Anti-viral conjugates

Abstract

This invention relates to the prevention or treatment of coronavirus infections and flavivirus infections, including ZIKV and DENV, using porphyrin-peptide conjugates. The conjugates comprise (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1, 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX. Methods of treatment and conjugates for use in methods of treatment are provided.

Description

[0001] Anti-Viral Conjugates

[0002] Field of the Invention

[0003] The present invention relates to porphyrin-peptide conjugates for prevention and treatment of viral infections. In particular, conjugates are provided for prevention and treatment of flavivirus infections (including dengue and Zika virus infections) and coronavirus infections (including SARS-CoV-2 infections). New porphyrin-peptide conjugates are also provided.

[0004] Background

[0005] Among human-targeting viruses, brain-penetrating ones such as flaviviruses pose formidable hurdles to pathogen targeting.

[0006] Flaviviruses are a family of positive-sense, single-stranded, enveloped RNA viruses that may infect humans and pose a significant threat to public health. In particular, flaviviruses are the causative agent of Zika fever, dengue fever, Japanese encephalitis, yellow fever and West Nile fever. Flaviviruses are known to be able to cross the blood-brain barrier (Chen et al., Reviews in Medical Virology 29(1): e2021 , 2019) and thus cause neurological symptoms.

[0007] Flaviviruses are arboviruses, meaning that they are transmitted by infected arthropod vectors such as mosquitos and ticks. The geographical distribution of flaviviruses is primarily determined by that of their arthropod vector. For the most part, the vectors are confined to tropical and sub-tropical regions, such as Southeast Asia and South America. However, climate change is broadening the distribution of some vectors, thereby increasing the population at risk of contracting flavivirus infections. Furthermore, the mosquito responsible for spreading Zika virus and yellow fever virus has been shown to be able to adapt to survive in high-density urban areas. It is therefore important to find effective methods for containing flavivirus infection.

[0008] While some flaviviruses (such as West Nile virus) only incidentally infect humans, other flaviviruses (such as yellow fever virus, Dengue virus and Zika virus) exist predominantly in an arthropod-human life cycle. Such flaviviruses grow well in the human host, and high viral titres allow infection to cycle back to arthropod vectors and onto new human hosts. In either case, vector-borne transmission and the ability to infect other species such as monkeys and birds means that flavivirus infections tend to spread quickly and easily. Controlling the spread of flavivirus infections is therefore challenging.

[0009] The structure of the flavivirus genome also contributes to the challenge of controlling spread. Few proof-reading and correction mechanisms exist for the replication of singlestranded RNA. Therefore, mutations arising in the course of replication frequently remain in the genome and are passed to the next generation. Flaviviruses therefore evolve quickly, which can make the development of effective antivirals challenging. Zika virus (ZIKV) is a particularly illustrative case in the challenge posed by brainpenetrating viruses: ZIKV, a member of the Flaviviridae family, causes the namesake Zika fever, a zoonosis transmitted by Aedes mosquito bites. The direst consequences of ZIKV infection concern pregnant women, as virus transmission to the foetus causes irreversible congenital brain abnormalities, including microcephaly, a predicament involving over 1400 cases during the 2016 outbreak in Brazil (Peiter et al., PLoS ONE 15: e0235010, 2020). ZIKV infection can also result in further complications such as foetal loss, preterm birth or other birth defects, and can also trigger Guillain-Barre syndrome (Parra et al., New England Journal of Medicine 375: 1513-1523, 2016) and encephalomyelitis and similar conditions (Niemeyer et al., European Neurology 77: 45-46, 2017) in adults. ZIKV was detected in a microcephalic foetus ca. 32 weeks after maternal exposure, suggesting long persistence in the foetal brain (Mlakar et al., New England Journal of Medicine 374: 951-958, 2016).

[0010] Dengue virus (DENV) is another medically challenging member of the Flaviviridae. It is the causative agent of dengue fever, which can progress into life-threatening dengue haemorrhagic syndrome or dengue shock syndrome. Neuroinvasion by dengue virus can cause encephalopathy, encephalitis and meningitis, and autoimmune conditions affecting the CNS such as acute disseminated encephalomyelitis, neuromyelitis optica, optic neuritis, myelitis and Guillain-Barre syndrome (Li etal., Frontiers in Cellular and Infection Microbiology 7: 449, 2017).

[0011] Moreover, brain penetration is not a capability limited to flaviviruses. SARS-CoV-2, the causative agent of COVID-19, has been found to be capable of neuroinvasion, causing a variety of symptoms from severe headaches to stroke (Dias de Melo et al., Nature Communications 14: 4485, 2023).

[0012] Effective vaccines have been developed for SARS-CoV-2, but COVID- 19 disease remains a substantial burden on global health systems. Flavivirus vaccine development is ongoing, with a single vaccine against DENV currently approved, but there is an unmet need for effective antivirals against these diseases. The need for antivirals to cross the blood-brain barrier (BBB) for effective treatment of these neuroinvasive viruses is a complicating factor. For treatment of ZIKV (and other infections caused by viruses capable of transmission from mother to foetus) antivirals must also be capable of crossing the blood-placenta barrier (BPB).

[0013] Porphyrins have been recently reported as displaying anti-viral activity against enveloped viruses (Cruz-Oliveira etal., Antimicrobial Agents and Chemotherapy 61 : e00053-17, 2017), a group that includes the aforementioned brain-penetrating ones. Porphyrins are tetrapyrrole macrocycles, linked by methine bonds, with the four pyrrole nitrogens defining a metal ion coordination site involved in biological processes such as respiration and photosynthesis, as well as in diagnostic and therapeutic applications. Their antiviral activity can be explained by two proposed mechanisms: (i) photoactivation, a task not simple to perform when treating brain-resident viruses; (ii) viral envelope targeting, a primary mechanism in the absence of light, where virus infectivity is blocked by porphyrin local accumulation and perturbation of the viral lipid bilayer, an approach suitable to inhibiting brain-resident viruses. Unfortunately, like other antivirals, most porphyrins are unable to cross the BBB, which hinders their use against viral brain infections.

[0014] However, conjugation of porphyrins to trans-BBB shuttle peptides has previously been found to successfully deliver the porphyrins across an in vitro model of the BBB (Mendonga et al., Bioconjugate Chemistry 32: 1067-1077, 2021). The porphyrin-peptide conjugates (PPCs) were found, in some cases, to display antiviral activity against HIV (Mendonga et al., supra) and ZIKV (Todorovski et al., Pharmaceutics 14: 738, 2022). However, the properties of a PPC, including whether it is capable of crossing the BBB or BPB, and whether it displays antiviral activity against a given target virus, and if so to what extent, have been found to be unpredictable. PPC properties have been found to depend not only on the exact combination of porphyrin moiety and trans-BBB shuttle peptide, but also on the orientation of the conjugation (i.e. whether the porphyrin moiety is conjugated to the N- or C-terminus of the peptide).

[0015] Summary of the Invention

[0016] Provided herein are porphyrin-peptide conjugates for use in the prevention or treatment of flavivirus infections, including ZIKV and DENV, and coronavirus infections, particularly SARS-CoV-2 infection. Certain porphyrin-peptide conjugates are also provided.

[0017] Thus in a first aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of a flavivirus infection, wherein the conjugate comprises:

[0018] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0019] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0020] Relatedly, the invention provides a method of treating or preventing a flavivirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising:

[0021] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0022] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0023] Also relatedly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0024] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0025] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a flavivirus infection in a subject. In a second aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of dengue virus infection, wherein the conjugate comprises:

[0026] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0027] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0028] Relatedly, the invention provides a method of treating or preventing dengue virus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising, comprising:

[0029] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0030] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0031] Also relatedly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0032] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0033] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a dengue virus infection in a subject.

[0034] In a third aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of a coronavirus infection, wherein the conjugate comprises:

[0035] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0036] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0037] Relatedly, the invention provides a method of treating or preventing a coronavirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising:

[0038] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0039] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0040] Also relatedly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0041] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0042] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a coronavirus infection in a subject. In a fourth aspect, the invention provides a porphyrin-peptide conjugate comprising mesoporphyrin IX conjugated to a peptide consisting of the amino acid sequence of SEQ ID NO: 3, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto, wherein a carboxylic acid group of the porphyrin moiety is conjugated to the N-terminal alpha-amino group of the peptide via a linker comprising a dimer of 8-amino-3,6-dioxaoctanoic acid.

