BACKGROUND OF THE INVENTION

Recombinant adeno-associated viruses (rAAVs) belong to the parvovirus family and are non-enveloped, single-stranded DNA viruses [17]. Over the past decades, rAAVs have become among the most promising vectors for gene therapy, which is in large part due to their reduced pathogenicity as compared with the wild-type virus, the ability to establish long-term transgene expression, and their ability to transduce both dividing and nondividing cells [1], However, complications of treatment related to immune responses against the vector represent serious obstacles for the broad use of rAAV vectors in gene therapy [2]. Antibodies against viruses may neutralize infectivity prior to viral attachment to host cell receptors, or post attachment; interfering with internalization or fusion at the cell surface, or during endosomal trafficking [3]. Natural exposure to AAV types 1, 2, 5, 6, 8, and 9 results in the production of antibodies of all four IgG subclasses [4],

Whereas the prevalence depends on the serotype, recent studies report that between 20% and 40% of the population has neutralizing antibody titers of >1/20 any given serotype [5]. For this reason significant proportion of the patients that harbors neutralizing antibodies against AAV should be excluded from clinical trials and cannot be treated using AAV vectors [6]. Recently empty AAV2 capsids were used to saturate neutralizing antibodies and thereby enhance transduction of the co-administration of vector in mice and non-human primates [7]. In a similar way, mixture of peptides that resemble AAV epitopes and blocked antibody binding in the ELISA was tested on their ability to block activity of neutralizing sera [8]. However, even if these peptides indeed blocked neutralization of AAV by serum poly antibodies, it appears undesirable to use peptides that are parts of AAV since their repeated administration can stimulate production of AAV-neutralizing antibodies, which in long term would decrease efficiency of gene therapy with particular AAV serotype. The document US 9,861,661 describes a T-cell epitope derived from a viral vector protein, functionalized with C-(X)2-[CST] or [CST]-(X)2-C motif (redox active cysteines encountered in thioreductases), which can be used for suppressing immune response against a viral vector induced by gene therapy. However, the identification of efficient T-cell epitopes is complex to design. The document WO 2018/158397 presents another strategy consisting in removing anti-AAV antibodies from a blood-derived composition by contacting said blood-derived composition with at least one support onto which is grafted one or more affinity ligand(s) that specifically bind to anti-AAV antibodies, wherein said one or more affinity ligand is an AAV particle of one or more serotypes. However, this type of column is expensive and is not totally safe since AAV particles risk to come off the support, consequently contaminating the blood-derived composition [9].

Therefore, there is still a need to find a new strategy to prevent and/or suppress immune responses against AAV vectors.

SUMMARY OF THE INVENTION

In a first aspect, the invention relates to peptide that specifically binds to an AAV capsid, said peptide comprising an amino acid sequence having at least 80% identity with an amino acid sequence selected from SEQ ID NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO : 6.

In a second aspect, the invention relates to a composition comprising one or more peptides that specifically bind to an AAV capsid selected from :

(i) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 1, (ii) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 2,

(iii) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 3,

(iv) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 4,

(v) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 5, (vi) a peptide comprising an amino acid sequence having at least 80% identity with SEQ ID NO : 6, or

(vii) at least two peptides selected from (i) to (vi). In a third aspect, the invention relates to a kit-of-parts comprising (i) the peptide according to the invention or the composition according to the invention and (ii) an AAV vector.

In a fourth aspect, the invention relates to the peptide according to the invention or the composition according to the invention for use in the prevention of an immune response against an AAV capsid in a subject receiving an AAV vector.

In a fifth aspect, the invention relates to a molecular imprinted polymer (MIP) particle that specifically binds to the peptide according to the invention.

The invention also relates to a support onto which are grafted one or more MIP particles according to the invention.

The invention also relates to a method for removing ex vivo anti-AAV antibodies from a blood material, comprising contacting said blood material with the support according to the invention.

The invention also relates to an extracorporeal device comprising the support according to the invention.

LEGENDS TO THE FIGURES

Figure 1 represents a scheme of solid phase synthesis of a peptide (with X meaning "temporary amino protecting group", Y meaning "permanent side-chain protecting group", and A meaning "carboxy activating group"). The step one consists in the deprotection of the X group from a first amino acid which is grafted on a resin via a linker. The second step consists in the coupling of the grafted amino acid with another amino acid. Steps one and two can be repeated. The last step consists in the deprotection of the X group and the cleavage from the resin.

Figure 2 is a schematic representation of the method of preparation of MIP particles according to the invention. The first step corresponds to the immobilization of a peptide on a solid phase, the second step corresponds to a co- polymerization of a MIP particle using UV, the third step corresponds to the wash- off of unspecific MIP particles and the fourth step corresponds to the elution of the MIP particles that specifically bind to the peptide (from left to right).

Figure 3 represents the binding inhibition between anti-AAV8-specific-antibodies immobilised on a sensor surface and AAV-8, in presence or in the absence of the peptides of the invention (Y axis RU Responses Units, X axis in minute). It was observed that presence of the peptides has inhibited binding between AAV8 capsids and virus-specific IgG immobilized on the sensor chip approximately by 65%.

Figure 4 is a schematic representation of a column comprising a support onto which MIP particle 1 (MIP 1) was grafted. The first step consists in the addition of IVIG solution on the column (left), which will lead to the binding of specific anti- AAV8 IgGs on MIP1 (right).

Figure 5 shows the elution profile over time obtained with 3 mL of IVIG injected in the column GE Healthcare NHS-Sepharose, on which MIP was grafted as described in example 4 (Y Axis is in milli absorbance units (mAU), X Axis is in mL). Especially, this profile represents three fractions: A fraction corresponds to the flow-through (FT), a fraction corresponds to the Wash, and a fraction corresponds to the elution. Figure 6 ELISA test shows the removal of anti-AAV8 antibodies from the IVIG sample by passage over column GE Healthcare NHS-Sepharose, on which MIP 1 was grafted as described in example 4 (0.1 mg of MIP grafted). Note: Y axis is in nanogram (ng) of specific anti-AAV8 IgG, X axis is the fraction tubes. Loading and washing represents fractions 4 at 10; elution with citrate buffer represents fractions 23 at 31. The Elution fraction represents 17% of retention, as measured by the ELISA test.

DETAILED DESCRIPTION OF THE INVENTION

Definitions The term "peptide" refers to an amino acid sequence, i.e. a chain of amino acids linked by peptide bonds.

The term "amino acid sequence" has its general meaning and is a sequence of amino acids that confers to a peptide its primary structure.

According to the present invention, the "identity" is calculated by comparing two aligned sequences in a comparison window. The alignment of the sequences makes it possible to determine the number of positions (nucleotides or amino acids) in common for the two sequences in the comparison window. The number of positions in common is therefore divided by the total number of positions in the comparison window and multiplied by 100 to obtain the percentage of identity. The determination of the percentage of sequence identity can be carried out manually or by means of well-known computer programs. In a particular embodiment of the invention, the identity or the homology corresponds to at least one substitution, for example 1, 2, 3, 4, 5 substitutions, of an amino acid residue, without appreciable loss of interactive binding capacity. Preferably, the at least one substitution is a conservative amino acid substitution. By "conservative amino acid substitution", it is meant that an amino acid can be replaced with another amino acid having a similar side chain. Families of amino acid having similar side chains have been defined in the art, including basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., glycine, cysteine, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine).

