Tailored Anti-fungal drugs and tools to diagnose invasive fungal infections

Abstract

The present application presents peptides that bind selectively to fungi, in particular opportunistic fungi, and not to human cells. These properties can be utilized to specifically target invading fungal pathogens, and via genetic engineering create peptibodies. Thus, the present application describes novel approaches for detection and treatment of infections caused by fungi.

Description

[0001] TAILORED ANTI-FUNGAL DRUGS AND TOOLS TO DIAGNOSE INVASIVE FUNGAL INFECTIONS

[0002] FIELD

[0003] The present application presents peptides that bind selectively to fungi, in particular opportunistic fungi, and not to human cells. These properties can be utilized to specifically target invading fungal pathogens, and via genetic engineering create peptibodies. Thus, the present application describes novel approaches for detection and treatment of infections caused by fungi.

[0004] The specific sequence could be a tool for targeted treatment of fungal infections. The sequence can be coupled to biological active components that provide a function to the peptide and that contributes to the physiochemical properties of the construct.

[0005] BACKGROUND

[0006] Fungal infections represent an increasing health problem worldwide. Patients undergoing high dose chemotherapy or stem cell / organ transplantation are at high risk of being infected with opportunistic fungal pathogens. Omnipresent, airborne fungal spores can reach the lungs and cause invasive infections. Despite prophylactic antifungal treatment, fungal infections still constitute a severe clinical problem associated with high mortality rates (30-90%). The problem is aggravated by the fact that drug-resistant strains of one of the dominating pathogens, Aspergillus fumigatus, appear more and more frequently.

[0007] Moreover, the fungicidal compounds currently used in the clinic have the disadvantage of making drug-drug interactions and giving rise to adverse and often systemic toxic reactions in the patient.

[0008] Rosbjerg et al. (Journal of Innate Immunity, vol. 13, no. 4, 1 January 2021, pages 211-24) relates to human MASP-1 and studies its direct binding to Aspergillus fumigatus, as well as other fungi. The authors suggested that the A. fumigatus- interacting binding site was at the CCP2-SP domain area.

[0009] WO 2021 / 231343 Al discloses means and methods for treating infections, including fungal pathogens, such as Aspergillus fumigatus. It is suggested to use a mimic of e.g. MASP-1 as binding moiety in the method of treating infections. J. Dobo et al. (The Journal of Immunology, vol. 183, no. 2, 29 June 2009, pages 1207-1214) describes MASP-1 and its domain structure as well as rMASP-1 consisting of only CCP1-CCP2-SP.

[0010] Ambrus G et al (The Journal of Immunology, Vol. 170, No. 3, 1 February 2003, pages 1374-1382) shows the MASP-1 structure.

[0011] W02015048748 discloses peptibodies.

[0012] Hence, to develop a targeted drug delivery system that circumvents systemic exposure and circumvents the increasing resistance problem is a desirable goal, as there is an unmet need in the treatment of opportunistic fungal infections.

[0013] SUMMARY OF THE INVENTION

[0014] This disclosure presents peptides that bind selectively to fungi, and which does not bind to human cells.

[0015] As also outlined further below, the inventing team has identified a small peptide element from MASP-1 capable of selective binding to fungi. Such peptide may find use in treatment of fungi infection, such as in a peptibody format.

[0016] Small peptides have several advantages compared to longer fragments from MASP-1, since MASP-1 domains have several functions such as:

[0017] - Avoid unwanted off-target proteolytic activity of the peptide o MASP-1 has many substrates and proteolytic activity can cause several unwanted effects such as activation of the coagulation system, complement system, kininogen, PAR4, fibrinogen etc.

[0018] - Avoid binding of pattern recognition molecules to the peptide o MASP-1 binds to mannose-binding lectin (MBL), ficolins and collectins that possible already sit in complexes with e.g. MASP-2 and therefore are able to be proteolytically active.

[0019] - Enable tissue penetrance of the peptide o Smaller peptides have the advantage that they more easily penetrate through tissue to the site of infection, e.g. brain tissue.

[0020] In the example section the following data is presented: - Example 1 shows that MASP-1, a serine protease from the complement system, binds directly to various pathogenic fungi.

[0021] - Example 2 and example 2A show that the amino acid sequence KLMAR from MASP-1 is necessary for the binding of MASP-1 to A. fumigatus.

[0022] - Example 3 shows that a KLMAR-comprising 30 AA MASP-1 peptide specifically binds to the different Aspergillus (A. fumigatus, A. niger, A. terreus and A. flavus) and Mucorales fungi Lichtemia corymbifera, Mucor circinelloides and Rhizopus arrhizus.

[0023] - Example 3A shows that three CCP2 peptides named CCP2 Ctrl. Pepl-3 did not bind to A. fumigatus conidia. The KLMAR-comprising MASP-1 peptide (SEQ ID NO: 1) bound A. fumigatus as previously shown. Hence, the binding is specific to this peptide and does not involve the CCP2 domain in general.

[0024] - Example 3B shows that CHS and C7S from SEQ ID NO: 1 are important (albeit not essential) for fungus binding.

[0025] - Example 3C shows that SEQ ID NO: 1 does not require calcium for interacting with A. fumigatus.

[0026] - Example 3D demonstrates that the A. fumigatus binding is unique for the MASP-1 derived peptide, since the corresponding peptide from MASP-3 did not bind to A. fumigatus.

[0027] - Example 3E demonstrates that the 30 AA peptide does not bind to human cells.

[0028] - Example 4 shows that the peptides of the invention are also functional in a Peptibody setting.

[0029] - Example 5 and Example 5A demonstrate that the Peptibody binds to A. fumigatus germ tubes, hyphae and conidia. Binding to the hyphal tip is also shown, which is of importance, because the hyphal tip is likely a desired target for a therapeutic drug. Hence this Peptibody binding feature could be important for its therapeutic function.

[0030] - Example 6 shows, in a mice model, that there is a therapeutic effect of the Peptibody against a pulmonary A. fumigatus infection. - Example 7 shows, in a mice model, that the increased chance of survival mediated by the Peptibody is due to elimination of living fungi in the lungs.

[0031] - Example 8 shows the design and test of an optimized peptibody containing the peptide of the invention.

[0032] - Example 9 shows that the optimized peptibody according to the invention can efficiently treat immunocompromised mice intranasally infected with Aspergillus fumigatus conidia, without detrimental side effects.

[0033] Overall, the presented data shows that an antifungal agent has been identified showing very efficient antifungal effects in mice model without detrimental side effects.

[0034] As also outlined above, Rosbjerg et al. suggests that the A. um / gatus-interacting binding site is at the CCP2-SP domain area. However, Rosbjerg et al. also suggests that the CCP2 domain "might have a central function in the interaction with the fungal cell surface. Moreover, the protease domain may stabilize the binding..." (p.220-221, bridging par.).

[0035] The present invention surprisingly discloses that a very small fragment of the CCP2-SP domain area can bind to fungi in the absence of the full protease domain, which in Rosbjerg et al. is indicated to be important for stabilizing the binding. The fact the protease domain can be avoided is very important from a technical point of view, since adding protease sites to a drug, can have several side effect (see also above).

[0036] As also outlined above, the selected fragment is by no means arbitrary, since other 30 amino acid fragments from the CCP2-SP domain does not bind to the fungi (see example 3A). That the fungus binding ability is unique for the peptide derived from MASP-1 can be seen in example 3E, wherein a corresponding 30 amino acid fragment from MASP-3 does not bind to fungi.

[0037] In one aspect, the present disclosure relates to an isolated fungus binding amino acid comprising a binding part having at least 70% sequence identity to SEQ ID NO: 1.

[0038] In another aspect, the present disclosure relates to an isolated fungus binding amino acid comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2. In another aspect, the present disclosure relates to fusion-protein constructs that contain the amino acid sequence as described herein and the amino acid sequence of an Fc-region from an immunoglobulin.

[0039] In another aspect, the present disclosure relates to a fungus binding peptibody comprising a fungus binding amino acid conjugated to an Fc-region from an immunoglobulin.

[0040] In another aspect, the present disclosure relates to an isolated nucleic acid encoding a fungus binding amino acid sequence as described herein or a nucleic acid construct encoding a fusion-protein as described herein.

[0041] In another aspect, the present disclosure relates to a host cell comprising a nucleic acid or nucleic acid construct as described herein.

[0042] In another aspect, the present disclosure relates to a method for producing a fungus binding amino acid sequence, the method comprising culturing the host cell as described herein, and recovering the fungus binding amino acid sequence.

[0043] As the skilled addressee knows, then the fungus binding amino acid sequence can also be made via chemical synthesis of peptides. Chemical synthesis of peptides can be carried out using classical solution-phase techniques, although these have been replaced in most research and development settings by solid-phase methods. However, solution-phase synthesis retains its usefulness in large-scale production of peptides for industrial purposes.

[0044] In another aspect, the present disclosure relates to a composition comprising a fungus binding amino acid sequence as described herein, and the use of such as a medicament, in therapy, prophylaxis and diagnostics.

[0045] In another aspect, the present disclosure relates to a method for detecting a fungus in a sample comprising providing a sample suspected of containing a fungus adding a fungus binding amino acid as described herein to the sample identifying the binding of the fungus binding amino acid to the fungus or fungal fragments within the sample. The compound can also be used to deliver site-directed anti-fungal compounds.

[0046] Another application for the peptide is in diagnostics. Invasive fungal infections are not easily diagnosed and often require several attempts using ELISA, PCR, cultivation, x-rays, CT or MRI scanning. The currently used assays lack sensitivity and new diagnostic tools are urgently needed. Our peptide would be a fast way to detect whether the patient has a fungal infection using plasma or lung fluid or by in vivo imaging.

[0047] DETAILED DESCRIPTION

[0048] As shown in the Examples below, MASP-1, a serine protease from the complement system, binds directly to various pathogenic fungi, for example, Aspergillus fumigatus. This discovery was utilized to design a targeted antifungal compound comprising the specific fungus binding amino acid sequences as disclosed herein.

[0049] MASP-1 binding was tested on the different growth stages of A. fumigatus. First, the A. fumigatus conidia were incubated on microscopy glass slides for 0, 4, 8 and 16 hours to obtain resting conidia, swollen conidia, germ tubes and hyphae. Recombinant MASP-1 was then added in a concentration of 5 pg / ml and binding was detected with a pan anti-MASP-l / -3 / MAP-l monoclonal antibody 8B3 and Alexa fluor 488-coupled goat anti-mouse antibody. Using fluorescence microscopy, recombinant MASP-1 binding was detected for all growth stages (fig. 1-8). This is very surprising since until now, MASP-1 is known only to interact with endogenous pattern recognition molecules (PRMs) and not so-called pathogen- associated molecular patterns (PAMPs). As exemplified below, then figures 13-15 demonstrate binding to the species Lichtemia corymbifera, Mucor circinelloides and Rhizopus arrhizus all belonging to the Mucorales order.

[0050] The novel aspect and the surprise element of the research disclosed herein lie in the unforeseen interaction of serine protease MASP-1, a protein typically implicated in the complement and coagulation systems, with the fungal species Aspergillus fumigatus. This previously unknown binding inspired a comprehensive exploration of its biological significance and possible dependence on the protein's activation state.

[0051] MASP-1 is produced as an inactive zymogen that undergoes activation during an immune response. Consequently, its interaction with Aspergillus fumigatus could be contingent on the state of the protein - active or inactive. To investigate this, we experimented with both the zymogen form of wild-type recombinant MASP-1 (rMASP-1 (444KLMAR448)) and the zymogen and active forms of a mutated variant (rMASP-1 (444DDDDK448)).

[0052] Unexpectedly, this investigation revealed that the mutated rMASP-1 (444DDDDK448) failed to bind to Aspergillus fumigatus conidia, regardless of the protein's activation state. In contrast, the wild-type rMASP-1 (444KLMAR448) demonstrated binding, emphasizing that the key determinant for the interaction was the alteration of the amino acid sequence rather than the protein's state of activation (see figure 28).

[0053] In the present context, the term "rMASP-1", refers to recombinant MASP-1.

[0054] In an unanticipated twist, the investigation into whether MASP-l's binding was state-dependent led to the unexpected discovery of a specific amino acid sequence essential for the fungus-MASP interaction. This finding has been instrumental in pinpointing the specific part of MASP-1 involved in interactions with Aspergillus fumigatus. It is this surprising revelation that underscores the novelty and significance of the findings disclosed herein, setting the foundation for further research into this unexpected fungal-protein interaction.

[0055] Fungus binding amino acid

[0056] In the present context, a fungus binding peptide relates to a peptide that binds to fungi belonging to the order of Mucorales, and in particular the genus Aspergillus.

[0057] Thus, in one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding amino acid comprising a binding part having at least 70% sequence identity to SEQ ID NO: 1.

[0058] The ability of such a peptide to bind to a fungus is exemplified for instance in figures 9-16 by a 30 amino acid peptide according to SEQ ID NO: 1 that binds Aspergillus fumigatus.

[0059] In an aspect the invention relates to an isolated fungus binding amino acid sequence comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2, preferably at least 90% sequence identity, more preferably at least 95% sequence identity, and having a length of at the most 70 amino acids, preferably, at the most 50 amino acids, more preferably at the most 40 amino acids, and even more preferably at the most 35 amino acids, such as being 30 amino acids long.

[0060] In an embodiment, the isolated fungus binding amino acid comprises a sequence differing from SEQ ID NO: 2 in at the most 1 amino acid position.

[0061] In another embodiment, the isolated fungus binding amino acid sequence comprises a fungus binding part having at least 80% sequence identity to SEQ ID NO: 1, and having a length of at the most 70 amino acids, preferably, at the most 50 amino acids, more preferably at the most 40 amino acids, and even more preferably at the most 35 amino acids.

[0062] In yet an embodiment, the isolated fungus binding amino acid sequence comprises a sequence differing from SEQ ID NO: 1 in at the most 6 amino acid positions, such as at the most 5 amino acids positions, such as at the most 4 amino acids positions, such as at the most 3 amino acids positions, preferably at the most 2 amino acids positions, and more preferably at the most 1 amino acids position.

[0063] In a preferred embodiment, the isolated fungus binding amino acid sequence comprises SEQ ID NO: 2.

[0064] In yet another preferred embodiment, the isolated fungus binding amino acid sequence comprises the cysteine at position 7 and / or the cysteine at position 11 of SEQ ID NO: 1, preferably comprising the cysteine at position 7 and the cysteine at position 11 of SEQ ID NO: 1.

[0065] In a further embodiment, the isolated fungus binding amino acid sequence has a length in the range 5-70 amino acids, such as 15-60 amino acids, such as 20-60 such as 20-60 amino acids, preferably 30-60 amino acids, more preferably 30-50 amino acids, even more preferably 30-40 amino acids, and even more preferably 30-35, such as being 31-35, 32-35, 33-35 amino acids long.

[0066] In yet an embodiment, the isolated fungus binding amino acid sequence comprises at least SEQ ID NO: 1.

[0067] The isolated fungus binding amino acid sequence may comprise part of the MASP- 1 sequence positioned directly N-terminal or C-terminal to SEQ ID NO: 1 in MASP- 1. Thus, in an embodiment, the isolated fungus binding amino acid sequence comprises or consists of the amino acid sequence 108-177 of SEQ ID NO: 10 (CCP1-CCP2-SP domain sequence), such as amino acids 118-177 of SEQ ID NO: 10, such as amino acids 128-177 of SEQ ID NO: 10, such as amino acids 128-167 of SEQ ID NO: 10, such as amino acids 128-157 of SEQ ID NO: 10, or such as amino acids 118-167 of SEQ ID NO: 10, such as amino acids 123-162.

[0068] One advantage of having identified a short peptide capable of binding to fungi, is that active domains which may cause side effects in vivo are omitted. Thus, in an embodiment, the isolated fungus binding amino acid is free of one or more of the following domains from MASP-1:

[0069] - CUB1 domain; and / or

[0070] - EGF domain; and / or

[0071] - CUB2 domain; and / or

[0072] - CCP1 domain; and / or

[0073] - full or functional CCP2 domain; and / or

[0074] - full or functional SP domain.

[0075] In a related embodiment, the isolated fungus binding amino acid sequence is free from one or more of the following functional sites of MASP-1:

[0076] • homodimerization site; and / or

[0077] • MBL binding site; and / or

[0078] • Ficolin-2 binding site; and / or

[0079] • One or more of the amino acids constituting the catalytic triad of the SP domain, preferably being from the catalytic site of the SP domain.

[0080] In an embodiment, the isolated fungus binding amino acid sequence according to any of the preceding claims, being at least 10 amino acids long, such as at least 15 amino acids long, preferably at least 20 amino acids long, more preferably at least 30 amino acids long.

[0081] Sequence identity

[0082] In the present context, proteins, homologues, derivatives, peptides and / or fragments thereof having an amino acid sequence at least, for example 70% identical to a reference amino acid sequence, is intended that the amino acid sequence of e.g., the peptide is identical to the reference sequence, except that the amino acid sequence may include up to 30 mutations per each 100 amino acids of the reference amino acid sequence.

[0083] In other words, to obtain a sequence at least 30% identical to a reference sequence, up to 30% of the amino acids or nucleotides in the reference sequence may be deleted or substituted with another amino acid / nucleotide, or several amino acids / nucleotides up to 30% of the total amount of the reference sequence.

[0084] These mutations of the reference sequence may occur at the amino or carboxy terminal positions of the reference amino acid sequence or anywhere between those terminal positions, interspersed either individually among amino acids in the reference sequence or in one or more contiguous groups within the reference sequence.

[0085] Methods to determine identity and similarity are codified in publicly available programs. Preferred computer program methods to determine identity and similarity between two sequences include, but are not limited to, the GCG program package, BLASTP, BLASTN, and FASTA.

[0086] The BI.ASTX program is publicly available from NCBI and other sources. Each sequence analysis program has a default scoring matrix and default gap penalties. In general, a molecular biologist would be expected to use the default settings established by the software program used.

[0087] Amino acid sequence identity

[0088] Thus, in one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding amino acid comprising a binding part having at least 70% sequence identity to SEQ ID NO: 1, such as for example 71% sequence identity to SEQ ID NO: 1, 72% sequence identity to SEQ ID NO: 1, 73% sequence identity to SEQ ID NO: 1, 74% sequence identity to SEQ ID NO: 1, 75% sequence identity to SEQ ID NO: 1, 76% sequence identity to SEQ ID NO: 1, 77% sequence identity to SEQ ID NO: 1, 78% sequence identity to SEQ ID NO: 1, 79% sequence identity to SEQ ID NO: 1, 80% sequence identity to SEQ ID NO: 1, 81% sequence identity to SEQ ID NO: 1, 82% sequence identity to SEQ ID NO: 1, 83% sequence identity to SEQ ID NO: 1, 84% sequence identity to SEQ ID NO: 1, 85% sequence identity to SEQ ID NO: 1, 86% sequence identity to SEQ ID NO: 1, 87% sequence identity to SEQ ID NO: 1, 88% sequence identity to SEQ ID NO: 1, 89% sequence identity to SEQ ID NO: 1, 90% sequence identity to SEQ ID NO: 1, 91% sequence identity to SEQ ID NO: 1, 92% sequence identity to SEQ ID NO: 1, 93% sequence identity to SEQ ID NO: 1, 94% sequence identity to SEQ ID NO: 1, 95% sequence identity to SEQ ID NO: 1, 96% sequence identity to SEQ ID NO: 1, 97% sequence identity to SEQ ID NO: 1, 98% sequence identity to SEQ ID NO: 1, 99% sequence identity to SEQ ID NO: 1 or 100% sequence identity to SEQ ID NO: 1.

[0089] Fungus binding peptide motif

[0090] Peptide motifs are amino acid patterns that are often conserved between functional peptide homologs, and which define structural peptide elements that are important for the specific functionality or bioactivity of a given polypeptide. Therefore, in the present context a fungus binding peptide motif is a motif that is required for a polypeptide or polypeptide construct to be able to bind a fungal cell or a fungal fragment.

