WO2025051955A1

WO2025051955A1 - Materials with anti-inflammatory and/or anti-microbial properties

Info

Publication number: WO2025051955A1
Application number: PCT/EP2024/074991
Authority: WO
Inventors: Artur Schmidtchen; Manoj PUTHIA
Original assignee: IN2CURE AB
Current assignee: IN2CURE AB
Priority date: 2023-09-06
Filing date: 2024-09-06
Publication date: 2025-03-13
Anticipated expiration: 2026-03-06

Abstract

The invention relates to materials comprising thrombin derived peptides and poly(lactic-co-glycolic acid) polymers. Said materials have anti-microbial, such as anti-bacterial, and/or anti-inflammatory properties. Also provided are medical products, such as sutures, comprising said materials, and methods for preventing and/or inhibiting inflammation and/or infection, or for treating wounds, in a subject using said medical products.

Description

Materials with anti-inflammatory and/or anti-microbial properties

Technical field

The present invention relates to materials comprising thrombin derived peptides and poly(lactic-co-glycolic acid) (PLGA) polymers. Said materials have anti-microbial, such as antibacterial, and/or anti-inflammatory properties and are useful when comprised in medical products, such as sutures.

Background

Of more than 300 million surgical procedures performed worldwide annually, almost 10% develop surgical site infections (SSIs). SSIs account for a substantial clinical and economic burden. Although many factors contribute to SSIs, preventive measures before, during, and after surgery can lower the SSI incidence. Causes of SSIs vary depending on anatomy, surgical procedure, and exogenous in addition to endogenous, patient-derived factors. Bacterial contamination is one cause that can be controlled. In hospitals, preventive measures such as hygiene routines are implemented but even under sterile surgical conditions, infections may occur due to the spread of bacteria from the patient’s own bacterial flora. For example, it has been reported that up to 60% of the bacteria recovered from infected surgical wounds developed antibiotic resistance.

In the case of medically implanted materials, such as sutures, these provide an ideal surface for bacteria to adhere to and grow and may be involved in the development of SSI pathogenesis. Moreover, in the case of bacterial adhesion, the release of bacterial products, such as lipoteichoic acid (LTA) and LPS, may stimulate inflammation in the vicinity of biomaterials. Therefore, efforts have been made to functionalize sutures to prevent bacterial colonization and consequent infection. Common anti-infective sutures containing antiseptic molecules, such as triclosan, have however shown inadequate efficacy and may induce bacterial resistance. In addition, triclosan can cause side effects, such as hormonal disruption. Another serious problem is the declining effectiveness of antibiotics and other antimicrobials due to antimicrobial resistance (AMR) development. The development of resistance is particularly important in surgical procedures in which the combination of extensive antibiotic use, high risk of systemic spread, and bacterial sepsis leads to a high frequency of infections for which “last resort” antibiotics need to be used.

Additionally, in the initial contact with tissues, it is plausible that a given biomaterial also induces an immune response perse, which may cause dysregulation of inflammatory responses, leading to inefficient host defense and causing a given biomaterial to become infection-prone.

There is thus an urgent need for materials for use in medical products to improve medical outcomes after surgery in relation to reducing or preventing both bacterial infection and the associated inflammation.

Summary

The present disclosure provides materials comprising thrombin derived peptides and poly(lactic-co-glycolic acid) (PLGA) polymers. Said materials have anti-microbial, such as anti-bacterial, and/or anti-inflammatory properties and are particularly useful when comprised in medical products, such as sutures.

Interestingly, the present invention discloses that PLGA polymers associate with thrombin derived peptides, which allow for slow, continuous release of the peptides in vivo. This in turn allows the materials of the invention to exert their anti-bacterial and/or anti-inflammatory properties over an extended time period. Additionally, by tuning the ratio between lactic acid and glycolic acid in the PLGA polymers, the release profile of said peptides from the polymers can be controlled. This thus allows for tailored release timing for specific applications.

Furthermore, the medical products of the invention are stable in storage and provide continuous and sufficiently high release of the peptide when the product is applied to the relevant parts of the body. This ensures effective provision of both the antibacterial and anti-inflammatory effects of the polymer to the tissue(s) of treatment.

In addition, PLGA provides a microenvironment promoting activity of thrombin derived peptides. Without being bound by theory, it is believed that PLGA provides an acidic microenvironment leading to protonation of thrombin derived peptides, which may promote their activity, e.g. it may promote anti-bacterial and/or anti-inflammatory activity.

In some aspects of the present disclosure is provided a material comprising a poly(lactic-co-glycolic acid) (PLGA) polymer and a peptide comprising or consisting of the amino acid sequence

X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein

X3, X7, X10, X11, X13, X15, X16 is any standard amino acid,

X8, X12 is any amino acid,

X1 , X6, X14 is G, A, V, L, I, P, F, M, Y or W, and

X2, X4, X5, X9 is R, K or H, wherein said peptide has a length of from 10 to 30 amino acid residues.

In some aspects of the present disclosure is provided a medical product comprising the material as described herein.

In some aspects of the present disclosure is provided a method of producing the medical product as described herein, the method comprising a step of submerging a starting material comprising or consisting of PLGA in a coating solution comprising a dissolved peptide as described herein, followed by drying said starting material.

In some aspects of the present disclosure is provided a method of preventing and/or inhibiting inflammation and/or infection in a subject in need thereof, said method comprising contacting said subject with or implanting in said subject the medical product as described herein and maintaining said contact for a period of time.

In some aspects of the present disclosure is provided a method of treating a wound in a subject in need thereof, said method comprising contacting said wound with or implanting in said wound the medical product as described herein.

In some aspects of the present disclosure is provided a medical product as described herein for use in the prevention and/or inhibition of inflammation and/or infection in a part of the body of a subject. In some aspects of the present disclosure is provided a medical product as described herein for use in the treatment of a wound in a subject in need thereof.

Description of Drawings

Figure 1. Coating conditions, antibacterial properties, and release profile of TCP-25 peptide-coated polyglactin suture. Effects of coating conditions on the peptide loading and antimicrobial activity of polyglactin sutures. Sutures were coated with TCP-25 under varying conditions of coating concentrations (a), coating times (b), and coating temperatures (c). The peptide was eluted from the coated sutures and protein concentrations were estimated (Left panels). To study effects on antimicrobial activity, elutions from sutures were used in an radial discussion assay (RDA) as shown in the right panel. Diameter of the clear zone (excluding the 4 mm well) is presented as the inhibitory effect of the released peptide (For a, b and c, mean values ± standard error of the mean [SEM] are presented, n=3). (d) The cumulative release of TCP-25 from sutures in vitro. As illustrated, a Transwell insert was used to obtain conditions mimicking a surgical wound, and TCP-25 release was estimated, (e) In vivo release of TCP-25 from suture. In SKH-1 mice, sutures coated with tetramethylrhodamine (TAMRA)-labeled TCP-25 were subcutaneously placed. At 1 , 6, 24, and 72 hours, peptide release was longitudinally monitored by quantifying fluorescence using I VIS imaging. Heat map overlays obtained from emitted light are shown. Bar chart shows the measured radiance emitted from the region of interest (mean values are presented, n=4). The dotted line shows the region of interest, (f) Scanning electron microscopy showing the surface of the polyglactin sutures before and after the TCP-25 coating. *P < 0.05; ***P < 0.001.

Figure 2. QCM-D, coarse-grained simulation, and O-PTIR analysis showing peptide- polyglactin interactions, (a) QCM-D monitoring of TCP-25 binding to Vicryl fibers. Finely cut fibers were dissolved in ethanol and drop-cast to poly-L-lysine-coated SiO2 to a confluent fiber mat. Peptide binding onto fiber-functionalized sensors was confirmed by frequency changes (AF) of -100 ± 27 Hz and dissipation changes (AD) of (+42 ± 11 )- 10’⁶ with respect to pure MQ water. In contrast, the possible adsorption of the peptide onto the underlying poly-L-lysine surface was ruled out in control experiments, in which the interaction of the peptide with fiber-free poly-L-lysine-functionalized SiO2 surfaces was monitored under the same experimental conditions, showing only a minute frequency shift of -4±1 Hz. Measurements were performed at room temperature (n=3). (b-e) Coarse-grained simulations of Vicryl assembly with TCP-25, (b) A 1 ps CG self-assembly simulation of 50 copies of the 100-mer polyglactin 910 model was performed to build a Vicryl polymer model. Subsequently, 10 copies of TCP-25 were added to the system and three independent 10 ps simulations were conducted at 320 K. The figure shows initial and final snapshots from one of the simulations, (c) (Top) Minimum distance between TCP-25 peptides and the surface of the Vicryl polymer for all three simulations. Thick lines show average over 10 peptides and the shaded areas indicate standard deviation. (Bottom) Solvent accessible surface area (SASA) of the Vicryl polymer comparing simulation with (grey) and without (black) TCP-25. Thick lines show average over three repeat simulations and shaded areas indicate standard deviation. Probe radius used for SASA calculation is 0.26 nm. (d) (Top) The average percentage of contacts made by each subunit of polyglactin with the peptide at three time points during the simulations. COO, carboxyl terminus; G1 to G9, glycolide subunits; L6, lactide subunit; G10-OH, glycolide with hydroxyl terminus. (Bottom) The same analysis performed for each residue of the peptide. Distance cut-off used for contact measurement is 0.6 nm. (e) (Top) An enlarged snapshot showing entanglement of the TCP-25 peptide with the polyglactin polymer from the end of one simulation. (Bottom) SASA of the peptides throughout the simulations, averaged over ten peptides and three simulations, (f) Normalized O-PTIR spectra acquired at 2 cm^-1 spectral data point spacing with 5 averages from uncoated (control, bottom line on graph), TCP-25 coated and washed TCP-25 coated sutures. Dashed line shows the band position characteristic for TCP-25 coated sutures at 1656 cm^-1, (g) O-PTIR ratio maps were derived from the images acquired at 1656 cm^-1 (ratio map nominator) and divided by the images acquired at 1760 cm^-1 (ratio map denominator). The light grey color shows the distribution of TCP-25 on the suture surface.

Figure 3. Structural analysis of TCP-25 eluted from coated sutures, (a) High- performance liquid chromatography (HPLC) analysis of fresh TCP-25 or TCP-25 eluted from coated sutures, (b) Circular dichroism (CD) spectra of TCP-25 from coated sutures either alone or after incubation with LPS (left panel). The a-helical content of TCP-25 estimated from molar ellipsometry at 222 nm (right panel) is displayed. Data are presented as the mean ± SEM (n=3). P values were determined using an unpaired t test. Top line in graph is TCP-25, the line beneath is +2, etc. (c) Intrinsic fluorescence spectra of 10 pM TCP-25 from the coated sutures, showing shifts in the emission maximum of the peptides after incubation with varying concentrations of LPS (left panel), (d) Fitting of the TCP-25’s emission maximum wavelength (A_max) in function of varying concentrations of LPS. Data are represented as the mean ± SEM (n=3). *P < 0.05.

Figure 4. In vitro and in vivo antibacterial effects of TCP-25 suture (a) Bioluminescent Staphylococcus aureus or Pseudomonas aeruginosa bacteria were incubated with TCP-25-coated or control suture and imaged using I VIS. Representative photos are shown (n = 3). (b) Bioluminescence emitted from bacteria after treatment with TCP-25 sutures. Bioluminescent versions of S. aureus or P. aeruginosa were incubated with TCP-25 sutures. Signals from bacteria were acquired using a luminometer. Data are presented as the mean ± SEM (n = 3). P values were calculated using a two-way analysis of variance (ANOVA) and Tukey’s post hoc test, (c) Representative images showing results from Bacterial live-dead assay. Staining was performed using a LIVE/DEAD Baclight bacterial viability kit and imaged using fluorescence microscopy. The white color shows live bacteria whereas the light grey color shows dead bacteria (n = 3). (d) Scanning electron microscopy (SEM) images showing the bacterial morphology after contact with TCP-25 sutures, (e) In vivo infection imaging showing anti-bacterial properties of TCP-25 sutures in a mouse model of suture infection.

Uncoated control sutures or TCP-25 sutures were subcutaneously implanted on the left or right side respectively and contaminated with bioluminescent S. aureus. Bacterial bioluminescence was non-invasively imaged using the I VIS imaging system, (f) The line chart shows the bacterial bioluminescence emission at 1 , 6, 24, 48, and 72 h postinfection. Data are shown as the mean ± SEM (n = 5). P values were calculated using a two-way ANOVA with Sidak’s test, (g) Bar-chart shows bacterial counts in the tissue surrounding control or TCP-25 sutures. After suture implantation and contamination with S. aureus, mice were sacrificed at 72 h and tissue adjacent to the suture was collected for CFU enumeration using viable count assay. Data are shown as the mean ± SEM (n=5 mice per group). P values were calculated using unpaired t tests, (h) In vivo infection and drug imaging by I VIS in mice. To image in vivo drug localization along with bioluminescent bacterial imaging, sutures were coated with fluorescently labeled TCP-25. Representative photos display bioluminescence (lum) and TCP-25 TAMRA fluorescence (flu) 6 h post-suture implantation, (n = 5). *P < 0.05; **P < 0.01 ;

***P < 0.001. Figure 5. In vitro and in vivo effects of TCP-25 sutures on endotoxin-induced inflammation, (a) NF-KB and AP-1 activation analysis by quantifying secreted alkaline phosphatase in THP1-Xblue™-CD14 cells (upper bar chart). Cell viability was determined with the MTT assay (lower bar chart). Lysed cells were included as positive control. Data are shown as the mean ± SEM (n=3). P values were calculated using a one-way ANOVA. (b) Longitudinal imaging of inflammation in NF-KB reporter mice. TCP-25 or control sutures were implanted on the left or right side respectively and contaminated with LPS. An I VIS Spectrum system was used for in vivo bioimaging of NF-KB reporter gene expression. Representative heat-map overlay photos show bioluminescent signals at 3 and 24 h after implantation. Bar charts show analysis of the emissions from the NF-KB reporter mice. Data are shown as the mean ± SEM (n = 5). P values were determined using unpaired t tests, (c) TNF-a and IL-6 cytokine levels in fluid extracted from implanted sutures after 24 h of implantation. Data are shown as the mean ± SEM (n = 5 mice per group). P values were calculated by unpaired t tests. *P<

O.05, **P < 0.01 , ***P < 0.001 , ***P < 0.001. NS, non-significant.

Figure 6. Antibiofilm effects of TCP-25-coated suture, (a) Representative fluorescence microscopy images after live/dead staining showing S. aureus or P. aeruginosa biofilm adhered to the suture surface (white, live bacteria; light grey, dead bacteria). TCP-25- coated or control sutures were added to the wells of the biofilm microtiter plate and biofilms were allowed to grow for 48 hours. Sutures were stained using LIVE/DEAD Baclight bacterial viability kit and imaged using fluorescence microscopy. Vicryl® Plus was used as a benchmark comparison. The bar chart shows the total number of live bacteria on suture-adhered biofilms estimated using a viable count analysis. Data are shown as the mean ± SEM (n=3). P values were analyzed using a one-way ANOVA. (b) Crystal violet staining showing measurement of biofilm mass. Vicryl® Plus was used as a benchmark comparison. Data are shown as the mean ± SEM (n=3). P values were determined using a one-way ANOVA. (c) Scanning electron microscopy of S. aureus or

P. aeruginosa biofilm grown on TCP-25-coated or control sutures. Representative SEM images showing the suture surface. Arrowhead shows bacterial biofilms formed on the suture surface, (d) Bacterial live-dead analysis showing the antibiofilm effects of the TCP-25-coated sutures. S. aureus and P. aeruginosa mature biofilms were treated with TCP-25-coated or control sutures. Staining was performed using LIVE/DEAD Baclight bacterial viability kit and followed by fluorescence microscopy. The white color shows live bacteria whereas the light grey color shows dead bacteria. Representative images are shown (n = 3). Vicryl® Plus was used as a benchmark comparison. *P< 0.05, **P < 0.01 , ***p< 0.001. NS, non-significant.

Figure 7. Human neutrophil elastase-induced TCP-25 fragmentation in TCP-25 sutures, (a) Peptide fragmentation pattern of TCP-25 after treatment of TCP-25 suture with human neutrophil elastase. TCP-25 suture was incubated with human neutrophil elastase and analyzed by nano LC-MS/MS. The figure shows the sequences of main peptides and the quantity of successful identifications by mass spectrometry at 0, 30, and 180 min (n = 2). (b) Illustration of main peptides obtained after human neutrophil elastase digestion of TCP-25 (SEQ ID NO: 1) from coated sutures. *These peptides have been reported to show antibacterial effects, (c) Antibacterial activity of peptide fragment products obtained after treatment of TCP-25 suture with human neutrophil elastase was analyzed by evaluating the antibacterial activity against E. coli by RDA. The bar chart shows quantification of the clear zone. Data are shown as the mean ± SEM (n = 3). (d) The anti-inflammatory activity of peptide fragment products obtained after treatment of TCP-25 suture with human neutrophil elastase. NF-KB and AP-1 activation was evaluated in THP1-Xblue™-CD14 reporter cells. Data are shown as the mean ± SEM (n=3). ***P < 0.001. NS, non-significant.

