Peptide for the treatment of degenerated periodontium

Abstract

The invention relates to peptides containing or consisting of a specific amino acid sequence for use in a method for treating and / or preventing the degeneration of the periodontium, and to specific peptides which can be used for such an application.

Description

[0001] Eisenführ Speiser

[0002] Munich, December 8, 2025

[0003] Our reference: RM 5252-02WO SOE / DRE / fmO

[0004] Publisher / owner: RWTH Aachen University, a public corporation

[0005] Official file number: Subsequent registration to DE102024136715.6

[0006] RWTH Aachen University, a public corporation, 52062 Aachen

[0007] Peptide for the treatment of degenerated periodontal tissues

[0008] The present invention relates to peptides containing or consisting of a specific amino acid sequence for use in a method for treating and / or preventing the degeneration of the periodontal tissues, as well as specific peptides that can be used for such an application.

[0009] Chronic kidney disease (CKD) is a recurring cause of death worldwide, with a prevalence of approximately 10%. CKD is defined as pathological functional or structural changes in the kidney that persist for extended periods, i.e., at least three months. These changes lead to a progressive, usually irreversible decline in excretory and endocrine kidney function, which can culminate in end-stage renal failure. Chronic renal insufficiency is a specific form of CKD in which the kidney's filtration capacity is pathologically impaired. CKD is typically measured using the glomerular filtration rate (GFR), albuminuria, and retention values, which particularly reflect creatinine clearance.

[0010] As a consequence of impaired blood filtration in patients with chronic kidney disease (CKD), waste products accumulate in the blood instead of being excreted in the urine. Electrolyte balance, blood pressure regulation via the renin-angiotensin-aldosterone system, and erythrocyte production can also be affected by CKD. Similarly, reduced renal production of calcitriol can lead to decreased blood calcium levels and hyperparathyroidism, which can result in abnormal bone mineralization. Both blood purification procedures, usually dialysis, and treatment of secondary diseases are typically necessary.

[0011] The degeneration of the body's own tissue as a result of CKD can affect both vascular tissue, which consists mainly of endothelial cells and comparatively large vessels, and tissue which mainly contains fibroblasts, fibers and comparatively small vessels.

[0012] Furthermore, degeneration of the periodontal tissues is frequently observed in CKD patients, in addition to the changes mentioned above. This degeneration of the periodontal tissues progressively weakens the stable anchorage of the teeth, which can ultimately lead to the loss of one or more teeth.

[0013] Chronic kidney disease (CKD) is considered a risk factor for periodontitis and promotes the degeneration of the periodontium. Other risk factors include tobacco use and nicotine exposure, as well as genetic predisposition and the patient's age. Furthermore, factors such as poor oral hygiene, pregnancy, vitamin deficiencies, and diabetes mellitus play a significant role in the progressive degeneration of the periodontium.

[0014] The degeneration of the periodontal apparatus described above as a consequence of CKD or (e.g. chronic) nicotine exposure particularly affects the periodontal ligament and / or the cementum and / or the fiber system radiating into the periodontal ligament and is therefore distinguishable from other more infection-related degenerative phenomena such as periodontosis or periodontitis.

[0015] Periodontitis is a widespread dental problem in Western countries and describes the recession or loss of the gums as a result of structural changes and the frequently accompanying chronic inflammation of the periodontium. This recession or loss is often caused by a systemic metabolic disorder such as chronic kidney disease (CKD). When chronic inflammation of the periodontium is caused by bacterial plaque, it is called periodontitis. A bacteria-containing biofilm in the dental area can release metabolic and breakdown products that trigger an immune response, releasing endotoxins. In addition to these, the body's own tissue can also be destroyed in the process. Degeneration is the result.

[0016] A link between periodontitis and other diseases such as arteriosclerosis and Alzheimer's disease, as well as possible premature births, is being discussed.

[0017] Currently, treatment for periodontal degeneration in practice focuses on eliminating inflammation, pro-inflammatory factors, plaque and tartar, or pathogenic bacteria. This typically includes mechanical removal and hygiene measures such as cleaning periodontal pockets, as well as, if necessary, antiseptic or antibiotic treatment.