[0091] The fungus binding activity of SEQ ID NO: 1 - a MASP-1 peptide fragment of 30 amino acids - tagged with biotin was demonstrated to bind several Aspergillus species by incubation together with several Aspergillus subspecies and detected by flow cytometry using FITC-coupled streptavidin as shown in figures 9-12.

[0092] Additionally, SEQ ID NO: 1 tagged with biotin also demonstrated binding to the species Lichtemia crymbifera, Mucor circinelloides and Rhizopus arrhizus all belonging to the Mucorales order. Binding was detected by flow cytometry using FITC-coupled streptavidin as shown in figures 13-15.

[0093] Another 30 amino acid sequence from the CUB1 domain and a random 30 amino acid peptide were both tagged with biotin and compared to the biotin tagged MASP-1 peptide according to SEQ ID NO: 1 for fungus binding activity in a flow cytometry assay using FITC-coupled streptavidin. However, only the MASP-1 peptide according to SEQ ID NO: 1 showed fungus binding activity (figures 16- 18).

[0094] In an effort to identify the peptide motif responsible for the fungus binding activity of MASP-1, i.e. the fungus binding peptide motif, the MASP-1 wild type sequence was mutated so that a 5 amino acid section444KLMAR448was substituted by the 5 amino acid section DDDDK.

[0095] The fungus binding ability of the wild type MASP-1 peptide and the DDDDK substituted MASP-1 was assayed by comparing binding of 5 pg / ml of each peptide to conidia from A. fumigatus. As shown in figure 26, The peptide having the KLMAR sequence has fungus binding activity while the mutated peptide with the DDDDK substitution does not bind to conidia of the tested fungi.

[0096] This result highlights the importance of the 5 amino acid fungus binding peptide motif KLMAR [SEQ ID NO: 2] for a peptide to possess fungus binding activity.

[0097] Therefore, in the present context, a fungus binding peptide motif is a motif that binds a fungal cell or fungal cell fragment via fluorescence microscopy using for example a secondary antibody, as exemplified below.

[0098] Thus, in one or more exemplary embodiments, the present disclosure relates to any fungus binding peptide motif that binds a fungal cell or fungal cell fragment.

[0099] Thus, in one or more exemplary embodiments, the present disclosure relates to a fungus binding peptide motif comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0100] Motif sequence identity

[0101] In line with the above defined sequence identity, the binding motif may vary, thus in one or more exemplary embodiments, the binding motif has at least 80% sequence identity to SEQ ID NO: 2, such as for example 81% sequence identity to SEQ ID NO: 2, 82% sequence identity to SEQ ID NO: 2, 83% sequence identity to

[0102] SEQ ID NO: 2, 84% sequence identity to SEQ ID NO: 2, 85% sequence identity to

[0103] SEQ ID NO: 1, 86% sequence identity to SEQ ID NO: 2, 87% sequence identity to

[0104] SEQ ID NO: 2, 88% sequence identity to SEQ ID NO: 2, 89% sequence identity to

[0105] SEQ ID NO: 2, 90% sequence identity to SEQ ID NO: 2, 91% sequence identity to

[0106] SEQ ID NO: 2, 92% sequence identity to SEQ ID NO: 2, 93% sequence identity to

[0107] SEQ ID NO: 2, 94% sequence identity to SEQ ID NO: 1, 95% sequence identity to

[0108] SEQ ID NO: 2, 96% sequence identity to SEQ ID NO: 2, 97% sequence identity to

[0109] SEQ ID NO: 2, 98% sequence identity to SEQ ID NO: 2, 99% sequence identity to

[0110] SEQ ID NO: 2 or 100% sequence identity to SEQ ID NO: 2.

[0111] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptide motif that is at least 7 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2.

[0112] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 7 amino acids and wherein said amino acid sequence includes a section of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0113] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 7 amino acids and wherein said amino acid sequence includes a section of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0114] In one or more exemplary embodiments, the isolated fungus binding amino acid sequence is at least 10 amino acids long and comprise a binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0115] Thus, in one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 10 amino acids, and wherein said amino acid sequence includes a section consisting of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0116] Thus, in one or more exemplary embodiments, the isolated fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 10 amino acids, and wherein said amino acid sequence includes a section consisting of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0117] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding peptide motif that is at least 15 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2.

[0118] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 15 amino acids and wherein said amino acid sequence includes a section of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0119] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 15 amino acids and wherein said amino acid sequence includes a section of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0120] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding peptide motif that is at least 20 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2. In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 20 amino acids and wherein said amino acid sequence includes a section of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0121] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 20 amino acids and wherein said amino acid sequence includes a section of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0122] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding peptide motif that is at least 25 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2.

[0123] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 25 amino acids and wherein said amino acid sequence includes a section of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0124] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 25 amino acids and wherein said amino acid sequence includes a section of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0125] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding peptide motif that is at least 30 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2.

[0126] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 30 amino acids and wherein said amino acid sequence includes a section of 5 amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0127] In one or more exemplary embodiments, the fungus binding peptide motif relates to an isolated amino acid sequence comprising at least 30 amino acids and wherein said amino acid sequence includes a section of 5 consecutive amino acids having at least 80% sequence identity to SEQ ID NO: 2.

[0128] In one or more exemplary embodiments, a fungus binding peptide motif as disclosed herein comprises an isolated fungus binding peptide motif having at least 60% sequence identity to an amino acid sequence according to SEQ ID NO: 2.

[0129] Fungi

[0130] The fungus binding amino acids and fungus binding peptide motifs disclosed herein binds selectively to fungi, in particular opportunistic pathogenic fungi, including fungi belonging to the order of Mucorales and the species Aspergillus.

[0131] Thus, as these fungi are all relevant from clinical perspective, The fungus binding amino acids and fungus binding peptide motifs disclosed herein could be used a tool for example in targeted treatment or diagnosis of fungal infections.

[0132] The fungus binding amino acids or motifs can be coupled to biological active components that provide a function to the peptide and that contributes to the physiochemical properties of the construct.

[0133] In the present context, a fungus is a eukaryotic organism that includes microorganisms such as yeasts and molds, as well as the more familiar mushrooms.

[0134] The fungal life cycle for most fungi and in particular for the filamentous fungi can be divided into four growth stages, resting conidia (spores), swollen conidia, germ tubes and hyphae. The isolated fungus binding amino acid and fungus binding peptide motifs disclosed herein binds to all these four fungal growth stages as shown in figures 1-8.

[0135] The binding of Peptibody to A. fumigatus conidia was tested by incubating 10 pg / ml of purified Peptibody with lxlO7heat-inactivated or live conidia / ml and the binding was detected with a rabbit anti-human IgG antibody and a FITC-coupled goat anti-rabbit antibody using flow cytometry. As shown in figure 27, the Peptibody binds to both the heat-inactivated and live A. fumigatus conidia.

[0136] A. fumigatus is a filamentous fungus, which means that the conidia grow into elongated structures called germ tubes and hyphae. The fungus expands in its natural environment by growing hyphae structures and these growth stages are also found in infected patients. Thus, binding to these growth stages is likely an important feature of an antifungal drug in case the fungus has already developed in the patient prior to diagnosis and treatment. Binding of the Peptibody to germ tubes and hyphae was tested using fluorescence microscopy. First, the A. fumigatus conidia were incubated on microscopy glass slides for 6 and 18 hours to develop germ tubes and hyphae. Afterward, the Peptibody was added in a concentration of 5 or 10 pg / ml and binding was detected with rabbit anti-human IgG antibody and an Alexa fluor 488-coupled goat anti-rabbit antibody. The Peptibody binding was detected on both germ tubes and hyphae as shown in figures 20-22.

[0137] In one or more exemplary embodiments, the fungus binding amino acids and fungus binding peptide motifs disclosed herein binds selectively to resting conidia (spores).

[0138] In one or more exemplary embodiments, the fungus binding amino acids and fungus binding peptide motifs disclosed herein binds selectively to swollen conidia.

[0139] In one or more exemplary embodiments, the fungus binding amino acids and fungus binding peptide motifs disclosed herein binds selectively to germ tubes.

[0140] In one or more exemplary embodiments, the fungus binding amino acids and fungus binding peptide motifs disclosed herein binds selectively to hyphae.

[0141] In one or more exemplary embodiments, the fungus is a cellular organism belonging to the order of the Mucorales.

[0142] Mucorales is the largest and best studied order of zygomycete fungi. Members of this order are sometimes called pin molds. The term mucormycosis is commonly used for infections caused by molds belonging to the order Mucorales.

[0143] Thus in one or more exemplary embodiments the present disclosure relates to a fungus belonging to any one of the genus selected from the list consisting of Actinomucor spp, Apophysomyces spp, Benjaminella spp., chaetocladium spp., Circinella spp., Cokeromyces spp., Dicranophora spp., Ellisomyces spp., Helicostylum spp., Hyphomucor spp., Kirkomyces spp., Mucor spp., Parasitella spp., Pilaira spp., Pliphora spp., Pirella spp., Rhizomucor spp., Lichtheimia spp., Rhizopodopsis spp., Rhizopus spp., Sporodinella spp., Syzygites spp., Thamnidium spp., Thermomucor spp., Zygorhynchus spp., Fusarium spp., Scedosporium spp., Penicillium spp., Aspergillus spp., Candida spp., and Cryptococcus spp.

[0144] In one or more exemplary embodiments, the present disclosure relates to a fungus belonging to any one of the genus Rhizopus spp., Mucor spp., and Lichtemia spp. In one or more exemplary embodiments, the present disclosure relates to a fungus belonging to the genus Rhizopus spp.

[0145] In one or more embodiments the fungus belonging to the genus Rhizopus spp. is Rhizopus arrhizus.

[0146] In one or more exemplary embodiments, the present disclosure relates to a fungus belonging to the genus Mucor spp.

[0147] In one or more embodiments the fungus belonging to the genus Mucor spp. is Mucor circinelloides.

[0148] In one or more exemplary embodiments, the present disclosure relates to a fungus belonging to the genus Lichtemia.

[0149] In one or more exemplary embodiments the fungus belonging to the genus Lichtemia spp. is Lichtemia corymbifera.

[0150] In one or more exemplary embodiments the present disclosure relates to a fungus belonging to any of the genus Aspergillus spp., Candida spp., and Cryptococcus spp-

[0151] In one or more exemplary embodiments the present disclosure relates to a fungus belonging to the genus Aspergillus spp.

[0152] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is any one of the species selected from the list consisting of Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus, Aspergillus niger and Aspergillus nidulans.

[0153] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus fumigatus.

[0154] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus flavus.

[0155] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus terreus.

[0156] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus niger.

[0157] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus nidulans. In one or more exemplary embodiments the present disclosure relates to a fungus belonging to the genus Candida spp.

[0158] In one or more exemplary embodiments, the fungus binding amino acid does not bind to a fungus belonging to the genus Candida spp.

[0159] In one or more exemplary embodiments the present disclosure relates to a fungus belonging to the genus Cryptococcus spp.

[0160] In one or more exemplary embodiments, the fungus binding amino acid does not bind to a fungus belonging to the genus Cryptococcus spp.

[0161] Opportunistic fungal pathogens

[0162] Opportunistic fungal pathogens are fungi that are nonpathogenic in the immunocompetent host. These fungi may cause opportunistic infections in weakened or immunocompromised hosts. Such fungi can for example be part of the upper respiratory tract flora and may cause pulmonary infection in hosts that become weakened or immunocompromised because of a different condition or disorder.

[0163] In one or more exemplary embodiments, the isolated fungus binding amino acid or motifs binds to an opportunistic fungus.

[0164] Opportunistic fungal infections are commonly caused by Aspergillus spp., Candida spp., Cryptococcus spp. with Aspergillus being the mold most associated with infection in patients with hematological malignancy and bone marrow disease.

[0165] Thus, in one or more exemplary embodiments, the opportunistic fungus is selected from the group consisting o Aspergillus spp., Candida spp., and Cryptococcus spp.

[0166] In one or more exemplary embodiments, the opportunistic fungus belongs to the Aspergillus genus.

[0167] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is any one of the species selected from the list consisting of Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus, Aspergillus niger and Aspergillus nidulans.

[0168] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus fumigatus. In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus flavus.

[0169] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus terreus.

[0170] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus niger.

[0171] In one or more exemplary embodiments of the present disclosure, the fungus belonging to the genus Aspergillus, is Aspergillus nidulans.

[0172] In one or more exemplary embodiments, the opportunistic fungus belongs to the Candida genus.

[0173] In one or more exemplary embodiments, the opportunistic fungus belongs to the Cryptococcus genus.

[0174] In one or more exemplary embodiments, the opportunistic fungus is Aspergillus fumigatus.

[0175] Fungal fragments

[0176] Fungi may reproduce asexually by fragmentation, budding, or producing spores. Fragments of hyphae can grow new colonies. Mycelial fragmentation occurs when a fungal mycelium separates into pieces with each component growing into a separate mycelium. Fungal fragments and undocumented conidia may function as aeroallergen sources.

[0177] Thus, in one or more exemplary embodiments the isolated fungus binding amino acids and fungus binding peptide motifs disclosed herein binds to fungal fragments. Within the context of the present disclosure a fungal fragment is fragments of fungal conidia, fungal germ tubes or fungal hyphae.

[0178] Fusion Proteins

[0179] Fusion proteins are proteins created through the joining of genetic sequences encoding for partial or whole sequence of different proteins. Such joining of genetic sequences results in the transcription and translation of a single continuous genetic sequence, thereby resulting in chimeric proteins comprising the protein equivalents of each of the genetic sequences are fused to each other through peptide bonds as part of the translation process. The peptide-coupled functional component, e.g., an immunoglobulin Fc region, can work therapeutically against fungal infections by recruiting the immune apparatus of the patient, thereby facilitating clearance of the pathogen. An Fc region enables fungal killing via phagocytosis, antibody-dependent cellular cytotoxicity and by activating the complement system mediating further opsonization and phagocytosis as well as anaphylatoxin release and recruitment of inflammatory cells and complement-mediated cytolysis. Gathering multiple peptides with Fc regions also enables agglutination, which can block the fungi from accessing the epithelium and contribute to fungal clearance by making the phagocytosis more efficient.

[0180] Thus, in one or more exemplary embodiments, the present disclosure relates to a fusion-protein construct that contains a fungus binding amino acid as disclosed herein and the amino acid sequence of an Fc-region.

[0181] In one or more exemplary embodiments, the present disclosure relates to a fusion-protein construct that contains a fungus binding amino acid having at least 70% sequence identity to SEQ ID NO: 1 and the amino acid sequence of an Fc- region.

[0182] In one or more exemplary embodiments, the present disclosure relates to a fusion-protein construct that contains a fungus binding peptide motif as disclosed herein and the amino acid sequence of an Fc-region.

[0183] In one or more exemplary embodiments, the present disclosure relates to a fusion-protein construct that contains a fungus binding motif having at least 80% sequence identity to SEQ ID NO: 2 and the amino acid sequence of an Fc-region.

[0184] In one or more exemplary embodiments, the present disclosure relates to a fusion-protein construct that contains a fungus binding motif that is at least 10 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2 and the amino acid sequence of an Fc-region.

[0185] The fungus binding amino acid and / or fungus binding peptide motif as disclosed herein can be conjugated to an Fc-region from a human immunoglobulin or a nonhuman immunoglobulin. In one or more exemplary embodiments, the Fc-region is from a human immunoglobulin selected from the group consisting of an IgG, IgA, IgM, IgE, and IgD.

[0186] In one or more exemplary embodiments, the human immunoglobulin Fc-region is one of the immunoglobulin isotypes selected from the list consisting of an IgG, IgA or IgD Fc-region.

[0187] In one or more exemplary embodiments, the human immunoglobulin Fc-region is one of the immunoglobulin isotypes selected from the list consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2 or IgD Fc-region.

[0188] In one or more exemplary embodiments, the Fc-region is from a human immunoglobulin selected from the group consisting of an IgG and IgM.

[0189] In one or more exemplary embodiments, the Fc-region is from a human immunoglobulin selected from the group consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, and IgD. in one or more exemplary embodiments, the Fc-region is from a human immunoglobulin selected from the group consisting of an IgGl, IgG2, IgG3, IgG4, and IgM

[0190] In one or more exemplary embodiments, the Fc-region is a human IgGl.

[0191] In one or more exemplary embodiments, the Fc-region is a human IgG2.

[0192] In one or more exemplary embodiments, the Fc-region is a human IgG3.

[0193] In one or more exemplary embodiments, the Fc-region is a human IgG4.

[0194] In one or more exemplary embodiments, the Fc-region is a human IgAl.

[0195] In one or more exemplary embodiments, the Fc-region is a human IgA2.

[0196] In one or more exemplary embodiments, the Fc-region is a human IgM.

[0197] In one or more exemplary embodiments, the Fc-region is a human IgE.

[0198] In one or more exemplary embodiments, the Fc-region is a human IgD.

[0199] As described above, the fungus binding amino acid and / or fungus binding peptide motif as disclosed herein can also be conjugated to an Fc-region from a nonhuman immunoglobulin. Thus, in one or more exemplary embodiments, the Fe region is a non-human Fc-region selected from the list consisting of canine, horse, bovine, sheep, swine, goat, mink, ferret, cat, rat, mouse, rabbit and guinea pig.

[0200] In one or more exemplary embodiments, the Fc-region is a non-human Fc-region selected from the list consisting of canine, horse, bovine, sheep, swine, goat, rat, mouse, rabbit and guinea pig.

[0201] In one or more exemplary embodiments, the Fc-region is a non-human Fc-region selected from the list consisting of, goat, rat, mouse, rabbit and guinea pig.

[0202] In one or more exemplary embodiments, the Fc-region is a non-human Fc-region selected from the list consisting of, goat, rabbit, and guinea pig.

[0203] In one or more exemplary embodiments, the Fc-region is a rat Fc-region.

[0204] In one or more exemplary embodiments, the Fc-region is a rat IgG Fc-region.

[0205] In one or more exemplary embodiments, the Fc-region is a mouse Fc-region.

[0206] In one or more exemplary embodiments, the Fc-region is a mouse IgG Fc-region.

[0207] In one or more exemplary embodiments, the Fc-region is a rabbit Fc-region.

[0208] In one or more exemplary embodiments, the Fc-region is a guinea pig Fc-region.

[0209] In one or more exemplary embodiments, the Fc-region is a guinea pig IgG Fc- region.

[0210] Fusion protein linkers

[0211] Fusion proteins may also contain linkers that are located between the protein constituent parts of the fusion protein and thereby affect how each protein constituent part can move in relation to the other protein constituent parts. A poly-glycine linker consisting of a series of glycine residues, for example, can be used to increase mobility of the protein constituent parts, by providing a region that can curl and form an adaptable shape depending on the environment the fusion protein is currently in. Such linkers and methods for introducing these into fusion proteins are well known in the prior art.

[0212] Thus, in one or more exemplary embodiments, the present disclosure relates to a fungus binding fusion protein as disclosed herein, that further comprises a linker connecting the fungus binding amino acid or the fungus binding peptide motif to the fusion partner e.g., the exemplified Fc-region.

[0213] In one or more exemplary embodiments, the linker is a hinge-region connecting the fungus binding amino acid or the fungus binding peptide motif to the fusion partner.

[0214] Peptibodies

[0215] A peptibody is a peptide construct comprising at least two peptide moieties, a biologically active peptide and an Fc-region, wherein the biologically active peptide is grafted onto an Fc region. This makes peptibodies both an attractive and flexible alternative to monoclonal antibodies in any process or method that traditionally makes use of monoclonal antibodies.

[0216] A peptide-coupled component, e.g., an Fc region, can improve the pharmacokinetics and increase the in vivo half-life of the compound. An Fc region can also be manipulated to further increase the effectiveness and further prolong the half-life of the construct. An Fc region can for example be designed to be recycled in the endocytic pathway in host cells in order to increase the half-life of the construct. The compound can also be pegylated to extent the half-life and bioavailability.