The term "AAV" refers to adeno-associated virus, a small, non-enveloped virus of the parvovirus family that packages a single-stranded linear DNA genome, approximately 5 kb long. The term "AAV capsid" refers to the capsid of an adeno-associated virus (AAV), which comprises epitope recognized by an anti-AAV antibody. Especially, the AAV capsid comprises AAV capsid proteins, in particular VP capsid proteins, namely selected from VP1, VP2 and/or VP3 capsid proteins, which are encoded by VP genes (VP1, VP2 and/or VP3). In particular, the AAV capsid is selected from AAV-1 capsid, AAV-2 capsid, AAV-3 capsid, AAV-4 capsid, AAV-5 capsid, AAV-6 capsid, AAV-7 capsid, AAV-8 capsid, AAV-9 capsid, AAV-10 capsid (such as AAV-cylO or AAV-rhlO), AAV-11 capsid, AAV-rti74 capsid or engineered AAV capsid variants such as AAV-2i8 capsid, AAV2G9 capsid, AAV-LK3 capsid, AAV-DJ capsid, AAV- Anc80 capsid, preferably AAV-8 capsid. According to a particular embodiment, the AAV capsid is selected from AAV-1, AAV-2, AAV-2 variants (such as the quadruple- mutant capsid optimized AAV-2 comprising an engineered capsid with Y44+500+730F+T491V changes, disclosed in Ling et al., 2016 [10], AAV-3 and AAV-3 variants (such as the AAV3-ST variant comprising an engineered AAV3 capsid with two amino acid changes, S663V+T492V, disclosed in Vercauteren et al., 2016 [11], AAV-3B and AAV-3 B variants, AAV-4, AAV-5, AAV-6 and AAV-6 variants (such as the AAV-6 variant comprising the triply mutated AAV-6 capsid Y731F/Y705F/T492V form disclosed in Rosario et al., 2016 [12]), AAV-7, AAV-8, AAV-9 and AAV-9 variants (such as AAVhu68), AAV-2G9, AAV-10 such as AAV- cylO and AAV-rhlO, AAV-rh39, AAV-rh43, AAV-rh74, AAV-dj, AAV Anc80, AAV LK03, AAV. PH P, AAV 2i8, porcine AAV such as AAV po4 and AAV po6, and tyrosine, lysine and serine capsid mutants of AAV serotypes. In addition, the AAV capsid is selected from other non-natural engineered variants (such as AAV- spark100), chimeric AAV or AAV serotypes obtained by shuffling, rationale design, error prone PCR, and machine learning technologies. In a particular embodiment, the Cap gene encodes VP capsid proteins derived from at least two different AAV serotypes, or encodes at least one chimeric VP capsid protein combining VP capsid protein regions or domains derived from at least two AAV serotypes. For example a chimeric AAV capsid can derive from the combination of an AAV8 capsid sequence with a sequence of an AAV serotype different from the AAV8 serotype, such as any of those specifically mentioned above. In another embodiment, the AAV capsid comprises one or more variant VP capsid proteins such as those described in W02015013313, in particular the RHM4-1, RHM15-1, RHM15-2, RHM15-3/RHM15-5, RHM15-4 and RHM15-6 capsid variants. In a particular embodiment, the AAV capsid is a hybrid between AAV serotype 9 (AAV-9) and AAV serotype 74 (AAV-rh74) capsid proteins. For example, the AAV serotype may be a -rh74-9 serotype as disclosed in WO2019/193119 (such as the Hybrid Cap rh74-9 serotype described in examples of WO2019/193119; a rh74-9 serotype being also referred to herein as "-rh74-9", "AAVrh74-9" or "AAV-rh74-9") or a AAV-9-rh74 serotype as disclosed in WO2019/193119 (such as the Hybrid Cap 9-rh74 serotype described in the examples of WO2019/193119; a AAV-9-rh74 serotype being also referred to herein as "-9-rh74", "AAV9-rh74", "AAV-9-rh74", or "rh74-AAV9"). In a particular embodiment, the AAV capsid is a peptide-modified hybrid between AAV serotype 9 (AAV-9) and AAV serotype 74 (AAV-rh74) capsid proteins, as described in PCT/EP2019/076958, such as an AAV9-rh74 hybrid capsid or AAVrh74-9 hybrid capsid modified with the PI peptide. In a preferred embodiment, the AAV capsid is selected from AAV-1 capsid, AAV-2 capsid, AAV-3 capsid, AAV-4 capsid, AAV-5 capsid, AAV-6 capsid, AAV-7 capsid, AAV-8 capsid, AAV-9 capsid, AAV-10 capsid (such as AAV-cylO or AAV-rhlO), AAV-11 capsid, AAV-rh74 capsid or engineered AAV capsid variants such as AAV-2i8 capsid, AAV2G9 capsid, AAV-LK3 capsid, AAV-DJ capsid, AAV- Anc80 capsid and AAV9-rh74 hybrid capsid discloses in WO 2019/193119, preferably AAV-8 capsid.

The term "AAV vector" according to the invention means an AAV vector suitable for protein expression, preferably for use in gene therapy, such as a single- stranded or double-stranded self-complementary AAV vector. An AAV vector comprises a nucleic acid molecule, which is inserted into a nucleic acid construct for expressing said nucleic acid molecule, which is preferably a therapeutic nucleic acid molecule appropriate for the treatment of a disease, such as for example, proliferative diseases (cancers, tumors, dysplasias, etc.), infectious diseases; viral diseases (induced, e.g., by the Hepatitis B or C viruses, HIV, herpes, retroviruses, etc.); genetic diseases (cystic fibrosis, dystroglycanopathies, myopathies such as Duchenne Muscular Myopathy; myotubular myopathy; hemophilias; diabetes; amyotrophic lateral sclerosis, motoneurones diseases such as spinal muscular atrophy, spinobulbar muscular atrophy, or Charcot-Marie-Tooth disease; arthritis; cardiovascular diseases (restenosis, ischemia, dyslipidemia, homozygous familial hypercholesterolemia, etc.), or neurological diseases (psychiatric diseases, neurodegenerative diseases such as Parkinson's or Alzheimer's, Huntington's disease addictions (e.g., to tobacco, alcohol, or drugs), epilepsy, Canavan's disease, adrenoleukodystrophy, etc.), eye diseases such as retinitis pigmentosa, Leber congenital amaurosis, Leber hereditary optic neuropathy, Stargardt disease; lysosomal storage diseases such as San Filippo syndrome; hyperbilirubinemia such as CN type I or II or Gilbert's syndrome, Pompe disease, etc. In the context of the present invention, the AAV vector comprises an AAV capsid able to express the transgene into the target cells or tissues of interest, in particular hepatocytes or muscles. In a particular embodiment, the AAV vector has an AAV capsid selected from the one described above. In a preferred embodiment, the AAV vector has an AAV capsid selected from AAV-1 capsid, AAV-2 capsid, AAV- 3 capsid, AAV-4 capsid, AAV-5 capsid, AAV-6 capsid, AAV-7 capsid, AAV-8 capsid, AAV-9 capsid, AAV-10 capsid (such as AAV-cylO or AAV-rhlO), AAV-11 capsid, AAV-rh74 capsid or engineered AAV capsid variants such as AAV-2i8 capsid, AAV2G9 capsid, AAV-LK3 capsid, AAV-DJ capsid, AAV- Anc80 capsid and AAV9- rh74 hybrid capsid discloses in WO 2019/193119, preferably AAV-8 capsid.

The term "Molecular Imprinted Polymer particle" or "MIP particle" refers to a polymer particle that has been processed using a molecular imprinting technique which leaves cavities in the polymer matrix having a specific affinity for a chosen "template" molecule. The MIP particle thus specifically binds to the molecule, for example the molecule is a peptide. A process for preparing a MIP particle that specifically binds to a peptide is disclosed in WO/2018178629, [13], [14], and

[15].

The term "affinity" refers to the non-covalent interaction strength between a molecule and its ligand, for example between a peptide of the invention and an AAV capsid, or between a MIP particle of the invention and a peptide of the invention. The affinity may be characterized by the dissociation constant (Kd). The Kd may be measured by well-known methods, such as FRET or in SPR (surface plasmon resonance).