[0018] Currently, there are no medications that directly counteract degeneration or treatments for pre-degenerative stages. In particular, no method for regenerating the degenerated periodontal tissues has yet been described. Besides slowing down the degeneration process, regeneration is especially desirable.

[0019] It is therefore an object of the present invention to provide a means of treating and / or preventing the degeneration of the periodontal apparatus and / or its precursors and / or early stages.

[0020] The task is solved by a peptide containing or consisting of an amino acid sequence selected from the group consisting of

[0021] SEQ ID NO: 1 ,

[0022] SEQ ID NO: 2,

[0023] SEQ ID NO: 3,

[0024] SEQ ID NO: 4,

[0025] SEQ ID NO: 5,

[0026] SEQ ID NO: 6, SEQ ID NO: 7,

[0027] SEQ ID NO: 8,

[0028] SEQ ID NO: 9,

[0029] SEQ ID NO: 10,

[0030] SEQ ID NO: 11 ,

[0031] SEQ ID NO: 12,

[0032] SEQ ID NO: 13,

[0033] SEQ ID NO: 14, for use in a procedure for the treatment and / or prevention of periodontal degeneration, wherein periodontal degeneration includes periodontal ligament and / or cementum degeneration.

[0034] Surprisingly, it was found that the peptide calcium blocking factor (CBF) can prevent cementum degeneration and thus counteract the progression of periodontal degeneration. CBF is a cleavage product of chromogranin A, which is cleaved intracellularly in the adrenal gland and secreted into the bloodstream upon adrenal stimulation. It has been found that CBF can halt the degeneration of the dental cementum and the fundamental disorganization of the periodontal tissue structure. Therefore, CBF is suitable for both the treatment and prevention of periodontal degeneration, as described herein.

[0035] Surprisingly, it was also found that CBF can even have a regenerative effect. Histological examination of tissue sections after CBF application revealed a significant morphological improvement in the fiber and cell structure, with the fibers of the periodontal ligament normalizing and resembling a healthy state in relation to the root cementum. This allows the primary causes of periodontal degeneration to be addressed and even initiate regeneration.

[0036] Preferably, the method for treating and / or preventing periodontal degeneration as described herein includes or is a periodontal regeneration procedure.

[0037] Surprisingly, even small fragments of CBF were found to be effective in treating periodontal degeneration and even in pursuing regenerative interventions. Due to their small molecular size, these peptides are suitable as small molecule drugs and exhibit therapeutically appropriate bioavailability.

[0038] Orth-Alampour, Basic Res Cardiol. 2021, shows that CBF can reduce the calcium content of cells and thoracic aortic rings under calcifying culture conditions, as well as in aortas of animals treated with vitamin D and nicotine (VDN animals). Thus, CBF can prevent vascular calcification by inhibiting osteogenic transdifferentiation. However, the described vascular tissue differs significantly from that of the periodontal tissues. While vascular tissue morphologically consists primarily of endothelial cells and comparatively large vessels, the periodontal tissues consist predominantly of fibroblasts and fibers.

[0039] Orth-Alampour does not indicate that any degeneration of the periodontal tissues can be halted, prevented, treated, or even reversed by CBF or a peptide fragment thereof.

[0040] A positive effect of CBF has also been described in cases of alveolar bone degeneration. For example, Shruti Bhargava, “Identification and characterization of the mediators of the calcification paradox”, 2022, shows that bone mineral density in the alveolar bone is reduced in VDN rats, and this reduction could be mitigated by the administration of CBF.

[0041] The alveolar bone, however, is morphologically distinct from the periodontal ligament, which also results in different pathogenesis and possible treatment options. The periodontal ligament is responsible for anchoring the teeth. It consists of the gingiva (gingiva propria), the cementum (cementum), the alveolus (tooth socket), and the periodontal ligament (periodontal membrane). The fibroblasts of the periodontal ligament primarily produce collagen fibers that connect the cementum to the alveolus.

[0042] The cells of the alveolar bone (osteoblasts) are characterized by a compact, angular morphology, which is typical for osteoblasts. Cells of the periodontal ligament (as part of the tooth-supporting apparatus), on the other hand, are predominantly fibroblasts and exhibit a primarily elongated morphology.