[0217] The Fc region can be manipulated to change the mode of interaction with components from the immune system to increase / decrease the inflammatory process by e.g., changing the interaction with Clq from the complement system.

[0218] An Fc region can benefit the production of the construct by improving the structural and biochemical stability and by easing the purification process.

[0219] The first formulation made with the peptide is a fusion protein containing the peptide, a hinge region and the Fc-region of a human IgGl. This so-called peptibody mimics the structure, function and biological half-life of natural immunoglobulins, but has the unique peptide binding region instead of the antigen-binding Fab fragment. An example of a generalized structure of a peptibody is shown in figure 19. The peptibody was produced recombinantly using Expi293 cells and purified using protein G sepharose. A fungus binding peptibody

[0220] Thus, in one or more exemplary embodiments, a fungus binding peptibody is a fungus binding amino acid as disclosed herein grafted onto an Fc-region from an immunoglobulin.

[0221] In one or more other exemplary embodiments, the fungus binding peptibody comprise a fungus binding peptide motif as disclosed herein grafted onto an Fc- region from an immunoglobulin.

[0222] As described above, the Fc-region can both be a human or a non-human immunoglobulin.

[0223] In one or more other exemplary embodiments, the peptibody do not bind to a human cell.

[0224] In one or more exemplary embodiments, the fungus binding peptibody comprise a fungus binding amino acid having at least 70% sequence identity to SEQ ID NO: 1 conjugated to an Fc-region.

[0225] In one or more exemplary embodiments, the fungus binding peptibody comprise a fungus binding motif having at least 80% sequence identity to SEQ ID NO: 2 and conjugated to an Fc-region.

[0226] In one or more exemplary embodiments, the fungus binding peptibody comprise a fungus binding motif that is at least 10 amino acids long, said binding motif having at least 80% sequence identity to SEQ ID NO: 2 conjugated to an Fc- region.

[0227] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, further comprising a hinge region.

[0228] In an embodiment, the linker has length in the range 5-15 amino acids, preferably in the range 7-12 amino acids.

[0229] In an embodiment, the hinge / linker is selected from the group consisting of EPKSCDKTHTCP (SEQ ID NO: 14) and DKTHTCP (SEQ ID NO: 15). These are the linkers / hinges in the peptibody constructs tested.

[0230] It may be an advantage that the linker of the peptibody does not contain unpaired cysteines, since such cysteines may cause aggregates / complexes of the peptibody. Thus, in an embodiment, the hinge / linker is free from unpaired cysteines, such as only comprising one pairing cysteine in the linker region. Thus, in a preferred embodiment, the linker / hinge is DKTHTCP (SEQ ID NO: 15).

[0231] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the human immunoglobulin Fc-region is selected from the list consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, or IgD Fc-region.

[0232] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the human immunoglobulin Fc-region is one of the immunoglobulin isotypes selected from the list consisting of an IgG, IgA, IgM, IgE or IgD Fc-region.

[0233] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the human immunoglobulin Fc-region is one of the immunoglobulin isotypes selected from the list consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE or IgD Fc-region.

[0234] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgGl.

[0235] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgG2.

[0236] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgG3.

[0237] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgG4.

[0238] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgAl.

[0239] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgA2.

[0240] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgM.

[0241] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgE. In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a human IgD.

[0242] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a non-human Fc-region.

[0243] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a non-human Fc-region is selected from the list consisting of list consisting of canine, horse, bovine, sheep, swine, goat, mink, ferret, cat, rat, mouse, rabbit and guinea pig.

[0244] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a non-human Fc-region is selected from the list consisting of list consisting of canine, horse, bovine, sheep, swine, goat, rat, mouse, rabbit and guinea pig.

[0245] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a non-human Fc-region is selected from the list consisting of list consisting of goat, rat, mouse, rabbit and guinea pig.

[0246] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a non-human Fc-region is selected from the list consisting of list consisting of goat, rat, and guinea pig.

[0247] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a rat Fc-region.

[0248] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a rat IgG Fc-region.

[0249] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a mouse Fc-region.

[0250] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a mouse IgG Fc-region.

[0251] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a rabbit Fc-region.

[0252] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a rabbit IgG Fc-region. In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a guinea pig Fc-region.

[0253] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody, wherein the Fc-region is a guinea pig IgG Fc-region.

[0254] Peptibody formulae I

[0255] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody having the structure defined in Formula (I);

[0256] Formula (I): X1-L1-F1 wherein,

[0257] -XI is a fungus binding amino acid selected from the group consisting of Pl, P2 and P3, where;

[0258] -Pl is a fungus binding amino acid comprising a binding part, the binding part having at least 70% sequence identity towards SEQ ID NO: 1,

[0259] -P2 is a fungus binding motif comprising a binding motif having at least 80% sequence identity towards SEQ ID NO: 2,

[0260] -P3 is a fungus binding motif comprising a binding motif being at least 10 amino acids long and having at least 80% sequence identity to SEQ ID NO: 2

[0261] -Fl is an Fc Region, and

[0262] -LI is a peptide bond, linker or hinge region connecting XI to Fl.

[0263] In one or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody having the structure defined in Formula (II);

[0264] Formula (II): Xl*y-Ll-Fl wherein,

[0265] -XI is selected from the group consisting of Pl or P2, wherein

[0266] -Pl is a first peptide according to the invention, -P2 is a second peptide according to the invention,

[0267] -Fl is an Fc Region, and

[0268] -LI is a peptide bond, linker or hinge region connecting XI to Fl

[0269] -y is the number of fungus binding peptides wherein y=l-10, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.

[0270] Fc-region

[0271] In the present context, an Fc-region is the tail region of an antibody [fragment crystallizable region (Fc region)] that interacts with cell surface receptors called Fc receptors and some proteins of the complement system. This property allows antibodies to activate the immune system.

[0272] The conjugation / covalent attachment of a peptide of interest to an Fc-region provides a number of characteristics to the peptide construct that are useful for therapeutical and diagnostic uses. For example, the Fc-region confers increased half-life in serum onto the peptibody and allows for easy separation / isolation of the peptide construct through a high affinity to Protein A and Protein G.

[0273] The addition of the Fc-region also facilitates detection of peptibody binding to a fungus in a sample or a patient by allowing for immunoblotting targeting the Fc- region using known anti-Fc antibodies.

[0274] A therapeutic advantage in the case of a human IgG Fc-region is that this Fc- region also provides direct activation of the specific immune response upon fungus binding when delivered to the bloodstream of a human patient in need thereof.

[0275] A diagnostic advantage in the case of non-human Fc-regions is that non-human Fc-regions will not cross-react with host / subject immunoglobulin Fc-regions during the detection steps.

[0276] Thus, in one or more exemplary embodiments, the present disclosure relates to a fungus binding amino acid, a fungus binding peptide motif, a fungus binding peptibody or a peptibody according to formula (I) as defined herein conjugated to an Fc-region from a human immunoglobulin, wherein the Fc-region is chosen from the list of IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, or IgD Fc-region. In an embodiment, the human immunoglobulin is IgGl. IgGl can opsonize, interact with fc receptors and complement activating capacity.

[0277] In another embodiment, the human immunoglobulin is IgG3. IgG3 can opsonize, interact with fc receptors and has highest complement activating capacity among the IgG subclasses.

[0278] In one or more exemplary embodiments, the present disclosure relates to a fungus binding amino acid, a fungus binding peptide motif, a fungus binding peptibody or a peptibody according to formula (I) as defined herein conjugated to an Fc-region from a non-human immunoglobulin.

[0279] In an embodiment, the peptibody according to the invention comprises an Fc region (CH2-CH3 domains) as defined for SEQ ID NO: 8 and / or SEQ ID NO: 9. See sequence list overview further below.

[0280] In an embodiment, the peptibody is a peptibody according to SEQ ID NO: 8 or SEQ ID NO: 9.

[0281] Attachment of bioactive compounds

[0282] A bioactive compound is any compound or peptide that has an effect in or on an organism, tissue or cell. It is commonly known that such bioactive compounds can be attached to other carrier molecules such as for example antibodies in order to either facilitate detection of a molecular target or facilitate delivery of the bioactive compound to a molecular target. Methods for attachment of bioactive molecules onto a carrier molecule such as by conjugation is well known in the prior art.

[0283] Thus, in one or more exemplary embodiments, the present disclosure relates to fungus binding amino acids, fungus binding peptide motifs, fungus binding fusion proteins and / or fungus binding peptibodies as disclosed herein that are modified by the attachment of bioactive compounds.

[0284] In one or more exemplary embodiments, a bioactive compound is selected from a list consisting of fluorophores, fluorescent / phosphorescent proteins or compounds, radiolabeled compounds, antibiotics, enzymatically active peptides, molecular probes, functional tags such as HIS / FLAG-tags and / or PEG.

[0285] In one or more exemplary embodiments, the peptibody is conjugated to an imaging agent or a radioligand. Antibiotic conjugates

[0286] In the present context, an antibiotic / antifungal conjugate, is an antibiotic / antifungal that is conjugated onto a carrier molecule. In one example of such a carrier molecule is an antibody. The antibody enables specific immunorecognition of the target organism and brings the conjugated antibiotic / antifungal into contact with the target organism, delivering the antibiotic / antifungal directly to where it is needed.

[0287] In the same way, the isolated fungus binding amino acids, isolated fungus binding peptide motifs, fungus binding fusion proteins and / or fungus binding peptibodies as disclosed herein can take on the same targeting role as the antibody and provide delivery of an antibiotic / antifungal directly to the desired target.

[0288] Thus, in one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding amino acid, fungus binding peptide motif, fungus binding fusion protein and / or a fungus binding peptibody as disclosed herein onto which an antibiotic / antifungal is attached.

[0289] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding amino acid, fungus binding peptide motif, fungus binding fusion protein and / or a fungus binding peptibody as disclosed herein that is conjugated to an antibiotic / antifungal.

[0290] In one or more exemplary embodiments the isolated fungus binding amino acids, isolated fungus binding peptide motifs, fungus binding fusion proteins and / or fungus binding peptibodies as disclosed herein are conjugated to antibiotics such as amphotericin B, triazoles and / or radionuclides.

[0291] In one or more exemplary embodiments the isolated fungus binding amino acids, isolated fungus binding peptide motifs, fungus binding fusion proteins and / or fungus binding peptibodies as disclosed herein are conjugated to cytotoxic drugs and toxic radionucleotides.

[0292] Thus, in one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding amino acid sequence comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1 that is conjugated to an antibiotic / antifungal. In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding motif comprising a binding motif having at least 80% sequence identity towards SEQ ID NO: 2 that is conjugated to an antibiotic / antifungal.

[0293] In one or more exemplary embodiments, the present disclosure relates to an isolated fungus binding motif comprising a binding motif being at least 10 amino acids long and having at least 80% sequence identity towards SEQ ID NO: 2 that is conjugated to an antibiotic / antifungal.

[0294] In one or more exemplary embodiments, the present disclosure relates to a fungus binding fusion peptide as defined herein that is conjugated to an antibiotic / antifungal.

[0295] In or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody as defined herein that is conjugated to an antibiotic / antifungal.

[0296] In or more exemplary embodiments, the present disclosure relates to a fungus binding peptibody according to Formulas (I) that is conjugated to an antibiotic / antifungal.

[0297] PEGylation

[0298] PEGylation is the process of attaching polyethylene glycol (PEG) polymers onto molecules and commonly peptides. The attachment of PEG can result in changes in the physiochemical properties of the target molecule and may include changes in for example conformation, electrostatic binding, and hydrophobicity. These changes can increase systemic retention of the target molecule, improved solubility, reduced dosage frequency requirements, reduced toxicity and enhanced protection from proteolytic degradation.

[0299] Thus, in the present context, the isolated fungus binding amino acids, isolated fungus binding peptide motifs, fungus binding fusion proteins and / or fungus binding peptibodies as disclosed herein can be chemically modified by the addition of PEG.

[0300] In one or more exemplary embodiments, the present disclosure relates to fungus binding amino acids that are PEGylated. Labelling or tagging

[0301] In one or more exemplary embodiments of the present disclosure an isolated fungus binding peptide, isolated fungus binding peptide motif or fungus binding peptibody as disclosed herein are further modified in order to achieve various beneficial properties.

[0302] In one or more exemplary embodiments conjugation of the isolated fungus binding amino acid sequence, isolated fungus binding peptide motif or fungus binding peptibodies as defined herein to signal molecules further enable easy detection of fungal cells or fungal cell fragments in a sample or in vivo by monitoring the signal molecule.

[0303] Thus, in one or more exemplary embodiments the isolated fungus binding peptide, isolated fungus binding peptide motif or fungus binding peptibodies as disclosed herein are conjugated to phosphorescent molecules, fluorescent molecules such as fluorescein or fluorescent proteins, e.g. GFP, YFP, radiolabeled molecules or affinity based systems such as biotin or streptavidin that are helpful in enabling detection. Such molecules and methods are generally known in the prior art and is further exemplified by the use of biotin tagged peptides in example 3 and as shown in figures 9-16.

[0304] In one or more exemplary embodiments the isolated fungus binding peptide, isolated fungus binding peptide motif or fungus binding peptibodies as disclosed herein are tagged with a molecular tag such as HIS8, HIS6, HIS4 or FLAG-tags, facilitating peptide purification and enabling antibody detection by antibody binding of the molecular tag.

[0305] Nucleic acid

[0306] The term nucleic acid in the present context is used to describe DNA and RNA, members of a family of biopolymers, and is synonymous with polynucleotide. Thus, the present disclosure relates to a nucleotide sequence which encodes the polypeptides described herein.

[0307] In one or more exemplary embodiments, the present disclosure relates to an isolated nucleic acid encoding a fungus binding amino acid sequence comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1. In one or more exemplary embodiments, the present disclosure relates to an isolated nucleic acid encoding a fungus binding motif comprising a binding motif having at least 80% identity to SEQ ID NO: 2.

[0308] In one or more exemplary embodiments, the present disclosure relates to an isolated nucleic acid encoding a fungus binding motif comprising a binding motif being at least 10 amino acids long and having at least 80% sequence identity towards SEQ ID NO: 2.

[0309] Nucleic acid construct

[0310] The present disclosure also relates to a nucleic acid construct comprising the polynucleotides operably linked to one or more control sequences. A nucleic acid construct within the present context is an artificial construct comprising a nucleic acid insert that is integrated into or borne by a vector. The vector can be delivered via transformation / transfection to a host cell by for example physical, chemical, or viral methods and allow the nucleic acid inserts to be replicated or expressed in the host cell. Such methods for delivering a vector into a host cell are well known to the skilled person.

[0311] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct encoding a fusion-protein, the fusion-protein comprising a fungus binding amino acid comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1 and the amino acid sequence of an Fc-region from a human immunoglobulin.

[0312] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct encoding a fusion-protein, the fusion-protein containing a fungus binding motif comprising a binding motif having at least 80% identity to SEQ ID NO: 2 and the amino acid sequence of an Fc-region from a human immunoglobulin.

[0313] In one or more exemplary embodiments, the present disclosure relates to nucleic acid constructs or vectors that facilitate transformation into a suitable host cell.

[0314] In one or more exemplary embodiments, the nucleic acid constructs or vectors of the present disclosure comprise one or more control elements (expression start / stop / transposable elements). In one or more exemplary embodiments, the present disclosure relates to nucleic acid constructs and vectors of use in producing fungus binding amino acid sequences, fungus binding peptide motifs, fungal binding fusion proteins or fungal binding peptibodies as disclosed herein.

[0315] Fungal binding amino acid construct

[0316] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding amino acid as disclosed herein.

[0317] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding amino acid comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1.

[0318] Fungal binding peptide motif construct

[0319] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptide motif as disclosed herein.

[0320] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptide motif comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0321] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding motif that is at least 5 amino acids, at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 25 amino acids, at least 30 amino acids long.

[0322] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptide motif comprising a binding motif that is at least 10 amino acids long, said binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0323] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding motif that is at least 5 amino acids, at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 25 amino acids, at least 30 amino acids long, wherein said fungus binding peptide further comprises a section consisting of 5 consecutive amino acids, the 5 consecutive amino acids being the amino acid sequence KLMAR [SEQ ID NO: 2].

[0324] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding motif that is at least 5 amino acids, at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 25 amino acids, at least 30 amino acids long, wherein said fungus binding peptide further includes a section consisting of 5 consecutive amino acids, the 5 consecutive amino acids having 80% sequence identity to the amino acid sequence KLMAR [SEQ ID NO: 2],

[0325] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding motif that is at least 5 amino acids, at least 10 amino acids, at least 15 amino acids, at least 20 amino acids, at least 25 amino acids, at least 30 amino acids long, wherein said fungus binding peptide further includes a section consisting of 5 consecutive amino acids, the 5 consecutive amino acids having 60% sequence identity to the amino acid sequence KLMAR [SEQ ID NO: 2],

[0326] Fungal binding fusion protein construct

[0327] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding fusion protein as disclosed herein.

[0328] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding fusion protein including an Fc-region and a fungus binding amino acid comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1 and an Fc-region.

[0329] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding fusion protein comprising an Fc-region and a fungus binding peptide motif having at least 80% sequence identity to SEQ ID NO: 2.

[0330] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding fusion protein comprising and Fc-region and a fungus binding peptide motif that is at least 10 amino acids long, said binding motif having at least 80% sequence identity to SEQ ID NO: 2. Fungal binding peptibody construct

[0331] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptibody as disclosed herein.

[0332] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptibody including a fungus binding amino acid comprising a binding part, the binding part having at least 70% sequence identity to SEQ ID NO: 1.

[0333] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes fungus binding peptibody comprising a fungus binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0334] In one or more exemplary embodiments, the present disclosure relates to a nucleic acid construct that encodes a fungus binding peptibody comprising a fungus binding motif that is at least 10 amino acids long, said binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0335] Host cells

[0336] To manufacture biotechnology products, cells can be engineered for the production of recombinant proteins including for example peptibodies. Within the context of the present disclosure a host cell is a cell that is used for harboring and expressing a nucleic acid or nucleic acid construct as defined herein. Methods for introducing nucleic acid and nucleic acid constructs into cells to provide host cells are well known in the prior art.

[0337] Thus, in one or more exemplary embodiments, the present disclosure relates to a host cell comprising a nucleic acid or nucleic acid construct encoding a fungus binding amino acid sequence according to any one of the fungi binding amino acids, fungi binding motifs, fungi binding fusion-proteins, fungi binding peptibodies and / or peptibodies of Formula (I) as disclosed herein.

[0338] In one or more exemplary embodiments, the host cell expresses a fungus binding amino acid sequence encoded by the nucleic acid or nucleic acid construct as disclosed herein. In one or more exemplary embodiments, the present disclosure relates to a host cell that is selected from a list comprising yeast cells, bacterial cells, mammalian cells and plant cells.

[0339] In one or more exemplary embodiments, the host cell is a yeast cell.

[0340] In one or more exemplary embodiments, the host cell is a bacterial cell.

[0341] In one or more exemplary embodiments, the host cell is an E. coli bacterial cell.

[0342] In one or more exemplary embodiments, the host cell is a mammalian cell.

[0343] In one or more exemplary embodiments, the host cell is a plant cell.

[0344] Method for polypeptide production

[0345] In the present context, a method for polypeptide production relates to a method wherein a host cell is cultured / cultivated to produce a polypeptide by expression of a nucleic acid or nucleic acid construct. Such methods for production of polypeptide in a host cell are well known in the prior art.

[0346] Thus, in one or more exemplary embodiment, the present disclosure relates to a method for producing a fungus binding amino acid sequence encoding fungus binding amino acids, fungus binding motifs, fungus binding fusion-proteins, fungus binding peptibodies and / or peptibodies of Formula (I) as disclosed herein, the method comprising: culturing a host cell as defined herein, and recovering the fungus binding amino acid sequence.

