D141-

D152 (1999); A Hoeben et al, "Vascular Endothelial Growth Factor and Angiogenesis" in Pharmacological Reviews, vol 56:4, pp 549-580 (Dec 2004). As such, in some embodiments of my invention, the VEGF polypeptide encoded by the polynucleotide agent comprises eight cysteine residues that are involved in inter- and intramolecular disulfide bonds. In some cases, of these eight cysteine residues, two are involved in inter-dimer disulfide bridging and six are involved in intra-monomer disulfide bridging.

Some VEGF variants having heparin-binding domains, which helps anchor them in the extracellular matrix for presentation to VEGF receptors. See Neufeld and Ortega citations. This heparin binding increases VEGF potency [i.e. the heparin-binding forms are more active). As such, in some embodiments of my invention, the VEGF polypeptide encoded by the polynucleotide agent comprises one or more heparin-binding domains. 2. Gene Therapy Using Polynucleotide Agent. The polynucleotide agent can be used in conjunction with various vector systems or materials for gene transfer. The vector system may be macromolecules or complex of molecules containing or carrying the polynucleotide agent for delivery into the target tissue cells. The polynucleotide agent may further contain components that influence expression (e.g. promoters, control sequences, etc.]. In some cases, the VEGF-coding sequence may be inserted into a plasmid, i.e. the polynucleotide agent is a DNA plasmid carrying the VEGF sequence. The plasmid may incorporate sequences that direct expression of the polypeptide. Any type of plasmid suitable for human gene therapy may be used.

In some embodiments, the polynucleotide agent is not carried by a vector system, e.g. the DNA agent may be administered as "naked" DNA. In some embodiments, the polynucleotide agent may be contained inside a carrier vector, which may be viral or non-viral. Examples of carrier vectors that can be used in gene therapy include liposomes, nanoparticles, and viral vectors. In some cases, the carrier vector is a viral vector derived from a virus, such as adenovirus, adeno -associated virus (AAV], retroviruses, lentivirus, or herpes viruses. These viral vectors may carry DNA or RNA

(e.g. retroviruses carrying RNA]. Carrier vectors based on the herpes simplex virus (e.g. HSV-1 or HSV-2] may be particularly useful because they are naturally neurotropic. Examples of non-viral carrier vectors include liposomes and other lipid-containing complexes, nanoparticles, and other macromolecular complexes capable of mediating delivery of a polynucleotide into a host cell.

The carrier vectors can also comprise other components or functionalities that influence gene delivery and/or gene expression. Such other components include, for example, components that influence binding or targeting to cells (including components that mediate cell-type or tissue-specific binding); components that influence uptake of the polynucleotide agent by the cell; and components that influence expression of the polynucleotide agent (e.g. promoters, control sequences, etc.].

3. Pharmaceutical Composition. The polynucleotide agent may be formulated into any suitable form of pharmaceutical composition, including liquid, solid, or semisolid forms. Examples of pharmaceutical compositions in liquid form include solutions, emulsions, suspensions, colloids, etc. In liquid form, the pharmaceutical composition can be administered as nasal drops or nasal spray. Examples of pharmaceutical compositions in semi-solid form include ointments, salves, gels, or creams. Examples of pharmaceutical compositions in solid form include powders or tablets. The

pharmaceutical composition may include components that influence uptake of the polynucleotide agent by the cell, such as calcium phosphate (calcium chloride in a solution of phosphate ions), cyclodextrin, and cationic polymers such as DEAE-dextran or polyethylenimine.

3(a). Permeation Enhancers. The polynucleotide agent may be mixed with a permeation enhancer, which are materials that increases penetration across the olfactory epithelium and/or mucosa. The polynucleotide agent can be mixed with various types of permeation enhancing materials that work through various different effects.

Examples of different types of permeation enhancers include mucoadhesives, ciliary beat inhibitors, mucous fluidizers, membrane fluidizers, and tight junction modulators. Examples of materials that act as membrane fluidizers or tight junction modulators include solvents such as propylene glycol, glycofurol, and glycerol. Further examples include surfactants or emulsifiers such as glycocholate, taurocholate, tauroursodeoxycholate, and bile salts. Further examples of surfactants include alkyl polyglycosides such as decyl maltosides, dodecyl maltosides, and tetradecyl maltosides. Further examples of surfactants include lipids such as lysophopsphatidylcholine, DDPC (l,2-didecanoyl-sn-glycero-3-phosphocholine), and fatty acids such as oleic acid.