[0043] The function of the alveolar bone also differs significantly from that of the periodontal ligament. While the alveolar bone represents the bony part of the jaw, the periodontal ligament forms the functional anchoring system for the teeth.

[0044] Bhargava does not suggest that any degeneration of the periodontal tissues could be halted, prevented, treated, or even reversed by CBF or a peptide fragment thereof, as he only describes effects on osteoblasts and alveolar bone, but not effects on fibroblasts and the fibers they produce, which are responsible for the degeneration of the periodontal tissues described therein.

[0045] As described herein, it was surprisingly found that the application of CBF leads to a significant reduction in periodontal degeneration and cementum degeneration, and that CBF is therefore suitable for the treatment and / or prevention of periodontal degeneration as described herein.

[0046] Example 2 shows that CBF has a direct influence on periodontal ligament fibroblasts, leading to lower inflammation levels (here: NFKB).

[0047] The amino acids of the SEQ ID NO. are identified by letters of the lUPAC code.

[0048] Preferably, the term "degeneration" is characterized by the fact that cells, fibers, or tissues are regressed, i.e., exhibit a reduced volume and / or reduced function. Preferably, the term "treatment of periodontal degeneration" describes the slowing of degeneration as described herein and / or the regeneration of the degenerated cells, fibers, or tissue. Preferably, precursors and / or early stages of degeneration are also treated and / or regeneration of the affected cells is induced.

[0049] Preferably, the term "regeneration" is characterized by the fact that the degeneration, as described herein, is reversed, in particular that damaged cells, fibers, or tissues are functionally and / or morphologically restored, preferably by the formation of new cells.

[0050] Preferably, the term "cells" refers to fibroblasts, osteoblasts, osteocytes, osteoclasts, myocytes, chondrocytes, erythrocytes, leukocytes, thrombocytes and / or their precursors. Preferably, the term "fibers" refers to collagen fibers and / or elastic fibers.

[0051] Preferably, the term "root membrane" describes the connective tissue of the periodontal apparatus, i.e., in particular collagen fibers and elastic fibers, preferably collagen fibers. Preferably, the terms "root membrane", "periodontal ligament" and "periodontium" are used interchangeably herein.

[0052] Preferably, the term "dental cementum" describes a structure comprising minerals, preferably hydroxyapatite, organic components such as mineralized fibers (especially collagen fibers) and cementocytes, as well as water. Dental cementum typically covers the tooth root and embeds the tooth in the alveolar socket. Typically, dental cementum is interwoven with Sharpey's fibers and provides a protective function for the tooth root. Dental cementum is typically subdivided into surface cementum, anchorage cementum, repair cementum, and cellular mixed-fiber cementum.

[0053] Preferably, the terms “root cement”, “dental cement” and “cementum” are used interchangeably herein.

[0054] Preferably, the degeneration of the periodontal ligament, in particular the degeneration of the periodontal ligament and / or cementum, involves or includes the degeneration of fibroblasts, preferably periodontal ligament fibroblasts. In particular, the degeneration of the periodontal ligament, in particular the degeneration of the periodontal ligament and / or cementum, is mediated by fibroblasts or their degeneration. Preferably, the degeneration of the periodontal ligament described herein, in particular the degeneration of the periodontal ligament and / or cementum, does not involve reduced bone mineralization. Conversely, it is therefore preferred that the treatment and / or prevention of the degeneration of the periodontal ligament, in particular the degeneration of the periodontal ligament and / or cementum, described herein, does not involve an increase in bone mineralization; however, this may preferably occur additionally.

[0055] Preferably, the peptide according to the invention is a peptide that contains or consists of an amino acid sequence according to SEQ ID NO: 15 or an amino acid sequence with a sequence identity of at least 89%, preferably at least 90%, preferably at least 91%, preferably at least 92%, preferably at least 93%, preferably at least 94%, preferably at least 95%, preferably at least 96%, preferably at least 97%, preferably at least 98%, preferably at least 99%, to the sequence according to SEQ ID NO: 15.