[0347] In one or more exemplary embodiments, the present disclosure relates to a method for protein production by culturing a host cell comprising a nucleic acid or nucleic acid construct as defined herein.

[0348] Compositions

[0349] In one or more exemplary embodiments, the present disclosure relates to compositions comprising a fungus binding amino acid, a fungus binding motif, a fungus binding fusion-protein, a fungus binding peptibody and / or peptibodies of formula (I) as disclosed herein.

[0350] In one or more exemplary embodiments, the composition can be used as an antimicrobial agent. In one or more exemplary embodiments, the composition may be the fungus binding amino acid sequences per se.

[0351] Using a Fc fusion technology, one can easily couple cytotoxic drugs and toxic radionucleotides to the constructs allowing this to be a novel antibiotic / antifungal without the need for endogenous immune assistance. This will be a significant advantage in neutropenic patients, who are usually prone to aspergillus infections.

[0352] Pharmaceutical compositions

[0353] In one or more exemplary embodiments, the present disclosure relates to pharmaceutical compositions having as an active ingredient a fungus binding amino acid sequence, a fungus binding motif, a fungus binding fusion-protein or a fungus binding peptibody as disclosed herein.

[0354] Pharmaceutical composition comprises in addition to the active ingredient, therapeutically inactive ingredients, such as a pharmaceutically acceptable or physiologically acceptable excipient, carrier and / or adjuvants, which are well- known to the person skilled in the art and may include, but are not limited to, solvents, emulsifiers, wetting agents, plasticizers, solubilizers (e.g. solubility enhancing agents) coloring substances, fillers, preservatives, anti-oxidants, antimicrobial agents, viscosity adjusting agents, buffering agents, pH adjusting agents, isotonicity adjusting agents, mucoadhesive substances, and the like. Examples of formulation strategies are well-known to the person skilled in the art.

[0355] In the present context a pharmaceutical composition is a mixture of ingredients suitable for administering to a subject that includes an active ingredient.

[0356] Medical use

[0357] The fungus binding amino acid could also be used to treat infections with bacteria (e.g., mycobacteria), other fungi, viruses (e.g., herpes viruses) or parasites (e.g., malaria parasites) that may bind the peptide sequence.

[0358] One medical composition developed is the fusion-protein construct that contains a fungus binding amino acid and the Fc-region of a human IgG. There are different reasons to include an IgG Fc region; a) to improve the structural and biochemical stability and increase the PK / in vivo half-life of the compound, b) to use the fusion-protein as a carrier of anti-fungal reagents, c) to mediate fungal killing by the immune system through phagocytosis and antibody-dependent cellular cytotoxicity as an additive effect to the target sequence, d) to activate the complement system via the classical pathway mediating further phagocytosis and complement mediated cytolysis, and e) to have a straightforward purification method as protein A and protein G binds strongly and selectively to IgG Fc regions.

[0359] The fusion protein can be made with different formulations where the ratio between peptides and Fc regions can be changed.

[0360] The constructs can also be made with different Fc subclasses (IgGl, IgG2, IgG3, IgG4) as the subclasses have different biological properties regarding Fc-receptor interactions and complement activation abilities.

[0361] Thus, the identification of the fungi binding amino acid sequences as disclosed herein opens many possibilities for new approaches for medical use of these sequences, such as but not limited to treatment of fungal infections. The fungi binding amino acids are useful in treatment of infections caused by fungal pathogens as exemplified in example 6.

[0362] The medical use within the present context relates to the use of alleviating symptoms of a disease condition or disorder or in the treatment of a disease, condition, or disorder.

[0363] Thus, in one or more exemplary embodiments, the present disclosure relates to a composition comprising a fungus binding amino acid sequence, a fungus binding motif, a fungus binding fusion-protein, a fungus binding peptibody and / or a peptibody of formula (I) as disclosed herein for use as a medicament, use in therapy and / or prophylaxis.

[0364] Using a Fc fusion technology, one can easily couple cytotoxic drugs and toxic radionucleotides to the constructs allowing this to be a novel antibiotic / antifungal without the need for endogenous immune assistance. This will be a significant advantage in neutropenic patients, who are usually prone to aspergillus infections.

[0365] In an embodiment, the isolated fungus binding amino acid sequence, the fusionprotein construct, the peptibody, the composition (according to the invention), is for use in the treatment of a human subject, such as a male subject or a female subject.

[0366] In one or more exemplary embodiments, the composition as described above is coupled to a radionucleotide.

[0367] In one or more exemplary embodiments, the composition as described above is coupled to a cytotoxic drug.

[0368] In one or more presently preferred exemplary embodiments, the composition described above is use in the treatment of an immunocompromised individual.

[0369] In yet an embodiment, the isolated fungus binding amino acid sequence according to the invention, the fusion-protein construct according to the invention, the peptibody according to the invention, the composition according to the invention, said immunocompromization is caused by a treatment, a condition and / or disorder selected from the group consisting of

[0370] - treatment for solid tumor and hematologic malignancies, such as treatment with chemotherapeutics or radiation;

[0371] - Solid tumor and hematologic malignancies associated with poor responses to vaccines regardless of current treatment status,

[0372] - immunosuppressive therapy;

[0373] Receipt of solid-organ transplant or an islet transplant and receiving immunosuppressive therapy;

[0374] - Receipt of chimeric antigen receptor (CAR)-T-cell therapy or hematopoietic stem cell transplant;

[0375] - an immunodeficiency, such as common variable immunodeficiency disease (CVID), severe combined immunodeficiency (SCID), DiGeorge syndrome, Bruton's agammaglobulinemia, and Wiskott-Aldrich syndrome;

[0376] HIV infection, such as an advanced or untreated HIV infection, CD4 cell counts less than 200 / mm3, history of an AIDS-defining illness without immune reconstitution, or clinical manifestations of symptomatic HIV;

[0377] - treatment with high-dose corticosteroids (i.e., 20 or more mg of prednisone or equivalent per day when administered for 2 or more weeks), alkylating agents, antimetabolites, transplant-related immunosuppressive drugs, cancer chemotherapeutic agents classified as severely immunosuppressive, tumor necrosis factor (TNF) blockers, and other biologic agents that are immunosuppressive or immunomodulatory;

[0378] - young age such as below 48 months;

[0379] - old age, such as 70 years or above, such as above 80, such as above 90;

[0380] - chronic Respiratory Diseases: Individuals with chronic respiratory conditions such as asthma, cystic fibrosis, and chronic obstructive pulmonary disease (COPD) are at higher risk. The damaged airways and mucus accumulation in these conditions provide a suitable environment for fungal growth; poorly controlled Diabetes is at higher risk: Diabetes can impair the immune system, making it more difficult for the body to fight off infections, including fungal infections. This impairment is particularly evident in cases of poorly controlled diabetes, where high blood sugar levels can hinder the function of immune cells;

[0381] - prolonged Use of Antibiotics: Long-term use of broad-spectrum antibiotics can disrupt normal microbial flora, which can increase susceptibility to fungal infections; hospitalization, Especially in Intensive Care Units: Patients in hospitals, particularly in intensive care units, are at risk due to weakened health, invasive procedures, and potential exposure to hospital-acquired pathogens; pre-existing Lung Cavities: People with pre-existing lung cavities, such as those caused by tuberculosis or sarcoidosis, are at risk as these cavities can become colonized by the fungus; and

[0382] - environmental Exposure: Occupational exposure to environments with high levels of fungal spores, such as farming, gardening, or construction sites, can increase the risk of inhaling Aspergillus spores and give rise to fungal colonization.

[0383] In an embodiment, the hematologic malignancy is selected from the group consisting of chronic lymphocytic leukemia, non-Hodgkin lymphoma, multiple myeloma, and acute leukemia. In an embodiment, the subject suffers from Allergic bronchopulmonary aspergillosis (APPA), Aspergilloma ("fungus ball"), Chronic pulmonary aspergillosis and / or invasive aspergillosis. These are the four most distinct clinical forms.

[0384] In one or more presently preferred exemplary embodiments, the composition described above is for use in the treatment of an opportunistic fungal infection.

[0385] The first formulation made is a so-called peptibody, where two peptides substitute the Fab region of a natural IgGl, which is exemplified in figure 19). This peptibody was produced recombinantly using Expi293 cells. This peptibody binds to A. fumigatus as shown in figures 20-22.

[0386] An in vivo mouse study discloses the therapeutic effect of the peptibody. C57BI / 6J B6 mice were continuously immunosuppressed cyclophosphamide to mimic the human immunocompromised situation. The mice were then intranasally infected with lxlO7A. fumigatus conidia and treated intranasally with the peptibody. The mice were divided into groups of 6 to test different dosages of peptibody; Pl) PBS, P2) 0,6 mg / kg peptibody, P3) 6 mg / kg peptibody and P4) 60 mg / kg peptibody. In the PBS group, all mice died within 8 days, whereas all mice survived the study period of 2 weeks with the highest peptibody dose (fig. 23). Moreover, there was a dose-dependent effect of the peptibody on survival (fig. 23). Accordingly, the fungal burden in the lungs was shown to be lowered / eliminated in the peptibody treated groups (fig. 24 and 25).

[0387] Medical conditions associated with fungal

[0388] In the present context, a medical condition which is associated with a potential fungal infection can be Aspergillosis, Aspergilloma, Blastomycosis, Bone Marrow Transplantation, Candida Urinary Tract Infection, Candidemia, Chromomycosis, Chronic Mucocutaneous Candidiasis, Coccidioidomycosis, Coccidioidomycosis, Meningitis, Cryptococcal Meningitis, Immunocompetent Host, Cryptococcal Meningitis, Immunosuppressed Host, Cryptococcosis, Cutaneous Fungal Infection, Dermatophytosis, Esophageal Candidiasis, Eumycetoma, Febrile Neutropenia, Fungal Infection Prevention, Fungal Infection Prophylaxis, Fungal Infection, Fungal Meningitis, Fungal Peritonitis, Fungal Pneumonia, Fusariosis, Histoplasmosis, Microsporidiosis, Mucormycosis, Ocular Fungal Infection, Onychomycosis, Oral Thrush, Paracoccidioidomycosis, Pseudoallescheriosis, Sporotrichosis, Systemic Candidiasis, Systemic Fungal Infection, Tinea Capitis. Tinea Versicolor, and Vaginal Yeast Infection.

[0389] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is pulmonary aspergillosis, cerebral aspergillosis, invasive aspergillosis, systemic aspergillosis, disseminated aspergillosis or aspergilloma.

[0390] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is aspergilloma.

[0391] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is disseminated aspergillosis.

[0392] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is systemic aspergillosis.

[0393] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is pulmonary aspergillosis.

[0394] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is cerebral aspergillosis.

[0395] In one or more presently preferred exemplary embodiments, a medical condition which is associated with a potential fungal infection is invasive aspergillosis.

[0396] Drug resistant strains

[0397] Certain strains of fungi have become more resistant to antifungal medicines. They're known as superbugs. These fungi continue to multiply and cause infections even when you take medication, such as the current three major classes of antifungal medicines: azoles, echinocandin and polyenes. The emergence of multidrug-resistant (MDR) fungal strains of Candida auris and azole-resistant Aspergillus fumigatus were highlighted in a recent CDC report: Antibiotic Resistance Threats in the United States. Thus, conventional antifungals used to treat fungal infections are no longer as effective, leading to increased mortality. Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains.

[0398] Combination treatment

[0399] The combination of two or more antifungal drugs with different modes of action in combination is recommended. The concept of combination therapy is based on the synergistic or additive potential of two or more drugs, to improve therapeutic efficacy. It also delays the development of the resistance form of the fungi.

[0400] Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with other antifungal drugs with different modes of action, such as but not limited to amphotericin B, flucytosine, and fluconazole, alone or in combination.

[0401] Polyene antifungal agent

[0402] Polyene antimycotics, sometimes referred to as polyene antibiotics, are a class of antimicrobial polyene compounds that target fungi. Amphotericin B, nystatin, and natamycin are examples of polyene antimycotics. They are a subgroup of macrolides.

[0403] Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with a polyene antifungal agent.

[0404] Azoles antifungal agent

[0405] Azoles function by disrupting ergosterol biosynthesis through inhibition of the cytochrome P-450-dependent enzyme lanosterol 14-o-demethylase. Azole antifungal agents have added greatly to the therapeutic options for treatment of systemic fungal infections. The azoles that are available for systemic use can be classified into two groups: the triazoles (fluconazole, itraconazole, voriconazole, posaconazole, and isavuconazole) and the imidazoles (ketoconazole).

[0406] Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with an azole antifungal agent.

[0407] Echinocandin antifungal agent

[0408] Echinocandins are a class of antifungal drugs that inhibit the synthesis of 0-glucan in the fungal cell wall via noncompetitive inhibition of the enzyme 1,3-0 glucan synthase. The class has been termed the "penicillin of antifungals," along with the related papulacandins, as their mechanism of action resembles that of penicillin in bacteria. 0-glucans are carbohydrate polymers that are cross-linked with other fungal cell wall components, the fungal equivalent to bacterial peptidoglycan. Caspofungin, micafungin, and anidulafungin are semisynthetic echinocandin derivatives with limited clinical use due to their solubility, antifungal spectrum, and pharmacokinetic properties. [5]. They are used to treat invasive fungal infections and show good activity against amphotericin B-resistant and fluconazole-resistant Candida guilliermondii. Caspofungin was the first drug in this class to be approved.

[0409] Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with an echinocandin antifungal agent.

[0410] Papulacandins

[0411] Papulacandins are antibiotics, particularly active against Candida albicans and several other yeasts, and was originally isolated from a strain of Papularia sphaerosperma. The fermentation, isolation, physico-chemical properties and biological activity of the five structurally related papulacandins A, B, C, D and E are reported. Papulacandin B, the main component, was assigned the formula of C47H64O17.

[0412] Thus, in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with a papulacandin antifungal agent.

[0413] Thus, in one or more presently preferred exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is particularly useful for the treatment of drug-resistant fungal strains in combination with Papulacandin B.

[0414] Delivery

[0415] Drug delivery refers to approaches, formulations, manufacturing techniques, storage systems, and technologies involved in transporting a pharmaceutical compound to its target site to achieve a desired therapeutic effect. Principles related to drug preparation, route of administration, site-specific targeting, metabolism, and toxicity are used to optimize efficacy and safety, and to improve patient convenience and compliance. Throughout history, oral administration has been regarded as the most convenient mode of drug delivery, as it requires minimal expertise and invasiveness. Although oral delivery works well for smallmolecule drugs, oral delivery of macromolecules (particularly proteins and peptides) has been limited by acidic conditions in the stomach and low permeability across the intestinal epithelium. Accordingly, the large numbers of biologic drugs that have become available may require administration by injection or infusion.

[0416] In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by nasal injection / delivery, subcutaneous injection / delivery, intravenous injection / delivery, inhalation or intratracheal injection / delivery, transdermal delivery, and / or oral delivery.

[0417] Nasal injection / delivery

[0418] In some situations, the nasal delivery route is preferred for systemic therapy because it provides an agreeable alternative to injection or pills. Substances can be assimilated extremely quickly and directly through the nose. Many pharmaceutical drugs exist as nasal sprays for systemic administration (e.g. sedative-analgesics, treatments for migraine, osteoporosis and nausea). Other applications include hormone replacement therapy, treatment of Alzheimer's disease and Parkinson's disease.

[0419] Nasal sprays are seen as a more efficient way of transporting drugs with potential use in crossing the blood-brain barrier. In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by nasal injection / delivery.

[0420] Subcutaneous injection / delivery

[0421] A subcutaneous injection is administered as a bolus into the subcutis, the layer of skin directly below the dermis and epidermis, collectively referred to as the cutis. The instruments are usually a hypodermic needle and a syringe. Subcutaneous injections are highly effective in administering medications such as insulin, morphine, diacetylmorphine and goserelin. Subcutaneous administration may be abbreviated as SC, SQ, subcu, sub-Q, SubQ, or subcut. Subcut is the preferred abbreviation to reduce the risk of misunderstanding and potential errors.

[0422] Subcutaneous tissue has few blood vessels and so drugs injected here are for slow, sustained rates of absorption, often with some amount of depot effect. Compared with other routes of administration, it is slower than intramuscular injections but still faster than intradermal injections. Subcutaneous infusion (as opposed to subcutaneous injection) is similar but involves a continuous drip from a bag and line, as opposed to injection with a syringe.

[0423] In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by subcutaneous injection / delivery.

[0424] In tra venous in jection / delivery

[0425] Intravenous therapy (abbreviated as IV therapy) is a medical technique that administers fluids, medications and nutrients directly into a person's vein. It may also be used to administer medications or other medical therapy.

[0426] The intravenous route is the fastest way to deliver medications and fluid replacement throughout the body as they are introduced directly into the circulatory system and thus quickly distributed.

[0427] Many therapies are administered as a "bolus" or one-time dose, but they may also be administered as an extended infusion or drip. The act of administering a therapy intravenously, or placing an intravenous line ("IV line") for later use, is a procedure which should only be performed by a skilled professional. In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by intravenous injection / delivery.

[0428] Inhalation or intratracheal injection

[0429] The use of the lungs for the systemic delivery of medicines is a very efficient tool. Inhaled insulin for diabetics made the way and, so far, the only available inhaled protein- based drug for systemic effect.

[0430] The high surface area and high permeability of the lungs make them an ideal site for rapid systemic delivery of macromolecules and small-molecule drugs. Small molecules are absorbed more rapidly through the lungs than through the gastrointestinal tract, with higher bioavailabilites and reduced first-pass metabolism by enzymes. The lungs are significantly permeable to many peptides and proteins, with the rate of absorption decreasing with increasing molecular mass.

[0431] In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by inhalation or intratracheal injection / delivery.

[0432] Oral delivery

[0433] Oral drug delivery is the most preferred route of drug delivery of pharmaceuticals encompassing number of diseases which have been successfully treated. Owing to its potential advantages including well-established delivery system, patient friendly, convenient, cost effective, and noninvasiveness, it has been the most favored drug delivery system in pharmaceutical field.

[0434] In one in one or more exemplary embodiments, the composition comprising a fungus binding amino acid sequence as described herein is delivered by oral delivery.

[0435] Transdermal delivery

[0436] Transdermal delivery is to apply the drugs or medicines to the skin. Transdermal delivery systems have become a successful alternative for a continuous drug delivery on demand. In the present context, transdermal delivery relates to the application of composition disclosed herein onto the epidermis.

[0437] In one or more exemplary embodiments, the composition penetrates through the epidermis and dermis and enters circulation in the body.

[0438] In one or more exemplary embodiments, nanoparticles may be used for delivery of the compositions.

[0439] Several formulations have been used for transdermal delivery and they include, but are not limited to nanoemulsions, dendrimers, lipid nanoparticles, polymeric nanoparticles carbon nanotubes and vesicular systems such as liposomes, niosomes, transfersomes, ethosomes.

[0440] Thus, in one or more exemplary embodiments, the compositions can be formulated for transdermal delivery using nanoemulsions, dendrimers, lipid nanoparticles, polymeric nanoparticles carbon nanotubes, vesicles, liposomes, niosomes, transfersomes and ethosomes.

[0441] In one or more exemplary embodiments, the compositions are formulated for transdermal delivery using nanoparticles.

[0442] In one or more exemplary embodiments, the compositions are formulated for transdermal delivery using nanoemulsions.

[0443] In one or more exemplary embodiments, the compositions are formulated for transdermal delivery using liposomes.

[0444] In one or more exemplary embodiments, the compositions are formulated for transdermal delivery using lipid nanoparticles.

[0445] In one or more exemplary embodiments, the transdermal formulations disclosed above is delivered onto the epidermis using for example a patch, drug reservoir, gel, ointment, or cream.

[0446] In one or more exemplary embodiments, the transdermal formulations disclosed above is delivered onto the epidermis using a patch.

[0447] In one or more exemplary embodiments, the transdermal formulations disclosed above is delivered onto the epidermis using a gel. In one or more exemplary embodiments, the transdermal formulations disclosed above is delivered onto the epidermis using a cream.