Further examples of permeation enhancers are listed in Table A and Table 3 of US 2009/0136505 (Bentz et al.), which is incorporated by reference herein. Further examples include the penetration enhancers described in M.M. Wen, "Olfactory Targeting Through Intranasal Delivery of Biopharmaceutical Drugs to the Brain— Current Development" in Discovery Medicine, vol 11:61, pp 497-503 (June 2011) (incorporated by reference herein), such as peppermint oil and N-tridecyl-beta-D- maltoside.

3(b). Retention Enhancers. The polynucleotide agent may be mixed with a retention enhancer, which are materials that increase the residence time of the polynucleotide agent on the nasal mucosa so that more of it can be absorbed. Examples of

mucoadhesives includes bioadhesive polymers, chitosan, pectin, cellulose derivatives (e.g. hydroxypropyl methylcellulose, hydroxypropyl cellulose, and

carboxymethylcellulose), or starch or lactose microspheres.

Further examples include the mucoadhesive formulations described in M.M. Wen, "Olfactory Targeting Through Intranasal Delivery of Biopharmaceutical Drugs to the Brain— Current Development" in Discovery Medicine, vol 11:61, pp 497-503 (June 2011) (incorporated by reference herein), such as polymers of polyacrylic acid, carbopol, carboxymethylcellulose, hydroxylpropylmethyl cellulose, and hyaluronan.

Retention can also be enhanced by vasoconstricting drugs such as phenylephrine hydrochloride, which constricts blood flow and reduces carry away of the

polynucleotide agent.

4. Vascular Permeability. In addition to having an angiogenic effect, VEGF is also a potent inducer of vascular permeability. This hyperpermeability effect of VEGF may not be desirable in the brain where it could potentially cause harmful cerebral edema. As such, in some embodiments, the method further comprises intranasally

administering an agent that is a counteractant of vascular permeability. The

counteractant agent may be co-administered with the polynucleotide agent

simultaneously or sequentially in any order. The two agents may be combined in one pharmaceutical carrier or they may be placed in separate carriers and administered to the patient at different times. The term "co -administer" as used herein means that the two agents are administered sufficiently close in time that there is temporal overlap in the vascular hypermeability effect of the polynucleotide agent and the vascular permeability counteracting effect of the counteractant agent. The term "counteractant of vascular permeability" as used herein means a bioactive agent (whether small compound, peptide/protein, lipid, saccharide, polynucleotide, etc.) that has the effect of counteracting the vascular permeability induced by VEGF. Examples of such counteractants include agonist ligands of the sphingosine-l-phosphate receptors, including sphingosine-l-phosphate (SIP) sphingolipid; agonist ligands of the Roundabout (Robo) transmembrane receptor, including Slit proteins of the Slit-Robo cell signaling pathway; Bbetal5-42 (also known as FX006), which is a fibrin-derived natural peptide that results from fibrin degradation by plasmin; angiopoietin-1 (Angl), which is a vascular growth factor; angiopoietin-like 4 (ANGPTL4); and the vasoinhibin family of peptides derived by proteolytic cleavage of prolactin.

4. Dosage Amount. The polynucleotide agent is administered in a therapeutically effective amount. This can be assessed physiologically as the amount sufficient to cause increase vascularization in the brain. This can also be determined clinically as the amount sufficient to prevent the progression or improve the symptoms of dementia.

The amount of the polynucleotide agent administered for human treatment can range from 100 - 8,000 μg of the polynucleotide agent for each nostril; in some cases, in the range of 500 - 5,000 μg for each nostril. But other amounts are also possible. The polynucleotide agent can be administered in divided doses.

For polynucleotides packaged inside carrier particles (e.g. viral vectors or liposomes), the dosage amount for human treatment can range from 5xl0⁷ - 5xl0¹⁴ particle units for each nostril (measuring the total number of particles, both active and inactive). But other amounts are also possible. The polynucleotide agent can be administered in divided doses.