[0056] Preferably, the peptide according to the invention is a peptide that contains or consists of an amino acid sequence according to SEQ ID NO: 15, or that contains or consists of an amino acid sequence that differs from the amino acid sequence according to SEQ ID NO: 15 by a maximum of 2 amino acids, preferably by a maximum of 1 amino acid.

[0057] The amino acid sequence according to SEQ ID NO: 15 describes the full-length peptide CBF.

[0058] Preferably, information on the percentage sequence identity refers to values ​​that can be calculated via a sequence alignment, for example via the EMBOSS Water Pairwise Sequence Alignments: (http: / / www.ebi.ac.uk / Tools / psa / emboss_water / ).

[0059] Local sequence matching tools, such as those from the European Molecular Biology Laboratory (EMBL) and the European Bion Informatics Institute (EBI), use a modified Smith-Waterman algorithm (http: / / www.ebi.ac.uk / Tools / psa / and Smith, TF & Waterman, MS "Identification of common molecular subsequences" Journal of Molecular Biology, 1981 147 (1):195-197).

[0060] When a sequence comparison is performed using such a modified Smith-Waterman algorithm, reference is made to the standard parameters currently specified by EMBL-EBI. These are, for amino acids: Matrix = BLOSUM62, Gap open penalty = 10, and Gap extend penalty = 0.5. Preferably, the peptide according to the invention is a peptide consisting of 4 to 10 amino acids, preferably 4 to 9 amino acids, more preferably 4 to 8 amino acids, particularly preferably 4 to 7 amino acids, more preferably 4 to 6 amino acids, and even more preferably 4 to 5 amino acids.

[0061] Shorter peptides, as described herein, are particularly advantageous because they are easier to synthesize than full-length peptides. Furthermore, shorter peptides, as described herein, have better bioavailability than full-length peptides.

[0062] Preferably, the peptide according to the invention is a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 3 to 12.

[0063] Preferably, the peptide according to the invention has a molecular weight of at most 1500 Daltons, preferably at most 1250 Daltons, more preferably at most 1000 Daltons, and particularly preferably at most 900 Daltons.

[0064] Preferably, the treatment and / or prevention as described herein includes oral, sublingual, buccal, intravenous, intra-arterial, intramuscular, subcutaneous, transdermal, intranasal and / or aural administration of the peptide according to the invention.

[0065] Preferably, the treatment and / or prevention as herein comprises the administration of at least 0.05 g, preferably at least 0.075 g, more preferably at least 0.1 g, particularly preferably at least 0.12 g of a peptide according to the invention per kg body weight per day.

[0066] Another aspect of the present invention relates to a pharmaceutical composition containing a peptide according to the invention for use in a method for treating and / or preventing the degeneration of the periodontal apparatus, wherein the degeneration of the periodontal apparatus includes degeneration of the periodontal ligament and / or the cementum.

[0067] What is described herein regarding the peptides according to the invention, or the treatment and prevention of periodontal degeneration, in particular preferred features, also applies accordingly to the pharmaceutical composition according to the invention. Pharmaceutical compositions according to the invention are preferably in the form of capsules or tablets (coated and uncoated tablets, e.g.,with gastro-resistant coatings or modified release tablets, effervescent tablets, orally disintegrating tablets, chewable tablets, lozenges, pastilles for swallowing or sucking, lyophilisates for oral use, tablets for preparing a solution or suspension for oral use), granules, pellets, solid mixtures, dispersions in liquid phases, as emulsions, as suspensions, as syrups, as powders, as solutions, as drops, as sprays, as pastes or as other preparations that can be swallowed or chewed, and are preferably used as prescription, pharmacy-only or other medicinal products or as dietary supplements.

[0068] A further preferred dosage form is a pharmaceutical composition selected from the group consisting of ointments, creams, pastes, gels, suspensions, crystal suspensions, emulsions, solutions, drops, powders, sprays and foams, preparations for injection, preparations for infusion, concentrates, preparations for inhalation, preparations for use on the ear and implants.

[0069] Preferably, the composition according to the invention contains one or more peptides according to the invention in a total amount in the range of 0.001 to 20 wt.%, preferably 0.005 to 15 wt.%, particularly preferably 0.01 to 10 wt.%, further preferably 0.05 to 7.5 wt.%, particularly preferably 0.1 to 5 wt.%, most preferably 0.5 to 4 wt.%, and even more preferably 1 to 3 wt.%, based on the total weight of the pharmaceutical composition.