[0043] In a fifth aspect, the invention provides a porphyrin-peptide conjugate of the fourth aspect for use in therapy.

[0044] Detailed Description of the Invention

[0045] As set out above, the present invention relates to porphyrin-peptide conjugates (PPCs). A porphyrin peptide conjugate comprises a porphyrin moiety conjugated to a peptide.

[0046] Porphyrin Moieties

[0047] The porphyrin moiety in the PPCs of the present invention may be mesoporphyrin IX (MPIX) or protoporphyrin IX (PPIX), as set out in more detail below. MPIX has the structure set forth in Formula (A), and PPIX has the structure set forth in Formula (B). Formula (B):

[0048] The peptide of the PPC is conjugated to the porphyrin moiety via a carboxyl group. The PPCs provided herein thus have a structure selected from the following:

[0049] Formula (A1): Formula (A2):

[0050] Formula (B1): Formula (B2):

[0051] Formulae (A1) and (A2) are mesoporphyrin IX (MPIX) conjugates and formulae (B1) and (B2) are protoporphyrin IX (PPIX) conjugates. As referred to herein, an MPIX conjugate may have the structure of formula A1 or A2. The structures of formulae A1 and A2 are interchangeable and any reference herein to a structure as set out in formula A1 or A2 should be understood as encompassing both the A1 and A2 structures. Similarly, as referred to herein a PPIX conjugate may have the structure of formula B1 or B2. The structures of formulae B1 and B2 are interchangeable and any reference herein to a structure as set out in formula B1 or B2 should be understood as encompassing both the B1 and B2 structures. In the formulae above, (L) represents an optional linker, and X represents a peptide. Peptides which may be used in the conjugates of the invention are described below.

[0052] Porphyrin moieties may be metalated or non-metalated. In metalated porphyrins, a metal ion displaces the protons from the pyrrole groups and is chelated by the nitrogen atoms. Metalated porphyrins may comprise e.g. Zn2+, Fe2+or Sn4+. Herein, the terms “porphyrin moiety”, “mesoporphyrin IX” and “protoporphyrin IX” may refer to metalated or non-metalated versions of the compounds. Preferably, the terms “porphyrin moiety”, “mesoporphyrin IX” and “protoporphyrin IX” refer to non-metalated versions of the compounds.

[0053] Peptides

[0054] The peptide used in the PPCs described herein are cell-penetrating peptides capable of crossing the blood-brain barrier (BBB), and optionally also capable of crossing the bloodplacenta barrier (BPB), as shown in the Examples below. Such cell-penetrating peptides may be referred to as BBB shuttles. Conjugation of a payload (such as a porphyrin moiety) to the BBB shuttles used herein enables delivery of the payload across the BBB (and optionally across the BPB as well, where the BBB shuttle is also capable of crossing the BPB).

[0055] The BBB shuttles of SEQ ID NOs: 1 (pepH1) and SEQ ID NO: 3 (pepH3) (see WO 2016 / 120843) are derived from the DENV type 2 capsid protein (DEN2C). Specifically, pepH1 corresponds to amino acids 26-35 of DEN2C and pepH3 corresponds to amino acids 63- 69 of DEN2C. The BBB of SEQ ID NO: 2 (pepNeg) was designed as the “negative image” of pepH3, by switching cationic residues (Lys, Arg) in pepH3 to anionic (Glu), thereby changing the overall positive charge in pepH3 to negative in pepNeg (Neves-Coelho et al., Molecules 22: 1753, 2017).

[0056] The BBB shuttles used herein may be conjugated to the porphyrin moiety via any suitable functional group, e.g. an amino, hydroxyl, thiol or carboxyl group. For example, the BBB shuttle may be conjugated to the porphyrin moiety via its N-terminal alpha-amino group, the side chain (E) amino group of a lysine residue, the hydroxyl group of a threonine or serine residue, the thiol group of a cysteine residue the side chain carboxyl group of an aspartic or glutamic acid residue or the C-terminal carboxyl group. Where the BBB shuttle is directly conjugated to a carboxyl group of the porphyrin moiety (i.e. without a linker), conjugation via an amino group forms an amide group, conjugation via a hydroxyl group yields an ester group and conjugation via a thiol group yields a thioester group. Conjugation of a carboxyl group of the BBB shuttle to a carboxyl group of the porphyrin moiety must take place via a linker.

[0057] Generally it is advantageous for conjugation to be made via an amide bond, as these are more resistant to enzymatic cleavage than ester bonds. Accordingly, in preferred embodiments, a carboxyl group of the porphyrin moiety is conjugated to an amino group of the BBB shuttle peptide. In one preferred embodiment, a carboxyl group of the porphyrin moiety is conjugated to the N-terminal amino group of the BBB shuttle. In another preferred embodiment, a carboxyl group of the porphyrin moiety is conjugated to the E-amino group of a lysine side chain.

[0058] Preferably, the porphyrin moiety is conjugated to either the N-terminal or the C-terminal amino acid of the BBB shuttle. When the porphyrin moiety is conjugated to the N-terminal amino acid, it is generally conjugated to the N-terminal amino group (i.e. the a-amino group). Where the porphyrin moiety is to be conjugated to the C-terminal amino acid of a BBB shuttle, generally a lysine residue is added to the C-terminus of the shuttle peptide, and the porphyrin moiety conjugated to the amino group of that lysine residue. For example, SEQ ID NO: 4 corresponds to SEQ ID NO: 3 with a C-terminal lysine residue for porphyrin conjugation.

[0059] The PPCs of the present invention, as described further below, comprise a porphyrin moiety conjugated to a BBB shuttle peptide of any one of SEQ ID NOs: 1 to 4, or a variant of any one of SEQ ID NOs: 1 to 4 comprising up to 2 amino acid substitutions, deletions and / or additions relative to one of SEQ ID NOs: 1 to 4. That is to say, a variant of one of SEQ ID NOs: 1 to 4 for use herein may comprise a single amino acid substitution, deletion or addition relative to one of SEQ ID NOs: 1 to 4. Alternatively, a variant of one of SEQ ID NOs: 1 to 4 for use herein may comprise two amino acid substitutions, deletions and / or additions relative to one of SEQ ID NOs: 1 to 4. Such a variant may comprise two of the same type of sequence modifications, i.e. two amino acid substitutions, two amino acid deletions or two amino acid additions relative to one of SEQ ID NOs: 1 to 4. Alternatively it may comprise two different types of sequence modification, e.g. one amino acid substitution and one amino acid deletion, or one amino acid substitution and one amino acid addition.

[0060] Where a variant of one of SEQ ID NOs: 1 to 4 comprises an amino acid substitution relative to one of SEQ ID NOs: 1 to 4, that substitution may be a conservative amino acid substitution. The term "conservative amino acid substitution", as used herein, refers to an amino acid substitution in which one amino acid residue is replaced with another amino acid residue having a similar side chain. Amino acids with similar side chains tend to have similar properties, and thus a conservative substitution of an amino acid important for the structure or function of a polypeptide may be expected to affect polypeptide structure / function less than a nonconservative amino acid substitution at the same position. Families of amino acid residues 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. asparagine, glutamine, serine, threonine, tyrosine), non-polar side chains (e.g. glycine, cysteine, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan) and aromatic side chains (e.g. tyrosine, phenylalanine, tryptophan, histidine). Thus a conservative amino acid substitution may be considered to be a substitution in which a particular amino acid residue is substituted for a different amino acid in the same family.

[0061] Where a variant of one of SEQ ID NOs: 1 to 4 comprises an amino acid addition relative to one of SEQ ID NOs: 1 to 4, that addition may be at the N- or C-terminus, or internal within the sequence (i.e. it may be an insertion).