The term "anti-AAV antibody" refers to an antibody which binds to an AAV vector, especially an AAV capsid. In particular, an anti-AAV antibody can also be a neutralizing antibody (for example an immunoglobulin IgG), and are therefore called anti-AAV neutralizing antibodies (NAbs). Neutralizing antibodies (NAbs) against AAV capsids can abolish AAV infectivity on target cells, reducing the transduction efficacy. The term "pharmaceutically acceptable" means approved by a regulatory agency of the Federal or a state government or listed in the U.S. or European Pharmacopeia or other generally recognized pharmacopeia for use in animals, and humans. The term "pharmaceutical composition" means a composition comprising pharmaceutically acceptable carrier. For example, a carrier can be a diluent, adjuvant, excipient, or vehicle with which the therapeutic is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is a preferred sterile liquids carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as sterile liquid, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene glycol, water, ethanol and the like. When the pharmaceutical composition is adapted for oral administration, the tablets or capsules can be prepared by conventional means with pharmaceutically acceptable excipients such as binding agents (e.g. pregelatinized maize starch, polyvinylpyrrolidone or hydroxypropyl methylcellulose); fillers (e.g., lactose, microcrystalline cellulose or calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc or silica);disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulphate). The tablets may be coated by methods well known in the art. Liquid preparations for oral administration may take the form of, for example, solutions, syrups or suspensions, or they may be presented as a dry product for constitution with water or another suitable vehicle before use. Such liquid preparations may be prepared by conventional means with pharmaceutically acceptable additives such as suspending agents (e.g., sorbitol syrup, cellulose derivatives or hydrogenated edible fats); emulsifying agents (e.g., lecithin or acacia); non-aqueous vehicles (e.g., almond oil, oily esters, ethyl alcohol or fractionated vegetable oils); and preservatives (e.g., methyl or propyl-p- hydroxybenzoates or sorbic acid). The preparations may also contain buffer salts, flavoring, coloring and sweetening agents as appropriate. The composition according to the invention is preferably a pharmaceutical composition.

The term "subject", , "patient" or "individual", as used herein, refers to a human or non-human mammal (such as a rodent (mouse, rat), a feline, a canine, or a primate) affected or likely to be affected with a disease. Preferably, the subject is a human, man or woman.

The term "blood material" refers to blood, plasma or serum and any fraction thereof, such as plasma precipitate, plasma supernatant, preferably blood, blood plasma or blood serum. The term "extracorporeal device" refers to a device which can be used outside the body, i.e. ex vivo. In particular, a blood material can circulate throw said device. Peptide of the invention

A first object of the present invention concerns a peptide that specifically binds to an AAV capsid, said peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity with an amino acid sequence selected from SEQ ID NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO : 6.

In some embodiments, the peptide of the invention comprises an amino acid sequence selected from SEQ ID NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO : 6. In some embodiments, the peptide of the invention consists in an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99%, identity with an amino acid sequence selected from SEQ ID

NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO : 6.

In some embodiments, the peptide of the invention consists in an amino acid sequence selected from SEQ ID NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO : 6. Advantageously, the peptide of the invention neutralizes the binding of an anti- AAV antibody to the AAV capsid, allowing to reduce immune response against said AAV capsid. Preferably, the peptide of the invention binds to the AAV capsid with a Kd comprised between 0.0001 and 20 nM, preferably between 0.001 and 15 nM, even more preferably between 0.001 and 10 nM, for example between 0.001 and 7 nM, between 0.002 and 6.5 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least

90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 1 or SEQ ID NO : 7, and binds to the AAV capsid with a Kd comprised between 0.0001 and InM, preferably between 0.001 and 0.1 nM, even more preferably between 0.002 and 0.01 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 2 or SEQ ID NO : 8, and binds to the AAV capsid with a Kd comprised between 0.0001 and 1 nM, preferably between 0.001 and 0.1 nM, even more preferably between 0.01 and 0.032 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 3 or SEQ ID NO : 9, and binds to the AAV capsid with a Kd comprised between 1 and 7 nM, preferably between 2 and 5 nM, even more preferably between 3.5 and 4.5 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least

85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least

90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 4 or SEQ ID NO : 10, and binds to the AAV capsid with a Kd comprised between 1 and 7 nM, preferably between 2 and 6 nM, even more preferably between 3 and 5 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least

90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 5 or SEQ ID NO : 11, and binds to the AAV capsid with a Kd comprised between 1 and 7 nM, preferably 3 and 6 nM, even more preferably between 4 and 5.6 nM.

In particular, the peptide comprises an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity, such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or more preferably 100% identity with SEQ ID NO : 6 or SEQ ID NO : 12, and binds to the AAV capsid with a Kd comprised between 1 and 7 nM, preferably 3 and 6.5 nM, even more preferably between 5 and 6.2 nM.

Preferably, the Kd is measured by SPR (surface plasmon resonance). The peptide of the invention can comprise a linker at its N-terminal end, consisting of at least one amino acid residue. This linker can, for example, be used for immobilizing the peptide onto a support. The at least one amino acid residue can be selected from cysteine and/or glycine. Classical linkers well known in the literature can be used and will be selected by the person skilled in the art depending on the final application of the peptide of the invention. An example of a peptide that further comprises a linker can be selected from SEQ ID NO : 7, SEQ ID NO : 8, SEQ ID NO : 9, SEQ ID NO : 10, SEQ ID NO : 11 or SEQ ID NO : 12. The peptide according to the invention can be synthesized by conventional methods, such as solid phase, as disclosed in Figure 1.

In a specific embodiment, the peptide of the invention specifically binds to an AAV-8 capsid and consists in an amino acid sequence selected from SEQ ID NO : 1, SEQ ID NO : 2, SEQ ID NO : 3, SEQ ID NO : 4, SEQ ID NO : 5 or SEQ ID NO :

6.

Composition of the invention A second object of the present invention concerns a composition comprising one or more peptides that specifically bind to an AAV capsid selected from: (i) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 1, more preferably 100% identity with SEQ ID NO 1, (II) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 2, more preferably 100% identity with SEQ ID NO 2,

(iii) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

3, more preferably 100% identity with SEQ ID NO 3,

(iv) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

4, more preferably 100% identity with SEQ ID NO 4,

(v) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

5, more preferably 100% identity with SEQ ID NO 5, (vi) a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 6, more preferably 100% identity with SEQ ID NO 6, or

(vii) at least two peptides selected from (i) to (vi) , for example at least three peptides selected from (i) to (vi), at least four peptides selected from (i) to (vi), at least five peptides selected from (i) to (vi) or the six peptides from (i) to (vi). In a particular embodiment, the composition of the invention comprises one or more peptides that specifically bind to an AAV capsid selected from :

(i) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%with SEQ ID NO : 1, more preferably 100% identity with SEQ ID NO : 1,

(ii) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%with SEQ ID NO :

2, more preferably 100% identity with SEQ ID NO : 2,

(iii) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

3, more preferably 100% identity with SEQ ID NO : 3,

(iv) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

4, more preferably 100% identity with SEQ ID NO : 4,

(v) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%with SEQ ID NO :

5, more preferably 100% identity with SEQ ID NO : 5, (vi) a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 6, more preferably 100% identity with SEQ ID NO : 6, or (vii) at least two peptides selected from (i) to (vi), for example at least three peptides selected from (i) to (vi), at least four peptides selected from (i) to (vi), at least five peptides selected from (i) to (vi) or the six peptides from (i) to (vi).

When the composition of the invention comprises (vii) at least two peptides selected from (i) to (vi), said peptides may be formulated in the same composition or separately, preferably in the same composition.

The composition of the invention can also comprise an AAV vector.