Because the polynucleotide agent is being dispensed intranasally, for liquid or semi-solid dosage forms, small volumes are preferred. In some embodiments, each dosage amount is contained in a volume of 50 μΐ - 2 ml. But other dosage volumes are also possible.

5. Method of Administering. The polynucleotide agent can be applied intranasally in any suitable manner. In some cases, the polynucleotide agent is applied as a nasal spray in which the liquid is atomized and dispersed into air. In some cases, the polynucleotide agent is applied as nasal drops (i.e. bulk liquid drops that are not atomized and dispersed into air).

The polynucleotide agent can be administered intranasally with the patient in any suitable position. In some cases, the patient is upright. In some cases, the

polynucleotide agent is administered with the patient in a supine position, and in some cases, supine at an angle with the patient's head pointing downward (e.g. on a tilt table). In some cases, it is administered with the patient's head tilted back (i.e. neck extension). Combinations of the preceding patient positions are also possible. These patient positions may be useful in allowing the polynucleotide agent to flow onto the olfactory mucosa at the roof of the nasal cavity, and may be particularly suitable for nasal drop administration.

5(a). Repeat Dosing. It is possible that the patient may need continued dosing to maintain cognitive improvements. In some embodiments, the patient is given another dose after a period of time. For example, a second dose may be given 2 weeks to 6 months after the first dose. The dosage amount may be the same or different from the first dosage amount. In some cases, the patient receives multiple repeat dosing, wherein each of the time intervals between dosing can range from 2 weeks to 6 months.

In some cases, the patient receives multiple repeat dosing at a periodic time P, wherein P is a time value selected from 2 weeks to 6 months. For example, the instructions for use may indicate that the patient should receive repeat dosing every 10 weeks (i.e. P = 10 weeks). In some cases, the patient receives multiple repeat dosing at a periodic time window W, wherein W is a time window selected from a time window that is within the range of 2 weeks to 6 months. For example, the instructions for use may indicate that the patient should receive repeat dosing every 6-8 weeks (i.e. W = 6-8 weeks).

In some cases, the patient receives multiple repeat dosing of the polynucleotide agent, but does not receive the treatment on occasions when the patient is having nasal discharge (e.g. runny nose from an upper respiratory infection or allergies).

5(b). Pre-screening. In some embodiments, the administration of the polynucleotide agent to the patient is preceded by the clinician performing a screening evaluation to determine whether the patient should not receive the treatment (e.g. contraindications). Examples of conditions that may preclude use of the treatment include retinopathy [e.g. diabetic retinopathy or neovascular macular degeneration), chronic glomerular kidney diseases, cancer or malignancy, and rheumatoid arthritis. The clinician may perform such a screening evaluation by obtaining a medical history (e.g. asking the patient or a family member, or reviewing the patient's chart for a prior diagnosis, examinations, laboratory tests, etc.). For example, in screening for retinopathy, the clinician may review the patient's medical chart, ask the patient or family member about vision loss or a prior diagnosis of retinopathy, and/or the clinician may perform a retinal exam (e.g. through an ophthalmoscope) or refer to the patient to a specialist for performing a retinal exam (with the clinician receiving a report about the retinal examination).

In some embodiments, the screening evaluation includes a nasal evaluation. The nasal evaluation assesses for conditions that may affect the decision about whether the patient should not receive the treatment. The nasal evaluation may involve obtaining a medical history (e.g. by asking the patient or a family member, or reviewing the patient's chart for a prior diagnosis, examinations, laboratory tests, etc.) to identify nasal conditions such as ongoing nasal discharge, nasal congestion, coughing/sneezing, nasal bleeding, allergic rhinitis, nasal anatomic deformities (e.g. deviated septum), etc. The nasal evaluation may involve performing a nasal examination to identify conditions such as those aforementioned. The clinican may perform the nasal examination (e.g. using a nasal speculum or otoscope) or the patient can be referred to a specialist for performing the nasal examination (with the clinician receiving a report about the nasal examination).