Figure 8. Determination of the tensile strength, hemolytic activity, and effects upon long-term storage of TCP-25 sutures, (a) Effect of TCP-25 coating on the tensile strength of the sutures. The tensile strengths of freshly coated non-implanted (left panel) or tissue-implanted (right panel) sutures were measured. To study the effect of TCP-25 coating on the tensile strength in vivo, sutures were subcutaneously implanted in mice for 4 days. Data are shown as the mean ± SEM (n = 5). A Mann-Whitney II test was used to calculate P values, (b) Hemolytic activity of control and TCP-25-coated sutures. A hemolysis assay was performed using human blood. The left bar chart shows the hemolytic activity of 1 cm long sutures and the right bar chart shows the hemolytic activity of 10 cm long sutures (mean values ± SEM are presented, n=3). NS, non-significant. (c) HPLC analysis of control (fresh TCP-25) or TCP-25 eluted from coated sutures after long-term storage. TCP-25 sutures were stored at room temperature for 18 months after which peptides were eluted for HPLC analysis (d) Western blot analysis of TCP-25 eluted from coated sutures (n=3) after 18 months of storage at room temperature. 1, 2, 3 denote different sutures, (e) Results showing CD spectra of TCP-25 from stored sutures with and without LPS. Fresh TCP-25 was used as a control for comparison, (f) Suture storage effect on the antimicrobial activity and release of TCP-25 as evaluated by radial diffusion assay using E. coli. Fresh TCP-25 was used as a control for comparison (con). Results show quantification of the clear zones. Data are shown as the mean ± SEM (n = 3-4). NS, non-significant.

Figure 9. TCP-25 suture targets human wound fluid-induced inflammation, (a) TCP-25 suture decreases human wound fluid’s pro-inflammatory ability in vitro. In a reporter assay, THP-1-XBIue™-CD14 cells were used. In the presence of TCP-25 suture or control suture, cells were stimulated by acute (AWF) and chronic wound fluid (CWF) derived from infected wounds from human patients, NF-KB and AP-1 activation was evaluated by determining the production of secreted alkaline phosphatase from the reporter cells. Data are presented as the mean ± SEM (n = 6). P values were calculated using paired t test, (b) TCP-25 suture decreases human wound fluid’s pro- inflammatory ability in vivo. TCP-25-coated sutures were contaminated with human chronic wound fluid and implanted on the back of NF-KB reporter mice. Non-invasive I VIS imaging was performed to visualize NF-KB activation. Data are shown as the mean ± SEM (n = 5). P values were computed using a Mann-Whitney II test. *P< 0.05, **P< 0.01.

Figure 10. SDS-PAGE analysis of PLGA particles coated with TCP-25. The results are further described in Example 2.

Figure 11. Detection and localization of TCP-25 on PLGA particles. Coated particles were imaged using fluorescence microscopy to detect TCP-25-TAMRA (light grey). PLGA is dark grey in the figure. The results are further described in Example 2.

Figure 12. Assay of bacterial growth on media comprising dried TCP-25-coated PLGA particles. Viable Count Assays (VCA) were made of different bacteria that had been incubated with TCP-25-coated PLGA particles (TCP-25 PLGA) or uncoated PLGA particles (control), and then plated on TH agar plates. The results are further described in Example 2.

Figure 13. Antimicrobial effects of dried TCP25-coated PLGA particles. Radial diffusion assays showing antibacterial effects of TCP25-coated PLGA particles on Gram-negative E. coli (left) and Gram-positive S. aureus (right). The results are further described in Example 2.

Detailed description Definitions

In this specification, unless otherwise specified, “a” or “an” means “one or more”.

As used herein the term “approximately” when used in relation to a numerical value refers to +/-10%, preferably +/- 5%, more preferably to +/- 1%.

The term “sequence identity” as used herein refers to the % of identical amino acids or nucleotides between a candidate sequence and a reference sequence following alignment. Thus, a candidate sequence sharing 80% amino acid identity with a reference sequence requires that, following alignment, 80% of the amino acids in the candidate sequence are identical to the corresponding amino acids in the reference sequence. Identity according to the present invention is determined by aid of computer analysis, such as, without limitations, the Clustal Omega computer alignment program for alignment of polypeptide sequences (Sievers et al. (2011 October 11) Molecular Systems Biology 7 :539, PMID: 21988835; Li et al. (2015 April 06) Nucleic Acids Research 43 (W1) :W580-4 PMID: 25845596; McWilliam et al., (2013 May 13) Nucleic Acids Research 41 (Web Server issue) :W597-600 PMID: 23671338, and the default parameters suggested therein. The Clustal Omega software is available from EMBL- EBI at https://www.ebi.ac.uk/Tools/msa/clustalo/. Using this program with its default settings, the mature (bioactive) part of a query and a reference polypeptide are aligned. The number of fully conserved residues are counted and divided by the length of the reference polypeptide. The MUSCLE or MAFFT algorithms may be used for alignment of nucleotide sequences. Sequence identities may be calculated in a similar way as indicated for amino acid sequences. Sequence identity as provided herein is thus calculated over the entire length of the reference sequence.

The term "standard amino acid" refers to any of the twenty genetically-encoded amino acids commonly found in naturally occurring peptides. The standard amino acids are referred to herein both by their IUPAC 1 -letter code and 3-letter code. The term “standard amino acid” is used to refer both to free standard amino acids, as well as standard amino acids incorporated into a peptide. For the peptides shown, each encoded amino acid residue, where appropriate, is represented by a single letter designation.

The term “any amino acid', as used herein refers to compounds chemically classified as amino acids. The term thus includes the twenty standard amino acids and their corresponding stereoisomers in the 'D' form (as compared to the natural ‘L’ form), omega-amino acids, other naturally-occurring amino acids, unconventional amino acids (e.g., a,a-disubstituted amino acids, N-alkyl amino acids, etc.) and chemically derivatised amino acids.

The term “treatment” as used herein refers to any type of treatment or prevention of a disorder, including improvement in the disorder of the subject (e.g., in one or more symptoms), delay in the progression of the disorder, delay the onset of symptoms or slowing the progression of symptoms. Treatment may also be ameliorating or curative treatment. As such, the term "treatment" also includes prophylactic treatment of the individual to prevent the onset of symptoms.

As used herein the term "hydrogel" refers to a continuous phase of an aqueous solution and a hydrophilic polymer that is capable of swelling on contact with water. The "hydrogel" comprises nanostructures formed of said polymer and water, and typically contain more than 90% water. Hydrogels are typically transparent or translucent, regardless of their degree of hydration. Hydrogels are generally distinguishable from hydrocolloids, which typically comprise a hydrophobic matrix that contains dispersed hydrophilic particles. Hydrogels typically have a flow point of at least 10 Pa, such as at least 15 Pa, for example in the range of 10 to 80 Pa, such as in the range of 40 to 60 Pa.

Material

The present disclosure relates to materials comprising a PLGA polymer as described elsewhere herein and a peptide as described elsewhere herein, said materials having anti-bacterial and/or anti-inflammatory properties, and this being particularly useful when comprised in medical products, such as sutures, microparticles or nanoparticles.

In some aspects is provided a material comprising a poly(lactic-co-glycolic acid) (PLGA) polymer and a peptide comprising or consisting of the amino acid sequence X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein

X3, X7, X10, X11 , X13, X15, X16 is any standard amino acid,

X8, X12 is any amino acid,

X1 , X6, X14 is G, A, V, L, I, P, F, M. Y or W, and

In some embodiments, said material is anti-inflammatory. In some embodiments, said material is anti-bacterial. In some embodiments, said material is anti-inflammatory and anti-bacterial.

In some embodiments, said material is capable of binding to lipopolysaccharides (LPS). In some embodiments, said LPS is a bacterial LPS.

In some embodiments said material is biodegradable and/or bioabsorbable.

In some embodiments, said peptide is associated with said PLGA polymer. In some embodiments, the peptide is coated on said PLGA polymer. In some embodiments, the peptide is dissolved, such as fully or partially dissolved, in said PLGA polymer. In some embodiments, the peptide is interspersed between the fibers of said PLGA polymer.

In some embodiments, the material comprises at least 10 pg/cm², such as at least 25 pg/cm², such as least 50 pg/cm², such as least 75 pg/cm², such as least 100 pg/cm², such as least 150 pg/cm², such as least 200 pg/cm², such as least 250 pg/cm², such as least 300pg/cm², such as at least 350 pg/cm² such as least 400 pg/cm², such as least 450 pg/cm², such as least 500pg/cm² of said peptide.

In some embodiments, the material comprises from 10 pg/cm² to 10000 pg/cm², such as from 25 pg/cm² to 10000 pg/cm², such as from 25 pg/cm² to 7500 pg/cm², such as from 25 pg/cm² to 5000 pg/cm², such as from 25 pg/cm² to 2500 pg/cm², such as from 50 pg/cm² to 2000 pg/cm², such as from 75 pg/cm² to 1500 pg/cm², such as from 100 pg/cm² to 1000 pg/cm², such as from 150 pg/cm² to 750 pg/cm², such as from 200 pg/cm² to 500 pg/cm², such as from 250 pg/cm² to 400 pg/cm² of said peptide. In some embodiments, for every 1 mg of PLGA in said material, the material comprises at least 1 pg of the peptide as described herein. In other words, each 1 mg of PLGA may associate with at least 1 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 1 to 200 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 5 to 200 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 1 to 100 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 5 to 50 pg of the peptide as described herein.

In specific embodiments, the material is a suture. In some embodiments, for every 1 mg of PLGA in said material, the material comprises at least 2 pg, such as at least 3 pg, such as at least 4 pg, such as at least 5 pg, such as at least 6 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 1 to 10 pg, such as from 1 to 8 pg, such as from 1 to 6 pg, such as from 3 to 6 pg, such as from 5 to 6 pg of the peptide as described herein.

In specific embodiments, the material is a particle, such as a nanoparticle or a microparticle. In some embodiments, for every 1 mg of PLGA in said material, the material comprises at least 20 pg, such as at least 30 pg, such as at least 40 pg of the peptide as described herein. In some embodiments, for every 1 mg of PLGA in said material, the material comprises from 20 to 60 pg, such as from 30 to 55 pg, such as from 40 to 50 pg of the peptide as described herein.

In addition to the polymer and the peptide as disclosed herein, the material according to the present disclosure may also comprise additional therapeutic agents, such as antibiotic, anti-inflammatory or antiseptic agents such as anti-bacterial agents, antifungicides, anti-viral agents, and anti-parasitic agents.

Peptide

The peptides comprised in the materials of present disclosure have useful anti-bacterial and/or anti-inflammatory properties. The inventors have shown that surprisingly the peptide must comprise hydrophobic residues, preferably at specific positions, which may more efficiently integrate into the polymer phase of the PLGA polymer. This particularly relates to the amino acids at positions X1 , X6, and X14. Glycine (G), alanine (A), valine (V), leucine (L), isoleucine (I), proline (P), phenylalanine (F), methionine (M), tyrosine (Y) and tryptophan (W) are examples of hydrophobic amino acids.

It is furthermore preferred that the peptide comprises one or more amino acids, which can be protonated, in particular it is preferred that X2, X4, X5, X9 are R, K or H.

In some aspects, the peptide comprises or consists of the amino acid sequence X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein X3, X7, X10, X11 , X13, X15, X16 is any standard amino acid, X8, X12 is any amino acid,

X1, X6, X14 is G, A, V, L, I, P, F, M. Y or W, and

X2, X4, X5, X9 is R, K or H, wherein said peptide has a length of from 10 to 40 amino acids, such as from 10 to 30 amino acid residues.

As is clear to the skilled person in the art, said peptide may comprise additional amino acids either N-terminally of X1 or C-terminally of X16.

In some embodiments, the peptide comprises or consists of the amino acid sequence X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein X1, X6, X14 is F or W, and

X2, 4, 5, 9 is R or K.

In some embodiments, the the peptide comprises or consists of the amino acid sequence F-R-X3-K-K-W-X7-X8-K-X10-X11-X12-X13-F-X15-X16.

In some embodiments, the the peptide comprises or consists of the amino acid sequence H-V-F-R-X3-K-K-W-X7-X8-K-X10-X11-X12-X13-F-X15-X16.

It is preferred that the peptide is capable of simultaneously binding both to lipopolysaccharides and to the LPS-binding hydrophobic pocket of CD14. In some embodiments, said peptide has a length of 10 amino acids. In some embodiments, said peptide has a length of 11 amino acids. In some embodiments, said peptide has a length of 12 amino acids. In some embodiments, said peptide has a length of 13 amino acids. In some embodiments, said peptide has a length of 14 amino acids. In some embodiments, said peptide has a length of 15 amino acids.

In some embodiments, said peptide has a length of 16 amino acids. In some embodiments, said peptide has a length of 17 amino acids. In some embodiments, said peptide has a length of 18 amino acids. In some embodiments, said peptide has a length of 19 amino acids. In some embodiments, said peptide has a length of 20 amino acids. In some embodiments, said peptide has a length of 21 amino acids.

In some embodiments, said peptide has a length of 22 amino acids. In some embodiments, said peptide has a length of 23 amino acids. In some embodiments, said peptide has a length of 24 amino acids. In some embodiments, said peptide has a length of 25 amino acids. In some embodiments, said peptide has a length of 26 amino acids. In some embodiments, said peptide has a length of 27 amino acids. In some embodiments, said peptide has a length of 28 amino acids. In some embodiments, said peptide has a length of 29 amino acids. In some embodiments, said peptide has a length of 30 amino acids. In some embodiments, said peptide has a length of 31 amino acids. In some embodiments, said peptide has a length of 32 amino acids. In some embodiments, said peptide has a length of 33 amino acids. In some embodiments, said peptide has a length of 34 amino acids. In some embodiments, said peptide has a length of 35 amino acids. In some embodiments, said peptide has a length of 36 amino acids. In some embodiments, said peptide has a length of 37 amino acids. In some embodiments, said peptide has a length of 38 amino acids. In some embodiments, said peptide has a length of 39 amino acids. In some embodiments, said peptide has a length of 40 amino acids.

In some embodiments, the peptide has a length of 10 to 40 amino acids. In some embodiments, the peptide has a length of 13 to 40 amino acids. In some embodiments, the peptide has a length of 16 to 30 amino acids. In some embodiments, the peptide has a length of 18 to 30 amino acids. In some embodiments, the peptide has a length of 18 to 25 amino acids.

In preferred embodiments the peptides is a fragment of thrombin or has as at least 90% sequence identity with a fragment of thrombin. In particular, the peptide may be the peptides set as set forth in SEQ ID NO: 1 or a fragment thereof or a peptide having at least 90% sequence identity with SEQ ID NO: 1 or said fragment thereof.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 1, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 1 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 2, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 2 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 3, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 3 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 4, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 4 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 5, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 5 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 6, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 6 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 7, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 7 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 8, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 8 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 9, or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto. In some embodiments, the peptide comprises or consists of a variant of the amino acid sequence as set forth in SEQ ID NO: 9 wherein any one amino acid has been altered for another amino acid, with the proviso that no more than 5 amino acids have been so altered, for example wherein 5, 4, 3, 2, or 1 amino acid has been so altered in said amino acid sequence.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 1 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 2 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 3 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 4 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 5 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 6 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises or consists of the amino acid sequence as set forth in SEQ ID NO: 7 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises the amino acid sequence as set forth in SEQ ID NO: 8 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In some embodiments, the peptide has a length of 18 to 30 amino acids, preferably 18- 25 amino acids, and comprises the amino acid sequence as set forth in SEQ ID NO: 9 or an amino acid sequence with at least 85% sequence identity thereto, such as at least 90% sequence identity thereto, such as at least 95% sequence identity thereto, such as at least 98% sequence identity thereto, such as at least 99% sequence identity thereto.

In one embodiment the peptide is the peptide as set forth in SEQ ID NO: 1. WO 2021/260164 discloses that SEQ ID NO: 1 can be cleaved into the multiple peptides including FYT21, GKY20 and HVF18, i.e. SEQ ID NO: 2, 3 and 4. Said peptides also include the amino acid sequences necessary for both lipopolysaccharide (LPS) binding and CD14 binding. Accordingly, the peptide may in some embodiments comprise or consists of any of these peptides (TCP-25FYT21 , GKY20 and HVF18). Preferably the peptide has a length of 18-25 amino acids but, as long as the peptide is based on any of these peptides, the peptide may be up to 30 amino acids long.

In some embodiments, the peptide comprises an internal covalent linkage between at least two amino acids. These peptides may have enhanced stability compared to similar peptides of identical sequence but not comprising said internal covalent linkage(s).

Such peptides are described in WO 2023/067167, the contents of which are hereby incorporated by reference in their entirety. In some embodiments, the peptide is as defined in any one of items 1 to 129 on p. 77- 98 of WO 2023/067167. In some embodiments, the peptide comprises or consists of an amino acid sequence as described in the section “Peptide sequence” on p. 28, I. 18 to p. 35, I. 16 of WO 2023/067167. In some embodiments, the peptide comprises a covalent linkage as described in the section “Internal covalent linkage” on p. 14, I. 20 to p. 27, I. 25 of WO 2023/067167.

In some embodiments, the peptide comprises or consists of the sequence: V-F-R-L-K-K-W-I-X1-K-V-I-X2-Z-F-G, wherein Xi and X2 are amino acids linked by a covalent linkage.

In some embodiments, said covalent linkage is a hydrocarbon staple.

In some embodiments, Xi and X2 are alkenylated amino acids. In some embodiments, Xi and X2 are two C-alkenylated amino acids. In some embodiments, Xi and X2 are two □-substituted alkenyl amino acids. In some embodiments, Xi and X2 are a,a- disubstituted alkenyl amino acids. In some embodiments, the covalent linkage is an olefin tether formed between said alkenyl residues.