[0448] Dosage regimens

[0449] In one in one or more exemplary embodiments, the present disclosure relates to a dosage regime comprising delivery of a composition comprising a fungus binding amino acid sequence as described herein comprising 3 deliveries at Tl = l hour, T2=8 hours, and T3=24 hours

[0450] In an embodiment, the isolated fungus binding amino acid sequence according to the invention, the fusion-protein construct according to the invention, the peptibody according to the invention the composition according to the invention is administered to a human subject in a dosage in the range 0.06 mg / kg bodyweight to 600 mg / kg bodyweight, preferably in the range 0.1 -100 mg / kg bodyweight, more preferably 1-100 mg / kg bodyweight, an even more preferably 1-50 mg / kg bodyweight.

[0451] Diagnostic use

[0452] The identification of the fungus binding amino acid sequence opens many possibilities for new approaches to detection and thus diagnostic identification of fungal infections. The fungus binding amino acid sequence could also be used for diagnosis of invasive fungal infections. In vitro detection can be done via different reagents conjugated to the fungus binding amino acid sequence e.g. biotin-tag, fluorescent tag or enzymes directly coupled to the peptide construct. In vivo detection can be done via radiolabeling the peptide and subsequent diagnostic imaging.

[0453] The fungus binding amino acid sequence could be used as a diagnostic tool to detect whether the patient has a fungal infection using plasma, lung fluid, spinal fluid or by in vivo imaging. Various detection systems could be coupled to the peptide e.g., a biotin-tag or fluorophore. The peptide could also be coupled to an Fc region which allows the peptide binding to be detected via the Fc region, which is the backbone in many well-established test systems. An Fc region or other stabilizing components could also work as a scaffold for radiolabeling or for carrying other detection reagents such as fluorophores. The fungus binding amino acid sequence could also be used to diagnose and follow infections with bacteria (eg mycobacteria), other fungi, viruses (e.g., herpes viruses) or parasites (e.g., malaria parasites) that bind the peptide sequence.

[0454] In the present context, a diagnosis is the process of determining which disease or condition explains a subject's symptoms and signs. An integrated part of this process relates to the collection of data and information about a symptoms or signs of disease in a subject as well as the identification via the fungus binding amino acid sequence. Diagnostic use in the present context therefore relates to the use of compounds, compositions, or equipment in the collection of medical data with regards to a subject that will assist the medical practitioner in establishing a clinical picture and a diagnosis.

[0455] Thus, in one or more exemplary embodiments, the present disclosure relates to a composition comprising a fungus binding amino acid sequence according to any one of the herein described fungus binding amino acids, fungus binding motifs, the fungus binding fusion-proteins, the fungus binding peptibodies and / or peptibodies of formula (I) for use in diagnosis of a fungal infection. The fungal infection may be an opportunistic fungal infection. Opportunistic fungi are defined above.

[0456] As described above, the isolated fungus binding peptides of the disclosure are particularly useful in detection of fungal infections caused by fungi belonging to the Aspergillus genus or the Mucorales order.

[0457] In one or more exemplary embodiments, the present disclosure relates to a method for detecting a fungus in a sample comprising identifying the binding of the fungus binding amino acid sequence as disclosed herein to a fungus or fungal fragments within the sample.

[0458] In one or more exemplary embodiments, the present disclosure relates to a method for detecting a fungus in a sample, where binding of the fungus binding amino acid sequence as disclosed herein to the sample can be identified by immunodetection or by measuring a signal from a radionuclide or a signaling molecule in the sample. Detection method

[0459] In one or more exemplary embodiments, the method of detection is based on immunodetection.

[0460] In one or more exemplary embodiments the immunodetection is performed using a primary antibody targeting either the fungus binding amino acid sequence, or a molecular tag, or Fc region attached to the fungus binding amino acid.

[0461] The primary antibody can then be targeted by a secondary antibody tagged with a dye.

[0462] In one or more exemplary embodiments, the method of detection is dye staining, in particular fluorescein isothiocyanate (FITC) staining as disclosed or other fluorophores exited by a blue laser e.g. R-phycoerythrin (PE). Also, fluorophores exited by a red laser e.g. Allophycocyanin (APC) and by a violet laser e.g. Pacific Orange.

[0463] Size separation-based methods that utilizes methods such as, but not limited to centrifugation or membrane separation techniques to separate a cellular fraction from a sample that has been treated with fungus binding amino acid sequences as disclosed herein, followed by washing, resuspension and detection of signals directly emitted by the fungus binding amino acids or by immunodetection.

[0464] Sample

[0465] Fungi are found everywhere in and on the human body. Thus, all types of samples obtainable from the human body can in principle by examined for the presence of fungus. Skin, hair and nail tissue are collected for microscopy and culture (mycology) to establish or confirm the diagnosis of a fungal infection.

[0466] Body fluids, bodily fluids, or biofluids are liquids within the human body and are often easy assessable. Thus, in one or more exemplary embodiments, the sample is a body fluid.

[0467] In the present context, a specific body fluid can be selected from the group consisting of blood, saliva, mucus, sputum, serum, plasma, bile, pus, urine, semen, breast milk, transudate, rheum, cerebrospinal and interstitial fluid.

[0468] In one or more exemplary embodiments, the sample is a blood sample. In one or more exemplary embodiments, the sample is a lung fluid sample.

[0469] Bronchoalveolar lavage is a diagnostic method of the lower respiratory system in which a bronchoscope is passed through the mouth or nose into an appropriate airway in the lungs, with a measured amount of fluid introduced and then collected for examination.

[0470] In one or more exemplary embodiments, the sample is bronchoalveolar fluid sample.

[0471] In one or more exemplary embodiments, the sample is a brain fluid sample.

[0472] In one or more exemplary embodiments, the sample is a spinal fluid sample.

[0473] Methods for delivery to target physiology

[0474] Intranasal delivery of droplets comprising / containing peptibody in PBS

[0475] Identification of fungus binding

[0476] The person skilled in the art can easily identify whether an amino acid sequence can be characterised as fungus binding by e.g., making an immunoblotting and dye staining (in particular fluorescein isothiocyanate (FITC) staining), as shown in the Examples.

[0477] In general, for any given peptide, it's binding activity against a fungus can be assessed in a number of standard methods, such as, but not limited to fluorescence microscopy, immunodetection, flow cytometry or other methods based on measuring a signal from a sample. This signal can either come from a tag, label or fusion added to the peptide or from a signal from a primary or secondary antibody that is tagged or labeled.

[0478] For non-microscopy methods, the signal intensity of a sample comprising a peptide of interest can be compared to e.g. a negative control sample (a peptide known to not bind a fungus) and a positive control sample (a peptide known to bind a fungus). Thus, if a given signal from a sample is stronger than the negative control, then the peptide has fungus binding activity.

[0479] This comparison will make it possible to identify whether a given peptide is capable of binding a fungus and with what relative intensity (see e.g., example 3 and 5; figs. 9-18 and 20-22). For microscopy methods, the detection of a signal can be compared directly to the brightfield view of the sample. Thus, if signal can be detected from the sample it can be directly mapped, whether the signal originated from the fungus or not. If the shape or localization of the signal corresponds directly to the shape or localization of the fungus in the brightfield view, then the peptide has fungus binding activity (see e.g. figures 1-8). This case can be distinguished from unspecific binding, wherein binding is scattered across the sample and does not specifically co-locate with the fungus as seen in the brightfield view. Again, negative and positive controls can be utilized to further distinguish whether a peptide binds to a fungus or not by comparison.

[0480] In one or more exemplary embodiments of the present disclosure, an increase in signal intensity in a sample by at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or at least 100 % when compared to a negative control is indicative of a fungus binding peptide sequence binding to a fungus.

[0481] Thus, virtually any method that utilizes a tag, dye, or signal that enables detection of a signal in a sample can be used to detect if a given peptide has fungus binding activity.

[0482] General

[0483] It should be understood that any feature and / or aspect discussed above in connection with fungus binding amino acid sequences, fungus binding peptide motifs, fungal binding fusion proteins, or fungal binding peptibodies according to the present disclosure apply by analogy to the methods described herein.

[0484] It should be understood that any feature and / or aspect discussed above in connection with fungus binding amino acid sequences, fungus binding peptide motifs, fungal binding fusion proteins, or fungal binding peptibodies according to the present disclosure apply by analogy to each other.

[0485] The terms peptide and fungus binding amino acid sequence are used interchangeably.

[0486] The following figures and examples are provided below to illustrate the present invention. They are intended to be illustrative and are not to be construed as limiting in any way. BRIEF DESCRIPTION OF THE FIGURES

[0487] Figure 1

[0488] Fig. 1 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the resting conidia growth stage incubated together with rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0489] Figure 2

[0490] Fig. 2 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the resting conidia growth stage incubated without rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0491] Figure 3

[0492] Fig. 3 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the swollen conidia growth stage incubated together with rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0493] Figure 4

[0494] Fig. 4 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the swollen conidia growth stage incubated without rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0495] Figure 5

[0496] Fig. 5 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the germ tube growth stage incubated together with rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody. Figure 6

[0497] Fig. 6 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the germ tube growth stage incubated without rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0498] Figure 7

[0499] Fig. 7 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the hyphae growth stage incubated together with rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488- conjugated secondary antibody.

[0500] Figure 8

[0501] Fig. 8 shows both a brightfield image and a fluorescence image of Aspergillus fumigatus in the hyphae growth stage incubated without rMASP-1. A monoclonal antibody that recognizes a MASP-1 epitope was applied and binding of rMASP-1 was assessed with fluorescence microscopy using an Alexa Fluor 488-conjugated secondary antibody.

[0502] Figure 9

[0503] Fig. 9 shows lxlO7conidia / ml of heat-inactivated conidia from Aspergillus fumigatus 293 incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0504] Figure 10

[0505] Fig. 10 shows lxlO7conidia / ml of heat-inactivated conidia from Aspergillus niger incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0506] Figure 11

[0507] Fig. 11 shows lxlO7conidia / ml of heat-inactivated conidia from Aspergillus terreus incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0508] Figure 12

[0509] Fig. 12 shows lxlO7conidia / ml of heat-inactivated conidia from Aspergillus flavus incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0510] Figure 13

[0511] Fig. 13 shows lxlO7conidia / ml of heat-inactivated conidia from Lichtheimia corymbifera incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0512] Figure 14

[0513] Fig. 14 shows lxlO7conidia / ml of heat-inactivated conidia from Mucor circinelloides incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0514] Figure 15

[0515] Fig. 15 shows lxlO7conidia / ml of heat-inactivated conidia from Rhizopus arrhizus incubated with a 30 amino acid MASP-1 derived peptide tagged with a biotin tag compared to a control comprising no peptide. The biotin tag was detected with a FITC-conjugated streptavidin using flow cytometry.

[0516] Figure 16

[0517] Fig. 16 shows binding data for 5 pg / ml and 10 pg / ml solutions of a MASP-1 derived 30 amino acid peptide that was expected to bind when incubated together with Aspergillus fumigatus. The peptide was tagged with a biotin tag and detected with FITC-conjugated streptavidin using flow cytometry.

[0518] Figure 17

[0519] Fig. 17 shows binding data for 5 pg / ml and 10 pg / ml solutions of a peptide from the CUB1 domain of MASP-1 that was not expected to bind when incubated together with Aspergillus fumigatus. The peptide was tagged with a biotin tag and detected with FITC-conjugated streptavidin using flow cytometry.

[0520] Figure 18

[0521] Fig. 18 shows binding data for 5 pg / ml and 10 pg / ml solutions of a random peptide that was not expected to bind when incubated together with Aspergillus fumigatus. The peptide was tagged with a biotin tag and detected with FITC- conjugated streptavidin using flow cytometry.

[0522] Figure 19

[0523] Fig. 19 shows a generalized view of a peptibody construct. A) Fc-peptide fusionprotein (peptibody) containing two peptides and one Fc region. The peptibodies are constructed as immunoglobulins with an Fc region (blue) and a hinge region, but with the 30 amino acid MASP-1 binding sequence instead of the Fab region (orange). Overall, the Fc region has the potential to activate and direct the immune system to kill the pathogenic fungus and moreover it increases the halflife of the molecule. Peptibodies with Fc regions from e.g. other IgG subclasses (B) or other immunoglobulin classes are also relevant.

[0524] Figure 20

[0525] Fig. 20 shows data for binding of peptibody at a concentration of 5 pg / ml to different growth stages of A. fumigatus. Fluorescent microscopy shows binding of the peptibody to germ tubes (top panel) and hyphae (bottom panel). The peptibody was detected with rabbit anti-human IgG antibody and an Alexa fluor 488-coupled goat anti-rabbit antibody.

[0526] Figure 21

[0527] Fig. 21 shows data for binding of peptibody at a concentration of 10 pg / ml to different growth stages of A. fumigatus. Fluorescent microscopy shows binding of the peptibody to germ tubes (top panel) and hyphae (bottom panel). The peptibody was detected with rabbit anti-human IgG antibody and an Alexa fluor 488-coupled goat anti-rabbit antibody.

[0528] Figure 22

[0529] Fig. 22 shows binding data for different growth stages of A. fumigatus without the addition of peptibody. Fluorescent microscopy shows the A. fumigatus sample after addition of rabbit anti-human IgG antibody and an Alexa fluor 488-coupled goat anti-rabbit antibody.

[0530] Figure 23

[0531] Fig. 23 shows survival curves of the peptibody efficacy testing in a murine model of pulmonary aspergillosis. Cyclophosphamide-treated C57BI / 6J mice (n = 6) were intranasally infected with 4xl06spores of A. fumigatus per mouse. Mice were either mock-treated with PBS or treated with 0.6 mg, 6 mg, or 60 mg human Peptibody (hPep) per kilogram mouse, which was intranasally applied 1 h, 8 h, and 24 h post- infection. The survival was monitored for 14 days. Asterisks (*) indicate the level of significance for a p-value resulting from a Mantel-Cox test analyzing pairs of survival curves. Mantel-Cox test: * ... p<0.05, ** ... p<0.01, *** ... p<0.001, **** ... p< 0.0001.

[0532] Figure 24

[0533] Fig. 24 shows fungal load in lung tissue in a murine model of pulmonary aspergillosis. Cyclophosphamide-treated (n = 6) C57BI / 6J mice were intranasally infected with 4xl06spores of A. fumigatus per mouse. Mice were either mock- treated with PBS or treated with 0.6 mg, 6 mg, or 60 mg human Peptibody (hPep) per kilogram mouse, which was intranasally applied 1 h, 8 h, and 24 h postinfection. Bronchoalveolar lavage (BAL) fluid was collected from the lungs of 2 of these animals and the fungal load was quantified by striking out the resulting suspension in duplicates on Sabouraud agar plates, incubating them overnight at 37°C, and counting the resulting colony-forming units (CFUs). The data is depicted as CFU per mL homogenized tissue (mean±SD).

[0534] Figure 25

[0535] Fig. 25 shows fungal load in lung tissue in a murine model of pulmonary aspergillosis. Cyclophosphamide-treated (n = 6) C57BI / 6J mice were intranasally infected with 4xl06spores of A. fumigatus per mouse. Mice were either mock- treated with PBS or treated with 0.6 mg, 6 mg, or 60 mg human Peptibody (hPep) per kilogram mouse, which was intranasally applied 1 h, 8 h, and 24 h postinfection. Lungs of 4 of these animals were homogenized and the fungal load in these organs was quantified by striking out the resulting suspension in duplicates on Sabouraud agar plates, incubating them overnight at 37°C, and counting the resulting colony-forming units (CFUs). The data is depicted as CFU per mL homogenized tissue (mean±SD).

[0536] Figure 26

[0537] Fig. 26 shows binding of rMASP-1 wt and rMASP-1 mut to A. fumigatus. 5 pg / ml recombinant wild-type MASP-1 (444KLMAR448) or mutated rMASP-1 (444DDDDK448) were incubated with inact. A. fumigatus conidia and the binding of rMASP-1 was detected using an anti-MASP-l / -3 / MAP-l monoclonal antibody 8B3 followed by a FITC-conjugated secondary antibody. Binding was measured using flow cytometry.

[0538] Figure 27

[0539] Fig. 27 shows peptibody binding to live and inactivated A. fumigatus. Live A. fumigatus conidia or heat-inactivated conidia were incubated with 5 pg / ml peptibody and the binding was detected with an anti-human IgG polyclonal antibody and measured using flow cytometry.

[0540] Figure 28

[0541] Fig. 28 shows binding of mutated active and zymogen rMASP-1 to A. fumigatus. Mutated zymogen and activated rMASP-1 (444DDDDK448) and wild-type rMASP-1 (444KLMAR448) were incubated with inactivated A. fumigatus conidia and the binding of rMASP-1 was detected using an anti-MASP-l / -3 / MAP-l monoclonal antibody 8B3 followed by a FITC-conjugated secondary antibody. Binding was measured using flow cytometry.

[0542] Figure 29

[0543] Fig. 29 shows the binding of modified MASP-1 peptides to A. fumigatus as measured in terms of mean fluorescence intensity (MFI). Substituting a cysteine with a serine at amino acid position 7 (C7S) from the N-terminal does not affect binding. Substitution at position 11 (CHS) or 7+11 (C7S + C11S) decreases the binding.

[0544] Figure 30

[0545] Fig. 30 shows the ability of the MASP-1 peptide to bind A. fumigatus conidia with or without calcium. Peptide binding to A. fumigatus was detected using a biotin- tagged peptide and a FITC-coupled streptavidin. The MASP-1 peptide binds A. fumigatus both in the presence and absence of calcium. Figure 31

[0546] Fig. 31 shows binding of the MASP-1 peptide and the corresponding MASP-3 peptide. The 30 aa MASP-3 peptide located at the same position as the MASP-1 peptide and having partly sequence overlap does not bind A. fumigatus as the MASP-1 peptide.

[0547] Figure 32

[0548] Fig. 32 shows binding of the MASP-1 and MASP-3 peptides to human cell lines. Neither of the 30 aa MASP peptides bind to the THP-1 monocytes (figure 32 B) or the HMEC-1 endothelial cells (figure 32 A).

[0549] Figures 33-40

[0550] Figs. 33-40 shows peptibody binding to the different A. fumigatus growth stages using the same microscope imaging parameters for all growth stages (with peptibody in figures 33-36 vs no peptibody in figures 37-40). The peptibody binds stronger to germ tubes (figure 35) and hyphae (figure 36) than conidia (figures 33-34). Moreover, the Peptibody preferably binds to the hyphal tip, which is the growth zone of the fungus (figure 35-36). No binding was detected in samples that were not treated with peptibody (negative control; figures 37-40).

[0551] Figure 41

[0552] Fig. 41 shows the ability of several peptide sequences of the MASP-1 CCP2 domain to bind A. fumigatus conidia in comparison d to the MASP-1 binding peptide (SEQ ID NO: 1). None of the CCP2 domain peptide sequences (CCP2 Ctrl.Pepl-3) demonstrated fungus binding capabilities as measured in terms of median fluorescence intensity (MFI).

[0553] Figure 42

[0554] Figure 42 shows a western blot analysis of the peptibody before (A) and after (B) sequence optimization. The peptibody molecular weight is expected to be around 60 kDa, but the non-optimized peptibody also assembles in higher molecular weight complexes, which is undesired. The optimized peptibody does not form complexes due to the removal of an unpaired cysteine in the hinge region. Figure 43

[0555] Figure 43 shows survival curves of the peptibody efficacy testing in a murine model of pulmonary aspergillosis. Cyclophosphamide-treated C57BI / 6J mice (n=9) were intranasally infected with 6xl06spores of A. fumigatus per mouse. Mice were either mock-treated with PBS or treated with 10 mg or 30 mg human Peptibody (hPep) per kilogram mouse, which was intranasally applied 1 h, 8 h, and 24 h post- infection. At the same time points PBS or 30 mg hPep per kilogram mouse was applied to uninfected (uninf) cyclophosphamide-treated (n=9) C57BI / 6J mice. The survival was monitored for 11 days. Asterisks (*) indicate the level of significance for a p-value resulting from a Mantel-Cox test analyzing pairs of survival curves.