6. Delivery Device. My invention also encompasses an intranasal treatment device that comprises an intranasal injector for extending into a nostril and a container that holds a liquid composition comprising the polynucleotide agent. In some embodiments, the concentration of the polynucleotide agent in the liquid composition is in the range of 50 ug/ml to 160 mg/ml. In some embodiments, when contained inside a carrier vector (e.g. virus, liposomes, etc.), the concentration of the polynucleotide agent is in the range of 2 χ 10⁷ particle units/ml to lxlO¹⁵ particle units/ml. In some embodiments, each activation of the nasal injector delivers 50 ul to 2 ml of the liquid composition into the patient's nose. In some embodiments, the container holds a dosage amount intended for a single administration to the patient through both nostrils, i.e. a unit dosage amount intended for one-time use only. In some embodiments, the container is replaceable, i.e. the container can be removed and then replaced with a new one. For example, the container may be a unit dose ampule that is removed and discarded after a single use. A replacement unit dose ampule can be inserted into the intranasal treatment device for the next administration.

In some embodiments, the container holds 100 ul to 4 ml of the liquid

composition. The intranasal injector may be a spray nozzle, introducer, spray tip, atomizier, pump, etc. or any configuration or mechanism to deliver a liquid composition into the nose by spray droplets or stream.

7. Animal Experiments. Experiments can be performed on animal models of degenerative brain diseases (e.g. Alzheimer's or Parkinson's disease). For example, animal models of Alzheimer's disease are described in G. Casadesus, Handbook of Animal Models in Alzheimer's Disease, IOS Press (2011). Rat and mice models of

Alzheimer's disease are reported in Sonia Do Carmo et al, "Modeling Alzheimer's disease in transgenic rats" in Molecular Neurodegeneration, vol 8:37 (2013); and Piotr Religa et al, "VEGF significantly restores impaired memory behavior in Alzheimer's mice by improvement of vascular survival" in Scientific Reports, vol 3:2053 (2013).

These animals can be treated with intranasal administration of the

polynucleotide agent (human or its animal counterpart) and tested for improvement in cognitive skills. The animal study can be performed using a viral carrier vector such as adeno-associated virus (AAV) or herpes simplex virus. Intranasal administration into rats and mice are described in L.R. Hanson et al, "Intranasal Administration of CNS Therapeutics to Awake Mice" in / Vis Exp (74), e4440, doi:10.3791/4440 (2013), with video available at [http://www.jove.com/video/4440/intranasal-administration-of- cns-therapeutics-to-awake-mice]; R.G. Thorne et al, "Delivery of Insulin-Like Growth Factor-I to the Rat Brain and Spinal Cord Along Olfactory and Trigeminal Pathways Following Intranasal Administration" in Neuroscience, vol 127, pp 481-496 (2004); I.K. Han et al, "Enhanced brain targeting efficiency of intranasally administered plasmid DNA: an alternative route for brain gene therapy" in J Mol Med, vol 85:75-83 (2007). The clinical effect of the treatment can be measured using any well-known animal testing methodology to measure spatial memory, movement control, and/or cognitive mapping. Examples of such tests includes water maze testing (e.g. Morris water maze, water T-maze, radial arm water maze, etc.] and dry maze testing (T-maze, radial arm maze, oasis maze, Barnes maze, etc.).

Protocols for such animal testing are described in the scientific literature: H Hodges, "Maze procedures: the radial-arm and water maze compared" in Cognitive Brain Research, vol 3 (3-4], pp 167-81 (June 1996); W Crusio, "Methodological considerations for testing learning in mice" in WE Crusio & RT Gerlai, Handbook of molecular-genetic techniques for brain and behavior research (1st ed), pp 638-651

(1999); GL Wenk, "Assessment of spatial memory using the radial arm maze and Morris water maze" in J Crawley et al, Current Protocols in Neuroscience, Ch 8, Unit 8.5A (May 2004); and D Morgan, "Water Maze Tasks in Mice: Special Reference to Alzheimer's Transgenic Mice" in Ch 14 of Methods of Behavior Analysis in Neuroscience (ed JJ Buccafusco), CRC Press (2nd ed 2009).

Any use of the word "or" herein is intended to be inclusive and is equivalent to the expression "and/or," unless the context clearly dictates otherwise. As such, for example, the expression "A or B" means A, or B, or both A and B. Similarly, for example, the expression "A, B, or C" means A, or B, or C, or any combination thereof.