[0062] When a variant of one of SEQ ID NOs: 1 to 4 is used, that variant is a functional variant of one of SEQ ID NOs: 1 to 4, i.e. it retains the functionality of the sequence of SEQ ID NOs: 1 to 4 upon which it is based. In particular, a PPG comprising that variant peptide retains the functionality displayed by the corresponding PPG comprising the original peptide (i.e. one of SEQ ID NOs: to 4) upon which the variant is based. A PPG comprising one of SEQ ID NOs: 1 to 4 corresponds to a PPG comprising a variant peptide if the modification in the peptide sequence is the only difference between the two PPCs, i.e. the porphyrin moiety and the means of conjugation are identical.

[0063] A PPG comprising a variant peptide can be seen to retain the functionality of the corresponding PPG comprising the original peptide if the PPG comprising the variant peptide retains both the BBB shuttle capability and antiviral activity of a PPC comprising the original peptide (and optional also BPB shuttle capability). A PPC comprising a variant peptide may retain at least 70, 75, 80, 85, 90, 95 or 100 % of the BBB shuttle capability of the corresponding PPC comprising the original peptide. In some cases, a PPC comprising a variant peptide may display enhanced BBB shuttle capability compared to the corresponding PPC comprising the original peptide. A PPC comprising a variant peptide may retain at least 70, 75, 80, 85, 90, 95 or 100 % of the antiviral activity of the corresponding PPC comprising the original peptide. The antiviral activity may be measured against at least one virus against which the corresponding PPC comprising the original peptide displays antiviral activity. In some cases, a PPC comprising a variant peptide may display enhanced antiviral compared to the corresponding PPC comprising the original peptide, e.g. it may display more potent or broader spectrum antiviral activity. A PPC comprising a variant peptide may retain at least 70, 75, 80, 85, 90, 95 or 100 % of the BPB shuttle capability of the corresponding PPC comprising the original peptide. In some cases, a PPC comprising a variant peptide may display enhanced BPB shuttle capability compared to the corresponding PPC comprising the original peptide.

[0064] Preferably, a PPC comprising a variant peptide retains at least 70, 75, 80, 85, 90 or 95 % of the BBB shuttle capability and antiviral activity compared to the corresponding PPC comprising the original peptide. Where the PPC comprising the original peptide displays BPB shuttle capability, a PPC comprising a variant peptide preferably retains at least 70, 75, 80, 85, 90 or 95 % of the BBB shuttle capability, BPB shuttle capability and antiviral activity compared to the corresponding PPC comprising the original peptide. BBB and BPB shuttle capability of PPCs may be assessed using the translocation assays across in vitro models of the BBB and BPB, as described in the examples below. PPC antiviral activity may be assessed by measuring the IC50 for viral particle inactivation, as also described in the examples below.

[0065] In the case of SEQ ID NO: 4, which comprises a C-terminal lysine residue for conjugation to a porphyrin moiety, any sequence modification leaves this residue unchanged, i.e. any variant of SEQ ID NO: 4 retains a C-terminal lysine residue.

[0066] In preferred embodiments, the peptide is amidated at the C-terminus, i.e. the C-terminal carboxyl group of the peptide is amidated. Thus the C-terminus of the peptide preferably has the following structure: where R indicates the rest of the peptide. Linkers

[0067] As set out above, in the conjugates of the invention the peptides may be either directly conjugated to a porphyrin moiety (such that a bond is formed between a carboxyl group of the porphyrin moiety and a functional group, such as an amino group, of the peptide), or may be conjugated via a linker. A linker is any molecular structure located in between the porphyrin moiety and the peptide in the conjugates of the invention, i.e. a linker forms bonds with both a carboxyl group of the porphyrin moiety and a functional group of the peptide, thus linking the porphyrin moiety and peptide.

[0068] In some embodiments, the linker is a diradical of formula -(Yi-Li-CO)z- or -(YI-LI-SC>2)Z-, wherein Yi is O or NH and is attached to a carboxyl group of the porphyrin moiety (i.e. the carbonyl group of Formula (A1), (A2), (B1) or (B2)); wherein the CO or SO2 group is attached to the peptide X; and wherein Li is a diradical of formula -(CH2-)i(CH2-CH2-O-)m(CH2)i- where I and I’ are each independently selected from the integers 0 to 3, m is an integer of from 1 to 6; or, alternatively, Li is a diradical deriving from a radical selected from the group consisting of a C1-C12 alkyl, a C3-C8 cycloalkyl and a C2-C12 alkenyl; and z is selected from the integers 1 to 3.

[0069] The term “C1-C12 alkyl” refers to a saturated, linear or branched, hydrocarbon chain having from 1 to 12 carbon atoms. Non-limiting examples of C1-C12 alkyl groups include methyl, ethyl, propyl, iso-propyl, n-butyl, tert-butyl, iso-butyl, sec-butyl, n-pentyl, tert-pentyl, iso-pentyl, sec-pentyl, neo-pentyl, 1-ethylpropyl, n-hexyl, 1-methyl-pentyl, 2-methyl-pentyl, 3-methyl-pentyl, 4-methyl-pentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1 ,1 -dimethylbutyl, 2,3-dimethylbutyl, 1 ,3-dimethylbutyl, 1 ,2-dimethylbutyl, 3-ethylbutyl, 2-ethylbutyl, 1 -ethylbutyl, n-heptyl, n-octyl, n-nonyl, n-decanyl, undecanyl, and dodecanyl.

[0070] The term “C2-C12 alkenyl” refers to a saturated, linear or branched, hydrocarbon chain having from 2 to 12 carbon atoms (as described above) and further comprising a carbon-carbon double bond.

[0071] The term “C3-C8 cycloalkyl” refers to a saturated monocyclic or bicyclic hydrocarbon having from 3 to 8 carbon atoms. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl.

[0072] In particular embodiments of the invention, L is a diradical of formula -(Yi-Li-CO)z- wherein Y1 is NH, Li is a diradical of formula -(CH2-)i(CH2-CH2-O-)m(CH2)i- where I is 0 and I’ is 1 , and z is 1.

[0073] In other embodiments of the invention, L is a diradical of formula -(Yi-Li-CO)z- wherein Y1 is NH and Li is a diradical of formula -(CH2-)i(CH2-CH2-O-)m(CH2)i- where I is 0, I’ is 1 and m is 2, and z is 1. In this case, the linker is derived from 8-amino-3,6-dioxaoctanoic acid, also referred to herein as O2OC. Such linkers are referred to herein as “8-amino-3,6-dioxaoctanoic acid linkers” or “O2OC linkers”. Such a linker may be said to be 8-amino-3,6-dioxaoctanoic acid (or O2OC). In other embodiments of the invention, L is a diradical of formula -(Yi-Li-CO)z- wherein Yi is NH and Li is a diradical of formula -(CH2-)i(CH2-CH2-O-)m(CH2)i- where I is 0, I’ is 1 and m is 2, and z is 2. In such embodiments, the linker is derived from two units of 8-amino-3,6- dioxaoctanoic acid, alternatively referred to as a dimer of 8-amino-3,6-dioxaoctanoic acid. Such linkers are referred to herein as “8-amino-3,6-dioxaoctanoic acid dimer linkers”, “O2OC dimer linkers” or “O2OC-O2OC linkers”. Such a linker may be said to be a dimer of 8-amino-3,6- dioxaoctanoic acid (or a dimer of O2OC).

[0074] In other embodiments of the invention, the linker is selected from t-Boc-PEG-acid or Boc-NH-PEG(2-8)-CH2CH2COOH.

[0075] PPC Manufacture

[0076] A PPC of the invention can be prepared by a method comprising the following steps:

[0077] (i) providing a peptide of any one of SEQ ID NOs: 1-4, or a variant thereof as described above (optionally wherein the peptide is amidated at the C-terminus);

[0078] (ii) providing MPIX or PPIX;

[0079] (iii) optionally, coupling the MPIX or PPIX provided in step (ii) with a linker compound as described above; and

[0080] (iv) reacting the peptide provided in step (i) with the compound provided in step (ii) or in step (iii) to produce said PPC.

[0081] Alternatively, and also preferably, a PPC of the invention may be prepared by a method comprising the following steps:

[0082] (I) providing a peptide of any one of SEQ ID NOs: 1-4, or a variant thereof as described above (optionally wherein the peptide is amidated at the C-terminus);

[0083] (II) providing MPIX or PPIX;

[0084] (III) optionally, coupling the peptide provided in step (I) with a linker compound as described above; and

[0085] (IV) reacting the PPIX or MPIX provided in step (II) with the peptide / peptide-linker compound provided in step (I) or in step (III) to produce said conjugate.