The composition of the invention is preferably a pharmaceutical composition. Thus, the composition can comprise one or more peptides of the invention and a suitable pharmaceutically acceptable vehicle, as defined above. The pharmaceutically acceptable vehicle can, for example, contribute to the solubility, stability or sterility of the composition or increase the efficiency of uptake into the body.

Kit-of-parts of the invention

The invention also relates to a kit-of-parts comprising (i) the peptide according to the invention or the composition according to the invention and (ii) an AAV vector.

Method of treatment of the invention

The invention also relates to the peptide according to the invention or the composition according to the invention, for use as a medicament.

The invention also relates to the peptide according to the invention or the composition according to the invention for use in the prevention of an immune response against an AAV capsid in a subject receiving an AAV vector. In particular, the prevention is a reduction or a suppression of an immune response against an AAV capsid. In particular, the immune response can be either a pre-existent immune response against an AAV capsid, or an immune response triggered by the administration of an AAV vector in a subject in need for a gene therapy.

In particular, the immune response against an AAV capsid comprises the production of neutralizing anti-AAV antibodies (Nabs) and/or the binding of said Nabs to AAV capsid.

The peptide or the composition according to the invention can be administered before, simultaneously and/or after the AAV vector. The treatment according to the invention enhances the efficacy of the gene therapy by preventing and/or decreasing the immune response against AAV capsid, thereby suppressing or reducing the production of neutralizing anti-AAV antibodies and/or the binding of said Nabs to AAV capsid.

In a preferred embodiment, the peptide or the composition according to the invention is administered before and/or simultaneously to the AAV vector. For example, the peptide of the invention can be administered at least 60 days or more before administering the AAV vector.

The subject may receive an AAV vector by a direct AAV vector gene therapy, in which the AAV vector is directly administered to the subject.

The administration of the AAV vector is for example but is not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, intrathecal and oral routes. In a particular embodiment, the administration is via the intravenous or intramuscular route. The AAV vector may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local. In a specific embodiment, it may be desirable to administer the AAV vector locally to the area in need of treatment. This may be achieved, for example, by means of an implant, said implant being of a porous, nonporous, or gelatinous material, including membranes, such as sialastic membranes, or fibers.

The amount of the AAV vector which will be effective depend on the type of and can be determined by standard clinical techniques. In addition, in vivo and/or in vitro assays may optionally be employed to help predict optimal dosage ranges. The precise dose to be employed in the formulation will also depend on the route of administration, and the seriousness of the disease, and should be decided according to the judgment of the practitioner and each patient's circumstances. The dosage of AAV vector administered to the subject in need thereof will vary based on several factors including, without limitation, the route of administration, the specific disease treated, the subject's age or the level of expression necessary to achieve the therapeutic effect. One skilled in the art can readily determine, based on its knowledge in this field, the dosage range required based on these factors and others. With a treatment comprising administering an AAV vector, to the subject, typical doses of the vector are for example of at least 1x10 ⁸ vector genomes per kilogram body weight (vg/kg), such as at least 1x10 ⁹ vg/kg, at least 1x10 ¹⁰ vg/kg, at least 1x10 ¹¹ vg/kg, at least 1x10 ¹² vg/kg at least 1x10 ¹³ vg/kg, or at least 1x10 ¹⁴ vg/kg.

In an aspect of the invention, the subject receives repeated administration of an AAV vector. In this aspect, said administration may be repeated at least once or more, and may even be considered to be done according to a periodic schedule, such as once per year. The periodic schedule may also comprise an administration once every 2, 3, 4, 5, 6, 7, 8, 9 or 10 year, or more than 10 years. In another particular embodiment, administration of each administration of an AAV vector is done using an AAV vector having different capsid for each successive administration, thereby avoiding a reduction of efficacy because of a possible immune response against a previously administered AAV vector, even if the use of the peptide according to the invention should help minimize this immune response.

The peptide or the composition of the invention may be administered by oral administration, nasal administration, transdermal administration or by parenteral injection such as intravenous infusion, subcutaneous injection, intraperitoneal injection, intrathecal injection, preferably by oral administration or intravenous infusion, more preferably by oral administration. The peptide or the composition of the invention may be orally administered continuously, such as daily, in order to maintain a constant level in the circulation. Such constant level will be one that has been determined to be non-toxic to the patient, and optimal regarding interaction with the AAV vector during the time of administration to confer a non-inhibitory, therapeutic effect.

In a particular embodiment of the method of the invention, an immunosuppressor can be administered to the patient before the administration of the peptide according to the invention or before the administration of the composition according to the invention. Any immunosuppressor suitable for improving AAV vector transfer in gene therapy can be used for carrying out the invention. For example, the immunosuppressor is selected from corticosteroids, such as prednisolone, rapamycin, tacrolimus, sirolimus, mycophenolate mofetil (MMF), or their mixture. Plasma derived polyclonal immunoglobulins are not considered as a suitable immunosuppressor for carrying out the invention.

The present invention may generally be applied for therapy of any disease that may be treated by expression of a therapeutic nucleic acid molecule in a cell or tissue of a subject mediated by an AAV vector. These include, for example, proliferative diseases (cancers, tumors, dysplasias, etc.), infectious diseases; viral diseases (induced, e.g., by the Hepatitis B or C viruses, HIV, herpes, retroviruses, etc.); genetic diseases (cystic fibrosis, dystroglycanopathies, myopathies such as Duchenne Muscular Myopathy; myotubular myopathy; hemophilias; diabetes; amyotrophic lateral sclerosis, motoneurones diseases such as spinal muscular atrophy, spinobulbar muscular atrophy, or Charcot-Marie-Tooth disease; arthritis; cardiovascular diseases (restenosis, ischemia, dyslipidemia, homozygous familial hypercholesterolemia, etc.), or neurological diseases (psychiatric diseases, neurodegenerative diseases such as Parkinson's or Alzheimer's, Huntington's disease addictions (e.g., to tobacco, alcohol, or drugs), epilepsy, Canavan's disease, adrenoleukodystrophy, etc.), eye diseases such as retinitis pigmentosa, Leber congenital amaurosis, Leber hereditary optic neuropathy, Stargardt disease; lysosomal storage diseases such as San Filippo syndrome; hyperbilirubinemia such as CN type I or II or Gilbert's syndrome, Pompe disease, etc.

Molecular imprinted polymer (MIP) particle

The invention also relates to a molecular imprinted polymer (MIP) particle that specifically binds to the peptide according to the invention. In particular, the MIP particle according to the invention specifically binds to a peptide selected from:

- a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 1, more preferably 100% identity with SEQ ID NO : 1,

- a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 2, more preferably 100% identity with SEQ ID NO : 2,

- a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 3, more preferably 100% identity with SEQ ID NO : 3,

- a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 4, more preferably 100% identity with SEQ ID NO : 4,

- a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 5, more preferably 100% identity with SEQ ID NO : 5, or - a peptide comprising an amino acid sequence having at at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%with SEQ ID NO : 6, more preferably 100% identity with SEQ ID NO : 6.

In a specific embodiment, the MIP particle according to the invention specifically binds to a peptide selected from:

- a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 1, more preferably 100% identity with SEQ ID NO : 1, - a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

2, more preferably 100% identity with SEQ ID NO : 2,

- a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

3, more preferably 100% identity with SEQ ID NO : 3, - a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 4, more preferably 100% identity with SEQ ID NO : 4,

- a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

5, more preferably 100% identity with SEQ ID NO : 5, or

- a peptide consisting of an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO :

6, more preferably 100% identity with SEQ ID NO : 6.

Thanks to the peptides of the invention, which specifically bind to AAV capsid, it is possible to obtain MIP particles which bind variable regions of anti-AAV antibodies. The MIP particles may therefore be considered as anti-idiotype particles. Said MIP particles are thus "AAV capsid like" and can advantageously replace expansive and non-safe AAV capsid in plasmapheresis devices which will be able to remove AAV- specific neutralizing antibodies from the blood material of patients. An AAV capsid like particle is a molecule that mimics AAV capsid, but are non-infectious because it does not contain viral genetic material [16].