[0070] If the composition comprises several peptides according to the invention, the total quantity preferably refers to the sum of the individual peptides according to the invention.

[0071] If the composition comprises several peptides according to the invention, the total amount preferably refers to each of the individual peptides according to the invention, each of which is present in an amount within the aforementioned ranges, i.e., preferably the composition according to the invention contains 0.001 to 20 wt.%, more preferably 0.005 to 15 wt.%, particularly preferably 0.01 to 10 wt.%, further preferably 0.05 to 7.5 wt.%, particularly preferably 0.1 to 5 wt.%, most preferably 0.5 to 4 wt.%, and even more preferably 1 to 3 wt.%, based on the total weight of the pharmaceutical composition, of a first peptide according to the invention, 0.001 to 20 wt.%, more preferably 0.005 to 15 wt.%, particularly preferably 0.01 to 10 wt.%, further preferably 0.05 to 7.5 wt.%, particularly preferably 0.1 to 5 wt.%, and most preferably 0.5 to 4 wt.%. wt.%, preferably 1 to 3 wt.%.-%, based on the total weight of the pharmaceutical composition, a second peptide according to the invention, etc.

[0072] Preferably, the composition according to the invention contains one or more pharmaceutically acceptable carrier substances.

[0073] A peptide or composition according to the invention is preferred for use, each as described herein, wherein the degeneration of the periodontal apparatus relates to a degeneration of the periodontal ligament and / or the cementum.

[0074] A peptide or composition according to the invention is preferred for use, each as described herein, wherein the degeneration of the periodontal apparatus is caused by or associated with reduced renal function, preferably by chronic renal insufficiency.

[0075] Preferably, reduced kidney function describes a disease according to ICD-11, GB61 ("Chronic kidney disease"), GB90.4 ("Renal tubular function disorders") or GC2Z ("Abnormal renal function").

[0076] Preferably, reduced kidney function is characterized by an increased concentration in the blood of substances that must be excreted in urine, such as creatinine, urea and uric acid.

[0077] Preferably, reduced kidney function is measured and classified by measuring the GFR and / or retention values, which reflect, among other things, the concentration of creatinine, urea and cystatin C.

[0078] Preferably, chronic kidney disease refers to a disease according to ICD-11, GB61 (“Chronic kidney disease”).

[0079] Additionally or alternatively, it is preferred that the degeneration of the periodontal tissues is caused by or associated with heart failure. Additionally or alternatively, it is preferred that the degeneration of the periodontal tissues is caused by or associated with nicotine exposure, preferably chronic nicotine exposure.

[0080] Preferably, the nicotine exposure refers to a disease according to ICD-11, 6C4A (“Disorders due to use of nicotine”).

[0081] Preferably, chronic nicotine exposure refers to a disease according to ICD-11, 6C4A.2 (“Nicotine dependence”).

[0082] Nicotine exposure typically affects the entire organism, as nicotine enters the bloodstream. In the nicotine exposure described here, the oral cavity, particularly the teeth and / or surrounding tissue, is most likely to come into contact with nicotine.

[0083] Another aspect of the present invention relates to a peptide consisting of 4 to 7 amino acids, preferably 4 to 6 amino acids, more preferably 4 or 5 amino acids, containing an amino acid sequence selected from the group consisting of SEQ ID NOs: 3 to 12.

[0084] Preferably, the peptide according to the present invention is a peptide consisting of an amino acid sequence selected from the group consisting of SEQ ID NOs: 3 to 12.

[0085] Figure 1 shows the effect of VDN and CBF treatment on the tooth root cementum observed in Example 1, shown on p-CT images.

[0086] (A) The white rectangle represents the relevant area for the qualitative assessment of the cementum;

[0087] From: alveolar bone,

[0088] D: Dentin,

[0089] Cm: Cement.

[0090] (B) In the control group (top, “Ctr”), intact cementum and portion of the periodontal ligament (PDL) are visible. Damaged cementum in the VDN group (middle, “VDN”) leads to the formation of channels connecting the PDL to the pulp chamber. The CBF group (bottom, “CBF”) exhibits a less distorted mineralized portion of the cementum.