[0086] Peptides for use in the PPCs of the invention may be prepared by solid phase synthesis following well-known procedures. As will become apparent to the skilled person, adequate protection of side chain residues may be necessary during the performance of steps (i) and (iv), or (I), (III) and (IV), in particular for the side chains of amino acids bearing hydroxyl, amino, guanidine or amide groups. Thus, said method may further include several protection and deprotection steps as well as the use of amino acids bearing protecting groups as reagents. In particular, the trityl (Trt) group is suitable for protecting amide functional groups located in the side chains of Gin and Asn. Also particularly, the tert-butyl group is suitable for protecting the hydroxyl groups located in the side chains of Ser and Thr. Also particularly, the tert-butylcarboxy (Boc) group is suitable for protecting the amine groups located in the side chains of Lys. Also particularly, the 2,2,4,6,7-pentamethyl-2,3-dihydrobenzofuran-5-sulfonyl group (Pbf) is suitable for protecting the guanidine groups located in the side chains of Arg. Procedures for introducing and removing the aforementioned protecting groups are well known in the art and will become apparent to the skilled person.

[0087] In optional step (iii), the porphyrin moiety may be coupled with a linker compound via the formation of an ester, i.e. by reaction of an acyl chloride deriving from PPIX or MPIX with a hydroxyl group located in the linker compound. The porphyrin moiety may alternatively be coupled with a linker compound via the formation of an amide, i.e. by reaction of an acyl chloride deriving from PPIX or MPIX with an amine group located in the linker compound or, alternatively, by reaction of PPIX or MPIX with an amine group located in the linker compound in the presence of reagents for peptidic coupling. A porphyrin moiety may alternatively be coupled with a linker compound via the formation of an alkene, i.e. by a metathesis reaction of PPIX or MPIX with an alkene group located in the linker compound. In particular, the linker compound of step (iii) may be a compound of formula H-(Yi-Li-CO)z-OH or H-(YI-LI-SC>2)Z-OH wherein Yi, Li and z are as defined above in the first aspect of the invention.

[0088] Similarly, in optional step (III), the peptide may be coupled with a linker compound via the formation of an ester through a hydroxyl group located in an amino acid side chain by reaction with an acyl chloride group located in the linker compound. Alternatively, the peptide may be coupled with a linker compound via the formation of an amide, i.e. by reaction of an acyl chloride or carboxylic acid group located in the linker compound with an amine group located in the peptide in the presence of reagents for peptidic coupling.

[0089] Step (iv) or (IV) may be carried out by peptidic coupling by contacting the peptide provided in step (i) with the compound provided in step (ii) or in step (iii), or by contacting the peptide provided in step (I) or the peptide-linker conjugate provided in step (III) with the compound provided in step (II), in the presence of reagents for peptidic coupling. This is particularly the case when the conjugation site in the peptide is an amino group.

[0090] Alternatively, when the conjugation site in the peptide is a hydroxyl group borne by a Ser or Thr amino acid side chain, step (iv) or (IV) may be carried out by contacting an acyl or sulfonyl chloride of the porphyrin moiety or porphyrin-linker conjugate with the peptide.

[0091] For example, step (iv) / (IV) of the synthesis may be represented by the following scheme (using as an example PPIX as the porphyrin moiety and pepH1, of SEQ ID NO: 1, as the peptide):

[0092] wherein R is OH, -(Yi-Li-CO)z-OH or -(YI-LI-SO2)Z-OH.

[0093] M-L-L-H3 Conjugate In one aspect of the invention, a porphyrin-peptide conjugate (PPC) is provided, the PPC comprising mesoporphyrin IX conjugated to a peptide consisting of the amino acid sequence of SEQ ID NO:3, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto, wherein a carboxylic acid group of the porphyrin moiety is conjugated to the N-terminal alpha-amino group of the peptide via a linker comprising a dimer of 8-amino-3,6-dioxaoctanoic acid.

[0094] In some embodiments, the PCC comprises a peptide consisting of the amino acid sequence of SEQ ID NO:3, or a variant thereof comprising a single amino acid substitution, deletion or addition relative thereto. In a preferred embodiment, the PPC comprises a peptide consisting of the amino acid sequence set forth in SEQ ID NO: 3. Preferably, the PPG has the formula set out below in Formula (7):

[0095] Formula (7):

[0096] The conjugate of Formula (7) is referred to herein as conjugate M-L-L-H3.

[0097] Also provided herein is the conjugate of this aspect of the invention for use in therapy.

[0098] Therapy for Flavivirus Infections

[0099] As shown in the examples below, the conjugates of the invention display antiviral activity against flaviviruses.

[0100] Thus in one aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of a flavivirus infection in a subject, wherein the conjugate comprises:

[0101] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0102] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0103] Similarly, the invention provides a method of treating or preventing a flavivirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising:

[0104] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0105] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0106] Similarly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0107] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0108] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a flavivirus infection in a subject.

[0109] The conjugates provided herein for prevention or treatment of a flavivirus infection may comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising a single amino acid substitution, deletion or addition relative thereto. Preferably, the conjugates for prevention or treatment of a flavivirus infection comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4.

[0110] Preferably, in the conjugates provided herein for prevention or treatment of a flavivirus infection, the porphyrin moiety is conjugated to:

[0111] (i) the N-terminal alpha-amino group of the peptide; or

[0112] (ii) the E-amino group of a lysine side chain of the peptide, preferably the E-amino group of the side chain of a lysine residue at the C-terminus of the peptide. Conjugation is generally via a carboxyl group of the porphyrin moiety.

[0113] In some such embodiments, the porphyrin moiety is directly conjugated to the peptide via an amide bond. Alternatively, the porphyrin moiety is conjugated to the peptide via a linker. Such a linker may be as described above. Preferably the linker is 8-amino-3,6-dioxaoctanoic acid.

[0114] In a preferred embodiment of this aspect, the PPC comprises protoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3.

[0115] In another preferred embodiment of this aspect, the PPC comprises protoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4.

[0116] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4.

[0117] In a preferred embodiment of this aspect, the PPC comprises protoporphyrin IX directly conjugated (i.e. without a linker) to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3. A PPC of this embodiment has the structure set forth in Formula (4) below:

[0118] Formula (4):

[0119] In a preferred embodiment of this aspect, the PPC has the structure of Formula (4). The conjugate of Formula (4) is referred to herein as conjugate P-H3. In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX directly conjugated (i.e. without a linker) to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4. A PPC of this embodiment has the structure set forth in Formula (1) below:

[0120] Formula (1):

[0121] In a preferred embodiment of this aspect, the PPC has the structure of Formula (1). The conjugate of Formula (1) is referred to herein as conjugate H3-M. In another preferred embodiment of this aspect, the PPC comprises protoporphyrin IX directly conjugated (i.e. without a linker) to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4. A PPC of this embodiment has the structure set forth in Formula (2) below:

[0122] Formula (2):

[0123] In a preferred embodiment of this aspect, the PPC has the structure of Formula (2). The conjugate of Formula (2) is referred to herein as conjugate H3-P.

[0124] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated via an 8-amino-3,6-dioxaoctanoic acid linker to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4. A PPC of this embodiment has the structure set forth in Formula (6) below.

[0125] In a preferred embodiment of this aspect, the PPC has the structure of Formula (6). The conjugate of Formula (6) is referred to herein as conjugate H3-L-M. Formula (6):

[0126] The conjugates provided herein for use in treatment or prevention of a flavivirus infection, as described above, may be used for the treatment or prevention of a Zika virus infection. Alternatively or additionally, they may be used for the treatment or prevention of a dengue virus infection. The conjugates may be used to treat or prevent infection with any dengue serotype (i.e. DEN-1 , DEN-2, DEN-3 or DEN-4). Other flaviviruses may also be treated with these conjugates. In another aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of dengue virus infection in a subject, wherein the conjugate comprises:

[0127] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0128] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX. Similarly, the invention provides a method of treating or preventing dengue virus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising, comprising:

[0129] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0130] Similarly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0131] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0132] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a dengue virus infection in a subject.