Thus, the MIP particle according to the invention also binds to one or more anti- AAV antibody, and is for example useful in a method according to the invention, i.e. for removing ex vivo anti-AAV antibodies from blood material. The MIP particle according to the invention can be obtainable by a process comprising the following steps:

- (i) contacting a plurality of monomers with a peptide according to the invention,

- (ii) polymerizing the plurality of monomers to create a peptide: polymer complex, and - (iii) isolating the peptide: polymer complex, to obtain a MIP particle.

The document WO 2018/178629 describes a method for obtaining said MIP particle and is herein incorporated by reference. The step of (i) contacting a plurality of monomers with a peptide according to the invention may be in any manner, or under any conditions, suitable for the formation of a molecularly imprinted polymer particle. The peptide according to the invention may be in solution or immobilized on a solid surface when brought into contact with the plurality of monomers. The solid surface for immobilization of the peptide of the invention may be of any composition or form as disclosed in the example. In particular, the peptide of the invention is immobilized on a solid phase before the step of (i) contacting a plurality of monomers with said peptide. An example of a peptide of the invention suitable for this step can be selected from SEQ ID NO : 7, SEQ ID NO : 8, SEQ ID NO : 9, SEQ ID NO : 10, SEQ ID NO : 11 or SEQ ID NO : 12. The immobilization is formed by grafting said peptide onto a designated solid surface. Grafting may comprise the attachment to any solid surface, for instance a surface comprising polysaccharide, silica, organic or inorganic polymer, metal, or a combination thereof. The solid surface may be of any form, for instance the form of the solid surface may be selected from beads, such as glass beads, magnetic beads, arrays, the surface of waveguides, fibres, membranes, or capillaries.

The plurality of monomers may be one type of monomer, or a mixture of different types of monomer. Document WO/2018178629 discloses a plurality of monomers suitable for obtaining the MIP particle according to the invention and is incorporated by reference. The polymerization of the monomers results in the creation of a molecularly imprinted polymer (MIP) particle. The plurality of monomers (also called "monomer mixture") may take any form or composition suitable for the formation of a molecularly imprinted polymer. The plurality of monomers may include monomers selected from the group: vinyl monomers, allyl monomers, acetylenes, acrylates, methacrylates, acrylamides, methacrylamides, chloroacrylates, itaconates, trifluoromethylacrylates, derivatives of amino acids, nucleosides, nucleotides, carbohydrates, or any combination thereof. The plurality of monomers may optionally include cross-linking monomers. Cross- linking monomers are optionally used to stabilize the structure of the resulting polymer, so that it remains bound to that of the peptide. Typical examples of cross-linkers suitable for the synthesis of polymer include, but are not limited to, ethylene glycol dimethacrylate, trimethylolpropane trimethacrylate, divinylbenzene, methylene bisacrylamide, ethylene bisacrylamide and Ν,Ν'- bisacryloylpiperazine. The function of cross-linking agents can be performed by particles or precursor polymers containing double bonds, or particles or polymers with multiple functionalities attached which can bind to functional monomers. Those skilled in the art could select monomers and cross-linkers suitable for a particular system. There are a wide range of possible concentrations of monomers and combinations of monomers that would be suitable for use with the presently disclosed method. While potential ranges and sub-ranges are disclosed herein, the skilled person would be aware that these ranges are merely guides, and that any plurality of monomers suitable for obtain the MIP particle of the invention would be appropriate. For instance, the plurality of monomers or monomer mixture may take any form or composition suitable for the formation of a moleculariy imprinted polymer particle. The monomers may be present in a polymerization mixture in an amount of from about 0.01 to 80% by weight, optionally from about 0.1 to 80% by weight. The plurality of monomers may be prepared in aqueous solution or in appropriate organic solvent. The solution or solvent may optionally comprise an initiator, and optionally buffer salts. The concentration of each type of monomer may be varied depending on the intended final concentration in the reaction mixture. The following concentrations described for N-Isopropylacrylamide (NIPAM), methylene bisacrylamide (MBAA), tert- butylacrylamide, acrylic acid and 3-aminopropyl methacrylate might, for example, be used to form a reaction mixture by the combination of 10 mL of monomeric mixture with 0.7 mL of a mixture comprising the target protein; this example is purely illustrative and is intended to indicate acceptable concentrations in final reaction volume.

The plurality of monomers may comprise N-isopropylacrylamide (NIPAm). The NIPAm may be present at 0.001 mg/mL to 100 mg/mL, 0.01 mg/mL to 10 mg/mL, 0.05 mg/mL to 5 mg/mL, or 0.1 mg/mL to 1 mg/mL.

The plurality of monomers may comprise methylene bisacrylamide (MBAA). The MBAA may be present at 0.0001 mg/mL to 10 mg/mL, 0.001 mg/mL to 5 mg/mL, 0.005 mg/mL to 0.5 mg/mL, or 0.01 mg/mL to 0.1 mg/mL.

The plurality of monomers may comprise tert-butylacrylamide (TBAm). The tert- butylacrylamide may be present at 0.001 mg/mL to 100 mg/mL, 0.01 mg/mL to 10 mg/mL, 0.05 mg/mL to 5 mg/mL, or 0.1 mg/mL to 1 mg/mL. The plurality of monomers may comprise acrylic or methacrylic acid. The acrylic or methacrylic acid may be present at 0.001 mg/mL to 100 mg/mL. The plurality of monomers may comprise 3-aminopropyl methacrylate. The 3-aminopropyl methacrylate may be present at 0.0001 mg/mL to 10 mg/mL, 0.001 mg/mL to 5 mg/ mL, 0.005 mg/ mL to 0.5 mg/mL, or 0.01 mg/mL to 0.1 mg/mL.

Preferably, the plurality of monomers includes monomers selected from N- isopropylacrylamide (NIPAm), Ν,Ν'-methylene-bis-acrylamide (MBAA), n-tert- butylacrylamide (TBAm), acrylic acid, 3-aminopropyl methacrylate or their mixture.

In an embodiment, the plurality of monomers are in a solution comprising phosphate buffered saline (PBS), ammonium formate, or acetate. This mixture may optionally be purged with nitrogen. The purging with nitrogen may be for 1 min to 10 hours, 2 min to 1 hour, or about 5 min to 20 min. In a particular embodiment, the plurality of monomers are included in a deoxygenated mixture of monomers which comprises NIPAM, methylene bisacrylamide (MBAA), tert- butylacrylamide, acrylic acid, 3-aminopropyl methacrylate, and PBS. Many possible mixtures and combinations of monomers are possible, and the invention is not limited to the aforementioned. Any plurality of monomers, and any combination of concentrations, that would be suitable for forming a molecularly imprinted polymer would be suitable for use with the invention. In some embodiments, the plurality of monomers preferably contains magnetic nanoparticles that are incorporated into the polymer.