[0091] (C) 3D reconstruction of the first molar (M1) confirms these assumptions of a distorted cementum after nicotine exposure and the possible recovery of the cementum after CBF treatment.

[0092] In wild-type Wistar Kyoto rats, the cementum forms a uniformly thin layer on the outer surface of the tooth root (A). In the apical region, the layer thickness increases significantly compared to the cervical region. VDN mice exhibit marked morphological variations. Pronounced defects within the cementum layer are visible near the apex. These defects appear as hypomineralized discontinuities in the cementum layer (B). Three-dimensional views of the region show that the defects are not limited to a few layers but affect the entire apex (C). In the CBF group, these cementum resorptions are reduced compared to the cementum morphology of the control animals.

[0093] Figure 2 shows the microscopic images of histological longitudinal sections of the proximal jawbone tissue of the rat obtained in Example 1, stained with hematoxylin and eosin (H&E), and represents representative sections of the respective experimental groups;

[0094] (A) Control group (Ctr)

[0095] (B) VDN Group

[0096] (C) CBF Group

[0097] The animals treated with CBF showed a shift in condition towards control compared to the VDN animals.

[0098] From: alveolar bone,

[0099] D: Dentin,

[0100] Cm: Cement,

[0101] PDL: periodontal ligament. The periodontal ligament (PDL) fibers of the control group were aligned parallel in a functional direction (A.1). In contrast, the PDL fibers in the VDN group appeared disturbed, with loss of orientation, no parallel alignment, and no attachment to the bone surfaces (B.1).

[0102] Furthermore, in the apical region of the VDN group, a basophil-rich region (A.2) with cell-rich cementum was observed in the area of ​​the basophilic zone between cementum and dentin (B.2) compared to the control group. Here too, the CBF group shows an intermediate stage of basophilic regions in this area compared to that of the control animals.

[0103] Further aspects and advantages of the invention will become apparent from the following description of preferred embodiments.

[0104] Examples

[0105] Example 1: CKD animal model and CBF treatment

[0106] A total of 18 wild-type rats were used (n = 5–7, 3 groups). The rats were fed standard rodent food and had ad libitum access to water. A model version of the protocol originally described by Niederhoffer et al. was used. The rats in the VDN group (n = 7) received vitamin D3 (300,000 IU / kg, IM) and nicotine (25 mg / kg, oral) to simulate a CKD state and thus served as a negative control. The rats in the control group (n = 6) received a daily injection of normal saline (IM) for 4 weeks. The rats in the CBF group (n = 5) received vitamin D3 (300,000 IU / kg, IM) and nicotine (25 mg / kg, oral) to simulate a CKD state and were stabilized to a plasma CBF concentration of 100 pmol / L. All rats were housed under standard conditions and provided with standard rodent food and water at all times.

[0107] Histological sections and staining

[0108] The right mandibles of wild-type rats were fixed in 3.7% paraformaldehyde solution for at least 24 hours. Subsequently, the samples were decalcified at room temperature for eight weeks using 10% Tris-buffered ethylenediaminetetraacetic acid (EDTA, pH 7.4) (Morphisto Offenbach, Germany). The buffer was renewed every other day. After decalcification, each sample was embedded in paraffin and sectioned for histological analysis. For hematoxylin and eosin (H&E) staining, 5 µm thick serial longitudinal sections of mandibular sections were prepared. Prior to staining, the sections were incubated at 60 °C for 30 minutes, followed by incubation in xylene (PNZ 09208831, Fischar). After hydrogenation by a descending alcohol series, the sections were stained with hematoxylin solution (721 1, Epredia). The sections were then counterstained under running warm water with 1% eosin G solution (3137.2 Roth).The sections were then dehydrated using an ascending alcohol series. After xylene treatment, they were covered with Eukitt (03989, Sigma-Aldrich). Two mandibular sections from each animal were randomly selected for histological evaluation. Statistical analysis was performed.

[0109] First, the data were tested for normal distribution, followed by an ANOVA analysis of the variance using Tukey's post-hoc test (Prism version 9.1.1; GraphPad software). A p-value < 0.05 was considered statistically significant.