[0133] The conjugates provided herein for prevention or treatment of dengue virus infection may comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising a single amino acid substitution, deletion or addition relative thereto.

[0134] Preferably, the conjugates for prevention or treatment of dengue virus infection comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2.

[0135] Preferably, in the conjugates for use according to this aspect of the invention, the porphyrin moiety is conjugated to the N-terminal alpha-amino group of the peptide. Conjugation is generally via a carboxyl group of the porphyrin moiety. Preferably, the porphyrin moiety is directly conjugated to the peptide via an amide bond.

[0136] In a preferred embodiment of this aspect, the PPC comprises protoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 1, or a variant thereof, preferably the peptide of SEQ ID NO: 1.

[0137] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 2, or a variant thereof, preferably the peptide of SEQ ID NO: 2.

[0138] In a preferred embodiment of this aspect, the PPC comprises protoporphyrin IX directly conjugated (i.e. without a linker) to the N-terminal amino group of the peptide of SEQ ID NO: 1 , or a variant thereof, preferably the peptide of SEQ I D NO: 1. A PPC of this embodiment has the structure set forth in Formula (3) below:

[0139] Formula (3): In a preferred embodiment of this aspect, the PPC has the structure of Formula (3). The conjugate of Formula (3) is referred to herein as conjugate P-H1.

[0140] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX directly conjugated (i.e. without a linker) to the N-terminal amino group of the peptide of SEQ ID NO: 2, or a variant thereof, preferably the peptide of SEQ ID NO: 2. A PPC of this embodiment has the structure set forth in Formula (5) below:

[0141] Formula (5):

[0142] In a preferred embodiment of this aspect, the PPC has the structure of Formula (5). The conjugate of Formula (5) is referred to herein as conjugate M-Neg.

[0143] The conjugates for use according to this aspect are for the treatment or prevention of dengue virus infection in a subject. As for those above, the conjugates may be used to treat or prevent infection with any dengue serotype (i.e. DEN-1 , DEN-2, DEN-3 or DEN-4).

[0144] Therapy for Coronavirus Infections

[0145] As shown in the examples below, the conjugates of the invention display antiviral activity against a coronavirus.

[0146] Thus in another aspect, the invention provides a porphyrin-peptide conjugate for use in the prevention or treatment of a coronavirus infection in a subject, wherein the conjugate comprises:

[0147] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0148] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

[0149] Similarly, the invention provides a method of treating or preventing a coronavirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate comprising:

[0150] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0151] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX. Similarly, the invention provides the use of a porphyrin-peptide conjugate comprising:

[0152] (i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and

[0153] (ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX in the manufacture of a medicament for the treatment or prevention of a coronavirus infection in a subject.

[0154] The conjugates provided herein for prevention or treatment of a coronavirus infection may comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising a single amino acid substitution, deletion or addition relative thereto.

[0155] Preferably, the conjugates for prevention or treatment of a flavivirus infection comprise a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4.

[0156] Preferably, in the conjugates provided herein for prevention or treatment of a coronavirus infection, the porphyrin moiety is conjugated to:

[0157] (i) the N-terminal alpha-amino group of the peptide; or

[0158] (ii) the E-amino group of a lysine side chain of the peptide, preferably the E-amino group of the side chain of a lysine residue at the C-terminus of the peptide. Conjugation is generally via a carboxyl group of the porphyrin moiety.

[0159] In some such embodiments, the porphyrin moiety is directly conjugated to the peptide via an amide bond. Alternatively, the porphyrin moiety is conjugated to the peptide via a linker. Such a linker may be as described above. Preferably the linker is a dimer of 8-amino-3,6-dioxaoctanoic acid.

[0160] In a preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 2, or a variant thereof, preferably the peptide of SEQ ID NO: 2.

[0161] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3.

[0162] In another preferred embodiment of this aspect, the PPC comprises protoporphyrin IX conjugated to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3.

[0163] In another preferred embodiment of this aspect, the PPC comprises protoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4.

[0164] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4.

[0165] In a preferred embodiment of this aspect, the PPG comprises mesoporphyrin IX directly conjugated (i.e. without a linker) to the N-terminal amino group of the peptide of SEQ ID NO: 2, or a variant thereof, preferably the peptide of SEQ ID NO: 2. A PPG of this embodiment has the structure set forth in Formula (5), above. In a preferred embodiment of this aspect, the PPG has the structure of Formula (5) (i.e. the conjugate is M-Neg).

[0166] In another preferred embodiment of this aspect, the PPG comprises protoporphyrin IX directly conjugated (i.e. without a linker) to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3. A PPG of this embodiment has the structure set forth in Formula (4), above. In a preferred embodiment of this aspect, the PPG has the structure of Formula (4) (i.e. the conjugate is P-H3).

[0167] In another preferred embodiment of this aspect, the PPG comprises mesoporphyrin IX directly conjugated (i.e. without a linker) to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4. A PPC of this embodiment has the structure set forth in Formula (1), above. In a preferred embodiment of this aspect, the PPC has the structure of Formula (1) (i.e. the conjugate is H3-M).

[0168] In another preferred embodiment of this aspect, the PPC comprises protoporphyrin IX directly conjugated (i.e. without a linker) to the E-amino group of the C-terminal lysine of the peptide of SEQ ID NO: 4, or a variant thereof (in which the C-terminal lysine residue is retained), preferably the peptide of SEQ ID NO: 4. A PPC of this embodiment has the structure set forth in Formula (2), above. In a preferred embodiment of this aspect, the PPC has the structure of Formula (2) (i.e. the conjugate is H3-P).

[0169] In another preferred embodiment of this aspect, the PPC comprises mesoporphyrin IX conjugated via an 8-amino-3,6-dioxaoctanoic acid dimer linker to the N-terminal amino group of the peptide of SEQ ID NO: 3, or a variant thereof, preferably the peptide of SEQ ID NO: 3. A PPC of this embodiment has the structure set forth in Formula (7), above. In a preferred embodiment of this aspect, the PPC has the structure of Formula (7) (i.e. the conjugate is M-L- L-H3).

[0170] The conjugates for use according to this aspect of the invention may be used to treat or prevent any coronavirus infection. In a particular embodiment, the conjugates may be used to treat infections caused by SARS-CoV-2, i.e. COVID-19 disease.

[0171] Means of Therapy

[0172] As set out above, the conjugates provided herein may be used in therapy, including for treatment or prevention of various viruses. The term “therapy” as used herein encompasses both treatment and prophylaxis / prevention of a disease or disorder. The term “treatment” as used herein in the context of treating a disease or disorder, pertains generally to treatment of a human, or alternatively of a non-human animal, in which some desired therapeutic effect is achieved. In particular, in the context of treatment for a viral infection, treatment may cure the subject of the infection, reduce the length of infection or severity of infection, e.g. by reducing viral load and / or reducing, ameliorating or preventing one or more symptoms of the infection. Particularly in the context of treating Zika virus, treatment of a pregnant woman infected with Zika may prevent transmission of Zika to her unborn child, or may prevent microcephaly in the unborn child.

[0173] The terms “prevention” and “prophylaxis” as used herein in the context of preventing a disease or disorder are interchangeable and refers to a measure which is administered in advance of detection of a condition, particularly a viral infection, with the aim of preserving health by helping to delay, mitigate or avoid that particular condition. For instance, prevention or prophylaxis of a viral infection may prevent a subject becoming infected with the virus in question, or cause any subsequent infection to be reduced in length and / or severity, or may reduce, ameliorate or prevent one or more symptoms of the infection.

[0174] The subject treated, or to whom the conjugates may be administered as a preventative or prophylactic measure, may be any human or non-human animal. Generally the subject is a mammal. Preferably the subject is a human. Human subjects including infants, children and adults may receive therapy according to the present invention. When used for treatment of a viral infection, the subject has been diagnosed with the infection to be treated, i.e. is currently infected with one of the viruses specified herein. Such a subject may be severely ill or may be at risk of severe illness, e.g. due to age or underlying conditions. In the case of Zika virus, the subject may be a pregnant woman whose baby is at risk of neurological damage due to the maternal infection. When used for prevention of a viral infection, the subject may be particularly at risk of infection, e.g. due to location, season, job or underlying condition. The subject may be particularly at risk of severe disease e.g. due to age or underlying conditions. For prevention of Zika virus infection, the subject may be a pregnant woman.