In some embodiments, the internal hydrocarbon staple is formed by linking two (S)-2- (4’-pentenyl)-alanines.

In some embodiments, one or more of the standard amino acids comprised in the peptide are modified or derivatised.

In some embodiments, one or more of the standard amino acids comprised in the peptide are PEGylated. In some embodiments, one or more of the standard amino acids comprised in the peptide are amidated. In some embodiments, one or more of the standard amino acids comprised in the peptide are acylated. In some embodiments, one or more of the standard amino acids comprised in the peptide are acetylated. In some embodiments, one or more of the standard amino acids comprised in the peptide are alkenylated. In some embodiments, one or more of the standard amino acids comprised in the peptide are alkylated.

In some embodiments, the C-terminal amino acid comprised in the peptide is PEGylated. In some embodiments, the C-terminal amino acid comprised in the peptide is amidated. In some embodiments, the C-terminal amino acid comprised in the peptide is acylated. In some embodiments, the C-terminal amino acid comprised in the peptide is acetylated. In some embodiments, the C-terminal amino acid comprised in the peptide is alkenylated. In some embodiments, the C-terminal amino acid comprised in the peptide is alkylated.

The peptide according to the present disclosure may also be a pharmaceutically acceptable acid or base addition salt of the peptide as disclosed herein above. The acids which are used to prepare the pharmaceutically acceptable acid addition salts of the peptides are those which form non-toxic acid addition salts, i.e. salts containing pharmacologically acceptable anions, such as the hydrochloride, hydrobromide, hydroiodide, nitrate, sulphate, bisulphate, acid, acetate, lactate, citrate, acid citrate, tartrate, bitartrate, succinate, maleate, fumarate, gluconate, saccharate, benzoate, methanesulphonate, ethanesulphonate, benzenesulphonate, p-toluenesulphonate and pamoate [i.e. 1 ,1'-methylene-bis-(2- hydroxy-3 naphthoate)] salts, among others.

Without being bound by theory, it may be beneficial for binding to the polymer that the peptide is also cationic, i.e. has a net positive charge, at neutral pH. Thus, in some embodiments, said peptide is cationic at pH 7.0.

Polymer

The polymer comprised in the materials of present disclosure comprise or consist of poly(lactic-co-glycolic acid) (PLGA), a copolymer. PLGA is a synthetic copolymer of lactic acid (a-hydroxy propanoic acid) and glycolic acid (hydroxy acetic acid).

The skilled person is aware of how to synthesize the PLGA polymer so as to acquire specific, desired properties. For example, a broad spectrum of performance characteristics, such as e.g. solubility, crystallinity, thermal stability, strength, toughness, elasticity, and degradation rate can be controlled by manipulating three key properties of the copolymer: composition (glycolic acid to lactic acid ratio), lactic acid stereoisomeric composition (L- or DL-lactide), and molecular weight of the peptide. Examples of these performance characteristics and associated values for these properties can be found in Avgoustakis, 2005, “Polylactic-Co-Glycolic Acid (PLGA)”, Encyclopedia of Biomaterials and Biomedical Engineering, doi:10.1081/E-EBBE- 120013950. The PLGA polymer may be synthesized as a random or a block copolymer.

In some embodiments, the PLGA polymer is synthesized as a block copolymer. In some embodiments, the monomer of said PLGA polymer is according to formula I:

wherein x is number of units of lactic acid and y is the number of units of glycolic acid.

As indicated above, the value of x and y may be chosen by the skilled person to give particular desired properties to said polymer. In some embodiments, x is 1 and y is 1. In some embodiments, x is 1 and y is 2. In some embodiments, x is 2 and y is 1. In some embodiments, x is 2 and y is 2.

Preferably, the PLGA polymer may be synthesized as a random copolymer by mixing specific molar ratios of the monomers glycolic acid and lactic acid and polymerising these.

By selecting specific ratios between lactic acid and glycolic acid in the PLGA polymers, the release profile from said polymers of the associated peptides as described herein can be controlled. This thus allows for tailored release timing for specific applications.

In particular, a higher ratio of glycolic acid to lactic acid may facilitate faster degradation of the polymer, and therefore faster release of the associated peptide as described herein to the site of application. This may be useful at sites of injury requiring rapid delivery of the associated peptide, such as acute wounds. Conversely, a higher ratio of lactic acid to glycolic acid may cause the polymer to degrade more slowly, resulting in prolonged release of the associated peptide over time. Additionally, a higher amount of lactic acid in the PLGA polymer may protect the associated peptide from exogenous proteases due to the degradation of the polymer being slower. This may be particularly useful in body environments that are characterized by high proteolytic activity, such as wounds. In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 5:95 to 95:5 (said ratio denoting the ratio of glycolic acid to lactic acid). A ratio of 5:95 denotes that for every 5 units of glycolic acid in the PLGA polymer, the polymer comprises 95 units of lactic acid.

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 80:20 to 95:5 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 90:10 to 15:85 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 95:10 (said ratio denoting the ratio of glycolic acid to lactic acid). In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 90:10 to 5:95 (said ratio denoting the ratio of glycolic acid to lactic acid). In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 90:10 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 50:50 (said ratio denoting the ratio of glycolic acid to lactic acid). This particular ratio may result in relatively fast degradation and release of the associated peptide, and may be used for applications where the peptide needs to be released over days to a few weeks.

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 30:70 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 25:75 (said ratio denoting the ratio of glycolic acid to lactic acid). This particular ratio may result in slower degradation and release of the associated peptide, and may be used for applications where the peptide needs to be released over several weeks to a few months.

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 15:85 (said ratio denoting the ratio of glycolic acid to lactic acid). This particular ratio may result in slow degradation and prolonged release of the associated peptide, and may be used for applications where the peptide requires longterm delivery, such as for implants and microspheres for chronic conditions.

Without being bound by theory, the peptide as described herein mainly binds to glycolic acid as compared to lactic acid within the polymer. In some embodiments, a ratio of glycolic acid to lactic of 50:50 is sufficient to achieve maximal binding of said peptide to the polymer, i.e. increasing the amount of glycolic acid compared to lactic acid in the polymer will lead to minimal increase in peptide binding.

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 50:50 to 5:95 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer is polyglactin 910, also known as Vicryl. Polyglactin 910 consists of glycolic acid and lactic acid in a ratio of 90:10 (said ratio denoting the ratio of glycolic acid to lactic acid).

In some embodiments, the PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 93:7 (said ratio denoting the ratio of glycolic acid to lactic acid).

The degradation rate of the polymer may also be influenced by the molecular weight of the PLGA.

In some embodiments, the molecular weight of said PLGA polymer is from 5 to 250 kDa. In some embodiments, the molecular weight of said PLGA polymer is from 5 to 200 kDa. In some embodiments, the molecular weight of said PLGA polymer is from 10 to 150 kDa. In some embodiments, the molecular weight of said PLGA polymer is from 20 to 100 kDa. In some embodiments, the molecular weight of said PLGA polymer is from 30 to 60 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 5 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 10 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 15 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 30 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 40 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 60 kDa. In some embodiments, the molecular weight of said PLGA polymer is approximately 100 kDa.

In preferred embodiments, the PLGA polymer is carboxyl-terminated. Thus, in preferred embodiments, the PLGA polymer is according to formula A:

, (Formula A)

In some embodiments, the PLGA polymer is ester-terminated. Thus, in some embodiments the PLGA polymer is according to formula B:

Without being bound by theory, ester termination adds hydrophobic characteristics to the polymer, making it less water-soluble and thus slowing down the absorption of water. This slow water uptake may affect how quickly the polymer breaks down in biological environments. Ester-terminated PLGA may thus exhibit longer degradation half-life.

Without being bound by theory, when PLGA is hydrolysed to lactic and glycolic acid during degradation e.g. at the site of a wound, this may lower the pH of the microenvironment of said site. The peptide according to the present invention may in some embodiments have increased affinity to bacteria and LPS in such a low pH environment due to protonation, thus providing a better therapeutic effect when delivered with a material comprising PLGA as described herein, such as when coated on a PLGA-comprising suture or PLGA-comprising particle. In addition, stabilization of said peptide during application to the area of treatment, such as a wound, may also be caused by protonation of the peptide due to pH reduction in the microenvironment effected by PLGA.

Binding to PLGA-comprising materials, such as a PLGA-comprising sutures or PLGA- comprising particles may also increase the stability during long-term storage of said peptide.

Medical product

In some aspects of the present disclosure is provided a medical product comprising the material as described elsewhere herein.

In some embodiments, said medical product is selected from the group consisting of a suture, a strip, a film, a stent, a graft, a hydrogel, a nanoparticle, a microparticle, and a dressing, such as a mesh, patch or a bandage.

In some embodiments, said medical product is a suture. In some embodiments, said medical product is a strip. In some embodiments, said medical product is a film. In some embodiments, said medical product is a stent. In some embodiments, said medical product is a graft. In some embodiments, said medical product is a hydrogel. In some embodiments, said medical product is a particle, such as a microparticle or a nanoparticle.

In some embodiments, the material comprising PLGA polymer and a TCP25 peptide according to the invention is a particle, such as a microparticle or a nanoparticle. In some embodiments, said particle has an average diameter of between 100 nm to 100 pm. In some embodiments, said particle has an average diameter of between 100 nm to 50 pm. In some embodiments, said particle has an average diameter of about 100 nm. In some embodiments, said particle has an average diameter of about 500 nm. In some embodiments, said particle has an average diameter of about 1 pm. In some embodiments, said particle has an average diameter of about 5 pm. In some embodiments, said particle has an average diameter of about 10 pm. In some embodiments, said particle has an average diameter of about 20 pm. In some embodiments, said particle has an average diameter of about 30 pm. In some embodiments, said particle has an average diameter of about 40 pm. In some embodiments, said particle has an average diameter of about 50 pm.

The size of the particle may be selected based on the release characteristic desired, allowing a further degree of control over the release profile. For example, smaller particles, with their larger surface-to-volume ratio, exhibit faster hydrolysis and greater surface binding, leading to more rapid modulation of the associated protein’s activity. Larger particles degrade more slowly, allowing for sustained activity of the associated protein.

Said PLGA polymer may be as described elsewhere herein. In some embodiments wherein the material is a particle, the PLGA polymer of said particle may comprise or consist of glycolic acid and lactic acid in a ratio of from 5:95 to 95:5, such as from 5:95 to 95:10, such as 30:70, such as 50:50 (said ratio denoting the ratio of glycolic acid to lactic acid).

The medical product may be any medical product comprising such microparticles or nanoparticles. In some embodiments, said medical product is a strip, a film, a stent, a graft, a spray, a powder, a fibrin glue, a hydrogel or a dressing comprising or coated with microparticles or nanoparticles. For example, the medical product may be a hydrogel comprising said particles or a spray or powder comprising or consisting of said particles. It is preferred that the medical product is compatible with application to a wound, such as at a site of surgery, or is compatible with the insertion of an implant comprising the medical product in the body.

In some embodiments, said medical product is a dressing. In some embodiments, said medical product is a mesh. In some embodiments, said medical product is a patch. In some embodiments, said medical product is a bandage.

The medical product is preferably pharmaceutically acceptable, i.e. not toxic. The medical product may be subjected to conventional pharmaceutical operations such as sterilisation and/or may contain conventional adjuvants such as preservatives, stabilisers, wetting agents, emulsifiers, buffers, fillers, etc. The medical products of the present disclosure are useful for combined treatment or prevention of inflammation and infection, for example for treatment of inflammation associated with an infection in an individual in need thereof. The infection may be caused by a micro-organism. Said microorganism may be selected from the group consisting of bacteria, fungi, virus and protozoa.

In some embodiments, said medical product is anti-microbial. In some embodiments, said medical product inhibits growth of bacteria. In some embodiments, said medical product prevents growth of bacteria.

Said bacteria may be any infectious bacteria. For example, the bacteria may be Gram, negative or Gram positive bacteria. Thus, the bacteria may for example be of a genus selected from the group consisting of Staphylococcus, Enterococcus, Streptococcus, Corynebacterium, Escherichia, Klebsiella, Stenotrophomonas, Shigella, Moraxella, Acinetobacter, Haemophilus, Pseudomonas and Citrobacter. In some embodiments, said bacteria are selected from the group consisting of Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli. In another embodiment, the bacteria are gram negative bacteria.

Said bacteria may even be multi-resistant bacteria. The medical products of the present disclosure are capable of providing an antibacterial effect against several multiresistant bacteria, i.e. bacteria which are resistant to several known antibiotics.

In some embodiments, said medical product is anti-bacterial and/or anti-inflammatory. In some embodiments, said medical product is anti-bacterial. In some embodiments, said medical product is anti-inflammatory. In some embodiments, said medical product is anti-bacterial and anti-inflammatory.

In some embodiments, said medical product inhibits or prevents growth or formation of biofilm. In some embodiments, said biofilm is a bacterial biofilm.

In some embodiments, said medical product reduces endotoxin-induced NF-KB and AP-1 activation and/or TNF-alpha induction. In some embodiments, said medical product reduces endotoxin-induced NF-KB and AP-1 activation. In some embodiments, said medical product reduces TNF-alpha induction. In some embodiments, said medical product reduces endotoxin-induced NF-KB and AP-1 activation and TNF-alpha induction.

The medical product of the present disclosure is capable of continuously releasing the peptide at useful concentrations to the relevant part of the body at which it is applied, in order to exert its anti-microbial, such as anti-bacterial, and/or anti-inflammatory effects.

In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 12 hours when said medical product is contacted with said part of the body. In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 24 hours when said medical product is contacted with said part of the body. In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 36 hours when said medical product is contacted with said part of the body. In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 48 hours when said medical product is contacted with said part of the body. In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 60 hours when said medical product is contacted with said part of the body. In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 72 hours when said medical product is contacted with said part of the body.

The rate of release of said peptide to said part of the body is preferably approximately constant during said period of time.

In some embodiments, the medical product is capable of continuously releasing said peptide to a part of a body for a period of from 12 hours to 100 days when said medical product is contacted with said part of the body.

In some embodiments, said part of the body is a wound.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 75 pg/mL over a period of approximately 24 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 100 pg/mL over a period of approximately 24 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at the most 200pg/mL over a period of approximately 24 hours.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative percentage of from 5% to 60%, such as from 10 to 50% of the theoretical maximum concentration (c_max) over a period of approximately 24 hours.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 100 pg/mL over a period of approximately 48 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 125 pg/mL over a period of approximately 48 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at the most 250 pg/mL over a period of approximately 48 hours.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative percentage of from 10% to 70%, such as from 15% to 60% of the theoretical maximum concentration (c_max) over a period of approximately 48 hours.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 125 pg/mL over a period of approximately 72 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 150 pg/mL over a period of approximately 72 hours. In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative concentration of at the most 300 pg/mL over a period of approximately 72 hours.

In some embodiments, said medical product is capable of continuously releasing said peptide to a cumulative percentage of from 20% to 80%, such as from 25 to 70% of the theoretical maximum concentration (c_max) over a period of approximately 72 hours.

Said cumulative concentration may be measured by a method comprising the steps of: 1. placing said medical product in a transwell insert in the apical chamber of said transwell;

2. adding a volume of elution buffer to the basolateral chamber of said transwell in order to contact the porous filter of said transwell and said medical product with said elution buffer;

3. sealing the surface of said transwell to prevent evaporation of said elution buffer;

4. heating said transwell to a physiologically relevant temperature while shaking;

5. measuring a concentration of said peptide in said elution buffer in said basolateral chamber at a set point in time after performing step 4;

6. optionally, repeating step 5 one or more times at subsequent points in time;

7. calculating a cumulative concentration of the release of said peptide from the concentrations measured in step 5, and optionally step 6.

In some embodiments, said elution buffer is 10 mM Tris at pH 7.4.

In some embodiments, said step of heating and shaking is performed at 37 °C with shaking at 60 rpm.

In some embodiments, said set point in time after after performing step 4 is 12 hours, such as 24 hours, such as 36 hours, such as 48 hours, such as 60 hours, such as 72 hours or more.

In some embodiments, the step of measuring the concentration of the peptide in the elution buffer of step 5 is performed by measuring the absorbance of said elution buffer. In some embodiments said absorbance is measured at 280 nm (A280).

The medical product of the present disclosure may preserve the stability of the comprised peptide during storage. Said storage may be wet or dry storage.

In some embodiments, said peptide is stable for at least 12 months during storage, such as wet or dry storage, of said medical product at room temperature.

In some embodiments, said peptide is stable for least 18 months during storage, such as wet or dry storage, of said medical product at room temperature.

In some embodiments, said peptide is stable for at least 24 months during storage, such as wet or dry storage, of said medical product at room temperature. In some embodiments, said peptide is classified as stable when said peptide is not degraded by more than 15% after storage, such as after wet or dry storage. In some embodiments, said peptide is classified as stable when said peptide is not degraded by more than 10% after storage, such as after wet or dry storage. In some embodiments, said peptide is classified as stable when said peptide is not degraded by more than 5% after storage, such as after wet or dry storage. In some embodiments, said peptide is classified as stable when said peptide is not degraded by more than 2% after storage, such as after wet or dry storage. In some embodiments, said peptide is classified as stable when said peptide is not degraded by more than 1% after storage, such as after wet or dry storage.