The step of (ii) polymerizing the plurality of monomers to create a peptide: polymer complex, may be initiated chemically, thermally, or optically. The thermal initiation may be the application of heat. The optical initiation may be a photo-initiation. The optical initiation may be the use of UV or visible light. The chemical initiation may be carried out by an initiator or an initiation solution. The polymer may be synthesized by free radical polymerization living polymerization, ionic polymerization, or polycondensation. Several different forms of polymerizations are suitable for use with the present invention including bulk polymerization, polymerization in suspension and emulsion, precipitation polymerization, living polymerization, grafting or any other form known in the art. The physical form of synthesized polymer can be solid particulate, layer or coating, powder or monolith, micro- or nanoparticles. The optimal composition of mixture of polymers and conditions of polymerization reaction are those that preserve native structure of peptide target. Some examples of polymerization include, but are not limited to: iniferter polymerisation, nitroxide-mediated polymerization, atom-transfer radical polymerization and reversible addition-fragmentation chain- transfer polymerization. The advantage of living polymerization in contrast to traditional radical polymerization is that the former proceeds with a low rate producing nanopartides with large contact area for interacting with protein. The reaction conditions suitable for use with the presently disclosed process for the polymerization reaction and leading to formation of MIP particle include, but are not limited to: (i) using stoichiometric ratio between initiator and plurality of monomers; (ii) cooling reaction or stopping UV or other irradiations; (iii) removal of the monomers from contact with the growing polymer chain by e.g. filtration or chromatography; (iv) adding inhibitors to the reaction; (v) performing polymerization in dilute solution.

Conventional free-radical generating polymerization initiators may be employed to initiate polymerization. Examples of suitable initiators include peroxides such as OO-t-amyl-0-(2ethylhexyl)monoperoxycarbonate, dipropyl peroxydicarbonate, and benzoyl peroxide, as well as azo compounds such as azobisisobutyronitrile, 2,2'- azobis(2- amidinopropane)dihydrochloride, 2,2'-azobis(isobutyramide)dihydrate and l,l'-azobis (cyclohexane carbonitrile). Examples of initiators include photo- iniferter bearing a dithiocarbamyl group, or azo compound. Other examples of suitable initiators include, but are not limited to: 2-bromopropionitrile with Cu(I)Br complexed with N,N,N',N",N"-pentamethyldiethylenetriamine, polystyrene bromo macroinitiator with Cu(I)CI/PMDETA; ethyl 2-bromoisobutyrate with CuCI/bi pyridine; i,4-bis(2,6-diisopropylphenyl)acenaphatenediiminenickel (II) dibromide; 2,2-dimethoxy-2-phenylacephenone in combination with tetraethylthiuram disulfide; tetraphenyl biphosphine; tertiary peroxides such as di- tert-butyl peroxide; SmMe(C ₅ Me ₅ ) ₂ (THF); styrene-based epoxides in conjunction with TiCI ₄ ; methylstyrene tetramer disodium; MoOCI ₄ -n-BuSn-EtOH; HCI/ZnCI ₂ ; methyl p-toluenesulphonate; 2,10,15,20-tetraphenylporphinato aluminium methyl; 3-methyl-i,i-diphenylpentyllithium; butyllithium in THF; molybdenum alkylidine compounds; bifunctional organolanthanide(in) ; Mo(CH-t-Bu)(NAr)(OCMe ₃ ) ₂ and Mo(CHCPhMe ₂ )(NAr)(OCMe(CF ₃ ) ₂ ) ₂ ; HI/I ₂ ; Zr, Ti and Hf complexes combined with either methylaluminoxane or phenyl borates; diimide complexes of Pd, Ni, Fe or Co; homogeneous Ta, Ti, Mo, W carbene complexes; rare earth metal complexes composed of metallocene type or non-metallocene type complexes; cationic monocydopentadienyl zirconium acetamidinate complexes; esterified fluorinated telomers with one or two hydroxyl group as initiators for copper mediated living polymerization; Yb[C(SiMe ₃ ) ₃ ] ₂ . The initiator can be present in the polymerization mixture in an amount of from about 0.01% to 20% by weight of the monomers, preferably from 0.05 to 15%, or more preferably from 0.1 to 10% by weight of the monomers. The initiator is preferably present in the polymerization mixture in an amount of from about 0.2 to 5% by weight of the monomers. In a particular embodiment, an initiation solution can be used to initiate step (ii) of polymerizing the plurality of monomers. This solution may comprise TEMED and / or ammonium persulfate (APS), preferably a mixture of TEMED and APS. TEMED may be at a concentration in an initiation solution of: 0.001 μg/mL to 1 mg/mL. APS may be at a concentration in an initiation solution of: 0.1 μg/μL to 1000 μg/μL.

Preferably, the step of (ii) polymerizing the plurality of monomers to create a peptide: polymer complex is initiated by chemical initiation, in particular using a mixture of TEMED and APS. The step of (ii) polymerizing the plurality of monomers can be carried out as long as needed to obtain the MIP particle, for example between 30 minutes to 5 hours, such as during 1 hour.

The initiation solution may comprise PBS or other buffer salts. The role of buffer is to maintain or control intended conformation of peptide target during polymerization. The polymer product may comprise particles with a mass of at least 100 Da, 500 Da, or 1000 Da. The polymer product may comprise particles with a mass of at least 10 kDa, 50 kDa, or 100 kDa. The polymer product may comprise particles with a size 10 nm to 100 μm, 1 μm to 100 μm, 5 μm to 50 μm, 10 μm to 30 μm, or 15 μm to 25 μm. The preferred form of the polymer product is particles with size 500 Da -20 μm which could exist in soluble or colloidal forms. In some embodiments, the polymer product may have a magnetic core. In some embodiments synthesized polymers are linear or cross-linked.

The step of (iii) isolating the peptide: polymer complex, to obtain MIP particle may involve an increase in the purity or the concentration of the peptide: polymer complex within a reaction mixture. Alternatively, the isolation step may include the separation of the peptide: polymer complex from unspecific polymer complex. In some embodiments, the isolation step (iii) may include a washing step or washing steps. The isolation step (iii) may involve the separation of the peptide: polymer complex from unspecific polymer complex, unreacted monomers and/or from free peptides. In some embodiments the complex of peptide with synthesized polymer is separated from unreacted monomers and other components or products of any preceding reaction by ultrafiltration, ultracentrifugation, electrophoresis, sonication, chromatographic separation, washing, adding urea, guanidine, by using magnetic forces, or any combination thereof. To facilitate separation of the peptide-polymer complex from unbound monomers, the peptide or polymer is preferably linked to a solid support (such as a bead or matrix, including a microarray or multi-well plate) and or any other agent known in the art. Linking may be covalent or non-covalent. Methods for linking polymer or peptide to solid support are well known in the art. The MIP particle may be released from the polymer-peptides complex using conventional elution conditions such as change of temperature, pH, ionic strength, detergents, or any combination thereof. Preferably, the MIP particle is released from the polymer-peptide complex by using an ethanol solution, such as an hot ethanol solution (for example about 60°C). Preparation of the MIP particles according to the invention is described in the Examples of the present description. Figure 2 illustrates the general steps for preparing MIP particles.

Support

The invention also relates to a support onto which are grafted one or more MIP particles according to the invention.

MIP particles of the invention which specifically bind to the peptide of the invention can be used to bind anti-AAV antibodies, because said MIP particles are able to bind variable regions of anti-AAV antibodies (i.e. MIP Particles are antiidiotype particles). The MIP particles are "AAV capsid like" and can be grafted on a support. Said support may therefore be used in a plasmapheresis device to remove AAV-specific neutralizing antibodies from the blood material of patients.

In particular, the support of the invention is grafted with one or more MIP particles selected from (i) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 1, more preferably 100% identity with

SEQ ID NO 1,

(ii) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 2, more preferably 100% identity with SEQ ID NO 2,

(ill) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 3, more preferably 100% identity with

SEQ ID NO 3,

(iv) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 4, more preferably 100% identity with SEQ ID NO 4,

(v) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 5, more preferably 100% identity with

SEQ ID NO 5,

(vi) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 6, more preferably 100% identity with

SEQ ID NO 6, or (vii) at least two MIP particles selected from (I) to (vi) , for example at least three MIP particles selected from (I) to (vi), at least four MIP particles selected from (I) to (vi), at least five MIP particles selected from (i) to (vi) or the six MIP particles from (i) to (vi).

In particular, the support is a support suitable for clinical use, i.e. a support complying with regulatory safety provisions for devices to be used in purification/preparation method of biopharmaceuticals for animal and human use.