[0175] When administered to a subject in the context of therapy, the PPCs for use herein may be in the form of a pharmaceutically acceptable salt or a pharmaceutically acceptable cocrystal. The PPCs may be administered in the context of a pharmaceutical composition, comprising a pharmaceutically acceptable diluent, excipient or carrier. The term “pharmaceutically acceptable,” as used herein, pertains to compounds, ingredients, materials, compositions, etc., which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of the subject in question (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio. Each carrier, diluent, excipient, etc. must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation. The carrier, diluent or excipient may one conventionally used in the art, such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinylpyrrolidone; fillers, for example lactose, sugar, maize starch, calcium phosphate, sorbitol or glycine; tabletting lubricants, for example magnesium stearate; disintegrants, for example starch, polyvinylpyrrolidone, sodium starch glycollate or microcrystalline cellulose; or pharmaceutically acceptable wetting agents such as sodium lauryl sulfate.

[0176] The composition may be formulated as a tablet, pill, capsule, granule, aerosol, syrup, emulsion, suspension, solution, ointment, cream, dermal patch or gel of any kind. Solid oral compositions may be prepared by conventional methods of blending, filling or tabletting. Repeated blending operations may be used to distribute the active agent throughout those compositions employing large quantities of fillers. Such operations are conventional in the art. The tablets may for example be prepared by wet or dry granulation and optionally coated according to methods well known in normal pharmaceutical practice, in particular with an enteric coating. The composition may be for parenteral administration, such as a sterile solution, suspension or reconstitutable dry preparation, an aerosol or spray.

[0177] The conjugate or composition comprising it may be administered by any suitable route, e.g. by topical or systemic application, particularly dermal, transdermal, subcutaneous, intramuscular, intraperitoneal, intravenous, intra-arterial, intranasal, rectal, vaginal, oral, epidural, intrathecal, intraventricular, intracerebral, intracerebroventricular, intracisternal, intraspinal, perispinal or intracranial administration. Delivery may be via a needle or catheter with or without a pump device. Any other administrative route or method may alternatively be used as desired by the skilled person.

[0178] For the avoidance of any doubt, any theoretical explanations provided herein are provided for the purposes of improving the understanding of a reader. The inventors do not wish to be bound by any of these theoretical explanations.

[0179] Any section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0180] Throughout this specification, including the claims which follow, unless the context requires otherwise, the word “comprise” and “include”, and variations such as “comprises”, “comprising”, and “including” will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps.

[0181] It must be noted that, as used in the specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. The invention may be further understood by reference to the figures and non-limiting examples below.

[0182] Figure Legends

[0183] Figure 1 shows the serum stability of N-terminal conjugates (A, B, C, D) versus corresponding C-terminal conjugates (E, F, G, H). The conjugates are as follows: A = M-H1, B = P-H1, C = M-Neg, D = P-Neg, E = H1-M, F = H1-P, G = Neg-M and H = Neg-P.

[0184] Figure 2 shows an in vitro BBB model consisting of a transwell system with an insert which divides the system into two separate chambers, and upon which bEnd.3 or HBEC-5i cells are grown. The insert, or apical side (top chamber), corresponds to the blood side, while the basolateral side (bottom chamber) corresponds to the brain side.

[0185] Figure 3 shows the translocation of the porphyrin-CPP conjugates across an in vitro model of the human blood-brain barrier (BBB) using the HBEC-5i cell line. The amount of each conjugate in the basolateral compartment was quantified by the measurement of fluorescence emission intensity. The values correspond to the percentage of barrier translocation ± standard deviation (SD).

[0186] Figure 4 shows the translocation of the porphyrin-CPP conjugates across an in vitro model of the human blood-placental barrier (BPB) using the JEG-3 cell line. The amount of each conjugate in the basolateral compartment was quantified by the measurement of fluorescence emission intensity. The values correspond to the percentage of barrier translocation ± standard deviation (SD).

[0187] Figure 5 shows FD4 permeability of the BBB (A) and BPB (B) after application of the PPCs to the in vitro barrier models (in the case of the BBB, using the HBEC-5i cell line). The amount of FD4 in the basolateral chamber was quantified by the measurement of fluorescence emission intensity. The values correspond to the percentage of FD4 permeability ± standard deviation (SD).

[0188] Figure 6 shows an evaluation of porphyrin-CPP conjugate internalisation mechanisms in HBEC-5i (A) and JEG-3 (B) cells. Cells were incubated with conjugates in the presence of endocytosis inhibitors for 1 h. Flow cytometry was then used to determine the cellular uptake of conjugates. Values represent the mean fluorescence intensity of cells incubated with conjugates and endocytosis inhibitors relative to the mean fluorescence intensity of cells incubated only with conjugates ± standard deviation (SD). (n = 3 independent experiments; **P < 0.01 ;

[0189] ***P < 0.001 ; ****P < 0.0001, One-way ANOVA test).

[0190] Figure 7 shows PPCs’ therapeutic potential. ZIKV replication kinetics and treatment schemes are shown for JEG-3 cells (A) and SY-SY5Y cells (B). Treatment of ZlKV-infected JEG-3 cells (C) and SH-SY5Y cells (D) with the selected PPCs, following the treatment schemes described in the A and B reduces the resultant viral titres. Released infectious virus particles for the different samples were quantified by plaque assay. **p <0.01 , ***p < 0.001 , ****p < 0.0001 against the untreated control sample.

[0191] Figure 8 shows cytokine modulation by PPCs in JEG-3 cells. Treatment of ZlKV-infected JEG-3 cells with the selected PPCs followed the treatment scheme described for Figure 7. Cytokine release in cell supernatant was evaluated by ELISA 48 h.p.i.

[0192] Figure 9 shows that P-H1 and P-H3 treatment reduced brain viral load and mortality induced by ZIKV in a mouse model. (A) A neonatal mouse model of ZIKV infection was used. (B) Brain viral load quantification by qPCR. Both P-H1 and P-H3 significantly reduced viral load. The results are presented as means ± SEM (*p = 0.0313, **p = 0.0082, One-way ANOVA, Dunnett’s multiple comparisons test). (C) Survival curves. Both P-H1 and P-H3 significantly reduced disease-induced mortality (p = 0.0293, Gehan-Breslow-Wilcoxon test).

[0193] Figure 10 shows plasma levels of various conjugates and free porphyrins over time after i.p. injection in mice.

[0194] Figure 11 shows brain levels at 60 min and 240 min post injection of various conjugates and free porphyrins over time after i.p. injection in mice.

[0195] Examples

[0196] Synthesis of porphyrin-peptide conjugates

[0197] The PPCs were synthesized by a state-of-the-art Fmoc solid-phase approach where 3 CPPs, (PepH1 (H1 - SEQ ID NOs: 1 & 5), PepNeg (Neg - SEQ ID NOs: 2 & 6) and PepH3 (H3 - SEQ ID NOs: 3 & 4)) were synthesized and the antiviral payloads protoporphyrin IX (P) and / or mesoporphyrin IX (M) coupled to an amino group (N-terminal or Lys side chain) of the CPP while it remained anchored to the solid support. SEQ ID NOs: 1, 2 and 3 were used for conjugates with the porphyrin moiety attached to the peptide N-terminus. SEQ ID NOs: 5, 6 and 4 corresponds to SEQ ID NOs: 1, 2 and 3, respectively, with an additional C-terminal lysine, and were used for conjugates with the porphyrin moiety attached to the peptide N-terminus.

[0198] Afterwards, the conjugates were released from the solid support by acidolytic deprotection and cleavage. All conjugates were purified by preparative HPLC and characterized for identity by electrospray or MALDI-MS.

[0199] To investigate how to improve antiviral activity and the ability to translocate the BBB and / or BPB, and decrease the cytotoxicity of PPCs, various strategies were attempted. These included the synthesis of conjugates containing linkers (O2Oc and O2Oc-O2Oc) between the CPP and the porphyrin. The conjugates that were synthesised are set out in Table 1 below. Serum Stability Analysis

[0200] The in vitro stability in human serum of some H1 and Neg conjugates and their constitutive nonconjugated CPPs was evaluated by LC and LC-MS. Altogether, the obtained data suggest that the porphyrin ring somehow protects cleavable peptide bonds from protease access, with N-terminal conjugates showing higher stability than C-terminal ones (Figure 1).