In some embodiments, said peptide is stable when stored in an aqueous solution, such as an aqueous buffer, such as 10 mM Tris buffer, pH 5 or 7.

In some embodiments, said peptide is stable when stored in a gel, such as a hydrogel.

In some embodiments, said peptide is classified as stable when said peptide after storage, such as after dry storage, has at least 85%, such as at least 90%, such as at least 95%, such as approximately the same antimicrobial activity as an identical peptide that has not been stored, such as an identical fresh, such as a freshly synthesized, peptide. Said antimicrobial activity may be evaluated by radial diffusion assay using a suitable micro-organism, such as E. coli.

In addition to the material comprising the polymer and the peptide as disclosed herein, the medical product according to the present disclosure may also comprise additional therapeutic agents, such as antibiotic, anti-inflammatory or antiseptic agents such as anti-bacterial agents, anti-fungicides, anti-viral agents, and anti-parasitic agents.

Method of treatment

The medical products of the present disclosure may be for use in a method of treatment. In particular, the medical products of the invention are useful for combined treatment or prevention of inflammation and infection, for example for treatment of inflammation associated with an infection in an individual in need thereof.

In some aspects of the present disclosure is therefore provided the materials as disclosed herein for use in a method of preventing and/or inhibiting inflammation and/or infection in a subject in need thereof, said method comprising contacting said subject with or implanting in said subject the medical product as described elsewhere herein and maintaining said contact for a period of time.

In some embodiments, said contact is maintained for a period of time of at least 1 week. In some embodiments, said contact is maintained for a period of time of at least 2 weeks. In some embodiments, said contact is maintained for a period of time of at least 3 weeks. In some embodiments, said contact is maintained for a period of time of at least 1 month. In some embodiments, said contact is maintained for a period of time of at least 2 months. In some embodiments, said contact is maintained for a period of time of at least 3 months. In some embodiments, said contact is maintained until said medical product is dissolved and/or absorbed.

In some embodiments, the method is for preventing inflammation and/or infection in a subject in need thereof. In some embodiments, the method is for inhibiting inflammation and/or infection in a subject in need thereof. In some embodiments, the method is for inhibiting inflammation and/or preventing infection in a subject in need thereof. In some embodiments, the method is for preventing inflammation and/or inhibiting infection in a subject in need thereof.

During or following surgery, medical products, such as sutures or meshes, are necessary for closing, ligating or approximating internal wounds, such as internal surgical wounds, i.e. deep closures, or for fixing or supporting weakened or damaged internal tissue, such as hernias. The term “internal” wound or damaged tissue denotes injuries on the inside of the body, i.e. not external injuries, such as wounds present on the surface of the skin. These medical products must usually be dissolvable or absorbable by the body of the subject wherein they are implanted, as removal from outside the body is not a possibility. The medical products according to the present disclosure are preferably biodegradable and/or bioabsorbable and exhibit several beneficial properties in regard to tissue healing and/or support, such as being antimicrobial and/or anti-inflammatory, and are therefore useful for these applications.

In some embodiments, the medical product is contacted with an internal part of the body of said subject. In some embodiments, said internal part of the body of said subject is a hernia. In some embodiments, said internal part of the body is intradermal. In some embodiments, said internal part of the body is subcutaneous. In some embodiments, said internal part of the body is intravenous.

In some aspects of the present disclosure is provided a medical product according as described elsewhere herein for use in the prevention and/or inhibition of inflammation and/or infection in a part of the body of a subject.

In some embodiments, said part of the body is a hernia. In some embodiments, said part of the body is intradermal. In some embodiments, said part of the body is subcutaneous. In some embodiments, said part of the body is intravenous.

In some embodiments, the medical product is for use in the prevention of inflammation and/or infection in said part of the body of said subject. In some embodiments, the medical product is for use in the inhibition of inflammation and/or infection in said part of the body of said subject. In some embodiments, the medical product is for use in the prevention of inflammation and/or inhibition of infection in said part of the body of said subject. In some embodiments, the medical product is for use in the inhibition of inflammation and/or prevention of infection in said part of the body of said subject.

In some aspects of the present disclosure is also provided a method of treating a wound in a subject in need thereof, said method comprising contacting said wound with or implanting in said wound the medical product as described elsewhere herein.

In some aspects of the present disclosure is provided a medical product as described elsewhere herein for use in the treatment of a wound in a subject in need thereof.

In some embodiments, the wound is a surgical wound. Surgical wounds are described in more detail in the section “Surgical wounds” herein below.

In some embodiments, the wound is a burn wound. In some embodiments, the wound is a non-healing ulcer. In some embodiments, the wound is a chronic wound. In some embodiments, the wound is selected from the group consisting of a radiation-induced wound, a laser-induced wound and a cryotherapy-induced wound. In some embodiments, the wound is caused by a disorder selected from diabetes, cancer and vasculitis.

In some embodiments, the subject is a mammal. In some embodiments, the subject is a human.

The present methods of treatment concern both humans and other mammal such as horses, dogs, cats, cows, pigs, camels, among others. Thus, the medical products of the present disclosure are for use in both human therapy and veterinary applications.

Surgical wounds

The medical products of the present invention are particularly useful for the prevention or treatment of inflammation and/or infection in surgical wounds.

In some embodiments, said medical product is a suture.

A surgical wound may be caused by a cut or incision in the skin be made by e.g. a scalpel during surgery, such as during e.g. laparoscopy or open surgery. A surgical wound may also be the result of a drain placed during surgery.

Surgical wounds can be classified into one of four categories. These categories depend on how contaminated or clean the wound is, the risk of infection, and where the wound is located on the body.

Class I wounds are categorized as clean wounds. These types of wounds are not infected, do not exhibit any signs of inflammation, and are typically closed. If drainage is required, a closed draining approach is recommended. It is worth noting that Class 1 wounds do not involve the respiratory, alimentary, genital, or urinary tracts. Examples of clean wounds include an inguinal hernia repair or a thyroidectomy.

Class II wounds are categorized as clean-contaminated, which means they have a low level of contamination. These types of wounds involve entry into the respiratory, alimentary, genital, or urinary tracts but only under controlled circumstances. Class III wounds are classified as contaminated and typically result from a breach in sterile techniques or leakage from the gastrointestinal tract. Incisions resulting from acute or nonpurulent inflammation are also considered Class 3 wounds.

Class IV wounds are considered to be dirty or infected. These injuries usually occur from inadequate treatment of traumatic wounds, gross purulence, and evident infections. When tissues lose vitality, it can lead to Class 4 wounds. This is often caused by surgery or microorganisms found in perforated organs.

In some embodiments, the surgical wound is a class I wound. In some embodiments, the surgical wound is a class II wound. In some embodiments, the surgical wound is a class III wound. In some embodiments, the surgical wound is a class IV wound.

Method of production

Methods for the production of peptides are well known in the art.

Peptides may be produced by recombinant methods well known in the art (see e.g. Sambrook & Russell, 2000, Molecular Cloning, A Laboratory Manual, Third Edition, Cold Spring Harbor, New York).

Alternatively, peptides may be chemically synthesized, e.g. by linking multiple amino acids via amide bonds. Typically, peptides are chemically synthesized by the condensation reaction of the carboxyl group of one amino acid to the amino group of another. Protecting group strategies may be used to prevent undesirable side reactions with the various amino acid side chains.

Well known liquid-phase or solid phase peptide synthesis techniques are known to the skilled person (such as standard f-Boc or Fmoc solid-phase peptide synthesis).

The covalently linkage of the side chains of two, non-neighbouring internal amino acids may be introduced by any method known to the skilled person, such as for example by any of the methods described by Li et al., 2020.

Peptides according to the invention can also be ordered from companies specialised in producing custom made peptides, for example from AmbioPharm Inc. (US). Similarly, methods for producing materials comprising peptides and PLGA polymer are also known in the art.

In some aspects, the present disclosure provides a method of producing the medical product as described elsewhere herein, the method comprising a step of submerging a starting material comprising or consisting of PLGA in a coating solution comprising a dissolved peptide as defined elsewhere herein, followed by drying said starting material.

In some embodiments, said starting material is selected from the group consisting of a suture, a strip, a film, a stent, a graft, a hydrogel, a nanoparticle, and a dressing, such as a mesh, patch or a bandage.

In some embodiments, the concentration of said peptide in said coating solution is at least 0.5%. In some embodiments, the concentration of said peptide in said coating solution is at least 1%. In some embodiments, the concentration of said peptide in said coating solution is at least 2%. In some embodiments, the concentration of said peptide in said coating solution is at least 3%. In some embodiments, the concentration of said peptide in said coating solution is at least 4%.

In some embodiments, the concentration of said peptide in said coating solution is from 0.5%-4%. In some embodiments, the concentration of said peptide in said coating solution is from 1%-4%. In some embodiments, the concentration of said peptide in said coating solution is from 2%-4%. In some embodiments, the concentration of said peptide in said coating solution is from 2%-3%.

In preferred embodiments, the concentration of said peptide in said coating solution is approximately 2%, such as 2%.

In some embodiments, the starting material is submerged in said coating solution for at least 10 minutes. In some embodiments, the starting material is submerged in said coating solution for at least 20 minutes. In some embodiments, the starting material is submerged in said coating solution for at least 30 min. In some embodiments, the starting material is submerged in said coating solution for at least 1 hour. In some embodiments, the starting material is submerged in said coating solution for about 2 hours, such as 2 hours.

In some embodiments, the starting material is submerged in said coating solution for from 10 minutes to 4 hours. In some embodiments, the starting material is submerged in said coating solution for from 20 minutes to 3 hours. In some embodiments, the starting material is submerged in said coating solution for from 30 minutes to 2 hours. In some embodiments, the starting material is submerged in said coating solution for from 1 hours to 2 hours.

In some embodiments, the starting material is submerged in said coating solution at a temperature of from 10-50°C. In some embodiments, the starting material is submerged in said coating solution at a temperature of from 15-40°C. In some embodiments, the starting material is submerged in said coating solution at a temperature of from 20- 37°C. In some embodiments, the starting material is submerged in said coating solution at a temperature of from 20-30°C. In some embodiments, the starting material is submerged in said coating solution at a temperature of from 20-25°C.

In preferred embodiments, the starting material is submerged in said coating solution at a temperature of approximately 21°C, such as 21°C.

In a specific embodiment,

• the concentration of said peptide in said coating solution is approximately 2%;

• the starting material is submerged in said coating solution for approximately 2 hours; and

• the starting is submerged in said coating solution at a temperature of approximately 21°C.

In preferred embodiments, the starting material is a suture.

Thus, in some embodiments, the peptide is coated onto the starting material. The term 'coated' as used herein refers to the peptide being applied to the surface of the starting material. Thus, the peptide may be painted or sprayed with a solution comprising the composition. Alternatively, the material may be dipped in a reservoir comprising the peptide. In some embodiments, the starting material is impregnated with a composition comprising the peptide. By 'impregnated' is meant that the composition is absorbed or adsorbed with the starting material.

Items

1. A material comprising a poly(lactic-co-glycolic acid) (PLGA) polymer and a peptide comprising or consisting of the amino acid sequence X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein X3, X7, X10, X11 , X13, X15, X16 is any standard amino acid,

X8, X12 is any amino acid, X1, X6, X14 is G, A, V, L, I, P, F, M. Y or W, and X2, X4, X5, X9 is R, K or H, wherein said peptide has a length of from 10 to 30 amino acid residues.

2. The material according to item 1, wherein the peptide comprises or consists of the amino acid sequence X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein

X1, X6, X14 is F or W, and X2, 4, 5, 9 is R or K.

3. The material according to any one of the preceding items, wherein the peptide comprises or consists of the amino acid sequence F-R-X3-K-K-W-X7-X8-K-X10-X11 -X12-X13-F-X15-X16.

4. The material according to any one of the preceding items, wherein the peptide comprises or consists of the amino acid sequence H-V-F-R-X3-K-K-W-X7-X8-K-X10-X11 -X12-X13-F-X15-X16.

5. The material according to any one of the preceding items, wherein the peptide has a length of 18 to 30 amino acids, preferably 18-25 amino acids, and comprises or consists of any one of the amino acid sequences selected from SEQ ID NO: 1 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 2 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 3 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 4 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 5 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 6 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 7 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 8 or an amino acid sequence with at least 90% sequence identity thereto, and SEQ ID NO: 9 or an amino acid sequence with at least 90% sequence identity thereto.

6. The material according to any one of the preceding items, wherein the peptide has a length of 18 to 30 amino acids, preferably 18 to 25 amino acids, and comprises or consists of SEQ ID NO: 1.

7. The material according to any one of the preceding items, wherein the peptide comprises or consists of the sequence:

V-F-R-L-K-K-W-I-X1-K-V-I-X2-Z-F-G wherein

Xi and X2 are amino acids linked by a covalent linkage.

8. The material according to item 7, wherein the covalent linkage is a hydrocarbon staple.

9. The material according to any one of items 7 to 8, wherein Xi and X2 are alkenylated amino acids, such as two C-alkenylated amino acids, such as two □-substituted alkenyl amino acids and/or a,a-disubstituted alkenyl amino acids, and the covalent linkage is an olefin tether formed between said alkenyl residues.

10. The material according to any one of items 7 to 9, wherein the internal hydrocarbon staple is formed by linking two (S)-2-(4’-pentenyl)-alanines.

11. The material according to any one of the preceding items, wherein one or more of the standard amino acids comprised in the peptide are modified or derivatised.

12. The material according to any one of the preceding items, wherein one or more of the standard amino acids comprised in the peptide are PEGylated, amidated, acylated, acetylated, alkenylated and/or alkylated.

13. The material according to any one of the preceding items, wherein said peptide is cationic at pH 7.0.

14. The material according to any one of the preceding items, wherein the monomer of said PLGA polymer is according to formula I:

15. The material according to item 14, wherein x is 1 and wherein y is 1.

16. The material according to item 14, wherein x is 1 and wherein y is 2.

17. The material according to item 14, wherein x is 2 and wherein y is 1.

18. The material according to any one of the preceding items, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 5:95 to 95:5 (glycolic acid : lactic acid).

19. The material according to any one of the preceding items, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 5:95 to 90:10 (glycolic acid : lactic acid).

20. The material according to any one of items 1 to 18, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 80:20 to 95:5 (glycolic acid : lactic acid). The material according to any one of items 1 to 19, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 15:85 to 90:10 (glycolic acid : lactic acid). The material according to any one of the preceding items, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 90:10 (glycolic acid : lactic acid). The material according to any one of the preceding items, wherein said PLGA polymer is polyglactin 910. The material according to any one of items 1 to 19, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 93:7 (glycolic acid : lactic acid). The material according to any one of items 1 to 19, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 25:75 or 30:70 (glycolic acid : lactic acid). The material according to any one of items 1 to 19, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 50:50 (glycolic acid : lactic acid). The material according to any one of the preceding items, wherein said PLGA polymer is carboxyl-terminated. The material according to any one of the preceding items, wherein said peptide is coated on, dissolved in, and/or interspersed between the fibers of said PLGA polymer. The material according to any one of the preceding items, wherein said material is in the form of a particle, such as a microparticle or a nanoparticle. The material according to item 29, wherein said particle has an average diameter of between 100 nm to 100 pm, such as between 100 nm to 50 pm.

31. The material according to any one of the preceding items, wherein said material comprises at least 25 pg/cm², such as least 50 pg/cm², such as least 75 pg/cm², such as least 100 pg/cm², such as least 150 pg/cm², such as least 200 pg/cm², such as least 250 pg/cm², such as least 300pg/ cm², such as at least 350 pg/cm² of said peptide.

32. The material according to any one of the preceding items, wherein said material comprises at least 1 pg of said peptide for every 1 mg of said PLGA polymer.

33. The material according to any one of the preceding items, wherein said material comprises from 1 pg to 200 pg, such as from 5 pg to 200 pg of said peptide for every 1 mg of said PLGA polymer.

34. The material according to any one of the preceding items, wherein said material comprises from 1 to 10 pg, such as from 1 to 8 pg, such as from 1 to 6 pg, such as from 3 to 6 pg, such as from 5 to 6 pg of said peptide for every 1 mg of said PLGA polymer.

35. The material according to any one of the preceding items, wherein said material comprises at from 20 to 60 pg, such as from 30 to 55 pg, such as from 40 to 50 pg of said peptide for every 1 mg of said PLGA polymer.

36. The material according to any one of the preceding items, wherein said material is anti-inflammatory and/or anti-microbial, such as anti-bacterial.

37. The material according to any one of the preceding items, wherein said material is capable of binding to lipopolysaccharides (LPS), such as bacterial LPS.

38. The material according to any one of the preceding items, wherein said material is biodegradable and/or bioabsorbable.

39. A medical product comprising the material according to any one of the preceding items. 40. The medical product according to item 39, wherein said medical product is selected from a suture, a strip, a film, a stent, a graft, a hydrogel, a nanoparticle, and a dressing, such as a mesh, patch or a bandage.

41 . The medical product according to any one of items 39 to 40, wherein said medical product is a suture.

42. The medical product according to item 39, wherein said medical product is a strip, a film, a stent, a graft, a hydrogel, or a dressing comprising the particle according to any one of items 29 to 30.

43. The medical product according to any one of items 39 to 41 , wherein said medical product is anti-microbial, such as anti-bacterial, and/or antiinflammatory.

44. The medical product according to any one of items 39 to 43, wherein said medical product is anti-microbial, such as anti-bacterial, and anti-inflammatory.