The support of the invention may correspond to any kind of support onto which one or more MIP particles can be bound, either covalently or non-covalently. The support may correspond to any type of support, such as a compressible (such as a compressible smooth gel, e.g. sepharose) or non-compressible support (such as a robust incompressible high porosity support), in particular a sepharose, sephadex, agarose, cellulose, modified cellulose, CPG, poros or monolith support. In a further particular embodiment, the support is a sepharose support. The attachment need not be covalent, but is at least of sufficient permanence to withstand any separation techniques (including washes and elution) that may be part of the method of the present invention. In a preferred embodiment, attachment to the support is via a covalent bond. In a particular embodiment, the one or more MIP particles are covalently linked to the support. The support may be, for example, a chomatographic column, particles or beads, monoliths, a membrane or a filter. The support may further be a hollow fiber cartridge. The support may be an activated support, comprising functional groups allowing covalent chemical conjugation of the one or more MIP particles. Such activated supports include, without limitation cyanogen bromide (CNBr)-activated supports, N- hydroxysuccinimide- activated supports, carbonyl diimidazole (CDI)-activated supports, and the like. Commercially available coupling materials that may be used as described herein include, for example, CNBr Sepharose Fast Flow, NHS Sepharose Fast Flow, Epoxy Sepharose 6B, Thiol Sepharose Fast Flow, EAH Sepharose Fast Flow, Epoxy Poros EP, Aldehyde Poros AL, Epoxy Poros EP, Hydroxylated Poros OH and CDI and epoxy Monolithic materials. In a particular embodiment, the support is a NHS Sepharose support. Coupling onto such supports is well known in the art, and is typically done following the manufacturer's instructions. In addition, depending on the support chemistry and/or its coupling chemistry, a protein cross-linking compound such as formaldehyde may be used. Such a cross-linking compound may in particular be implemented on supports that activated with more labile functional groups. Accordingly, in a particular embodiment, the support functional groups are epoxy groups, and formaldehyde is used to cross-link the MIP particles of the present invention.

According to the present invention, the MIP particle density on the support may vary to a large extent. For example, for a volume of NHS Sepharose support of 1 mL, from 0.01 to 30 mg of MIP particles may be grafted, such as for example 30 mg of MIP partides/mL. In particular, for a volume of NHS Sepharose support of 5 mL, from 0.05 to 150 mg of MIP particles may be grafted, such as 150 mg of MIP particles.

Method for removing anti-AAV antibodies from Mood material

The invention also relates to a method for removing ex vivo anti-AAV antibodies from blood material, comprising contacting said blood material with at least one support according to the invention.

According to the method of the invention, the at least one support of the invention is grafted with one or more MIP particles selected from:

(i) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 1, more preferably 100% identity with SEQ ID NO : 1,

(II) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 2, more preferably 100% identity with SEQ ID NO : 2,

(ill) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 3, more preferably 100% identity with SEQ ID NO : 3,

(iv) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 4, more preferably 100% identity with SEQ ID NO : 4, (v) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 5, more preferably 100% identity with

SEQ ID NO : 5,

(vi) a MIP particle which specifically binds to a peptide comprising an amino acid sequence having at least 80% identity, such as at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, preferably at least 90% identity such as at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% with SEQ ID NO : 6, more preferably 100% identity with SEQ ID NO : 6, or

(vii) at least two MIP particles selected from (i) to (vi) , for example at least three MIP particles selected from (i) to (vi), at least four MIP particles selected from (i) to (vi), at least five MIP particles selected from (i) to (vi) or the six MIP particles from (i) to (vi).

The blood material can be treated once or several times through the at least one support of the invention. The blood material can be passed through two supports, three supports, four supports, five supports or six supports. The at least one supports may be grafted with MIP particles which specifically bind to one peptide of the invention or with different MIP Particles which specifically bind different peptides of the invention. The supports may be serially arranged. For example, the method of the invention may use a first support onto which is grafted a MIP particle which specifically binds to a peptide according to the invention, and a second support onto which is grafted another MIP particle which specifically binds to another peptide according to the invention, i.e. the MIP particle grafted onto the first support is different from the MIP particle grafted onto the second support. In an alternative example, the method of the invention may use a first support onto which is grafted a first MIP particle of the invention which binds to a first peptide of the invention and a second MIP particle of the invention which binds to a second peptide of the invention, and a second support onto which is grafted a third MIP particle of the invention which binds to a third peptide of the invention. In some embodiments, the blood material is contacted with the support at a flow rate between 0.01 and 1 mL/min, such as between 0.05 and 0.7 mL/min, for example between 0.1 and 0.5 mL/min (or 0.02 and 0.1 CV/min, wherein "CV" stands for "column volume"). In a particular embodiment, the flow rate is about 0.1 mL/min (or 0.02 CV/min). Thanks to the method of the invention, anti-AAV antibodies, such as anti-AAV NAbs are removed or depleted from the blood material. According to the invention, total depletion of anti-AAV antibodies, in particular NAbs, is not required, as long as the reduction in antibody titers allows an improvement in AAV vector transduction when AAV-based gene therapy is carried out after implementation of the above method. For example, antibody titer reduction may be of at least 1 log, 2 logs, 3 log, or at least 4 logs. For example, with the method of the invention, an anti-AAV neutralizing antibody titer of 1: 3.160 can be reduced to 1 : 1 (negative titer).

Device

The invention also relates to an extracorporeal device comprising the support according to the invention. The extracorporeal device can be used in a purification/preparation method of biopharmaceuticals for animal and human use. In particular, the extracorporeal device could be a classical plasmapheresis device with specific column grafted with MIP particles of the invention.

EXAMPLES

Example 1: Affinity test of the peptides of the invention to AAV-8 capsid Materials and Methods

Six peptides (PI to P6) were synthesized, immobilized onto Biacore gold chips 3000 and tested for their ability to bind AAV8 capsid. The peptides comprised small linkers at their N-terminal end consisting of cysteine or cysteine-glycine for the purpose of easy immobilization on the gold chips.

Results

The Results are presented on table 2. All synthesized peptides have shown strong binding to AAV8 with Kd at nM level. The percentage of binding to AAV is also presented in table 2 and was obtained by the Surface Plasmon resonance optical sensor platform Biacore 3000 (GE Healthcare Life Sciences, UK) at 25 °C using PBS (0.01 M phosphate buffer, 0.0027 M potassium chloride and 0.137 M sodium chloride, pH 7.4) as the running buffer at flow 35 μL min ^-1 . The self-assembled gold sensor chip (SA, GE Healthcare Life Sciences, UK) has been cleaned using plasma and placed in the ethanol solution of mercaptododecanoic acid (2.2 mg mL ^-1 ) where they were stored until use. The peptides were immobilized on the surface of the carboxylated sensor chips using thiol coupling using a protocol recommended by manufacturer (GE Healthcare Life Sciences, UK). The AAV-8 particles were diluted in PBS in the range of concentrations between 1.4 nM and 44.8 nM and injected sequentially using KINJECT mode (injection volume- 100 μL and dissociation time- 120 sec). Dissociation constants (Kd) were calculated from plots of the equilibrium biosensor response using the BiaEvaluation v4.1 software using a 1:1 binding model with drifting baseline (DB) fitting. The calculation of the dissociation constant was also done using Langmuir Blodgett (LB) algorithm using the AB (absorption) component of the SPR response, which was obtained after the subtraction of the drift and bulk effect. Table 1: sequence of peptides PI to P6 tested on their affinity to AAV8 (P1-P6) Example 2: Binding inhibition between oolv Antibodies and AAV-8. in presence of the peptides of the invention Preparation of the peptide solution :

Aqueous solutions of the peptides of SEQ ID NO : 7 (P1), SEQ ID NO : 8 (P2), SEQ ID NO : 9 (P3), SEQ ID NO : 10 (P4), SEQ ID NO : 11 (P5) and SEQ ID NO : 12 (P6) with concentration of 1 mg mL ^-1 were prepared (as described in table 2).