[0201] Table 1 . Final yield and HPLC purity of synthesized conjugates. L = O2OC linker. L-L = O2OC-O2OC linker. Translocation Efficacy Testing

[0202] Translocation of the conjugates across two in vitro models of the BBB was quantified. BBB models consist of transwell systems with an insert in which brain endothelial cells grow, separating two chambers (Figure 2). This system has been previously described in Mendonca et al. (supra). In the experimental procedure, the insert, or apical side / apex (top chamber), corresponds to the blood side, while the base (bottom chamber) corresponds to the brain side. The cells used are human brain endothelial cells (HBEC-5i cell line) and mouse brain endothelial cells (bEnd.3 cell line). The translocation studies are performed between 8 and 11 days post-seeding. The conjugates are added to the top chamber and incubated for 24h. After incubation, conjugates are collected from apex and base and fluorescence emission intensity and / or UV-Vis. absorbance measured. The percentage of translocation is calculated relative to the initial concentration in the apex. Porphyrin-peptide conjugates with translocation above 30 % were selected for continued analysis (Figure 3).

[0203] Similarly, the above-described transwell system was modified using the JEG-3 choriocarcinoma cell line, a widely-used model for human placental trophoblasts, and optimized to mimic the human BPB for in vitro evaluation of mother-to-foetus exchange (apical to basolateral) of PPCs. The results are shown in Figure 4, with translocation of at least 10 % being deemed the cut-off for promising BPB translocators.

[0204] Although the translocation values for the BPB model were considerably lower than those obtained using the BBB models, overall PPCs having better BBB crossing ability were also those with higher trans-BPB potential. Importantly, the BPB translocation incapacity was not an exclusion criterion in itself because drug leads traversing the brain but not the placenta are still valid for specific applications. The most promising PPCs were selected by ranking the conjugates based on their translocation efficacy across the BBB (two cell lines) and across the BPB.

[0205] Cytotoxicity Evaluation

[0206] Using the BBB and BPB in vitro models described above, the effect of the conjugates on the barriers’ integrity was evaluated based on their permeability to 4 kDa fluorescently labeled dextran (FD4). After recovering the apical and basolateral compartments of BBB and BPB models treated with PPCs, FD4 was added to the apical chamber and incubated for two hours. FD4 permeability was quantified by measuring the fluorescence emission intensity in the basolateral compartment (Figure 5). Conjugates showing an FD4 permeability lower than 25 % and 2.5 % in the BBB and BPB models, respectively, were considered not to cause fenestration and paracellular leakage and were thus capable of maintaining the barriers’ integrity. These PPCs were eligible for continued evaluation. In addition to the evaluation of the PPCs’ effect on BBB / BPB disruption, cell viability assays on brain endothelial (bEnd.3 and HBEC-5i), neuronal (SH-SY5Y), lung (A549), kidney (HREC) and liver (Huh-7) cell lines and red blood cells (RBC) were performed to estimate the appropriate dose ranges for the antiviral activity assays and to assess the toxic effects of the conjugates. The CC50 values (concentration of the conjugates required to reduce cell viability by 50 %) were calculated (Table 2).

[0207] Table 2. Cytotoxicity evaluation of conjugates in human (HBEC-5i) and mouse (bEnd.3) endothelial cells of the blood-brain barrier (BBB) model, choriocarcinoma JEG-3 cells of the blood-placental barrier (BPB) model, as well as in the neuronal (SH-SY5Y), lung (A549), kidney (HREC) and liver (Huh-7) cells lines and red blood cells (RBC). Values (pM) correspond to the CC50 ± standard error. NT = not tested. In all assays, the conjugates were considered suitable for continued analysis when presenting CC50 values at least two-fold greater than the highest admissible IC50 value (25 pM), i.e. when presenting CC50 values of at least 50 pM.

[0208] Conjugates which did not show cytotoxicity against mammalian cell lines were then subjected to a screen for anti-viral activity against dengue virus, Zika virus, HIV and SARS- CoV-2. Activity was assessed by determination of IC50 values for the various PPCs against each virus. Briefly, to determine the inactivation of DENV, ZIKV or SARS-CoV-2 by PPCs, 105PFU (plaque forming units) / ml were treated with different PPC concentrations for 1 h at 37°C, in the dark. After treatment, viral particle infectivity was assessed by plaque assay in Vero cells. For HIV inactivation, viral supernatants (100 TCID50 / well) were treated for 1 h with different PPC concentrations. Following the PPC treatment, viral particle infectivity was evaluated by single-cycle viral infectivity assays in TZM-bl reporter cells. The results of the initial screen are presented below in Table 3.

[0209] Table 3. Results of initial screen of antiviral activity of non-cytotoxic PPCs. The anti-viral activity is provided as IC50 values ± SD. NT = not tested. ND = not determined.

[0210] Based on the above results, 7 conjugates were selected for further analysis (Table 4).

[0211] Only the best performing conjugate in any given family (conjugates with the same peptide- porphyrin core structure, disregarding linkers) was included. Table 4. List of the best performing conjugates according to their cytotoxicity and efficacy to translocate BBB and BPB models.

[0212] According to their translocation ability, antiviral efficacy and cytotoxicity, a preliminary classification of the selected conjugates into 3 different groups has been established: i) braintargeting PPCs, with activity against at least 3 viruses (H3-M, H3-P and H3-L-M) and reasonable trans-BBB activity, ii) PPCs targeting only the flaviviruses (DENV and ZIKV), with the capacity to cross BBB and BPB (P-H1), and iii) universal, with BBB and BPB-translocating antiviral PPCs (P-H3, M-Neg and M-L-L-H3).

[0213] Mechanistic Investigations of Barrier Translocation

[0214] A systematic analysis of intracellular transport pathways based on the effects of inhibiting one or more individual routes was conducted using a set of chemical inhibitors of endocytosis: methyl-p-cyclodextrin (M CD) to inhibit the cholesterol-dependent endocytic pathway; dynasore to inhibit dynamin; chlorpromazine (CPZ) to restrict clathrin-dependent endocytosis; 5-(N-ethyl- N-isopropyl) amiloride (El PA) to block micropinocytosis; phloretin to restrict the reduction of membrane dipole potential; heparinase I to abrogate heparan sulfate proteoglycan (HSPG)- dependent endocytic pathways; brefeldin A to inhibit the formation of Golgi vesicles; and primaquine to inhibit Rab11. A temperature block (4°C) targeting so-called energy-dependent uptake mechanisms was also included.

[0215] Briefly, human brain endothelial cells (HBEC-5i cell line) or human placental choriocarcinoma cells (JEG-3 cell line) were treated with endocytosis inhibitors for 1 h and analysed by flow cytometry to determine the extent of cellular internalization of PPCs.

[0216] The results suggested that PPC internalisation in HBEC-5i cells was inhibited by low temperature (4 °C). Among the endocytosis inhibitors, brefeldin A and CPZ significantly decreased the internalisation of the selected conjugates, suggesting that Golgi vesicle trafficking and the clathrin-dependent endocytic pathway may be required for PPC uptake. Internalisation of the conjugate P-H3 seems also to be decreased by treatment with heparinase I and primaquine, implying that HSPG-dependent endocytic pathways and Rab11 may play an active role in P-H3 translocation. Additionally, phloretin may inhibit the uptake of the conjugate H3-M into HBEC-5i cells, suggesting the involvement of membrane dipole potential (Figure 6A).

[0217] Similarly, PPC internalisation in JEG-3 cells was also reduced by low temperature (4°C). Brefeldin A inhibited the uptake of the conjugates into JEG-3 cells, suggesting the involvement of Golgi vesicles. CPZ also reduced the internalisation of P-H1 and phloretin may inhibit the uptake of the conjugate M-Neg, suggesting the involvement of the clathrin-dependent endocytic pathway and membrane dipole potential, respectively. Interestingly, dynasore enhanced the uptake of P-H3, M-Neg and M-L-L-H3 conjugates while CPZ increased the internalisation of M-Neg and M-L-L-H3 (Figure 6B).