45. The medical product according to any one of items 39 to 44, wherein said medical product inhibits growth of bacteria.

46. The medical product according to item 45, wherein said bacteria are selected from Staphylococcus aureus, Pseudomonas aeruginosa and Escherichia coli.

47. The medical product according to any one of items 39 to 46, wherein said medical product inhibits or prevents growth or formation of biofilm, such as bacterial biofilm.

48. The medical product according to any one of items 39 to 47, wherein said medical product reduces endotoxin-induced NF-KB and AP-1 activation and/or TNF-alpha induction.

49. The medical product according to any one of items 39 to 48, wherein said medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 24 hours, such as at least 48 hours, such as at least 72 hours, when said medical product is contacted with said part of the body. The medical product according to item 49, wherein said part of the body is a wound. The medical product according to any one of items 39 to 50, wherein said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 75 pg/mL, such as at least 100 pg/mL, over a period of approximately 24 hours. The medical product according to any one of items 39 to 51 , wherein said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 100 pg/mL, such as at least 125 pg/mL, over a period of approximately 48 hours. The medical product according to any one of items 39 to 52, wherein said medical product is capable of continuously releasing said peptide to a cumulative concentration of at least 125 pg/mL, such as at least 150 pg/mL, over a period of approximately 72 hours. The medical product according to any one of items 39 to 53, wherein said peptide is stable for at least 12 months, such as for at least 18 months, such as for at least 24 months, during dry storage of said medical product at room temperature. The medical product according to item 54, wherein the peptide is stable when said peptide is not degraded by more than 10%, such as no more than 5%, after dry storage. A method of producing the medical product according to any one of items 39 to 55, the method comprising a step of submerging a starting material comprising or consisting of PLGA in a coating solution comprising a dissolved peptide according to any one of items 1 to 13, followed by drying said starting material. 57. The method according to item 56, wherein said starting material is selected from a suture, a strip, a film, a stent, a graft, a hydrogel, a particle, such as a microparticle or a nanoparticle, and a dressing, such as a mesh, patch or a bandage.

58. The method according to any one of items 56 to 57, wherein said starting material is a suture.

59. The method according to any one of items 56 to 57, wherein said starting material is a PLGA-comprising particle.

60. The method according to any one of items 56 to 59, wherein the concentration of said peptide in said coating solution is at least 0.5%, such as from 0.5-4%, such as at least 1%, such as at least 2%, such as from 2-4%, preferably 1% or 2%.

61. The method according to any one of items 56 to 60, wherein the starting material is submerged in said coating solution for at least 10 minutes, such as at least 20 minutes, such as at least 30 min, such as at least 1 hour or such as at least 2 hours.

62. A method of preventing and/or inhibiting inflammation and/or infection in a subject in need thereof, said method comprising contacting said subject with or implanting in said subject the medical product according to any one of items 39 to 55 and maintaining said contact for a period of time.

63. The method according to item 62, wherein said medical product is contacted with an internal part of the body of said subject, such as a hernia of said subject.

64. A method of treating a wound in a subject in need thereof, said method comprising contacting said wound with or implanting in said wound the medical product according to any one of items 39 to 55.

65. The method according to item 64, wherein the wound is a surgical wound. 66. The method according to any one of items 62 to 65, wherein said subject is a mammal, such as a human.

67. The method according to any one of items 62 to 66, wherein said contact is maintained for a period of time of at least 1 month, such as at least 2 months, or until said medical product is dissolved and/or absorbed.

68. A medical product according to any one of items 39 to 55 for use in the prevention and/or inhibition of inflammation and/or infection in a part of the body of a subject.

69. The medical product for use according to item 68, wherein said part of the body is a hernia.

70. The medical product for use according to item 68, wherein said part of the body is intradermal.

71. The medical product for use according to item 68, wherein said part of the body is subcutaneous.

72. A medical product according to any one of items 39 to 55 for use in the treatment of a wound in a subject in need thereof.

73. The medical product for use according to item 72, wherein said wound is a human wound.

74. The medical product for use according any one of items 72 to 73, wherein said wound is a surgical wound.

Example 1 - TCP-25-coated sutures

Materials and methods

Materials: TCP-25 (SEQ ID NO: 1) peptide was synthesized by Ambiopharm (Spain). The tetramethylrhodamine (TAMRA)-labeled TCP-25 was synthesized by Biopeptide Co. (San Diego, CA, USA). The purity of the peptide was 95% as confirmed by mass spectral analysis (MALDI-ToF Voyager).

Microorganisms: Bacterial strains used included Escherichia coli (ATCC 25922), Pseudomonas aeruginosa PAO1 , and Staphylococcus aureus (ATCC 29213). Bioluminescent P. aeruginosa Xen41 (PerkinElmer, Akron, OH), and S. aureus SAP229 were used for experiments requiring infection imaging. In some experiments, clinical isolates of S. aureus (2404, 2278, 2405, 2528, 1779), P. aeruginosa (27.1 , 23.1 , 13.2, 10.5, 62.1), S. epidermidis (2282), and E. faecalis (2374) were also used. These isolates were obtained either from skin or wound infections at the Department of Bacteriology, University Hospital, Lund, Sweden.

Preparation of TCP-25-coated suture: A Vicryl suture (3-0, ETHICON, Johnson & Johnson International, Belgium) was cut into 10 cm pieces. A 2% TCP-25 solution was prepared by solubilizing the peptide in sterile water. The suture pieces were coated in TCP-25 solution (5 mL) on a shaker for 1 h at room temperature. After coating, the sutures were air-dried in a class II biosafety cabinet at room temperature (20°C ± 1 °C) for 1 hour. Control sutures were coated with only sterile water.

To examine the effects of TCP-25 coating concentration on antibacterial properties, sutures were coated with 0.1%, 0.5%, 2.0%, or 4.0 % TCP-25 solution for 2 h. To examine the effects of coating time on antibacterial properties, sutures were coated with 2% TCP-25 solution for 1 , 2, 4, 8, and 24 h at room temperature. To examine the effects of coating temperature on antibacterial properties, sutures were coated with 2% TCP-25 solution at 21 °C, 37 °C, or 50 °C for 2 h. Coated sutures were kept at -80 °C in moisture-free conditions until further use. To elute peptide, coated sutures were added in Tris (10 mM, pH 7.4) on a shaker for 30 min at room temperature. Eluted solutions were stored at -80 °C. Suture TCP-25 loading was determined as described previously (Champeau et al., 2015). Briefly, sutures were weighed before and after coating with TCP-25. Drug loading was calculated using the following equation:

Drug loading (%) = ((Weight after coating - weight before coating)/weight before coating) x 100. Protein estimation: For the estimation of protein concentration, Nanodrop method (ND 1000, Thermo Scientific) was used at 280 nm using TCP-25 extinction coefficient (8 480 M'¹cnv¹) and molecular weight (3088,62 Da).

Radial diffusion assay (RDA): Ten mL of tryptic soy broth (TSB) from was used to grow E. coli to the mid-logarithmic phase. Afterwards, the bacteria were washed in 10 mM Tris (pH 7.4). Bacteria (4 x 10⁶ CFU) were added to 15 mL of the underlay agarose gel comprising of 0.03% (w/v) TSB, 1% (w/v) low electro endosmosis type (EEC) agarose (Sigma, St Louis MO, USA), and 0.02% (v/v) Tween 20 (Sigma). Underlay gel was added to a 144 mm diameter petri dish. Once the gel became solid, using a biopsy punch, 4 mm-diameter wells were cut into the underlay gel. A sample containing 6 pL of the test sample (solution eluted from the sutures) was added to each well.

Afterwards, plates were incubated at 37 °C for 3 h and 15 mL of overlay gel (6% TSB and 1% low-EEO agarose in distilled H2O) was added to cover underlay gel. Plates were incubated at 37 °C for 18 h. The antimicrobial activity was determined as a clear zone-to-well diameter (excluding the 4 mm well).

SDS-PAGE: To study the effects of storage of TCP-25-coated suture, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was employed to study eluted peptide. Briefly, sample (5 pl) was loaded on a 10%-20% Tris-Tricine gel and run for 90 min at 100 V. Two micrograms of TCP-25 solubilized in 10 mM Tris (pH 7.4) was used as control. Coomassie Brilliant blue (Invitrogen, Rockford, IL, USA) was used to stain the gel.

Western blotting: Immediately following electrophoresis, western blotting was performed as described before (Saravanan et al., 2018).

Peptide release assay: For the peptide in vitro release assay with some modifications, a wound scenario was simulated. A Transwell insert system (VWR® Tissue Culture Plate Insert - 6 well , 0.4 pm pore size, VWR International) was used in combination with a 6-well plate. Twenty 3 cm long pieces of TCP-25-coated sutures were placed on porous filter in the apical compartment. Two mL of elution buffer (10 mM Tris, pH 7.4) was added to the basolateral compartment. The elution buffer stayed in contact with the porous filter and thus with the sutures (as illustrated in Figure 1d). The plate was covered and sealed with parafilm to prevent evaporation of the solution. The plate was kept at 37 °C on a shaker at 60 rpm. A 20 pL sample was taken from the basolateral part at the designated times and instantaneously replaced with 20 pL of new Tris buffer. A spectrophotometer (Nanodrop, Thermofisher) was employed to measure absorbance at 280 nm (A280). The results were represented as cumulative TCP-25 release (mg/mL).

Scanning electron microscopy (SEM): Briefly, the bacteria were incubated for 15 min at 37 °C with coated and uncoated sutures after which they were transferred to new tubes. Sutures were washed two times in 0.1 M Sorensen’s phosphate buffer (pH 7.4). Samples were further processed for SEM as described previously (Stromdahl et al., 2021). For SEM of sutures with biofilm, after 48 h of microtiter plate incubation and suture removal (as described in the biofilm section), sutures were washed in Sorensen's buffer and processed for SEM as mentioned above.

Quartz Crystal Microbalance with dissipation monitoring (QCM-D): QCM-D measurements were performed using a QSense E4 system from Biolin Scientific (Gothenburg, Sweden) with four standard flow modules (QSense, Biolin Scientific), each equipped with silicon dioxide surfaces (QSense, Biolin Scientific, QSX 303 SiO2, 4.95 ± 0.05 MHz, 14 mm diameter, 0.3 mm thickness, 17.7 ng cm^-2 mass sensitivity). Prior to use, cells, tubings and o-rings were thoroughly cleaned with a 2% Hellmanex solution and multiple Milli-Q (MQ) water rinses, combined with bath sonication, followed by rinsing in pure ethanol and subsequent drying by N2 flow. The SiO2 surfaces were sequentially washed in 2% Hellmanex, MQ water and ethanol, then dried with N2, and plasma cleaned (Model PDC-32G, Harrick Plasma, USA) in residual air for 5 minutes. Right after, SiO2 surfaces were incubated in a 0.1 % (w/w) Poly-L-lysine (MW 150-300 kDa, Sigma-Aldrich, Merck, New Jersey, USA) solution in MQ water (pH 9.5), for 30 minutes at room temperature. After that, SiO2 surfaces were thoroughly rinsed with MQ water and dried with nitrogen. One hundred mg of finely cut fiber (50-100 pm length) in 1 mL ethanol was subsequently deposited on poly-L-lysine-functionalized SiO2 surfaces and left incubating overnight at room temperature. This led to solvent evaporation and subsequent fiber immobilization onto the positively charged poly-L- lysine-functionalized surfaces. Finally, fiber-functionalized surfaces were thoroughly rinsed in MQ water and mounted in QCM-D measurement cells. After reaching a stable baseline of frequency (AF) and dissipation (AD) shifts under 0.1 mL min^-1 MQ water flow (controlled through a peristaltic pump), a 0.001% (w/w) peptide solution in MQ water was injected into the measurement chamber at 0.1 mL min^-1 flow rate. Peptide adsorption onto fiber-functionalized sensors was confirmed by frequency changes (AF) of -100 ± 27 Hz and dissipation changes (AD) of (+42 ± 11 )- 10’⁶ with respect to pure MQ water. This was followed by rinsing with MQ water in the absence of peptide. The possible adsorption of the peptide onto the underlying poly-L-lysine surface was ruled out with a control experiment, in which the interaction of peptide with fiber-free Poly-L- lysine-functionalized SiC>2 surfaces was monitored under the same experimental conditions. Measurements were performed at room temperature.

In silico modelling and simulations of Vicryl with TCP-25 (Coarse-grained parameterization of Vicryl): A coarse-grained (CG) model of Vicryl polymer was developed using the Martini 2.2 forcefield (Marrink et al., 2007). Vicryl or polyglactin 910 is a copolymer made of 90% glycolide and 10% lactide. The molecular weight is around 80 kDa, which translates to approximately 600-mer (Chandrasekhar 2017). To simplify the system, we first built an atomic model of a 10-mer polyglactin containing 9 subunits of glycolide and 1 subunit of lactide using the CHARMM-GUI Polymer Builder (Choi et al., 2021) and the CHARMM36m all-atom forcefield (Huang and MacKerell, 2013). The polymer was solvated in a box of TIP3P water molecules and neutralized with 0.15 M NaCI salt. We then performed energy minimization and equilibration following the CHARMM-GUI standard protocol (Jo et al., 2008). A 1 ps production simulation was performed at 310 K and 1 atm using temperature coupling to Nose- Hoover thermostat (Nose, 1984) and isotropic pressure coupling to Parrinello-Rahman barostat (Parrinello and Rahman, 1981). The electrostatic interactions were computed using the particle mesh Ewald (PME) method (Essmann et al., 1995) while the van der Waals interactions were cut off at 1.2 nm with a force-switch smoothing function applied between 1.0 and 1.2 nm. A simulation time-step of 2-fs was used.

To generate the equivalent model for polyglactin in the CG Martini forcefield, we initially followed the mapping scheme derived in a previous study (Pannuzzo et al., 2022).

Each glycolide and lactide subunit is mapped into one bead and represented by the Na bead type. Unlike the previous study, our polyglactin model has a negatively charged carboxyl group at one end of the polymer and a hydroxyl group at the other end. The former is mapped into two small beads of types SQa and SNa, while the latter is mapped into one bead of type P3. The resultant CG model of polyglactin was then solvated with Martini water molecules and neutralized with 0.15 M NaCI salt. Energy minimization was performed using the steepest descent method and a short 100 ps equilibration simulation was conducted. Then, a 1 ps production simulation was conducted using a 10-fs time step. The electrostatic interactions were computed using the reaction field method with a 1.1 nm cut-off, while the van der Waals interactions were cut off at 1.1 nm with a potential shift Verlet scheme. Temperature and pressure were maintained at 310 K and 1 atm by a velocity rescaling thermostat (Bussi et al., 2007) and an isotropic coupling to a Parrinello-Rahman barostat, respectively. The distribution of bonds, angles, and dihedrals from the CG simulation was then compared to the all-atom simulation. The bonded parameters were iteratively modified to match the distribution of the all-atom simulation.

We multiplied the final model to generate 50-mer and 100-mer polyglactin polymers with the same 9:1 ratio of glycolide: lactide. These longer polymers were subject to a similar protocol of energy minimization, equilibration and a 1 ps production run described above. The structure of the polymers at the end of these simulations were then used for a self-assembly simulation to build a large Vicryl polymer aggregate. Thus, we solvated 50 copies of the polyglactin polymers with Martini water molecules and neutralized the system with 0.15 M NaCI salt. The systems were subject to the same energy minimization and equilibration protocols. We then performed a 1 ps simulation at 320 K using parameters described above to allow the polymer to selfaggregate. We then extracted the structure of the polymer aggregates from these simulations for subsequent simulations with TCP-25.

Coarse-grained simulation of Vicryl with TCP-25: The NMR structure of HVF18 (PBD: 5Z5X) (Saravanan et al., 2018) was used as a template to model the TCP-25 peptide. The missing N-terminal residues (GKYGFYT) were built using Modeller version 9.21 (Sali and Blundell, 1993), and the model with the lowest discrete optimized protein energy was chosen. The TCP-25 model was converted to CG representation using the Martini 2.2 forcefield with the standard EINeDYn elastic network model to preserve the secondary structure (Periole et al., 2009). We added 10 copies of TCP-25 peptide into a box containing the Vicryl polymer aggregate made of 100-mer polyglactin generated in the previous step (Figure 2a). The TCP-25 peptides were placed at least 2 nm away from the surface of the polymer and from each other. The system was then solvated with Martini water molecules and neutralized with 0.15 M NaCI. Energy minimization was performed using the steepest descent method. A 100 ns equilibration simulation was conducted whereby positional restraints with force constants of 500 kJ mol^-1 nm^-2 applied to every bead of the polymer and 1000 kJ mol^-1 nirr² applied to every backbone bead of the peptide. Three independent 10 ps production simulations were then performed with different initial velocities. The temperature was maintained at 320 K using the velocity rescaling thermostat (Bussi et al., 2007) and the pressure was maintained at 1 atm using an isotropic pressure coupling to a Parrinello-Rahman barostat (Parrinello et al., 1981). The electrostatic interactions were calculated using the reaction field protocol with a 1 .1 nm distance cut-off, while the van der Waals interactions were truncated at 1.1 nm with a potential shift Verlet scheme. A 10-fs integration time step was used. Using the same protocols, similar simulations were performed for a system with a higher concentration of TCP-25 peptides (20 copies), a shorter chain of polyglactin (50-mer), and lower temperatures (310 and 298 K). All simulations were performed using GROMACS 2022 (Abraham et al., 2015) and visualized using VMD (Humphrey et al., 1996).