[0556] Figure 44

[0557] Figure 44 shows body weight as a clinical parameter for inflammation in a murine model of pulmonary aspergillosis. Cyclophosphamide-treated (n=9) C57BI / 6J mice were intranasally infected with 6xl06spores of A. fumigatus per mouse. Mice were either mock-treated with PBS or treated with 10 mg or 30 mg human Peptibody (hPep) per kilogram mouse, which was intranasally applied 1 h, 8 h, and 24 h post- infection. At the same time points PBS or 30 mg hPep per kilogram mouse was applied to uninfected (uninf) cyclophosphamide-treated (n=9) C57BI / 6J mice. Body weight was measured twice a day and is shown here as a timeline (A) and for day 4 (B) after the infection. The data is depicted in percentage and the day of infection was set to 100 % (mean±SD). After the oneway ANOVA or the Kruskal-Wallis test showed significance, a Tukey or a Dunn post-hoc test respectively was conducted to compare pair of groups. Asterisks (*) indicate the degree of significance for an adjusted p-value.

[0558] Figure 45

[0559] Domain structure of Human MASP-1, including cleavage site and location of 30 amino acid peptide (SEQ ID NO: 1).

[0560] Figure 46

[0561] Figure 46 shows binding of the peptibody to heat-inactivated conidia from a range of Aspergillus fungi. A) Aspergillus flavus; B) Aspergillus terreus; C) Aspergillus niger. lxlO7conidia / ml were incubated with 2.5, 5 and 10 pg / ml peptibody and 2, 4 and 8 pg / ml Fc fragment as a negative control. The peptibody was detected with a rabbit anti-human IgG antibody and a FITC-coupled goat anti-rabbit antibody using flow cytometry. The data represents the mean of 3 repeats + / - SD.

[0562] Figure 47

[0563] Figure 47 shows binding of the peptibody to heat-inactivated conidia from Mucormycetes. A) Mucor circinelloides; B) Rhizopus arrhizus. lxlO7conidia / ml were incubated with 2.5, 5 and 10 pg / ml peptibody and 2, 4 and 8 pg / ml Fc fragment as a negative control. The peptibody was detected with a rabbit antihuman IgG antibody and a FITC-coupled goat anti-rabbit antibody using flow cytometry.

[0564] Figure 48

[0565] Figure 48 shows binding of Clq to the peptibody. ELISA plates were coated with 2 pg / ml peptibody before adding a titration of purified Clq. Clq was detected with a biotin-conjugated rabbit a-Clq polyclonal antibody followed by HRP-coupled streptavidin. Coated Fc fragments and BSA were applied as positive and negative controls, respectively. The data represents the mean of 3 repeats + / - SD.

[0566] Figure 49

[0567] Figure 49 shows activation of the complement system through the peptibody. There is no complement activation on the A. fumigatus conidia in 2% umbilical cord serum with an MBL inhibitor. However, when the peptibody is present on the conidia, complement activation products C4b, C3b and TCC are deposited in a Clq-dependent manner. The data represents the mean of 3 repeats + / - SD.

[0568] Figure 50

[0569] Figure 50 shows the phagocytosis of A. fumigatus conidia. There is no phagocytosis of non-opsonized conidia, whereas opsonization with the peptibody mediated the uptake of conidia by neutrophils. Peptibody-mediated complement activation on the A. fumigatus conidia in 2% umbilical cord serum with an MBL inhibitor also facilitated phagocytosis, which was only partly inhibited by a C3 inhibitor. Hence, phagocytosis was mediated through a combination of direct peptibody opsonization and indirectly through C3b deposition. The data represents the mean of 3 repeats.

[0570] EXAMPLES Example 1 - MASP-1 binding

[0571] MASP-1, a serine protease from the complement system, binds directly to various pathogenic fungi, for example, Aspergillus fumigatus and this knowledge can be utilized to design a targeted antifungal compound. MASP-1 binding was tested on the different growth stages of A. fumigatus. First, the A. fumigatus conidia were incubated on microscopy glass slides for 0, 4, 8 and 16 hours to obtain resting conidia, swollen conidia, germ tubes and hyphae. Recombinant MASP-1 was then added in a concentration of 5 pg / ml and binding was detected with a pan anti- MASP-1 / -3 / MAP-1 monoclonal antibody 8B3 and Alexa fluor 488-coupled goat anti-mouse antibody. Using fluorescence microscopy, recombinant MASP-1 binding was detected for all growth stages (fig. 1-8).

[0572] Example 2 - binding of wt vs. a mutated recombinant MASP-1 to opportunistic fungi

[0573] Background

[0574] Opportunistic fungi cause severe disease in immunocompromised patients and treatment can be challenging as mortality rates are high and treatment is further complicated by drug-drug interactions, adverse effects and resistance development. Therefore, it is important to create new targeted drugs against fungal infections. MASP-1, a serine protease from the complement system, binds directly to various pathogenic fungi e.g., it binds to all growth stages (resting and swollen conidia, germ tubes and hyphae) of Aspergillus fumigatus. The binding motif could be utilized in targeted treatment, however, the MASP-1 binding motif is not known.

[0575] Methods

[0576] A. fumigatus conidia were heat-inactivated (15 min 121 °C) to eliminate contamination in the laboratory. lxlO7conidia / ml were incubated with 5 pg / ml recombinant wild-type MASP-1 (444KLMAR448) or mutated rMASP-1 (444DDDDK448). The binding of rMASP-1 was detected using 10 pg / ml of a pan anti-MASP-1 / - 3 / MAP-l monoclonal antibody 8B3 followed by a FITC-conjugated goat anti-mouse polyclonal antibody. Binding was measured as the mean fluorescence intensity using flow cytometry (Beckman Coulter, Gallios). Resu / ts / conc / usion

[0577] The study showed that wild-type rMASP-1 (444KLMAR448) (SEQ ID NO: 2) binds to A. fumigatus conidia, whereas the mutated rMASP-1 (444DDDDK448) (SEQ ID NO: 4) did not bind. Hence, the amino acid sequence KLMAR is necessary for the binding of MASP-1 to A. fumigatus as shown in figure 26.

[0578] Example 2A - zymogen vs. activated rMASP-1 (444DDDDK448) binding to A. fumigatus

[0579] Background

[0580] The activated version of the mutated rMASP-1 (444DDDDK448) (SEQ ID NO: 4) does not bind to A. fumigatus. In the same experiment, it was also tested whether a zymogen version of the mutated rMASP-1 bound.

[0581] Methods

[0582] The experiment in this example 2A was performed using the method as described in example 2.

[0583] Results / conclusion

[0584] Neither the active nor zymogen mutated rMASP-1 (444DDDDK448) (SEQ ID NO: 4) bound to A. fumigatus conidia (Figure 28). Hence, this confirms that the amino acid sequence KLMAR is crucial for binding MASP-1 to A. fumigatus.

[0585] Example 3 - a 30 amino acid MASP-1 peptide binds to opportunistic fungi

[0586] Background

[0587] The MASP-1 amino acid sequence444KLMAR448(SEQ ID NO: 2) is flanked by amino acids on both the N-terminal and C-terminal side of the protein and possibly an amino acid sequence specifically covering the KLMAR is capable of binding to opportunistic fungi (see also figure 45).

[0588] Methods

[0589] A 30 amino acid peptide (SEQ ID NO: 1) comprising the KLMAR sequence was generated with an N-terminal biotin-tag. Besides, two control peptides were made; one with a random 30 amino acid sequence (SEQ ID NO: 16) and one corresponding to a sequence from the MASP-1 CUB1 domain that does not contain the KLMAR sequence (SEQ ID NO: 17). 5 pg / ml of the N-term biotin-tagged peptides were incubated with lxlO7heat-inactivated conidia / ml from different Aspergillus species and from fungi belonging to the Mucorales order. The peptide binding was subsequently detected with a FITC-coupled streptavidin using flow cytometry (Beckman Coulter, Gallios).

[0590] The KLMAR-comprising 30 AA MASP-1 peptide (SEQ ID NO: 1) successfully bound to the different Aspergillus (A. fumigatus, A. niger, A. terreus and A. flavus) and Mucorales fungi Lichtemia corymbifera, Mucor circinelloides and Rhizopus arrhizus (figs. 9-16) and the binding was specific for this peptide as A. fumigatus did not bind the random 30 amino peptide or the 30 amino acid MASP-1 CUB1 domain peptide (figs. 17-18).

[0591] Example 3A - binding comparisons

[0592] Background

[0593] The 30 aa MASP-1 peptide binding to A. fumigatus is partly covering the CCP2 domain of MASP-1. Hence, it was tested whether other 30 aa peptides from the same CCP2 domain of MASP-1 would also bind to A. fumigatus.

[0594] Methods

[0595] Three 30 amino acid peptides from the CCP2 domain of MASP-1 were generated with an N-terminal biotin-tag.

[0596] CCP2 Ctrl.Pepl: VDCRAPGELEHGLITFSTRNNLTTYKSEIK (SEQ ID NO: 5), CCP2 Ctrl.Pep2: STRNNLTTYKSEIKYSCQEPYYKMLNNNTG (SEQ ID NO: 6), CCP2 Ctrl.Pep3: GELEHGLITFSTRNNLTTYKSEIKYSCQEP (SEQ ID NO: 7).

[0597] 5 and 10 pg / ml of the N-term biotin-tagged peptides were incubated with lxlO7heat-inactivated A. fumigatus conidia / ml and the peptide binding was detected with a FITC-coupled streptavidin using flow cytometry (BD, Celesta).

[0598] The three CCP2 peptides named CCP2 Ctrl. Pepl-3 did not bind to A. fumigatus conidia (figure 41). The KLMAR-comprising MASP-1 peptide (SEQ ID NO: 1) bound A. fumigatus as previously shown. Hence, the binding is specific to this peptide and does not involve the CCP2 domain in general.

[0599] Example 3B - Binding of modified MASP-1 peptides

[0600] Background

[0601] The 30 amino acid MASP-1 peptide (SEQ ID NO: 1) has two cysteines at positions 7 and 11 from the N-terminal. Cysteines have the potential to form disulfide bridges, which affect the tertiary structure of the protein. The question is whether the MASP-1 peptide binding depends on a disulfide bridge between the two cysteines.

[0602] Methods

[0603] 5 pg / ml of the N-term biotin-tagged peptides with C - S mutations at position 7 (C7S), position 11 (CHS) and both positions (C7S + C11S) were incubated with lxlO7heat-inactivated A. fumigatus conidia / ml. The peptide binding was subsequently detected with a FITC-coupled streptavidin using flow cytometry (BD, Celesta).

[0604] Results

[0605] The peptide C7S binds in the same manner as the wild-type peptide. Hence, the MASP-1 peptide does not seem to be dependent on a tertiary structure mediated by the cysteines as a mutation in only one of the cysteines should hinder disulfide bond formation. To our surprise, the cysteines still seem important for the binding as CHS and C7S + CHS bind less to the fungus (Figure 29). The cysteines are located outside the444KLMAR448sequence and thus have an unforeseen impact on the binding.

[0606] Conclusion

[0607] These data indicate that CHS and C7S are important for fungus binding.

[0608] Example 3C - Binding of the MASP-1 peptide with or without calcium

[0609] Background The binding of MASP-1 to other proteins like collectins and ficolins is calciumdependent. Also, the dimerization of MASPs is calcium-dependent. It is unknown whether the MASP-1 peptide binding to A. fumigatus depends on calcium.

[0610] Methods

[0611] 5 pg / ml of the N-term biotin-tagged MASP-1 peptide were incubated with lxlO7heat-inactivated A. fumigatus conidia / ml in barbital buffer (containing Ca2+) with or without 20 mM EDTA that chelates the calcium. The peptide binding was subsequently detected with a FITC-coupled streptavidin using flow cytometry (Beckman Coulter, Gallios).

[0612] The MASP-1 peptide bound equally well to A. fumigatus with and without 20 mM EDTA. Hence, unlike the interaction with its binding partners from the immune system, MASP-1 does not require calcium for interacting with A. fumigatus (figure 30).

[0613] Example 3D - Corresponding MASP-3 peptide does not bind A. fumigatus

[0614] Background

[0615] MASP-1 and MASP-3 are splice variants of the common MASP1 gene and they both bind to A. fumigatus. The two serine proteases have the exact same heavy chain. Hence, it is expected that the two proteins bind to fungi with the same residues. In fact, it would seem unlikely if two nearly identical proteins binding to the same microbe do not share binding domains.

[0616] Methods

[0617] The binding of the MASP-1 peptide was compared to the binding of a MASP-3 peptide covering the same protein region and thus having partly overlapping amino acid sequences. MASP-1 peptide: GRSLPTCLPVCGLPKFSRKLMARIFNGRPA (SEQ ID NO: 1). MASP-3 peptide: GRSLPTCLPECGQPSRSLPSLVKRIIGGRN (SEQ ID NO: 3). 5 pg / ml of the N-term biotin-tagged peptides were incubated with lxlO7heat-inactivated A. fumigatus conidia / ml. The peptide binding was subsequently detected with a FITC-coupled streptavidin using flow cytometry (Beckman Coulter, Gallios). Results

[0618] The MASP-3 peptide did not bind to A. fumigatus as the MASP-1 peptide (figure 31). So, despite the high degree of similarity between the two proteases, they have developed unequal ways of binding to fungi and the binding sequence thus is highly unpredictable.

[0619] Conclusion

[0620] In sum, these results demonstrate that the A. fumigatus binding is unique for the MASP-1 derived peptide, since the corresponding peptide from MASP-3 did not bind to A. fumigatus.

[0621] Example 3E - The MASP-1 peptide does not bind to human cells

[0622] Background

[0623] The 30 aa MASP-1 peptide (SEQ ID NO: 1) binds to various fungi, which makes it suitable for targeted treatment. It is, however, important that targeted antifungal treatment is specifically targeted towards fungi and not to human cells, as this increases the risk of unwanted drug effects. Therefore, it is convenient to test the binding of the MASP-1 peptide to human cells.

[0624] Methods

[0625] 5 pg / ml of the N-term biotin-tagged MASP-1 peptide was incubated with THP-1 and HMEC-1 cells (lxlO7cells / ml). The peptide binding was subsequently detected with a FITC-coupled streptavidin using flow cytometry (Beckman Coulter, Gallios).

[0626] Results

[0627] The monocytic cell line THP-1 (figure 32 B) and the endothelial cell line HMEC-1 (figure 32 A) did not show any binding to the MASP-1 peptide.

[0628] Conclusion

[0629] These results demonstrate, although binding to other cell types or primary cells cannot be ruled out, the peptide does not generally bind to human cells. Example 4 - Peptibody binding to A. fumigatus conidia

[0630] A peptide-coupled functional component, e.g., an immunoglobulin Fc region, can work therapeutically against fungal infections by recruiting the immune apparatus of the patient, thereby facilitating clearance of the pathogen. An Fc region enables fungal killing via phagocytosis, antibody-dependent cellular cytotoxicity and by activating the complement system mediating further opsonization and phagocytosis as well as anaphylatoxin release and recruitment of inflammatory cells and complement-mediated cytolysis. Gathering multiple peptides with Fc regions also enables agglutination, which can block the fungi from accessing the epithelium and contribute to fungal clearance by making the phagocytosis more efficient. A peptide-coupled component can also work as a carrier of other therapeutic compounds such as antifungal agents e.g., azoles or as a carrier of radionuclides e.g.,213Bi to destroy the invading fungi via local radiation.

[0631] A peptide-coupled component, e.g., an Fc region, can improve the pharmacokinetics and increase the in vivo half-life of the compound. An Fc region can also be manipulated to further increase the effectiveness and further prolong the half-life of the construct. An Fc region can for example be designed to be recycled in the endocytic pathway in host cells in order to increase the half-life of the construct. The compound can also be pegylated to extent the half-life and bioavailability.

[0632] The Fc region can be manipulated to change the mode of interaction with components from the immune system to increase / decrease the inflammatory process by e.g., changing the interaction with Clq from the complement system.

[0633] An Fc region can benefit the production of the construct by improving the structural and biochemical stability and by easing the purification process.

[0634] Methods

[0635] The first formulation made with the peptide is a fusion protein containing the 30 AA peptide (SEQ ID NO: 1), a hinge region and the Fc-region of a human IgGl. This so-called Peptibody (SEQ ID NO: 8) mimics the structure, function and biological half-life of natural immunoglobulins, but has the unique peptide binding region instead of the antigen-binding Fab fragment. An example of the structure of a peptibody construct is shown in fig. 19. The Peptibody was produced recombinantly using Expi293 cells and purified using protein G Sepharose. Binding of the Peptibody to A. fumigatus conidia was tested by incubating 5 pg / ml of purified Peptibody with lxlO7heat-inactivated or live conidia / ml and the binding was detected with a rabbit anti-human IgG antibody and a FITC-coupled goat anti-rabbit antibody using flow cytometry.

[0636] Results

[0637] As shown in figure 27, the Peptibody (SEQ ID NO: 8) binds to both the heat- inactivated and live A. fumigatus conidia.

[0638] Conclusion

[0639] These data indicate that the peptides of the invention are also functional in a peptibody setting.

[0640] Example 5 - Peptibody binding to A. fumigatus germ tubes and hyphae

[0641] Background

[0642] A. fumigatus is a filamentous fungus, which means that the conidia grow into elongated structures called germ tubes and hyphae. The fungus expands in its natural environment by growing hyphae structures and these growth stages are also found in infected patients. Thus, binding to these growth stages is likely an important feature of an antifungal drug in case the fungus has already developed in the patient prior to diagnosis and treatment.

[0643] Methods

[0644] Binding of the Peptibody (SEQ ID NO: 8) to germ tubes and hyphae was tested using fluorescence microscopy. First, the A. fumigatus conidia was incubated on microscopy glass slides for 6 and 18 hours in RPMI media to develop germ tubes and hyphae. Afterward, the Peptibody was added in a concentration of 5 or 10 pg / ml and binding was detected with rabbit anti-human IgG antibody and an Alexa fluor 488-coupled goat anti-rabbit antibody. Images were acquired with BioTek LIONHEART FX Automated Microscope using Gen5 software.

[0645] Results The Peptibody binding to A. fumigatus was visualized with fluorescence microscopy. A fluorescent signal appeared on A. fumigatus germ tubes and hyphae when the fungus was incubated with both 5 and 10 pg / ml Peptibody and the detection antibodies (figs. 20-21) and it did not appear when the Peptibody was omitted (fig. 22).

[0646] Conclusion

[0647] These data demonstrate that the Peptibody binds to A. fumigatus germ tubes and hyphae.

[0648] Example 5A - Variation of Peptibody binding to the different growth stages

[0649] Background

[0650] The cell wall of A. fumigatus changes composition during growth. Resting conidia have a resilient rodlet layer, which is lost when the conidia get swollen and start elongating into germ tubes. Hence, the binding pattern of the Peptibody may change during growth.

[0651] Methods

[0652] Comparison of the Peptibody (SEQ ID NO: 8) binding to conidia, germ tubes and hyphae was made with fluorescence microscopy. A. fumigatus conidia were incubated on microscopy glass slides for 0, 2 6 and 17 hours in RPMI media to develop resting conidia, swollen conidia, germ tubes and hyphae, respectively. Afterward, the Peptibody was added in a concentration of 10 pg / ml and binding was detected with a rabbit anti-human IgG antibody and an Alexa fluor 488- coupled goat anti-rabbit antibody. Images were acquired with BioTek LIONHEART FX Automated Microscope using Gen5 software and imaging parameters were the same for all growth stages.

[0653] Results

[0654] It seems that the binding capacity of the Peptibody to the different A. fumigatus growth stages vary. As illustrated in fig. 33-40, the Peptibody binds stronger to germ tubes (fig. 35) and hyphae (fig. 36) than conidia. Moreover, the Peptibody preferably binds to the hyphal tip (fig. 35-36), which is the growth zone of the fungus.