The peptides P2 and P3, which had lower solubility than the others, were first solubilized in DMSO and then added to water (10:90 ratio). In order to prepare the mixture of the peptides, equal aliquots were mixed.

Preparation of the Agile chip: The COOH-Agile chip was docked into the instrument Agile R100 (Nanomedical diagnostics, USA) and conditioned with 50 mM MBS buffer, pH 6 for 15 min followed by activation using EDC/NHS mixture according to the manufacturer's protocol. This step of conditioning has allowed to equilibrate and/or stabilize the conditions before beginning the experiment. Then, AAV8-specific poly antibodies (pAbs) were immobilized onto the chip by adding a solution of AAV8-specific poly antibodies (pAbs) onto the chip surface and incubating for 15 min. After immobilization of pAbs the chip was washed with MBS. The surface of the chip was blocked using 1 M ethanolamine at pH 8.5 for 15 min and washed with phosphate buffer saline (PBS).

Virus testing:

A solution of a mixture of the six peptides PI to P6 (10 μg of each peptide) was mixed with a solution of AAV8 GFP particles (50:50 ratio), added to the sensor surface with immobilized AAV8-specific pAbs and incubated for 5 min. The dissociation was conducted for 5 min using PBS. Similarly, a solution of AAV8 GFP particles without peptides (mixed 50:50 with water instead of peptide solution) was added to the sensor surface and incubated for 5 min. Dissociation was conducted for 5 min using PBS. Results:

The results are presented in Figure 3 (in Response Units (RU), showing the sensor response to the binding) and show that the peptides inhibit the binding of AAV8 to the AAV8-specific pAbs. In particular, Figure 3 shows the binding between the anti-AAV8-specific antibodies immobilized on the sensor surface and the complex AAV8/peptides. The dissociation part of the sensor gram shows the dissociation between the specific anti-AAV8 antibodies immobilized on the surface and the complex AAV8/peptide. When AAV-8 is incubated without peptides (see «injections of AAV8»), the binding is higher and the dissociation is slower, which means that the association between anti-AAV8 antibodies and AAV8 is inhibited in the presence of the peptides. It was observed that the peptides have inhibited binding between AAV8 capsids and virus-specific IgG immobilized on the sensor chip approximately by 65%.

Example 3: Preparation of MIP particles 1 to 6. which bind to peptides PI to P6 respectively·

The peptides described in table 2 were used for preparing the MIPs. Figure 2 illustrates the general steps for preparing MIP particles.

Solid-phase synthesis of MIP particles specific for P1-P6: Glass beads were activated using 4 M sodium hydroxide followed by washing with water and neutralization using PBS. The glass beads were then silanized using N- (3-(trimethoxysilyl)propyl)ethylenediamine (see below).

Iodoacetic acid N-hydroxysuccinimide ester (SIA) (10 mg) was added to silanized glass beads (120 g) in anhydrous acetonitrile (50 mL) and incubated for 2 h under exclusion of light, before washing with acetonitrile (5 x 50 mL). Thiol buffer (pH 8.2), consisting of phosphate buffered saline (PBS) (50 mL) and ethylenediaminetetraacetic acid (EDTA) (74 mg, 500 μmol, 5mM), was degassed and purged with nitrogen prior to addition of the peptide (5 mg).

All peptides were synthetized with a cysteine amino acid at their N-terminal end which was used for immobilization through thiol coupling. The peptides PI to P6 (as described in Table 3) were immobilized respectively onto different solid phase (One solid phase by peptide, i.e. 6 solid phases), forming therefore six different columns. Incubation with SIA-linked glass beads (120 g) was conducted overnight with exclusion of light, followed by washing with water (1.5 L) and drying under vacuum (see below). Each solid phase with immobilized peptide was then used for the synthesis of MIP particles.

Synthesis of MIP particles:

The following mixture of monomers was prepared: 19.5 mg of N- isopropylacrylamide (NIPAm), 3 mg of N,N'-methylene-bis-acrylamide (MBAA), 15 mg of n-tert-butylacrylamide (TBAm) dissolved in 200 pi of absolute ethanol, 50 μL of the solution of 22 μL of acrylic acid in 1 ml. of water, 3 mg of 3-aminopropyl methacrylate were dissolved in 50 mL of phosphate buffered saline (PBS) and purged with nitrogen for 20 min.

50 mL of the mixture of monomers was added to 60 g of the solid phase with immobilized peptides and purged with nitrogen for 5 minutes. The polymerization was initiated by adding 500 μL of freshly prepared solution of 24 mg potassium persulfate (APS) and 12 μL of TEMED dissolved in 800 μL of PBS. The polymerization was carried out for 1 hour at a room temperature of 22°C.

After polymerization, the glass beads were washed with 100 mL of water at room temperature and with 100 mL of hot water (60 °C). The elution of the MIP particles was conducted using 100 mL of hot ethanol (60 °C). The ethanol solution of MIP particles was concentrated and reconstituted with water followed by dialysis for 3 days. The MIP particles were stored in the fridge at 4 °C until their use. The MIP particles obtained and their concentrations are presented in table 3 below.

Table 2: characteristics of the Pl-P6-specific MIP nanoparticles

Example 4: Binding of anti-AAV8 IaG onto a column grafted with MIP particles.

0.1 mg of MIP particles obtained in example 3 (MIP 1 particles, which specifically binds to peptide PI of SEQ ID NO: 7, also namedP1705418) were grafted on a GE Healthcare NHS-Sepharose column. Grafting with N-HydroxySuccinimide consists in using N-hydroxysuccinimide coupling which forms a chemically stable amide bond with a primary amino group of MIP particles. A commercial solution of IVIG (TEGELINE obtained from LFB BIOMEDICAMENTS) was used and prepared from a pool of a thousand donors, some of which being immunized against AAV-8 capsid. The IVIG solution was reformulated in PBS with a PD-10 column (GE). The immunoaffinity column was first loaded with 3 ml of IVIG solution (see Figure 4) with a flow rate of 0.1 mL min ^-1 . The flow-through (FT) corresponds to the IVIG solution that passed through the immunoaffinity column without binding to the grafted MIP particles. Then, the immunoaffinity column was washed with 25 mL of PBS running buffer to eliminate impurities, which were not bound to the MIP particles. Then, a citrate buffer (50 mM) having a pH of 3 was used for eluting the IVIG bound to the MIP particles. Three fractions were obtained: the FT fraction, the WASH fraction and the ELUTION fraction. The antibody titer (anti-AAV8 antibodies IgG) retained by the column was determined with an ELISA test. Briefly, wells were coated overnight at 4°C with AAV8 capsid ( 1x10 ¹⁰ capsid particles mL ^-1 , 50 μL per well), blocked with PBS and 6% non-fat dry milk for 2 h at room temperature, and serum or plasma fractions were added to the coated wells in duplicate. Goat anti-human IgG-HRPs (horse radish peroxidase) were used as detecting antibodies. IgG concentration was determined against standard curves made with serial dilution of IVIG commercial solution.

The IgG fraction retained on the column by MIP particles was determined with the following calculation:

Results

The results are presented in figures 5 and 6. The Y axis is in nanogram (ng) of specific anti-AAV8 IgG and the X axis represents the fraction tubes. Loading and washing represents fractions 4 at 10; elution with citrate buffer represents fraction 23 at 31. The elution fraction was 17% of retention, as measured by the ELISA test. The retained IgG fraction of 17%is a good retention percent considering that only one type of MIP particles was used. This experiment confirms that MIPs 1 particles allow the binding of anti-AAV8 IQG.

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