Optical photothermal infrared microspectroscopy: Optical photothermal infrared microspectroscopy (O-PTIR) is an analytical technique based on the photothermal effect induced by scanning infrared laser measured by scattered probe light (Klementieva et al., 2020). O-PTIR were performed at the SMIS beamline, SOLEIL synchrotron (France). Sutures were deposited directly on the glass slide and used for measurements. The photothermal effect was detected by modulating the CW 532 nm laser intensity induced by an infrared laser. The IR source was a pulsed, tunable quantum cascade laser, set to 22 % of laser intensity, scanning from 1800 to 1300 cm^-1, at an 80-kHz repetition rate. Further details about the fundamentals of the technique and the instrument itself can be found in previous work (Paulus et al., 2022).

Cryosectioning: To facilitate fluorescence imaging of TCP-25, Vicryl sutures (3-0, ETHICON, Johnson & Johnson International, Belgium) were coated as described before (section 4.3) except that TCP-25 (spiked with 5% TCP-25-Cy3) was used for the coating. Coated sutures were then mounted in OCT compound for cryosectioning. Suture cryosections (8 pm thick) were prepared using a cryostat (Leica Biosystems). Slides were washed in PBS (5 min, RT), dried, and mounted with antifade mounting medium (PermaFluor, ThermoFisher Scientific). Sections were then imaged using fluorescence microscopy (AxioScope.AI , Carl Zeiss, Germany). High-Performance Liquid Chromatography: One or two g of TCP-25 eluted from the suture was analyzed by reverse-phase high performance liquid chromatography (HPLC) as reported by Petruk et al, 2020. Two micrograms of TCP-25 freshly dissolved in 10 mM Tris at pH 7.4 was used as a control. Samples from three different elutions were analyzed.

Analysis of intrinsic fluorescence: The binding of LPS to TCP-25 was analyzed by determining the intrinsic fluorescence of the peptide as reported previously (Stromdahl et al., 2018). Ten pM TCP-25 eluted from the suture was titrated with increasing LPS concentrations (2-100 pg mL^-1). Kd was computed using GraphPad Prism v9 assuming a single binding site.

Circular dichroism (CD) spectroscopy: The secondary structure of TCP-25 eluted from suture was evaluated by Jasco J-810 spectropolarimeter (Jasco, USA). The spectropolarimeter had a Jasco CDF-426S Peltier set to 25 °C. The cell path length was 0.2 cm. The spectrum was acquired between 190 and 260 nm (20 nm min^-1 scan speed). TCP-25 (10 pM, 200 uL) alone or with 100 pg mL^-1 LPS was used for the experiment. Acquired spectra were corrected for buffer contribution with or without LPS and converted to mean residue ellipticity, Q (mdeg cm ² dmol^-1). Content of cr-helical structure was calculated as reported previously (Morrisett et al., 1973). The experiment was performed three times using a new elution of TCP-25 each time.

Viable-count assay: Viable-count assay was performed as reported previously (Saravanan et al., 2018). Briefly, bacteria were incubated at 37 °C for 5 min and 1 and 2 h with sutures coated in TCP-25 or control sutures under various conditions. Buffers used in this assay were 10 mM Tris with 5 mM glucose, pH 7.4 and 10 mM Tris with 1.3% glycerol, pH 7.4 supplemented with 20% human plasma or 20% acute wound fluid. Samples obtained after serial dilutions of were plated on TH broth agar and kept overnight in incubator at 37 °C.

Antibacterial effects of TCP-25 suture on bioluminescent bacteria: Bioluminescent bacteria were grown to in TH medium until OD 0.4. Bacteria were then washed in 10 mM Tris pH 7.4. and suspended in 10 mM Tris with 5 mM glucose, pH 7.4. or 10 mM Tris with 1.3% glycerol. The bacterial suspension (10⁷ CFU in 200 ml) and 1 cm long TCP-25-coated or control sutures were placed in a white polystyrene 96-well plate. The Plate was kept at 37 °C in an incubator. The bioluminescent signals were longitudinally imaged by I VIS (PerkinElmer, USA) and quantified by a luminometer.

Live-dead assay: Live-dead bacterial staining was performed as described previously (Puthia et al., 2020). Briefly, a 1 cm long piece of TCP-25-coated or control suture was added to 200 pL bacterial P. aeruginosa (PAO1) or S. aureus (ATCC 29213) suspension and incubated for 30 min at 37 °C. Fifty pL of mixture of component A and B were added to the samples and then incubated at room temperature for 15 min in the dark. Suture pieces were taken out of the stained suspension, placed on a slide and analyzed by fluorescence microscopy.

NF- KB/AP-1 assay: THP1-Xblue TM - CD14 reporter cells (InvivoGen, San Diego, USA) were used to study nuclear factor kappa beta/activate protein (NF-KB/AP-1 ) activation. Assay was performed as reported previously (Saravanan et al., 2018). Briefly, 180 pL of 1 x10⁶ cells were seeded into 96-well plates and treated with 10 pM TCP- 25 eluted form suture, with LPS (from E. coli O111 :B4, Sigma-Aldrich), or with 5 pL of human wound fluid. After incubation for 20 h at 37 °C in 5% CO2, 20 pL of media from each well was moved in a new 96-well plate having 180 pL QUANTI-Blue reagent (InvivoGen). Plates were then incubated at 37 °C for 1 to 2h. The amount of secreted embryonic alkaline phosphatase (SEAP) was measured at OD 600 nm.

Cell viability assay: The viability of THP-1 cells from the abovementioned assay was assessed as described previously (Saravanan et al., 2018).

Cytokine assay: Mice from suture-induced inflammation model were euthanized 24 h after suture implantation. Implanted sutures were recovered from the mouse, added to a pre-chilled Eppendorf tube. Fifty microliters of Tris buffer was added to the suture and vortexed for 10 min for elution. Finally, the tube was centrifuged (2000 *g at 4 °C, 5 min), and the supernatant was collected for cytokine analysis. Tumor necrosis factor alpha and interleukin 6 (TNF-a and IL-6, respectively) were assessed using the Mouse Inflammation Kit (Becton Dickinson AB, Franklin Lakes, NJ) as described by the manufacturer.

Hemolytic activity: Fresh venous blood was collected from healthy donors in lepirudin tubes (50 pg mL^-1). One or 10 cm of suture coated with TCP-25 were placed in tubes containing 0.5 or 1 mL, respectively, of 25% of human blood diluted with RPMI-1640- GlutaMAX-l without phenol red (Gibco). The hemolytic activity of only the suture was analyzed by putting 1 or 10 cm suture coated with only buffer. Blood (25%) in RPMI was used as a negative control. The positive control was obtained by mixing 75 pL of blood solution with 225 pL 5% Tween-20 in RPMI-1640- GlutaMAX-l without phenol red. Samples were incubated for 1 h at 37 °C (5% CO2) and tubes were centrifuged at 800 xg. One hundred pL of each sample were transferred to a flat-bottom 96-wells plate, and absorbance was measured at 450 nm. The percentage of hemolysis was calculated as reported previously (Stromdahl et al., 2021).

In vivo peptide release in mice: SKH-1 hairless 10-12-week-old male mice were used for the TCP-25 in vivo release study. Mice were anesthetized using 4% isoflurane (Baxter). All procedures were performed under aseptic conditions. Sutures coated with TAMRA-labeled TCP-25 were used for fluorescence bioimaging. With the help of a needle, a 2 cm piece was subcutaneously implanted into the back of hairless SKH-1 mice. TCP-25 release was longitudinally observed by acquiring fluorescence intensity using an I VIS imaging system (Perkin Elmer). Data were analyzed with Living Image 4.0 Software (PerkinElmer).

Experimental mouse model of suture infection: SKH-1 hairless mice (8-10 weeks old females) were used for the experimental model of suture infection. Anesthesia was achieved using a mixture of isoflurane (Baxter) for induction (4%) and maintenance (2%). All procedures were performed under aseptic conditions. The dorsum of the mouse was cleaned with an ethanol wipe and wiped with sterile gauze. A 5 mm incision was made on the dorsum’s skin, and the tip of the scissors was used to make a small pocket. A 2 cm long piece of TCP-25 suture or control suture was deposited in the pocket with the help of a needle on each side of the dorsum. With the help of a pipette, in the pocket, sutures were contaminated with bioluminescent S. aureus (SAP229) or Escherichia coli (ATCC 25922) (10⁵CFUs in 20 .L Tris buffer). The incision was closed with the help of tissue glue.

In some experiments, in addition to the bacterial visualization, to monitor the tissue distribution of TCP-25, sutures coated with TAMRA-labeled TCP-25 were used followed by I VIS imaging in both bioluminescence (for bacteria) and fluorescence (for TCP-25) modes. NF-KB reporter mouse model of suture-induced inflammation: BALB/c tg(NFi<p-RE- Luc)-Xen reporter mice (Taconic Biosciences, Albany, NY, USA) were used to evaluate the anti-inflammatory effects of TCP-25-coated sutures. Male mice (8-10 weeks old) were used in this study. As described above for the suture infection model, under aseptic conditions, a 5 mm incision was created on the skin of the mouse’s dorsum, and a small pocket was made. A 2 cm long piece of TCP-25 suture or control suture was deposited in the pocket with the help of a needle on each side of the dorsum. Using a pipette, in the pocket, sutures were contaminated with 2 pg LPS (in 20 pL Tris buffer). The incision was closed with the help of tissue glue. For inflammation analysis, longitudinal in vivo imaging with I VIS was employed to determine NF-KB activation. One hundred pL of D-luciferin (PerkinElmer, 150 mg kg^-1 body weight) was intraperitoneally injected 15 min before I VIS imaging. Bioluminescence were quantified using Living Image 4.0 Software (PerkinElmer). In the experiment where TCP-25- coated sutures were contaminated with human chronic wound fluid, the abovementioned procedures were followed except that instead of LPS, 10 pL of wound fluid was used to contaminate sutures in the subcutaneous pocket.

Nano LC-MS/MS analysis: For peptide digestion, 1cm long piece of TCP-25-coated suture was incubated with HNE (0.1 pg, in 20 pL 10 mM Tris, pH 7.4) at 37°C for 30 min and 3 h. HNE-digested TCP-25 peptides were separated with nanoflow reversed- phase chromatography using an Evosep One liquid chromatography (LC) system (Evosep) after loading the samples on Evosep tips. Separation was performed with the 60 SPD method (gradient length 21 min) employing an Evosep column (8 cm x 150 pm ) packed with ReproSil-Pur C18-AQ particles (1.5 pm). The Evosep One system was coupled to a capacitive source mounted on a timsTOF Pro mass spectrometer (Bruker Daltonics). The mode used to run the instrument was DDA PASEF mode. The raw files were searched against the Human Uniprot database (release 2021-03-09) using PEAKS Pro version with the following settings, MS tolerance 30 ppm, MSMS 0.02 Da, no enzyme, oxidation of methionine (variable), and maximum one post-translational modification per peptide.

Biofilm studies: To study the antibiofilm activity of TCP-25-coated sutures, biofilms of S. aureus (ATCC 29213) and P. aeruginosa PAO1 were used. To study the direct effect of TCP-25 suture on mature biofilms, S. aureus biofilms were grown on 96-well round- bottomed vinyl flexible plates (Corning, Kennebunk, USA). Growth media (100 pL),

O.5% Tryptic soy broth, and TBS, supplemented with 0.2% glucose, was added to each well. Five pl of 1 xio⁸ CFU mL^-1 bacteria were added to the growth medium. Similarly,

P. aeruginosa were grown in M63 growth medium supplemented with 0.5% casamino acids, 0.2% glucose and 1 mM MgSC>4, on the flat bottom 96 well microplates (Greiner Bio-One, Frickenhausen, Germany). For both bacterial strains, 5 pL of 1 xio⁸ CFU mL^-1 bacteria were added to the growth medium. After bacterial addition, plates were sealed with microplate seals and placed in moist containers to prevent evaporation. The containers were then incubated at 37 °C undisturbed for 48 hours to achieve mature biofilms. The mature biofilm in the well was washed (100 pL of Tris) twice to remove planktonic cells, after which an additional hundred pL of Tris was added to the well. For treatment of the biofilm, 1 cm long pieces of TCP-25 suture or control suture were then added to the wells. The plate was sealed, placed in the moist container, and incubated at 37 °C for an additional 2 hours. After treatment, to count viable bacteria, biofilm was disrupted by scraping using a pipette tip. A 10 pL aliquot was then removed from each well, serially diluted, and plated for CFU determination. For visualization of the treatment effects on biofilm-associated bacteria, biofilms were removed from the wells by scraping and stained with the Live/Dead bacterial viability Kit (ThermoFischer Scientific). Mixture of component A and B (0.3 pL each) were added to the biofilm samples and mixed. Samples were incubated in the dark (15 min, room temperature) and 10 pL were placed on a glass slide and viewed by fluorescence microscopy.

To evaluate the impact of TCP-25 suture on biofilm growth in the microtiter plate and on the suture itself, 30 min after the addition of bacteria in the growth medium for biofilm formation, a 1 cm piece of TCP-25 suture was added to the bottom of the well. As mentioned above, this was followed by incubation (37 °C, 48 h). Suture pieces were removed after incubation, and wells were washed (100 pL of Tris) twice to remove planktonic cells. Extracted suture pieces were processed further for either staining with the Live/Dead bacterial viability kit, viable count for CFU determination, or SEM. CFU counts of the suture-associated biofilm were conducted by placing the suture into tubes containing 100 pL of Tris buffer. Samples were then sonicated (1 min x 3 times) to disrupt the biofilm. Samples were taken and processed according to the previously mentioned procedure for viable count assay. For the Live/Dead analysis, the above- mentioned protocol was used with the modification of the sutures being placed into the premixed stain to avoid disruption of the biofilm by vortexing. To assess the biomass of the wells, the plate was further processed by washing each well twice with distilled water, before adding 150 pL of 1% crystal violet. The plate was incubated for 15 minutes after which the wells again were rinsed in distilled water. 200 pL of 96% ethanol was then added to the wells for an additional 15 min before 120 pL from each well was transferred to a fresh microtiter plate and analyzed at ODeoo to determine the absorbance in the wells.

Tensile strength determination: The tensile strength of the sutures was analyzed using Instron® 8511.20 (Instron Corp). Ten cm long suture pieces were mounted on the sample holder by turning the thread around a cylindrical bolt three times followed by three single knots to secure the suture. The same process was repeated on both ends of the testing jig, and the instrument was moved using a hydraulic controller to ensure that no relaxation had occurred in the string, and at the same time care was taken to not load the threads to a force of > 1 N. The distance from the center of the cylindrical bolts was measured using a Vernier caliper and assigned as L0 or the original length. The samples were then axially pulled at a predefined ramp speed of 0.25 mm/sec, and the failure force and displacement (L1) were recorded digitally using a 250N load cell. Maximum force at breaking was obtained from the force vs. displacement data.

Furthermore, % elongation was calculated using the formula: %Elongation = (L1-L0)/L0 x 100, where L1 = displacement at failure, and L0 = Original length of the thread before the test was started (measured as the distance between the center of the holding bolts). Data acquisition was done with MTS FlexTest 40 Controller, MTS TestSuite Multipurpose Elite Software.

Wound fluid from patients: Wound fluids from chronic venous leg ulcers and acute wounds were used in this study and the collection method has been described previously (Lundqvist et al., 2004). Wound fluids used for the experiments were collected from patients with positive P. aeruginosa and S. aureus cultures. Briefly, using a tabletop centrifuge, wound fluids were centrifuged at 10,000 rpm. Aliquots were prepared and stored at -20 °C until further used.

Data analyses: For normally distributed data, Student’s t test was used to determine differences in the mean between two groups and Mann-Whitney test was used otherwise. For more than two groups, means were compared using a one-way ANOVA with post hoc (Tukey) for normally distributed data or Kruskal-Wallis test otherwise. Data are presented as means ± SEM. Individual figure legend includes details of statistical analysis used for the experiment. Data analysis was accomplished by using GraphPad Prism software v8. P values <0.05 were considered to be statistically significant.

Ethics statement: Experiments included in this study were accomplished according to Swedish Animal Welfare Act SFS 1988:534. Approval was obtained from the Animal Ethics Committee of Malmd/Lund, Sweden (permit numbers M252-11, M 131 -16, M88- 91/14, M5934-19, 8871-19, M5935-19, 8643-20). Human wound fluid material use was approved by the Ethics Committee at Lund University (LU 708-01 and LU 509-01). All donors have given informed consent. Human blood use was approved by the Ethics Committee at Lund University (permit no. 657-2008).

Results and discussion

Characterization of peptide release from TCP-25-coated polyglactin sutures

In order to retain the antimicrobial and anti-inflammatory effects of TCP-25 we decided to use a reductionist approach, avoiding added formulation components for this proof- of-concept work. Hence, TCP-25 was used for coating of polyglactin sutures using a simple method involving submerging the suture in TCP-25 peptide solution followed by drying. To determine optimal coating conditions, sutures were kept in a 0.1%, 0.5%, 2.0%, and 4.0% TCP-25 solution for 2 h and subsequently air dried. The peptide was then eluted from the sutures, and peptide concentration was determined. Sutures coated with 2 % TCP-25 solution showed significantly higher peptide recovery (Figure 1a, left panel). Sutures coated with 4% TCP-25 solution did not exhibit better TCP-25 recovery than those coated with a 2% solution, possibly due to oligomerization of TCP- 25 molecules at high concentrations, as reported previously (Petruk et al., 2020).