[0655] Conclusion

[0656] The peptibody binding to A. fumigatus germ tubes and hyphae is superior to conidia binding. This was surprising as the initial discovery was made on inactivated resting conidia and not on live growing fungi. The hyphal tip is likely a desired target for a therapeutic drug. Hence this Peptibody binding feature could be important for its therapeutic function.

[0657] Example 6 - Therapeutic effect of the Peptibody in a pre-clinical model

[0658] The Peptibody (SEQ ID NO: 8) binds to A. fumigatus conidia, germ tubes and hyphae in vitro, however the in vivo effect of the Peptibody has to be tested in a pre-clinical animal model.

[0659] Methods

[0660] The Peptibody (SEQ ID NO: 8) was tested in a pre-clinical mouse model of an invasive A. fumigatus infection. C57BI / 6J B6 mice were immunosuppressed with 150 mg / kg cyclophosphamide to mimic the immunocompromised human situation. The mice were then intranasally infected with 4xl06A. fumigatus conidia (clinical strain A22) and treated intranasally with the Peptibody at different timepoints post- infection (see below). Nasal injections were performed under mild anesthesia with isoflurane. The mice were divided into groups of 6 mice to test different dosages of Peptibody (see below). The study period was 14 days, and the survival was monitored each day.

[0661] Cyclophosphamide and A. fumigatus injection schedule

[0662] Day -11: 150 mg / kg cyclophosphamide

[0663] Day -8: 150 mg / kg cyclophosphamide

[0664] Every 3rdday: 100 mg / kg cyclophosphamide

[0665] Day 0: Intranasal A. fumigatus injection Day 14: End of experiment

[0666] Treatment timepoints

[0667] 1 hour post-infection

[0668] 8 hours post-infection

[0669] 24 hours post-infection

[0670] Treatment groups

[0671] Pl) PBS

[0672] P2) 0.6 mg / kg

[0673] P3) 6 mg / kg

[0674] P4) 60 mg / kg

[0675] Results

[0676] The survival curves obtained from this animal model showed a clear dosedependent amelioration of the disease outcome when treated with the peptibody (23). While all animals in the control group (mock-treated with PBS) had a lethal outcome of pulmonary aspergillosis within eight days, even the lowest peptibody dosage (0.6 mg / kg) was already able to rescue 33.3 % of the mice challenged with A. fumigatus. The survival further improved when using 6 mg per kg mouse and 60 mg per kg mouse to 83.3 % and 100 % respectively.

[0677] Conclusion

[0678] Overall, this suggests that there is a therapeutic effect of the Peptibody against a pulmonary A. fumigatus infection. These results are shown in figure 23.

[0679] Example 7 - Peptibody effect on the fungal burden in a pre-clinical model

[0680] Background

[0681] In the pre-clinical model described above, the A. fumigatus conidia were given intranasally, which means that the fungal infection starts in the lungs. It is therefore of interest to test whether the Peptibody relieves the fungal burden, meaning whether the Peptibody mediates killing of the fungi in the pulmonary cavities. Methods

[0682] Post-death (either due to the infection or due to the end of the study period), the lungs of four mice were flushed with PBS to collect the bronchoalveolar lavage fluid (BALF) and the lungs of 2 mice were homogenized The BALF and the homogenized tissue were plated on Sabouraud Dextrose Agar in order to count the number of colony-forming units (CFU).

[0683] Results

[0684] In both tests, the CFU count and thus the fungal burden, was eliminated in the P3 group (6 mg / kg) and the P4 (60 mg / kg) group as shown in fig. 24-25. In these experiments, there were also a dose-dependent effect of the Peptibody since detectable CFUs was observed in the P2 group (0.6 mg / kg) and even more in the Pl (PBS) group.

[0685] Conclusion

[0686] These data suggest that the increased chance of survival mediated by the Peptibody is due to elimination of living fungi in the lungs.

[0687] Example 8 - Optimization of the peptibody

[0688] Background

[0689] It is important for the production and the mechanism of action of the peptibody that the formulation of the peptibody is optimized to give less aggregation / complex formation and higher yields.

[0690] Materials and methods

[0691] To avoid aggregation / complex formation a peptibody was designed which was free of an unpaired cysteine in the hinge region of the peptibody, giving rise to a peptibody according to SEQ ID NO: 9. The unpaired cysteine can cause unwanted aggregates / complexes of peptibody proteins by cysteine pairing between individual proteins. The peptibodies from SEQ ID NO: 8 and SEQ ID NO: 9 (both produced in Expi293 cells) were evaluated in western blots to check for aggregates / complexes. 0.1 pg protein was run on NuPAGE 4 -12% Bis-Tris polyacrylamide gels (Invitrogen) and blotted onto nitrocellulose membranes. Detection of peptibody was done using Rabbit a-human IgG HRP (P0214). Results

[0692] By comparing the western blot of the non-optimized peptibody (SEQ ID NO: 8) and the optimized (SEQ ID NO: 9) in figure 42 is clear that the optimization leads to a more uniform protein production resulting in a protein with a molecular weight around 60 kDa corresponding to the expected peptibody molecular weight. The non-optimized peptibody has higher molecular weight bands corresponding to protein aggregates / complexes. The production of the optimized peptibody gave high protein yield (~ 100 mg / L), high purity (>99%) after purification and low endotoxin levels (<0.38 EU / mg).

[0693] Conclusion

[0694] Optimizing the peptibody sequence to not contain an unpaired cysteine in the hinge region resulted in a uniform protein expression free of unwanted aggregates / complexes. This also resulted in a more high-yield peptibody production.

[0695] Example 9 - Therapeutic effect of the optimized Peptibody in a pre-clinical model

[0696] The therapeutic effect of the peptibody was previously assessed in a pre-clinical mouse model using the non-optimized peptibody (example 6). Hence, it is important to evaluate the effect of the optimized peptibody (SEQ ID NO: 9) in the same animal model. Again, the mice were immunosuppressed to mimic the human situation.

[0697] Materials and methods

[0698] The mouse model was performed similar to the model in example 6. Mice were immunocompromised using cyclophosphamide (100 mg / kg every third day) to mimic this impaired state of the immune system. Blood analyses showed that the immune suppression with this application regime is successful and provided a good simulation of immunosuppressed patients. The mice were then intranasally infected with 6xl06A. fumigatus conidia (clinical strain A22) and treated intranasally with the Peptibody at different timepoints post- infection (see below). Nasal injections were performed under mild anesthesia with isoflurane. The mice were divided in groups of 9 mice to test different dosages of Peptibody (see below). The study period was 11 days, and the survival was monitored each day as well as the body weight.

[0699] Day -11: 150 mg / kg cyclophosphamide

[0700] Day -8: 150 mg / kg cyclophosphamide

[0701] Every 3rdday: 100 mg / kg cyclophosphamide

[0702] Day 0: Intranasal A. fumigatus injection

[0703] Day 11: End of experiment

[0704] Treatment timepoints

[0705] 1 hour post-infection

[0706] 8 hours post-infection

[0707] 24 hours post-infection

[0708] Treatment groups

[0709] PBS

[0710] 10 mg / kg peptibody

[0711] 30 mg / kg peptibody

[0712] Results

[0713] Survival data:

[0714] The survival curves obtained from this study using the optimized peptibody showed the same significant difference in treated versus mock-treated groups challenged with Aspergillus fumigatus conidia as the first animal model using the non-optimized peptibody. When mock-treating the infected animals with PBS, 88.8 % of the mice met humane end points. This high lethality was significantly reduced with the application of both peptibody dosages to either 33.3 % if treated with 10 mg per kg or 22.2 % when treated with 30 mg per kg. Between those two selected peptibody dosages no significant difference in survival was observed, which means that both dosages are most likely within the desired therapeutic window (Figure 43). Moreover, the mice were unaffected by the peptibody in itself as seen on the curve representing uninfected mice treated with 30 mg / kg peptibody.

[0715] Besides the survival, the body weight, which can be used as a good clinical indicator of inflammatory reaction due to pro-inflammatory cytokines resulting in a lack of aDoetite, was evaluated.

[0716] The body weight of all animals, whether infected or not, decreased on average at least by 5 % compared to the start of the experiment (figure 44A-B). The uninfected animals lost the least amount of body weight, which can be expected. Looking at the infected animals, initially all groups lose weight drastically due to the infection and resulting inflammation. However, the mock-treated animals lose more weight from d3 post-infection onwards compared to the treated animals, independently of the applied dosage, showing the positive effect of this antifungal drug on the course of infection.

[0717] Conclusion

[0718] The peptibody can efficiently treat immunocompromised mice intranasally infected with Aspergillus fumigatus conidia.

[0719] Besides the survival, the body weight, which can be used as a good clinical indicator of inflammatory reaction due to pro-inflammatory cytokines resulting in a lack of appetite, was also positively affected by the application of the peptibody tested. The further the animal experiment progressed, the more body weight was lost by the control group, while the body weight in the peptibody-treated groups positively correlated with the dosage of the peptibody.

[0720] Common conclusion on both mice experiments

[0721] In conclusion, both animal experiments (example 6+7 and 9) show the impressive potential of this new antifungal drug as a salvage therapy for pulmonary aspergillosis. Considering the state-of-the-art therapy with daily administered liposomal Amphotericin B therapy leads to a 14-day-mortality of at least 40 % in neutropenic patients, which are mainly affected by mycoses like aspergillosis. Treatment with only three doses of the peptibody already showed to be superior leading to a better outcome in both animal models, although the animals were not only neutropenic but pancytopenic leaving them even more prone to a fatal outcome of the disease.

[0722] Furthermore, unlike other antifungal therapies, the peptibody utilizes an innate immunity target. This is usually a highly conserved molecule on the pathogen responsible for virulence or viability of the fungus, which makes it highly unlikely that any resistances will appear over time.

[0723] In addition, when comparing the efficiency of the mice data presented in example 6+7 with the mice data presented in example 9, it can be seen that it appears as if the optimized peptibody (SEQ ID NO: 9) is more efficient than the first tested peptibody (SEQ ID NO: 8). Without being bound by theory, the removal of the unpaired cysteine in the hinge region of the optimized peptibody appears to be an important feature.

[0724] Example 10 - Binding to different fungi

[0725] Background

[0726] The 30 amino acid MASP-1 sequence was shown to bind various opportunistic fungi (example 3), which led to the development of MASP-1 derived peptibody which was shown to successfully bind A. fumigatus. To confirm binding to a broad range of opportunistic fungi further tests were performed.

[0727] Materials and methods

[0728] The Peptibody was produced recombinantly using a stable CHO cell line and purified using protein G Sepharose. Binding of the Peptibody was tested on heat- inactivated conidia of the following fungi: A. fumigatus strain 6871, A. fumigatus strain A22, A. fumigatus strain A293, A. fumigatus strain BE8 (environmental strain), A. flavus strain FL15, A. flavus strain SP1598, A. terreus strain T44, A. terreus resistant to Ampotericin B, A. niger N32, Mucor circinelloides strain MAL- D3, Rhizopus arrhizus strain RA. The conidia were tested by incubating 2.5, 5 and 10 pg / ml of purified Peptibody with lxlO7heat-inactivated conidia / ml. Fc fragments were included as a negative control and added in a concentration of 2, 4 and 8 pg / ml. The binding was detected with a rabbit anti-human IgG antibody and a FITC-coupled goat anti-rabbit antibody using flow cytometry.

[0729] Results

[0730] The peptibody (SEQ ID NO: 8) successfully bound to the tested Aspergillus species / strains (A. fumigatus strain 6871, A. fumigatus strain A22, A. fumigatus strain A293, A. fumigatus strain BE8 (environmental strain), A. flavus strain FL15,

[0731] A. flavus strain SP1598, A. terreus strain T44, A. terreus resistant to Ampotericin

[0732] B, A. niger N32) (figure. 46A-C) and to the tested Mucormycetes (Mucor circinelloides strain MAL-D3, Rhizopus arrhizus strain RA) (figure 47A-B). The binding was mediated by the MASP-1 peptide portion of the peptibody as the Fc fragment did not bind.

[0733] Conclusion

[0734] The peptibody targets a broad range of opportunistic fungi, which is in line with the binding pattern of the isolated MASP-1 peptide (SEQ ID NO: 1).

[0735] Example 11 - Clq binding to the Fc portion of the peptibody.

[0736] Background

[0737] One of the reasons to combine the MASP-1 peptide with an Fc region, is to engage the complement system as a contributor the effector functions of the peptibody. The classical pathway of complement is activated by the Cl complex containing the pattern recognition molecule Clq, which can bind to Fc regions of various immunoglobulins including IgGl. We therefore investigated whether Clq could bind to the Fc portion of the peptibody.

[0738] Materials and methods

[0739] Binding of Clq to the peptibody was measured by titrating purified Clq on an ELISA plate coated with 2 pg / ml peptibody. Coated Fc fragments and BSA were applied as a positive and negative control, respectively. Clq was detected with 1 pg / ml biotin-conjugated rabbit a-Clq polyclonal antibody followed by HRP-coupled streptavidin. The optical density (OD) was measured on an ELISA plate reader at 450 nm.

[0740] Results

[0741] Clq was shown to bind to the peptibody and there was also binding to the Fc fragments but not to BSA (Figure 48).

[0742] Conclusion

[0743] As Clq binds to the peptibody, the peptibody likely activates the complement system.

[0744] Example 12 - peptibody activation of the complement system

[0745] The peptibody contains an Fc fragment and thus is expected to have antibody-like properties. One of the effector functions of antibodies is the activation of the complement system through the classical pathway. Hence, we investigated the ability of the peptibody to activate the complement system.

[0746] Materials and methods

[0747] A.fumigatus conidia (lxlO7conidia / ml, strain 6871) were incubated with 10 pg / ml peptibody. As a complement source, 2% umbilical cord serum (UCS) + an MBL inhibitor (5 pg / ml, 3F8) were added, allowing complement activation to mainly run through the peptibody. A Clq inhibitor (10 pg / ml, Clq85) was applied to test whether complement activation was specifically mediated through the classical pathway. Complement activation products C4b, C3b and TCC were measured with flow cytometry using specific antibodies and fluorophore-coupled secondary antibodies.

[0748] Results

[0749] Peptibody-driven complement activation was measured in umbilical cord serum (UCS) because the low "background" immunoglobulin-mediated classical pathway activation in UCS enables us to see the peptibody effect. The MBL inhibitor eliminated the residual activation. Hence, there was no deposition of complement activation product in UCS + MBL inhibitor, whereas the presence of bound peptibody on the conidia facilitated the deposition of C4b, C3b and TCC. The activation was abolished when adding the Clq inhibitor (figure 49).

[0750] Conclusion

[0751] The complement system was activated by the conidia-bound peptibody, and the activation was specifically driven by Clq.

[0752] Example 13 - peptibody mediated phagocytosis

[0753] Background

[0754] The peptibody contains an Fc fragment and thus is expected to have anti body- 1 ike properties. One of the effector functions of antibodies is direct opsonization leading to phagocytosis and opsonization indirectly mediated through C3b deposition. Hence, we investigated the ability of the peptibody to mediate phagocytosis.

[0755] Materials and methods

[0756] FITC-labeled A.fumigatus conidia (lxlO7conidia / ml, strain 6871) were incubated with 10 pg / ml peptibody. As a complement source 2% umbilical cord serum (UCS) + an MBL inhibitor (5 pg / ml, 3F8) were added, allowing complement activation and, thus, opsonization to mainly run through the peptibody. A C3 inhibitor (compstatin Cp40) was applied to test whether phagocytosis was mediated through Fc-Fc receptor interactions and / or C3b deposition. Neutrophils (2xl06neu / ml) isolated from human EDTA blood using PolymorphPrep was added to the conidia to allow phagocytosis. The level of phagocytosis was measured with flow cytometry and the phagocytic index was defined as % FITC-positive neutrophils x the mean fluorescence intensity of the FITC positive neutrophils.

[0757] Results

[0758] The presence of peptibody alone on the conidia increased the level of phagocytosis. This was also the case when complement activation was preceding the addition of neutrophils. The C3 inhibitor lowered, but did not abolish the level of phagocytosis (figure 50), meaning that phagocytosis was mediated through a combination of direct peptibody opsonization and indirectly through C3b deposition. SEQENCE LISTING

[0759] SEQ ID NO: 1 - 30 amino acid fragment

[0760] GRSLPTCLPVCGLPKFSRKLMARI FNGRPA

[0761] SEQ ID NO: 2 - 5 amino acid fragment

[0762] KLMAR

[0763] SEQ ID NO: 3 - MASP-3 peptide

[0764] GRSLPTCLPECGQPSRSLPSLVKRI IGGRN

[0765] SEQ ID NO: 4 - 444-DDDDK-448 mutation

[0766] DDDDK

[0767] SEQ ID NO: 5 - CCP2 Ctrl. Pep 1

[0768] VDCRAPGELEHGLITFSTRNNLTTYKSEIK

[0769] SEQ ID NO: 6 - CCP2 Ctrl.Pep2

[0770] STRNNLTTYKSEIKYSCQEPYYKMLNNNTG

[0771] SEQ ID NO: 7 - CCP2 Ctrl.Pep3

[0772] GELEHGLITFSTRNNLTTYKSEIKYSCQEP

[0773] SEQ ID NO: 8 - Peptibody 1 - with 30 AA fragment

[0774] GRSLPTCLPVCGLPKFSRKLMARI FNGRPA- EPKSCDKTHTCP-

[0775] PCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNA

[0776] KTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAK-

[0777] Peptide (bold)-Hinge (Italic)-CH2 (underlined)-CH3 (grayscale)

[0778] SEQ ID NO: 9 - Peptibody 2 - Fc part with optimized linker sequence - 30 AA fragment

[0779] GRSLPTCLPVCGLPKFSRKLMARI FNGRPA- DKTHTCP-

[0780] PCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVWDVSHEDPEVKFNWYVDGVEVHNA

[0781] KTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAK-

[0782] Peptide (bold)-Hinge (Italic)-CH2 (underlined)-CH3 (grayscale)

[0783] SEQ ID NO: 10 - CC1P-CCP2-SP domains

[0784] ASMTGNECPELQPPVHGKIEPSQAKYFFKDQVLVSCDTGYKVLKDNVEMDTFQIECLKDGTWS

[0785] NKI PTCKI VDCRAPGELEHGLITFSTRNNLTTYKSEIKYSCQEPYYKMLNNNTGIYTCSAQGV

[0786] Underline: CCP1

[0787] Italic: CCP2

[0788] Bold: 30 aa peptide (SEQ ID NO: 1)

[0789] Gray: Protease domain (SP)

[0790] Bold + underline: 5 aa peptide (SEQ ID NO: 2)

[0791] SEQ ID NO: 11 - MASPl_human sequence (Full length - Heavy chain + light chain)

[0792] MRWLLLYYALCFSLSKASAHTVELNNMFGQIQSPGYPDSYPSDSEVTWNITVPDGFRIKLYFM HFNLESSYLCEYDYVKVETEDQVLATFCGRETTDTEQTPGQEWLSPGSFMS ITFRSDFSNEE RFTGFDAHYMAVDVDECKEREDEELSCDHYCHNYIGGYYCSCRFGYILHTDNRTCRVECSDNL FTQRTGVITSPDFPNPYPKSSECLYTIELEEGFMVNLQFEDI FDIEDHPEVPCPYDYIKIKVG PKVLGPFCGEKAPEPI STQSHSVLILFHSDNSGENRGWRLSYRAAGNECPELQPPVHGKIEPS QAKYFFKDQVLVSCDTGYKVLKDNVEMDTFQIECLKDGTWSNKI PTCKIVDCRAPGELEHGLI TFSTRNNLTTYKSEIKYSCQEPYYKMLNNNTGI YTCSAQGVWMNKVLGRSLPTCLPVCGLPKF SRKLMARI FNGRPAQKGTTPWIAMLSHLNGQPFCGGSLLGSSWIVTAAHCLHQSLDPEDPTLR DSDLLSPSDFKI ILGKHWRLRSDENEQHLGVKHTTLHPQYDPNTFENDVALVELLESPVLNAF VMPICLPEGPQQEGAMVIVSGWGKQFLQRFPETLMEIEIPIVDHSTCQKAYAPLKKKVTRDMI CAGEKEGGKDACAGDSGGPMVTLNRERGQWYLVGTVSWGDDCGKKDRYGVYSYIHHNKDWIQR VTGVRN