The foregoing description and examples have been set forth merely to illustrate my invention and are not intended to be limiting. Each of the disclosed aspects and embodiments of my invention may be considered individually or in combination with other aspects, embodiments, and variations of my invention. In addition, unless otherwise specified, the steps of the methods of my invention are not confined to any particular order of performance. Modifications of the disclosed embodiments incorporating the spirit and substance of my invention may occur to persons skilled in the art, and such modifications are within the scope of my invention.

Claims

I claim:

\. A method of treating dementia in a patient, comprising intranasally

administering to the patient a DNA agent that contains a DNA sequence that encodes a vascular endothelial growth factor (VEGF) polypeptide, wherein the DNA agent is deposited onto the olfactory mucosa inside the patient's nose.

2. The method of claim 1, wherein the dementia is Alzheimer's disease.

3. The method of claim 1, wherein the DNA agent is administered as nasal drops or nasal spray.

4. The method of claim 1, further comprising administering another dose of the DNA agent at a time from 2 weeks to 6 months after the first dose.

5. The method of claim 1, further comprising administering multiple repeat dosing of the DNA agent, wherein each of the time intervals between dosing can range from 2 weeks to 6 months.

6. The method of claim 1, further comprising, prior to administering the treatment, performing a screening evaluation to determine whether the patient should not receive the treatment.

7. The method of claim 6, wherein the screening evaluation includes determining whether the patient has experienced vision loss or has a prior diagnosis of retinopathy.

8. The method of claim 6, wherein the screening evaluation includes performing a retinal exam on the patient or referring the patient to a specialist for undergoing a retinal exam.

9. The method of claim 1, wherein the VEGF polypeptide has a cystine knot motif comprising eight cysteine residues.

10. The method of claim 9, wherein the VEGF polypeptide has a heparin-binding domain.

11. The method of claim 1, wherein the DNA agent is contained inside a viral carrier vector.

12. The method of claim 11, wherein the DNA agent is administered in a dosage amount of 5*10⁷ - 5*10¹⁴ particle units for each nostril.

13. The method of claim 1, wherein the DNA agent is provided as a liquid or semisolid dosage form, and wherein each dosage amount is 50 μΐ - 2 ml of volume.

14. The method of claim 13, wherein the DNA agent is administered with the patient in a supine position or with the patient's head tilted back.

15. A method of treating dementia, comprising intranasally receiving a DNA agent that contains a DNA sequence that encodes a vascular endothelial growth factor (VEGF) polypeptide, wherein the DNA agent is deposited onto the olfactory mucosa inside the nose.

16. The method of claim 15, further comprising receiving another dose of the DNA agent at a time from 2 weeks to 6 months after the first dose.

17. The method of claim 15, further comprising receiving multiple repeat dosing of the DNA agent, wherein each of the time intervals between dosing can range from 2 weeks to 6 months.

18. The method of claim 1, further comprising, prior to receiving the treatment, receiving a screening evaluation to determine whether the treatment cannot be received.

19. The method of claim 18, wherein the screening evaluation includes determining experience of vision loss or prior diagnosis of retinopathy.

20. The method of claim 18, wherein the screening evaluation includes receiving a retinal exam.

21. The method of claim 15, wherein the DNA agent is contained inside a viral carrier vector and the DNA agent is received in a dosage amount of 5^χ10⁷ - 5^χ10¹⁴ particle units for each nostril.

22. The method of claim 15, wherein the DNA agent is received in a supine position or with the head tilted back.

23. An intranasal treatment device comprising:

an intranasal injector for extending into a patient's nostril;

a container that holds 100 ul to 4 ml of a liquid composition comprising a DNA agent that contains a DNA sequence that encodes a vascular endothelial growth factor (VEGF) polypeptide;

wherein each activation of the intranasal treatment device delivers 50 ul to 2 ml of the liquid composition into the patient's nose.

24. The intranasal treatment device of claim 15, wherein the concentration of the DNA agent in the liquid composition is in the range of 50 ug/ml to 160 mg/ml or in the range of 2 ^χ 10⁷ particle units/ml to lxlO¹⁶ particle units/ml.

25. The intranasal treatment device of claim 15, wherein the container holds only a single unit dosage amount.

26. The intranasal treatment device of claim 25, wherein the container is replaceable.