Radial diffusion assay (RDA) was used to analyze the release and antimicrobial activity of the eluted peptide, and the results showed that sutures coated with 2 % TCP-25 showed the highest antimicrobial activity (Figure 1a, right panel). A TCP-25 coating concentration of 2% was therefore found to be optimal. Next, the influence of coating time and temperature on the suture peptide content and antibacterial capacity was investigated. Coating times of 1 , 2, 4, 8, and 24 h all yielded similar TCP-25 recovery from the sutures (Figure 1b, left panel). Correspondingly, using RDA, the zones of inhibition for 1, 2, 4, and 8 h coating times were determined. Although the differences between the time points were small, the zone of inhibition for the 2-hour coating time was relatively higher than other coating times (Figure 1b, right panel). Sutures coated at temperatures of 21 °C showed relatively higher TCP-25 recovery than the sutures coated at 37 and 50 °C (Figure 1c, left panel).

All coating temperatures showed no apparent effect on the antibacterial activity as similar zones of inhibition were observed in RDA (Figure 1c, right panel). Upon SDS- PAGE analysis of the peptide eluted from the sutures, no degradation of TCP-25 was observed for the various coating times and temperatures. Moreover, suture TCP-25 loading was investigated at coating concentrations of 0.5%, 2.0%, and 4.0%. A coating concentration of 0.5% TCP-25 solution resulted in low loading efficiency, while a 2% coating concentration showed significantly higher suture TCP-25 loading efficiency. Interestingly, a 4% coating concentration exhibited suture TCP-25 loading efficiency similar to that of a 2% coating concentration. Based on the results, a 2% coating concentration for 2 hours at 21 °C was selected as the final coating condition for the experiments in this project.

Next, to assess peptide release from the suture, we first used an in vitro model (Del Amo et al., 2019). A Transwell filter insert system was employed, and TCP-25 sutures were kept on the porous filter in the apical compartment. Elution buffer was added to fill the basolateral compartment. The elution buffer was in contact with the filter and the TCP-25 suture (as illustrated in Figure 1d). After a quick release for the initial 24 h, continuous TCP-25 release was observed for 72 h (Figure 1d). Hairless SKH-1 mice were used to further study the in vivo release of TCP-25 from sutures. Sutures were coated with TAMRA-labeled TCP-25 and subcutaneously implanted. Longitudinal fluorescent bioimaging was then performed using the I VIS spectrum in vivo imaging system. Release of the peptide was observed in the immediate implant surroundings and most of the peptide appeared to be retained locally in and around the suture (Figure 1e). TCP-25 TAMRA fluorescent intensity was observed after 1 h and which gradually decreased over time, but significant signals were acquired even 72 h after the implantation (Figure 1e, bar chart). Finally, surface morphological characteristics of the uncoated control and TCP-25-coated sutures were studied by scanning electron microscopy (SEM). A typical braided structure of polyglactin sutures was observed in both control and coated sutures (Figure 1f). No apparent differences were observed on the surface of TCP-25-coated sutures in comparison to the surface of control sutures.

Characterization of peptide-polyg lactin interactions using QCM-D, in silico modelling and simulation, and O-PTIR analysis

To investigate the interaction of TCP-25 with Vicryl suture fibers, Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D) was employed. Finely cut suture fibers (50-100 pm) were dispersed into ethanol, and subsequently drop-cast onto SiO2- based substrate for QCM-D measurements. To allow precise monitoring of peptide binding to the suture fibers, the underlying SiO2 surface was modified by poly-L-lysine under low ionic strength, resulting in a thin and net positively charged surface (Ringstad et al., 2006), which has previously been demonstrated to display low adsorption of a range of host defense peptides (Malekkhaiat Haffner et al., 2019).

Indeed, the cationic poly-L-lysine surface coating was efficient in suppressing the adsorption also for TCP-25, despite the peptide concentration corresponding to plateau in the binding isotherm for other surfaces (Singh et al., 2013) (Figure 2a). In stark contrast, the fiber-coated surfaces displayed very pronounced peptide binding, as seen from the strongly decreasing frequency change (AF). Thus, a frequency shift of -100 ± 27 Hz was observed for the fiber-coated surfaces, whereas a mere -4 ± 1 Hz was observed for the underlying poly-L-lysine surface. While a quantitative interpretation of these results in terms of the amount of peptide bound to the fiber is not possible due to the uncertainty of the fiber surface area, as well as of the amount of solvent oscillating with the crystal during measurement, it is nevertheless clear that the peptide binding to, and affinity for, the suture fibers is high, much higher than typically found for host defense peptides binding to bare SiO2 surfaces (Lozeau et al., 2018).

Finally, we note that the kinetics for peptide binding is relatively slow (in line with the release results discussed above), and that initiation of rinsing after peptide loading does not result in an immediate peptide release; both effects suggest that peptide binding occurs not only on the outer surface of the fibers but also in pores and defects.

To understand the molecular mechanism underlying the interaction between TCP-25 with polyglactin sutures, we built a coarse-grained (CG) model of polyglactin 910 copolymer comprised of 90% glycolide and 10% lactide subunits; parameters were developed within the Martini forcefield framework based on atomic-resolution sampling. We simulated the spontaneous self-aggregation of the polymers made of 100-mer chains, and then added 10 copies of TCP-25 peptides.

The peptides rapidly adsorbed to the surface of the polyglactin polymer (Figure 2b). In less than 200 ns, all peptides in the system were interacting with the polymer (Figure 2c, top) leading to progressive burial of the polymer’s solvent accessible surface area (SASA) (Figure 2c, bottom). Contact analysis indicates that the negatively charged carboxyl termini of the polyglactin polymer chains interact the most with the cationic peptides (Figure 2d, top). Interestingly, we observed that once the peptides have adsorbed to the outer surface of the polymer, they become entangled with the polymer chains and gradually integrate into the polymer phase (Figure 2e), in agreement with the QCM-D results described above. This is evidenced by the dramatically reduced SASA of the TCP-25 peptides during the simulations. Polar (Y3, Y6, H8) and hydrophobic (F5, F10, W15, F23) residues play an important role in peptide integration into the polymer phase (Figure 2d, bottom).

At a higher peptide concentration, we saw slower overall peptide adsorption, likely due to peptide aggregation in solution prior to binding, which aligns with the experimental results obtained using different coating concentrations above. At shorter chain length (50-mer), we found no difference in the ability of the peptides to integrate into the polymer phase. At physiological temperature (310 K) and room temperature (298 K), we found marginally slower rates of peptide absorption; however, the peptides reached a similar SASA value by the end of the simulations, indicating that lower temperatures do not significantly affect the ability of the peptide to be integrated into the polymer. We also found that the binding of peptides to the polyglactin polymer displaced Na⁺ ions on the polymer surface, further highlighting the interaction between the peptides and the carboxyl groups of the polymer. While we observed aggregation of peptides in solution and on the surface of the polymer, the peptides disaggregated once integrated into the polymer phase.

Collectively, our results agree with previous experimental studies of poly-lactic-co- glycolic acid (PLGA) interaction with cationic peptides, whereby the peptides not only bind on the surface, but can also be integrated and distributed within the polymer phase (Giles et al., 2013). This may explain the retained bioactivity upon long-term storage of TCP-25 sutures.

To further validate the interaction of TCP-25 with Vicryl sutures, Optical Photothermal Infrared (O-PTIR) spectroscopy measurements were performed on uncoated and TCP- 25-coated sutures. Spectral analysis shows the presence of a new band at 1665 cm^-1 in coated sutures indicating the presence of TCP-25 on the fibrils (Figure 2f and g). Importantly, 30 min washing of sutures in water does not significantly affect the newly formed band further supporting the interaction of TCP-25 with the suture. Finally, fluorescence imaging of cryosections of sutures coated with TCP-25-Cy3 revealed the distribution and localization of TCP-25 on suture fibrils. TCP-25 appears to be intensely distributed on the surface and in between Vicryl fibrils.

Peptide structure and function analysis

Interaction of peptides with other materials might have deleterious effects on their structure and function. To study if the coating of polyglactin suture with TCP-25 affects peptide structure and activity, we analyzed TCP-25 eluted from the coated sutures using high-performance liquid chromatography (HPLC). No difference in the main peptide elution pattern was observed, indicating that the coating procedure had no apparent effect on peptide stability (Figure 3a). The capability of TCP-25 to bind to LPS and neutralize it is paramount to its anti-inflammatory activity. We further investigated whether TCP-25 retains its capability to interact with and bind to LPS after coating. The peptide was eluted from the coated suture, and in the presence of LPS, structural changes in the presence of LPS studied with circular dichroism (CD) analysis. Results indicated that TCP-25 eluted from the coated sutures showed a similar change in a- helix induction as observed for the control peptide, a finding that indicates compatibility with the interaction between TCP-25 and LPS (Figure 3b). Upon excitation at 280 nm, alteration in peptide’s intrinsic fluorescence was determined by the LPS-induced structural change. LPS-peptide binding was observed and indicated by a blue-shift in emission maximum (X_max) (Figure 3c) as determined by fitting the A_max of TCP-25 in the function of varying concentrations of LPS. A Kd of 11 .15±1 .76 .g/mL was observed which is in agreement with previous results (Figure 3d) (Stromdahl et al., 2021). Antibacterial properties and efficacy of TCP-25-coated sutures in vitro and in an experimental mouse model of suture infection

We used several in vitro assays to determine the antimicrobial efficacy of the TCP-25- coated sutures. Bioluminescent S. aureus and P. aeruginosa were used to demonstrate antibacterial efficacy. Compared to control sutures, bacteria treated with TCP-25-coated sutures showed a significant reduction in bioluminescence as visualized by IVIS bioimaging. This effect was already observed 5 minutes after addition of the TCP-25 sutures, showing the release and rapid antibacterial effects of TCP-25 (Figure 4a).

In another experiment using bioluminescent bacteria, with a luminometer, luminescence was measured after the addition of sutures. A rapid and consistent decrease in bacterial luminescence was observed after the addition of TCP-25-coated sutures (Figure 4b). Next, a live-dead assay was employed to study membrane integrity. The analyses showed that only the TCP-25-coated sutures yielded significant permeabilization of S. aureus and P. aeruginosa bacteria (Figure 4c). SEM was used to further study the effects of TCP-25-coated sutures on bacterial morphology. Bacteria adhering to TCP-25-coated sutures showed significant morphological changes, such as bacterial lysis and clumping, and the presence of debris was observed (Figure 4d). In contrast, smooth and normal cell wall surfaces were observed on bacteria in the control suture group. Overall, these results show that TCP-25 holds its rapid antibacterial activity even after being coated on the polyglactin sutures.

Next, to answer whether TCP-25-coated sutures maintain antibacterial efficacy in vivo, we employed an experimental mouse model of suture infection mimicking a situation of clinical SSIs. S. aureus was chosen for in vivo experiments due to its significant relevance to SSIs (Saleh and Schmidtchen, 2015). Control or TCP-25-coated sutures were subcutaneously implanted into the left or right side of the BALB/c mice and contaminated with bioluminescent S. aureus. IVIS spectrum was used for non-invasive longitudinal in vivo bioimaging of the infection. A significant decrease in bioluminescence intensity was observed at the TCP-25-coated suture side when compared with the control side (Figure 4e). Bioluminescence measurement showed that bacterial infection persisted around the control suture site, whereas TCP-25- coated sutures kept the infection significantly low throughout the experiment and even until the last observed time point of 72 h (Figure 4f). At the 72 h end-point, analysis of the tissue around the suture site showed that TCP-25-coated sutures caused a significant reduction in numbers of bacteria (Figure 4g).

To visualize the distribution of TCP-25 and further confirm that antibacterial effects were indeed due to the TCP-25-coated suture, fluorescently-labeled TCP-25 (TCP-25- TAMRA) was used for the coating, then subcutaneously implanted, and contaminated with S. aureus. We observed that TCP-25-coated sutures inhibited bacterial growth and TCP-25 fluorescence co-localized with the suture site (Figure 4h).

To investigate further if TCP-25 coated sutures exhibit in vivo antibacterial efficacy against Escherichia coli, sutures were contaminated with CLSI control strain E. coli (ATCC 25922), and tissue around the suture site was analyzed at 72 h. CFU determination showed that TCP-25-coated sutures caused a significant reduction in number of bacteria.

In vitro and in vivo effects of TCP-25-coated suture on endotoxin responses We further investigated the anti-inflammatory efficacy of TCP-25-coated sutures employing in vitro and in vivo models. A reporter assay employing THP1-XBIue™- CD14 cells was used. LPS with or without TCP-25 from coated sutures was used to stimulate reporter cells. A significant reduction in endotoxin-induced NF-KB and AP-1 activation was observed in the presence of TCP-25 eluted from sutures (Figure 5a, upper panel). Results from MTT assay showed that these concentrations of TCP-25 were not toxic to the cells (Figure 5a, lower panel).

Next, it was important to know whether TCP-25-coated sutures show similar antiinflammatory efficacy in vivo. To study this effect, we used an NF-KB reporter mouse model in which the suture was subcutaneously implanted and contaminated with LPS followed by longitudinal in vivo inflammation bioimaging using I VIS spectrum. A high level of local NF-KB activation was observed at the side at which the non-coated control suture was implanted. In contrast, TCP-25-coated sutures led to a significant reduction in this NF-KB-driven inflammation at 3 and 24 h after suture implantation (Figure 5b). At the end of the experiment, we recovered the sutures from the mice and eluted absorbed proteins for cytokine analysis. A significant reduction in TNF-a and IL-6 cytokine levels in fluid extracted from implanted TCP-25-coated sutures was observed compared to the control sutures (Figure 5c). Effects of TCP-25-coated sutures on bacterial biofilm

To investigate whether the TCP-25-coated sutures possess anti-biofilm effects, we used two different experimental approaches.

In the first approach, our aim was to evaluate the impact of TCP-25 coating of the suture on the growth of biofilms in the microtiter plate wells and on the sutures themselves. To achieve this, 30 minutes after the addition of bacteria, control or TCP- 25-coated sutures were added to the wells of the microtiter plate during the biofilm growth. Biofilms were allowed to grow at 37 °C. After 48 h, live/dead staining, bacterial count of the suture-attached biofilm, and crystal violet staining of the microtiter plate attached biofilm were performed. Under a fluorescence microscope after live-dead staining, bacterial biofilm formation (white staining) on the sutures was visible for both S. aureus and P. aeruginosa (Figure 6a). On the TCP-25-coated sutures, less biofilm formation and light grey staining, indicating the presence of dead bacteria, was visible. A viable count assay was used to assess the total number of live bacteria on suture- adhered biofilm. TCP-25-coated sutures showed significantly lower bacterial numbers (Figure 6a, bar chart).

Likewise, crystal violet staining of the plate showed a significantly lower amount of biofilm formation in the wells containing TCP-25-coated sutures. Importantly, comparison with a benchmark antibacterial suture (Vicryl® Plus) containing triclosan showed similar antibiofilm activity against S. aureus, but its activity against P. aeruginosa biofilm formation was not significant (Figure 6a and b). Finally, SEM of the sutures showed a significantly lower amount of adhered biofilm on the TCP-25-coated sutures (Figure 6c).

In the second approach, the aim was to study the impact of TCP-25-coated sutures on mature biofilms. S. aureus or P. aeruginosa biofilms were grown on the plate and exposed to TCP-25-coated sutures for 2 h. Live/dead staining showed a significant increase in dead bacterial cells (red-stained) in the samples exposed to TCP-25-coated sutures (Figure 6d and e). As expected, the viable count assay of the biofilms exposed to TCP-25-coated sutures showed a significant reduction in bacterial numbers (Figure 6d and e, bar charts). Benchmark triclosan-containing sutures showed comparable antibiofilm activity against S. aureus, whereas the activity against P. aeruginosa biofilm was not significant.

Effects of neutrophil elastase on TCP-25-coated suture

Human neutrophil elastase (HNE) is an important enzyme that is produced during wound healing, infection, and inflammation. We have previously demonstrated that HNE digests TCPs in vitro and generates various fragments (Puthia et al., 2020). It was also shown that HNE can produce active TCP-25 fragments in a TCP-25 hydrogel (Puthia et al., 2020). We wanted to investigate if HNE could produce bioactive fragments of TCP-25 from a TCP-25-coated suture. TCP-25-coated sutures were treated directly with HNE after which material was eluted, and the fragmentation pattern was analyzed by nano LC-MS/MS. HNE digestion of TCP-25 that had been coated on the sutures produced numerous peptides at different time points (Figure 7a).

In addition to fragments, the presence of intact TCP-25 (SEQ ID NO: 1) was observed for all the studied time periods. Many of these fragments are known to be bioactive and exert antimicrobial and anti-inflammatory activity (Puthia et al., 2020). Interestingly, the CD14 binding region was preserved in many of the fragments (Figure 7b). The peptide fragment “cocktail” produced by HNE digestion retained its antibacterial activities for digestion periods of up to 6 h as assessed by RDA (Figure 7c). Additionally, the generated peptide fragments retained their anti-inflammatory activity as observed in THP-1 cell model system (Fig. 7d).

Overall, the results demonstrated that HNE digestion of TCP-25 from coated sutures produces multiple TCP-fragments and that many of them are bioactive.