[0218] In Vitro Evaluation of Therapeutic Potential

[0219] PPC therapeutic potential was evaluated in vitro by treatment of ZIKV target cells, such as JEG-3 and SH-SY5Y. For this purpose, ZIKV infection kinetic curves in those cells were first determined (Figure 7A & B). The results suggest that ZIKV reaches its viral titre maxima at 48 h and 72 h post-infection (h.p.i.) on JEG-3 cells and SH-SY5Y, respectively. Thus, the PPCs were added to the infected cells at 8 h.p.i., and cell supernatants collected at 48 and 72 h.p.i. for JEG-3 and SH-SY5Y, respectively. As observed in Figure 7C & D, all the selected PPCs were able to significantly inhibit ZIKV on-going infection, proving their inhibitory capacity and therapeutic potential.

[0220] The PPCs’ effect on the pro-inflammatory response was also assessed, in infected JEG-3 cells. Briefly, JEG-3 cells were infected with ZIKV (MOI of 0.01), and at 8 h.p.i. PPCs were added to the cells. Supernatants were collected at 48 h.p.i. Cytokine modulation by PPCs was evaluated in cell supernatants by ELISA. PPCs were able to reduce secretion of IFN-y and IL-6 by infected cells, but did not have any effect in modulating IP-10 (Figure 8).

[0221] In Vivo Evaluation of Therapeutic Potential

[0222] The flavivirus-targeting BBB / BPB-translocating peptide P-H1 and P-H3 were tested in the neonatal model of ZIKV infection in vivo (Figure 9A), to determine the impact on viral load and disease severity. 3-day old mice were subcutaneously infected with ZIKV PRVABC59 (104PFU). Starting on day 1 post-infection, infected mice received daily i.p. injections of the PPCs (7 mg / kg). For viral load determination, mice received PPC doses for 9 days, then were euthanized and dissected at day 10 post-infection. Viral load was determined by qPCR (Figure 9B). Cycle threshold values were used to calculate the equivalents of PFU / mg RNA (Eq. PFU / pg RNA) after conversion using a standard-curve with 10-fold serial dilutions of RNA from a ZIKV stock with known titre. For survival analysis, mice received PPC doses for 18 days, and mortality monitored until day 28 post-infection (Figure 9C). Daily treatment with either P-H1 or P-H3 significantly reduced viral load and mortality

[0223] Safety Profiling A series of pharmacokinetic studies were performed in which the compounds were administered to mice and plasma (Figure 10) and brain (Figure 11) exposure was measured over time.

[0224] Substances were applied intraperitonealy (i.p.) in saline and blood samples and brain samples were taken as indicated over time to generate a time course for the distribution to, and elimination from, the brain. The data shows that the highest plasma levels are for conjugates with the porphyrin moiety at the N-terminus of the peptide, and free porphyrins have lower plasma levels than PPCs. the highest brain levels are for PPCs with the porphyrin moiety at the C-terminus of the peptide, and free porphyrins were found to have ca. 9x lower levels than conjugates in the brain.

[0225] Sequence Listing

[0226] All sequences are amino acid sequences:

[0227] AGILKRWK

[0228] SEQ ID NO: 5- with C-terminal

[0229] VQQLTKRFSLK

[0230] SEQ ID NO: 6- with C-terminal

[0231] SGTQEEYK

Claims

Claims1. A porphyrin-peptide conjugate for use in the prevention or treatment of a flavivirus infection, wherein the conjugate comprises:(i) a peptide consisting of the amino acid sequence of SEQ ID NO: 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and(ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

2. The conjugate for use according to claim 1 , wherein the peptide consists of the amino acid sequence set forth in SEQ ID NO: 3 or 4.

3. The conjugate for use according to claim 1 or 2, wherein a carboxyl group of the porphyrin moiety is conjugated to:(i) the N-terminal alpha-amino group of the peptide; or(ii) the E-amino group of a lysine side chain of the peptide.

4. The conjugate for use according to claim 3, comprising:(i) protoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 3;(ii) protoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of SEQ ID NO: 4; or(iii) mesoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of SEQ ID NO: 4.

5. The conjugate for use according to claim 4, wherein the porphyrin moiety is:(i) directly conjugated to the peptide via an amide bond; or(ii) conjugated to the peptide via a linker, preferably wherein the linker is 8-amino-3,6-dioxaoctanoic acid.

6. The conjugate for use according to claim 5, having the structure set out in:(a) Formula (1):(c) Formula (4):

7. The conjugate for use according to any one of claims 1 to 6, wherein the flavivirus infection is Zika virus infection or dengue virus infection.

8. A porphyrin-peptide conjugate for use in the prevention or treatment of dengue virus infection, wherein the conjugate comprises:(i) a peptide consisting of the amino acid sequence of SEQ ID NO: 1 or 2, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and(ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

9. The conjugate for use according to claim 8, wherein the peptide consists of the amino acid sequence set forth in SEQ ID NO: 1 or 2.

10. The conjugate for use according to claim 8 or 9, wherein a carboxyl group of the porphyrin moiety is conjugated to the N-terminal alpha-amino group of the peptide.

11. The conjugate for use according to claim 10, comprising:(i) protoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 1; or(ii) mesoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 2.

12. The conjugate for use according to claim 11, wherein the porphyrin moiety is directly conjugated to the peptide via an amide bond.

13. The conjugate for use according to claim 12, having the structure set out in:(a) Formula (3):(b) Formula (5):

14. A porphyrin-peptide conjugate for use in the prevention or treatment of a coronavirus infection, wherein the conjugate comprises:(i) a peptide consisting of the amino acid sequence of SEQ ID NO: 2, 3 or 4, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto; and(ii) a porphyrin moiety selected from mesoporphyrin IX or protoporphyrin IX.

15. The conjugate for use according to claim 14, wherein the peptide consists of the amino acid sequence set forth in SEQ ID NO: 2, 3 or 4.

16. The conjugate for use according to claim 14 or 15, wherein a carboxyl group of the porphyrin moiety is conjugated to:(i) the N-terminal alpha-amino group of the peptide; or(ii) the E-amino group of a lysine side chain of the peptide.

17. The conjugate for use according to claim 16, comprising:(i) mesoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 2;(ii) mesoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 3;(iii) protoporphyrin IX conjugated to the N-terminal amino group of SEQ ID NO: 3;(iv) protoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of SEQ IDNO: 4; or(v) mesoporphyrin IX conjugated to the E-amino group of the C-terminal lysine of SEQ ID NO: 4.

18. The conjugate for use according to claim 17, wherein the porphyrin moiety is:(i) directly conjugated to the peptide via an amide bond; or(ii) conjugated to the peptide via a linker, preferably wherein the linker is a dimer of 8-amino-3,6-dioxaoctanoic acid.

19. The conjugate for use according to claim 18, having the structure set out in: (a) Formula (1):(c) Formula (4):

20. The conjugate for use according to any one of claims 14 to 19, wherein the coronavirus infection is SARS-CoV-2 infection.

21. A method of preventing or treating a flavivirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate as defined in any one of claims 1 to 6, optionally wherein the flavivirus infection is as defined in claim 7.

22. A method of preventing or treating a dengue virus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate as defined in any one of claims 8 to 13.

23. A method of preventing or treating a coronavirus infection in a subject, comprising administering to the subject a porphyrin-peptide conjugate as defined in any one of claims 14 to 19, optionally wherein the coronavirus infection is as defined in claim 20.

24. A porphyrin-peptide conjugate comprising mesoporphyrin IX conjugated to a peptide consisting of the amino acid sequence of SEQ ID NO:3, or a variant thereof comprising up to 2 amino acid substitutions, deletions or additions relative thereto, wherein a carboxylic acid group of the porphyrin moiety is conjugated to the N-terminal alpha-amino group of the peptide via a linker comprising a dimer of 8-amino-3,6-dioxaoctanoic acid.

25. The conjugate of claim 24, wherein the peptide consists of the amino acid sequence of SEQ ID NO:3.

26. The conjugate of claim 25, having the structure set out in Formula (7):

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