[0793] SEQ ID NO: 12 - MASPl_human sequence (Heavy chain)

[0794] MR W L L L Y Y AL C F S L S KAS AHTVELNNMFGQIQSPGYPDSYPSDSEVTWNITVPDGFRIKLYFM HFNLESSYLCEYDYVKVETEDQVLATFCGRETTDTEQTPGQEWLSPGSFMS ITFRSDFSNEE RFFGFDAWfMAVDVDECKEREDEELSCDHYCHNYIGGYYCSCRFGYILHTDNRTCRVE C S DNL FTQRTGVITSPDFPNPYPKSSECLYTIELEEGFMVNLQFEDI FDIEDHPEVPCPYDYIKIKVG PKVLGPFCGEKAPEPI STQSHSVLILFHSDNSGENRGWRLSYRAAGNECPELQPPVHGKIEPS QAKYFFKDQVLVSCDTGYKVLKDNVEMDTFQIECLKDGTWSNKI PTCKIVDCRAPGELEHGLI TFSTRNNLTTYKSEIKYSCQEPYYKMLNNNTGI YTCSAQGVWMNKVLGRSLPTCLPVCGLPKF SRKLMAR

[0795] Bold: start of 30 aa peptide (SEQ ID NO: 1)

[0796] Bold + underline: 5 aa peptide (SEQ ID NO: 2)

[0797] Underline CUB1 domain

[0798] Grey: CUB2 domain

[0799] SEQ ID NO: 13 - MASPl_human sequence (light ch a in / p rotease domain)

[0800] IFNGRPAQKGTTPWIAMLSHLNGQPFCGGSLLGSSWIVTAAHCLHQSLDPEDPTLRDSDLLSP SDFKI ILGKHWRLRSDENEQHLGVKHTTLHPQYDPNTFENDVALVELLESPVLNAFVMPICLP EGPQQEGAMVIVSGWGKQFLQRFPETLMEIEI PIVDHSTCQKAYAPLKKKVTRDMICAGEKEG GKDACAGDSGGPMVTLNRERGQWYLVGTVSWGDDCGKKDRYGVYSYIHHNKDWIQRVTGVRN

[0801] Underline: Protease domain

[0802] Bold: End of 30 aa peptide (SEQ ID NO: 1) SEQ ID NO: 14 - Linker / hinge sequence 1

[0803] EPKSCDKTHTCP

[0804] SEQ ID NO: 15 - Linker / hinge sequence 2 - no unpaired cysteine DKTHTCP

[0805] SEQ ID NO: 16 - Random peptide sequence

[0806] MWVRRQDGYGHKEVYRSCGHQTNKCTSLKE

[0807] SEQ ID NO: 17 - CUB1 peptide: GRETTDTEQTPGQEWLSPGSEMS ITFRSD

[0808] ITEMS

[0809] 1. An isolated fungus binding amino acid comprising a binding part having at least 70% sequence identity to SEQ ID NO: 1

[0810] 2. An isolated fungus binding amino acid comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2.

[0811] 3. An isolated fungus binding amino acid sequence according to item 2 being at least 10 amino acids long.

[0812] 4. A fusion-protein construct that contains an amino acid sequence according to any of items 1-3 and the amino acid sequence of an Fc-region from a human immunoglobulin.

[0813] 5. A fusion-protein construct according to item 4, wherein the human immunoglobulin is selected from the list consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, or IgD.

[0814] 6. A fusion-protein construct according to item 4, wherein the human immunoglobulin is IgGl.

[0815] 7. A fungus binding peptibody comprising a fungus binding amino acid conjugated to an Fc-region from a human immunoglobulin or a non-human immunoglobulin.

[0816] 8. A fungus binding peptibody according to item 7, wherein the fungus binding amino acid sequence is according to any of claims 1-3.

[0817] 9. A fungus binding peptibody according to any of items 7-8, wherein the peptibody further comprises a hinge region.

[0818] 10. A fungus binding peptibody according to any of items 7-9, wherein the human immunoglobulin is selected from the list consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, or IgD.

[0819] 11. A fungus binding peptibody according to item 10, wherein the human immunoglobulin is IgGl.

[0820] 12. A fungus binding peptibody, wherein the structure is defined as in Formula (I)

[0821] Formula (I): XI- Ll-Fl wherein,

[0822] -XI is a fungus binding amino acid according to any of claims 1-3, -Fl is an Fc Region, and

[0823] -LI is a peptide bond, linker or hinge region connecting XI to Fl.

[0824] 13. An isolated fungus binding amino acid sequence according to claims 1-3 or a fungus binding peptibody according to items 7-12 that is conjugated to an antibiotic.

[0825] 14. An isolated fungus binding amino acid sequence according to claims 1-3 or a fungus binding peptibody according to items 7-12 that is PEGylated.

[0826] 15. An isolated nucleic acid encoding a fungus binding amino acid sequence according to claim 1-3.

[0827] 16. A nucleic acid construct encoding a fusion-protein according to claim 4-6.

[0828] 17. An isolated nucleic acid comprising a nucleotide sequence which encodes a polypeptide according to any of items 1-12.

[0829] 18. An isolated nucleic acid construct comprising an isolated nucleic acid according to claim 17 operably linked to one or more control sequences.

[0830] 19. A host cell comprising a nucleic acid or nucleic acid construct according to claims 15-18.

[0831] 20. A host cell according to claim 19, wherein said host cell expresses a fungus binding amino acid sequence encoded by a nucleic acid or nucleic acid construct according to items 15-18.

[0832] 21. A method for producing a fungus binding amino acid sequence, the method comprising culturing the host cell according to item 20, and recovering the fungus binding amino acid sequence.

[0833] 22. A composition comprising a fungus binding amino acid sequence according to any one of items 1-14.

[0834] 23. A composition according to item 22 for use as a medicament.

[0835] 24. A composition according to item 22 for use in therapy.

[0836] 25. A composition according to item 22 for use in prophylaxis. 26. A composition according to item 22 for use in treating an immunocompromised individual.

[0837] 27. A composition according to item 22 for use in treating an opportunistic fungal infection.

[0838] 28. A composition according to item 27, wherein the opportunistic fungus belongs to the order of Mucorales.

[0839] 29. A composition according to item 27, wherein the opportunistic fungus is a species belonging to the genus Aspergillus.

[0840] 30. A composition according to item 22-29, wherein the composition is administrated nasal injection, subcutaneous injection, intravenous injection, inhalation or intratracheal injection.

[0841] 31. A composition according to item 22 for use in diagnostics of a fungal infection.

[0842] 32. A method for detecting a fungus in a sample comprising providing a sample suspected of containing a fungus adding a fungus binding amino acid according to any one of items 1-14 to the sample identifying the binding of the fungus binding amino acid to the fungus or fungal fragments within the sample.

[0843] 33. A method according to item 32, where binding of the fungus binding amino acid to the sample is identified by immunodetection or a radionuclide or a signalling molecule.

Claims

CLAIMS1. An isolated fungus binding amino acid sequence comprising a binding motif having at least 80% sequence identity to SEQ ID NO: 2, preferably at least 90% sequence identity, more preferably at least 95% sequence identity, and having a length of at the most 70 amino acids, preferably, at the most 50 amino acids, more preferably at the most 40 amino acids, and even more preferably at the most 35 amino acids, such as being 30 amino acids long.

2. The isolated fungus binding amino acid sequence according to claim 1, comprising a sequence differing from SEQ ID NO: 2 in at the most 1 amino acid position.

3. An isolated fungus binding amino acid sequence according to claim 1 or 2, comprising a fungus binding part having at least 80% sequence identity to SEQ ID NO: 1, and having a length of at the most 70 amino acids, preferably, at the most 50 amino acids, more preferably at the most 40 amino acids, and even more preferably at the most 35 amino acids.

4. The isolated fungus binding amino acid sequence according to claim 3, comprising a sequence differing from SEQ ID NO: 1 in at the most 6 amino acid positions, such as at the most 5 amino acids positions, such as at the most 4 amino acids positions, such as at the most 3 amino acids positions, preferably at the most 2 amino acids positions, and more preferably at the most 1 amino acids position.

5. The isolated fungus binding amino acid sequence according to any of the preceding claims, comprising SEQ ID NO: 2.

6. The isolated fungus binding amino acid sequence according to any of claims 3-5 comprising the cysteine at position 7 and / or the cysteine at position 11 of SEQ IDNO: 1, preferably comprising the cysteine at position 7 and the cysteine at position 11 of SEQ ID NO: 1.

7. The isolated fungus binding amino acid sequence according to any of the preceding claims having a length in the range 5-70 amino acids, such as 15-60 amino acids, such as 20-60 such as 20-60 amino acids, preferably 30-60 amino acids, more preferably 30-50 amino acids, even more preferably 30-40 amino acids, and even more preferably 30-35, such as being 31-35, 32-35, 33-35 amino acids long.

8. The isolated fungus binding amino acid sequence according to any of the preceding claims, comprising at least SEQ ID NO: 1.

9. The isolated fungus binding amino acid sequence according to any of the preceding claims, comprising or consisting of the amino acid sequence 108-177 of SEQ ID NO: 10, such as amino acids 118-177 of SEQ ID NO: 10, such as amino acids 128-177 of SEQ ID NO: 10, such as amino acids 128-167 of SEQ ID NO: 10, such as amino acids 128-157 of SEQ ID NO: 10, or such as amino acids 118-167 of SEQ ID NO: 10, such as amino acids 123-162.

10. The isolated fungus binding amino acid sequence according to any of the preceding claims, being free of one or more of the following domains from MASP- 1:- CUB1 domain; and / or- EGF domain; and / or- CUB2 domain; and / or- CCP1 domain; and / or- full or functional CCP2 domain; and / or full or functional SP domain.

11. The isolated fungus binding amino acid sequence according to any of the preceding claims, being free from one or more of the following functional sites of MASP-1:• homodimerization site; and / or• MBL binding site; and / or• Ficolin-2 binding site; and / or• One or more of the amino acids constituting the catalytic triad of the SP domain, preferably being from the catalytic site of the SP domain.

12. The isolated fungus binding amino acid sequence according to any of the preceding claims, being at least 10 amino acids long, such as at least 15 amino acids long, preferably at least 20 amino acids long, more preferably at least 30 amino acids long.

13. A fusion-protein construct that contains an amino acid sequence according to any of the preceding claims and the amino acid sequence of an Fc-region from a human immunoglobulin.

14. A fusion-protein construct according to claim 4, wherein the human immunoglobulin is selected from the group consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, and IgD.

15. A fusion-protein construct according to claim 13 or 14, wherein the human immunoglobulin is IgGl.

16. The fusion-protein construct according to any of claims 13-14, wherein the human immunoglobulin is IgG3.

17. The fusion protein according to any of claims 13-16, having binding affinity for fungus.

18. A fungus binding peptibody comprising a fungus binding amino acid sequence according to any of claims 1-12 conjugated to an Fc-region from a human immunoglobulin or a non-human immunoglobulin.

19. A fungus binding peptibody according to claim 18, wherein the peptibody further comprises a hinge / linker region.

20. The fungus binding peptibody according to claim 19, wherein the hinge / linker is free from unpaired cysteines, such as only comprising one pairing cysteine in the linker region.

21. The fungus binding peptibody according to any of claims 18-20, wherein the human immunoglobulin is selected from the group consisting of an IgGl, IgG2, IgG3, IgG4, IgAl, IgA2, IgM, IgE, and IgD.

22. A fungus binding peptibody according to any of claims 18-21, wherein the human immunoglobulin is IgGl or IgG3.

23. A fungus binding peptibody, wherein the structure is defined as in Formula (I)Formula (I): X1-L1-F1 wherein,XI is a fungus binding amino acid sequence according to any of claims 1-12,LI is a peptide bond, linker or hinge region connecting XI to Fl; andFl is an Fc Region.

24. An isolated fungus binding amino acid sequence according to any of claims 1-12, a fusion protein according to any of claims 13-17 or a fungus bindingpeptibody according to any of claims 18-23 that is modified by the attachment of a bioactive compounds, such as selected from the group consisting of fluorophores, fluorescent / phosphorescent proteins or compounds, radiolabeled compounds, antibiotics, enzymatically active peptides, molecular probes, functional tags such as HIS / FLAG-tags, imaging agents, radioligands and / or PEG.

25. An isolated nucleic acid sequence encoding a fungus binding amino acid sequence according to claim 1-12.

26. A nucleic acid construct encoding a fusion-protein according to any of claims 13-17 and / or encoding a fungus binding peptibody according to any of claims 18- 23.

27. An isolated nucleic acid construct comprising an isolated nucleic acid sequence according to claim 25 or a nucleic acid construct according to claim 26, operably linked to one or more control sequences.

28. A host cell comprising a nucleic acid or nucleic acid construct according to any of claims 25-27.

29. The host cell according to claim 28, wherein said host cell expresses a fungus binding amino acid sequence encoded by a nucleic acid sequence or nucleic acid construct according to any of claims 25-27.

30. A method for producing a fungus binding amino acid sequence, the method comprising culturing the host cell according to claim 28 or 29, and recovering the fungus binding amino acid sequence.

31. A composition comprising a fungus binding amino acid sequence according to any one of claims 1-12, the fusion-protein construct according to any of claims 13-17, and / or the fungus binding peptibody according to any of claim 18-23.

32. The composition according to claim 31, being a pharmaceutical composition.

33. The composition according to claim 31 or 32, further comprising a pharmaceutical acceptable, carrier, diluent and / or adjuvant.

34. The composition according to any of claims 31-33, comprising a buffering system, such as PBS, acetate, phosphate, citrate, and / or glutamate.

35. The composition according to any of claims 27-30, having a pH in the range 5- 9, preferably 6-8, more preferably 6.5-8, or 7-8.

36. The isolated fungus binding amino acid sequence according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35 for use as a medicament.

37. The isolated fungus binding amino acid sequence according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35 for use in therapy.

38. The isolated fungus binding amino acid sequence according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35, for use in prophylaxis, such as prophylaxis treatment for a fungus infection.

39. The isolated fungus binding amino acid sequence according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35, for use in the treatment of a human subject, such as a male subject or a female subject.

40. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, wherein said subject suffers from cancer.

41. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, for use in treating an immunocompromised individual, preferably a human subject.

42. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, wherein said immunocompromised individual is a human subject.

43. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, wherein said immunocompromization is caused by a treatment, a condition and / or disorder selected from the group consisting of- treatment for solid tumor and hematologic malignancies, such as treatment with chemotherapeutics or radiation; hematologic malignancies associated with poor responses to COVID-19 vaccines regardless of current treatment status,- immunosuppressive therapy;- receipt of solid-organ transplant or an islet transplant and receiving immunosuppressive therapy;- receipt of chimeric antigen receptor (CAR)-T-cell therapy or hematopoietic stem cell transplant;- an immunodeficiency, such as common variable immunodeficiency disease (CVID), severe combined immunodeficiency (SCID), DiGeorge syndrome, Bruton's agammaglobulinemia, and Wiskott-Aldrich syndrome;HIV infection, such as an advanced or untreated HIV infection, CD4 cell counts less than 200 / mm3, history of an AIDS-defining illness without immune reconstitution, or clinical manifestations of symptomatic HIV;- treatment with high-dose corticosteroids (i.e., 20 or more mg of prednisone or equivalent per day when administered for 2 or more weeks), alkylating agents, antimetabolites, transplant-related immunosuppressive drugs, cancer chemotherapeutic agents classified as severely immunosuppressive, tumor necrosis factor (TNF) blockers, and other biologic agents that are immunosuppressive or immunomodulatory;- young age such as below 48 months;- old age, such as 70 years or above, such as above 80, such as above 90;- chronic Respiratory Diseases: Individuals with chronic respiratory conditions such as asthma, cystic fibrosis, and chronic obstructive pulmonary disease (COPD) are at higher risk. The damaged airways and mucus accumulation in these conditions provide a suitable environment for fungal growth; poorly controlled Diabetes is at higher risk: Diabetes can impair the immune system, making it more difficult for the body to fight off infections, including fungal infections. This impairment is particularly evident in cases of poorly controlled diabetes, where high blood sugar levels can hinder the function of immune cells;- prolonged Use of Antibiotics: Long-term use of broad-spectrum antibiotics can disrupt normal microbial flora, which can increase susceptibility to fungal infections;hospitalization, Especially in Intensive Care Units: Patients in hospitals, particularly in intensive care units, are at risk due to weakened health, invasive procedures, and potential exposure to hospital-acquired pathogens; pre-existing Lung Cavities: People with pre-existing lung cavities, such as those caused by tuberculosis or sarcoidosis, are at risk as these cavities can become colonized by the fungus; and- environmental Exposure: Occupational exposure to environments with high levels of fungal spores, such as farming, gardening, or construction sites, can increase the risk of inhaling Aspergillus spores and give rise to fungal colonization.

44. The composition for use according to claim 43, wherein the hematologic malignancy is selected from the group consisting of chronic lymphocytic leukemia, non-Hodgkin lymphoma, multiple myeloma, and acute leukemia.

45. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, for use in treating an opportunistic fungal infection.

46. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, for use according to claim 45, wherein the opportunistic fungus belongs to the order of Mucorales.

47. The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35, for use according to claim 45, wherein the opportunistic fungus is a species belonging to the genus Aspergillus.

48. The isolated fungus binding amino acid sequence for according to any of claims 1-9, the fusion-protein construct according to any of claims 10-13, the peptibody according to any of claims 14-18, and / or the composition according to any of claims 27-31, for use according to any of claims 36-47, wherein the composition is administrated nasal injection, subcutaneous injection, intravenous injection, inhalation or intratracheal injection.

49. The isolated fungus binding amino acid sequence according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35, for use according to any of claims 36-47, for use according to any of claims 36-48, being administered to a human subject in a dosage in the range 0.06 mg / kg bodyweight to 600 mg / kg bodyweight, preferably in the range 0.1 -100 - mg / kg bodyweight, more preferably 1-100 mg / kg bodyweight, an even more preferably 1-50 mg / kg bodyweight.

50. A combination for use in the treatment, prevention and / or alleviation of a fungal infection comprising- The isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13- 17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35; and- an additional antifungal compound, different from the isolated fungus binding amino acid sequence for use, according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31-35.

51. The combination for use according to claim 50, wherein the additional antifungal compound is selected from the group consisting of polyene antifungal agents, azoles antifungal agent, echinocandin antifungal agent, pyrimidine analogantifungal agent, allylamine antifungal agent, Papulacandins and combinations thereof.

52. The combination for use according to claim 50 or 51, wherein the additional antifungal compound is selected from the group consisting of otrimazole (Canesten), econazole, miconazole, terbinafine (Lamisil), fluconazole (Diflucan), ketoconazole (Daktarin), nystatin (Nystan), voriconazole (Vfend), Amphotericin B, flucytosine, and combination thereof.

53. An isolated fungus binding amino acid sequence, according to any of claims 1- 12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, the composition according to any of claims 31- 35, for use in diagnostics of a fungal infection.

54. A method for detecting a fungus in a sample comprising providing a sample suspected of containing a fungus; adding the isolated fungus binding amino acid sequence according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, the peptibody according to any of claims 18-23, and / or the composition according to any of claims 31-35 to the sample; identifying the binding of the isolated fungus binding amino acid sequence according to any of claims 1-12, the fusion-protein construct according to any of claims 13-17, and / or the peptibody according to any of claims 18-23 to the fungus or fungal fragments within the sample.

55. The method according to claim 48, wherein binding is identified by immunodetection of a radionuclide or a signaling molecule.

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