Tensile strength and hemolytic effects of TCP-25-coated suture

We next wanted to investigate whether the TCP-25 coating of polyglactin sutures produced any adverse effect on its tensile strength. Mechanical testing of sutures was done using an Instron tensile strength testing system. No change in the tensile strength of the suture was observed after coating with TCP-25 (Figure 8a, left panel). Host tissue reaction leads to degradation and dissolution of the polyglactin sutures (Reul, 1977). We also wanted to examine whether the TCP-25 coating would adversely affect the degradation of the sutures in the tissue. TCP-25-coated or control sutures were subcutaneously implanted in the mice and removed after four days. Mechanical testing of the TCP-25-coated or control sutures recovered from the mice tissue showed no differences in the tensile strengths, suggesting that TCP-25 coating does not adversely affect the degradation of the sutures in the tissue (Figure 8b, right panel). After subcutaneous implantation, a similar loss of tensile strength was noted for both coated and control sutures compared to the sutures that were not implanted. Furthermore, we investigated the hemolytic activity of the TCP-25 suture. Results using a 1 or 10 cm long suture showed less than 10% hemolysis in human blood, suggesting hemocompatibility (Figure 8b).

Effects of storage of TCP-25-coated suture

Furthermore, we wanted to investigate whether the peptide is degraded upon long-term storage of TCP-25-coated sutures. TCP-25 sutures were stored at room temperature for 18 months after which the peptides were eluted for analysis. HPLC analysis showed no apparent storage-related effects, and the peptide eluted from the stored TCP-25 sutures showed peaks similar to the peaks of fresh TCP-25 control peptide (Figure 8c). Non-significant storage-related effects were noticed as a few small extra peaks were observed in the chromatogram of TCP-25 eluted from stored sutures compared to freshly prepared TCP-25 control peptide.

To confirm that the extra peaks in the chromatogram of eluted TCP-25 did not correspond to significant degradation of peptide, we applied Western blotting analysis. Western blotting analysis of the eluted peptide showed multiple bands of higher molecular weight confirming oligomerization (Figure 8d) of the peptide as also reported before (Petruk et al., 2020). Next, we analyzed whether long-term storage of the TCP- 25-coated suture produced any negative effect on the LPS binding capability of the TCP-25. Circular dichroism analysis showed that even after long-term storage, the peptide retained its LPS binding capacity (Figure 8e). Importantly, RDA analysis of the eluted peptide showed that no loss of antimicrobial activity upon long-term storage of TCP-25 sutures had occurred (Figure 8f). Oligomerization or aggregation of TCP-25 on suture upon storage might explain its retained bioactivity. TCP-25 from coated sutures targets clinical bacterial isolates from human wounds and inflammation induced by human wound fluid

After showing anti-microbial and anti-inflammatory efficacy in vitro and in animal models, we finally explored whether TCP-25-coated sutures could also target clinical bacterial isolates from wounds and inflammation pertaining to human wounds. When incubated with various clinically derived human wound isolates, clear zones of inhibition were observed around the TCP-25-coated suture. Furthermore, acute wound fluid (AWF) and chronic wound fluids (CWF) derived from patients with wounds colonized by bacteria such as S. aureus and P. aeruginosa were used to activate inflammation in THP-1 reporter cells. TCP-25 eluted from the coated suture caused a reduction in the human wound fluid-induced inflammation in THP-1 cells (Figure 9a, upper panel). Results from the MTT assay showed that TCP-25 at these concentrations was not toxic to the cells (Figure 9a, lower panel).

Finally, in an in vivo setting, TCP-25-coated sutures were contaminated with human chronic wound fluid and implanted on the back of NF-KB reporter mice. Non-invasive I VIS imaging was performed to visualize NF-KB activation. Control sutures contaminated with human chronic wound fluid induced significantly more inflammation than the TCP-25-coated sutures (Figure 9b).

Altogether, results suggest that TCP-25-coated sutures have the potential to reduce infection and inflammation in a complex human wounding situation.

Conclusion

Using a combination of in silico molecular modeling studies, in vitro antimicrobial, biochemical, and biophysical assays combined with in vivo models, we here show a proof-of-concept demonstrating the possibility of functionalizing a polyglactin suture material with a dual action host defense peptide that can target both bacteria and the excessive inflammatory response. This broad capability may be of biological and clinical importance. In the initial contact with tissues, it is plausible that a given biomaterial also induces an immune response perse, which may cause dysregulation of inflammatory responses, leading to inefficient host defense and causing a given biomaterial to become infection-prone (Busscher et al., 2012). Moreover, in the case of bacterial adhesion, the release of bacterial products, such as lipoteichoic acid (LTA) and LPS, may stimulate inflammation in the vicinity of biomaterials. The possible mechanisms underlying suture-associated infections have only been partially explained, mainly with research focused on bacterial adhesion and biofilm formation. Though, applying the above reasoning to sutures, it is plausible that creating a local suture environment that counteracts bacteria but controls immune responses could be a new attractive strategy for optimizing and boosting infection control. Another serious problem is the declining effectiveness of antibiotics and other antimicrobials due to antimicrobial resistance (AMR) development. The development of resistance is particularly important in surgical procedures in which the combination of extensive antibiotic use, high risk of systemic spread, and bacterial sepsis leads to a high frequency of infections for which “last resort” antibiotics need to be used.

Considering the above issues, an unmet need for new biologically-oriented strategies exists that is based on the multi-pronged action of TCP-25 which targets bacteria in a new way, hence minimizing resistance problems and also controlling the excessive inflammatory process.

At present, multiple antimicrobials are in use or under development for use on sutures (Chua et al., 2022). Based on its wide use, we therefore decided to use triclosan as a benchmark antimicrobial in the studies (Schweizer 2001). As mentioned in the Introduction, increasing concerns with the use of this antiseptic exist. Thus, triclosan resistance is frequently reported in S. aureus (Suller and Russell, 2000). Moreover, Nadafpour et al., 2021 studied the bacterial colonization on various suture materials used in oral implantology. Vicryl® Plus sutures showed the highest buildup of E. coli and S. aureus when compared with Vicryl alone, and triclosan-coated Vicryl® Plus sutures exhibited no benefit over the commonly used silk sutures in terms of causing a decrease in the number of bacteria. Elsolh et al. performed a meta-analysis (Elsolh et al., 2017) to determine if antibiotic-impregnated sutures, including triclosan, could prevent post-operative surgical site infections and complications in abdominal surgery. No evidence was found by them to support routine use of these sutures. Recently, a systematic review and meta-analysis of randomized controlled trials (S. National Institute of Health Research Unit on Global, Lancet Infect Dis 2022, 22, 1242) showed no benefits from the use of triclosan-coated sutures. The authors state that global and national guidance should be reevaluated for the recommendations for routine use of triclosan-coated sutures. In this context, it should be mentioned that in 2016, the United States Food and Drug Administration (FDA) banned the triclosan incorporation in household products and then prohibited triclosan usage in over-the-counter antiseptic products without premarket review. The European Commission, in 2016, did not approve triclosan as an active substance for use in biocidal products for product-type 1. Given the questionable risk-benefit of triclosan, multiple other antimicrobial sutures are in use or under development (Chua et al., 2022). These sutures incorporate, among others, classical antiseptic products, such as chlorhexidine, polyhexamethylene biguanide (PHMB), octenidine, and povidone-iodine. Other substances used are derived from natural products, such as chitosan, aloe vera, silver nanoparticles, or different antibiotics. Notably, octenidine and chlorhexidine have been applied on Vicryl by dip coating as also used in this study.

In contrast to the above-mentioned antimicrobials, the present work adds a previously unexplored pharmacological functionality to Vicryl sutures that is based on the capability of TCP-25 not only to kill bacteria but also to scavenge multiple bacterial products, and concurrently inhibiting downstream CD14/TLR-mediated inflammatory responses (Saravanan et al., 2018). In this context, it is notable that the findings of our study highlight the specific interaction between TCP-25 and Vicryl suture fibers. Through the utilization of QCM-D, we demonstrated peptide binding and affinity to the suture fibers which was further substantiated by molecular modeling and simulations using a coarse-grained model of TCP-25 and polyglactin 910 copolymer. Moreover, O- PTIR spectroscopy measurements confirmed the presence of TCP-25 on the fibrillar surface of coated sutures, and fluorescence imaging demonstrated intense distribution of TCP-25 on the surface and between Vicryl fibrils. Taken together, these findings provide a comprehensive characterization of the TCP-25-polyglactin interaction supporting the in vitro and in vivo data.

It should be mentioned that the dual action concept based on TCP-25 has already shown promise in the setting of wound healing and infections (Puthia et al., 2020; Stromdahl et al., 2021), and that a TCP-25 hydrogel is currently in clinical development, undergoing phase I safety studies on humans. By utilizing a clinically approved absorbable suture and a peptide with established safety profiles, our study therefore paves the way for the expedited clinical translation of TCP-25-coated Vicryl sutures, addressing the urgent need for effective coated biomaterials that combat infections and inflammation in surgical settings. Moving forward, future development of the final suture product should include further evaluation and standardization of the coating procedure, peptide stability determination, analysis of optimal peptide release kinetics, assessment of biocompatibility, and comprehensive preclinical and clinical studies.

Example 2 - TCP-25-coated particles

Coating of PLGA particles with TCP-25

PLGA particles (1 mg, 50 pm average diameter, lactic acid to glycolic acid ratio of 50/50 (Sigma, No 805122)) were suspended in 400 pL of 1% TCP-25 solution. The mixture was incubated for 2 hours at room temperature under continuous shaking at 1000 rpm. Following incubation, the particles were centrifuged at 15,000 rpm for 20 minutes at 20°C to pellet the coated particles. The supernatant was carefully removed and the concentration of TCP-25 in the supernatant was measured to assess coating efficiency. The pelleted particles were then resuspended in 500 pL of 10 mM Tris buffer, vortexed for 5-10 seconds, and centrifuged again at 15,000 rpm for 15 minutes at 20°C. The washing step was repeated twice more to remove unbound peptides. After the final wash, the PLGA particles were dried using a SpeedVac for 1 hour at 30°C.

SDS-PAGE analysis of PLGA particles coated with TCP-25

For SDS-PAGE analysis, the dried PLGA pellet was resuspended in 500 pL of 10% SDS solution and incubated at 99°C for 5 minutes. The mixture was then centrifuged for 2 minutes, and 10 pL of the resulting supernatant was collected for analysis. Additionally, 8 pL of the supernatant from the initial coating step and 10 pL of each supernatant from the three wash steps were loaded onto a 10-20% Tricine gel. Electrophoresis was performed at 100 V for 1 hour and 40 minutes to assess the presence of TCP-25 on the particles.

The results as shown in Figure 10 demonstrate that after washing steps, TCP-25 is bound to the particles and released by the SDS solution.

Detection and localization of TCP-25 on PLGA particles

To detect TCP-25 on PLGA particles, particles were coated with TCP-25 spiked with TAMRA-TCP25 (1% of total TCP-25). Coating conditions were similar to as mentioned above. Coated particles were imaged using fluorescence microscopy to detect TCP-25- TAMRA as shown in Figure 11 (TCP-25-TAMRA is light grey in the figure).

The results from fluorescence microscopy show that TCP-25 is bound to the PLGA particles.

It is expected that binding to PLGA particles decreases TCP-25 degradation and thus enables retention of TCP-25 LPS binding capacity even after long-term storage, just as was observed for binding to the sutures described in Example 1 , above.

Viable Count Assay (VCA) with dried TCP-25-Coated PLGA Particles

To evaluate the antibacterial activity of the TCP25-coated PLGA particles, a viable count assay (VCA) was performed. Pseudomonas aeruginosa PA01 (50,000 CFU/tube) or E. coli ATCC 25922 were added to the dried TCP-25-coated or uncoated PLGA particles. The bacteria and particles were incubated in 10 mM Tris buffer (pH 7.4) for 2 hours at +37°C, 5% CO2. After incubation, serial dilutions were prepared in Tris buffer at 10X, 100X, 1 ,000X, 100,000X, and 1,000,000X. The diluted samples were then spread onto TH-agar plates and incubated o/n at +37°C, 5% CO2to determine bacterial viability by counting the resulting colonies.

The results showed that the TCP-25 coated particles reduced bacterial growth by over 99%. A typical example of the bacterial growth on TH agar is shown in Figure 12, where no bacterial colonies were detected after TCP-25-PLGA treatment.

Antimicrobial Effects of Dried TCP25-Coated PLGA Particles

For the radial diffusion assays (RDA), dried TCP25-coated PLGA particles were scraped off from the bottom of the tube and spread onto an underlay gel seeded with Escherichia coli or Staphylococcus aureus. The particles were evenly distributed on the gel surface, and the assay was conducted to evaluate the antimicrobial activity of the coated particles by demonstrating the zone of inhibition formed around the particles.

TCP-25 coated PLGA particles yielded antibacterial effects as seen from the absence of bacterial growth of both the Gram-negative E. coli and Gram-positive S. aureus around the PLGA particles as shown in Figure 13. Example 3 - Amount of TCP-25 absorbed by PLGA sutures and particles

After measurements, it was found that 1 mg PLGA sutures absorbs approximately 5-6 g TCP-25 and 1 mg PLGA particles absorbs approximately 40-50 pg TCP-25. Sequence overview

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Claims

X8, X12 is any amino acid, X1, X6, X14 is G, A, V, L, I, P, F, M. Y or W, and X2, X4, X5, X9 is R, K or H, wherein said peptide has a length of from 10 to 40 amino acid residues, such as between 10 to 30 amino acid residues.

2. The material according to claim 1 , wherein the peptide comprises or consists of the amino acid sequence

X1-X2-X3-X4-X5-X6-X7-X8-X9-X10-X11-X12-X13-X14-X15-X16, wherein X1, X6, X14 is F or W, and X2, 4, 5, 9 is R or K.

3. The material according to any one of the preceding claims, wherein the peptide has a length of 18 to 30 amino acids, preferably 18-25 amino acids, and comprises or consists of any one of the amino acid sequences selected from the group consisting of SEQ ID NO: 1 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 2 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 3 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 4 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 5 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 6 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 7 or an amino acid sequence with at least 90% sequence identity thereto, SEQ ID NO: 8 or an amino acid sequence with at least 90% sequence identity thereto, and SEQ ID NO: 9 or an amino acid sequence with at least 90% sequence identity thereto.

4. The material according to any one of the preceding claims, wherein the peptide comprises or consists of the sequence: V-F-R-L-K-K-W-I-X1-K-V-I-X2-Z-F-G wherein

Xi and X₂ are amino acids linked by a covalent linkage.

5. The material according to any one of the preceding claims, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 5:95 to 95:5 (glycolic acid : lactic acid).

6. The material according to any one of the preceding claims, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of from 5:95 to 90:10 (glycolic acid : lactic acid).

7. The material according to any one of claims 1 to 6, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 25:75 (glycolic acid : lactic acid), such as 30:70 (glycolic acid : lactic acid), such as 50:50 (glycolic acid : lactic acid).

8. The material according to any one of the preceding claims, wherein said PLGA polymer comprises or consists of glycolic acid and lactic acid in a ratio of 90:10 (glycolic acid : lactic acid), preferably wherein said PLGA polymer is polyglactin 910.

9. The material according to any one of the preceding claims, wherein said peptide is coated on, dissolved in, and/or interspersed between the fibers of said PLGA polymer.

10. The material according to any one of the preceding claims, wherein said material is in the form of a particle, such as a microparticle or a nanoparticle.

11. The material according to claim 10, wherein said particle has an average diameter of between 100 nm to 100 pm, such as between 100 nm to 50 pm.

12. The material according to any one of the preceding claims, wherein said material is anti-inflammatory and/or anti-microbial, such as anti-bacterial.

13. A medical product comprising the material according to any one of the preceding claims.

14. The medical product according to claim 13, wherein said medical product is selected from the group consisting of a suture, a strip, a film, a stent, a graft, a hydrogel, a particle, such as a microparticle or a nanoparticle, and a dressing, such as a mesh, patch or a bandage, preferably wherein said medical product is a suture.

15. The medical product according to claim 14, wherein said medical product is a strip, a film, a stent, a graft, a hydrogel, or a dressing comprising the particle according to any one of claims 10 to 11.

16. The medical product according to any one of claims 13 to 15, wherein said medical product is anti-microbial, such as anti-bacterial, and/or antiinflammatory.

17. The medical product according to any one of claims 13 to 16, wherein said medical product is capable of continuously releasing said peptide to a part of a body for a period of at least 24 hours, such as at least 48 hours, such as at least 72 hours, when said medical product is contacted with said part of the body.

18. The medical product according to any one of claims 13 to 17, wherein said peptide is stable for at least 12 months, such as for at least 18 months, such as for at least 24 months, during dry storage of said medical product at room temperature.

19. A method of producing the medical product according to any one of claims 13 to 18, the method comprising a step of submerging a starting material comprising or consisting of PLGA in a coating solution comprising a dissolved peptide according to any one of claims 1 to 12, followed by drying said starting material.

20. The method according to claim 19, wherein said starting material is a PLGA- comprising particle.

21. The method according to any one of claims 19 to 20, wherein said medical product is a suture.

22. A medical product according to any one of claims 13 to 18 for use in the prevention and/or inhibition of inflammation and/or infection in a part of the body of a subject, preferably wherein said part of the body of the subject is a wound, such as a surgical wound.