[0110] Results

[0111] The inhibitory and preventive effect of CBF on the periodontal tissues was investigated in a CKD animal model (vitamin D-nicotinamide CKD / calcification model). In animals with induced CKD in the vitamin D-nicotinamide (VDN) group, significant degeneration of the cementum (Figure 1) and fundamental disorganization of the periodontal tissue structure (Figure 2) were observed compared to untreated controls.

[0112] The application of CBF led to a significant reduction in the observed CKD-associated changes of the periodontal tissues. The morphology of the periodontal fiber organization and the root cementum showed a marked reduction in the observed morphological changes following CBF administration.

[0113] Histological and tomographic analysis of the animals revealed significant morphological and cellular changes in the periodontal tissue of the examined mandibles. Degeneration of the cementum and disorganization of the periodontal ligament tissue were observed. Treatment with CBF resulted in improved alignment of the periodontal fibers and less deformed cementum tissue in the animals. Administration of CBF largely prevented degeneration of the periodontal apparatus.

[0114] Histological evaluation of nicotine / vitamin D-induced chronic kidney disease (CKD) in rats (VDN group) reveals significant morphological and cellular changes in the periodontal tissue of the examined mandibles. In addition to deterioration of the cementum, disorganization of the periodontal ligament (PDL) tissue was observed under CKD conditions. Treatment with calcium blocking factor (CBF) resulted in less deformed cementum tissue in the treated rats. It was found that CBF administration could even sufficiently restore the PDL tissue. Cell culture

[0115] Human periodontal ligament fibroblasts (PDLFs) were acquired from Lonza Bioscience (catalog # CC-7049). PDLFs were cultured in DMEM growth medium (Sigma Aldrich, D5796-500ML) with 10% fetal calf serum (FCS) (Bio-Sell, FCS.LE.0500), 100 units ml. -1 Penicillin and streptomycin (ThermoFisher Scientific, 15140122), and 50 mg L' 1Ascorbic acid (Sigma Aldrich, APO456787373-500G). PDLFs were incubated at 37 °C and 5% CO2, with regular medium changes every two to three days.

[0116] For quantitative real-time PCR (RT-PCR) and Western blotting, the PDLFs were seeded onto 24-well plates at a density of 2x10 5 Cells per well. Cells were maintained in DMEM growth medium until 80% confluence was reached. Subsequently, the growth medium was replaced with control and induction medium with and without calcification blocking factor (CBF).

[0117] The control medium consists of the growth medium with an additional 0.1% DMSO and water, as well as 0.12% NaCl solution. This was added as a control because the uremic toxins were dissolved in DMSO, water, or NaCl.

[0118] The induction medium consists of the growth medium including a cocktail of uremic toxins (see table), which were selected based on the EUTox database (Baaten, C., et al., Impaired Secondary Platelet Response in Chronic Kidney Disease as a Consequence of Prior Platelet Activation. JACC Basic Transl Sei, 2025. 10(9): p. 101355). An additional induction group received 100 ng ml of 1 CBF.

[0119] Phenyleacetic acid 3.5 mM (0.48 g L -1 )

[0120] Hippuric acid 0.61 mM (110 mg L -1 )

[0121] Indoxyl sulfate 0.15 mM (37 mg L' 1 )

[0122] Kynurenic acid 0.79 pM (37 mg L' 1 ) p-cresyl sulfate 0.1 mM (23 mg L' 1 )

[0123] Methylguanidine 2.1 pM (226 pg L' 1 )

[0124] Guanidinosuccinic acid 17.1 pM (3 pg ml' 1 ) Quantification of gene expression

[0125] To quantify the immediate effect of CBF under uremic conditions, PDLFs were stimulated with uremic toxins with and without CBF for 30 min. The cells were then lysed, the RNA isolated using a Quick-RNA Microprep Kit (Zymo Research, R1050), and transcribed to cDNA. The relative gene expression of NFKB was determined. β-Actin was chosen as the reference gene. Results were normalized to a control (control medium only, no CBF).

[0126] Figure 3A shows the NFKB gene expression in human periodontal ligament fibroblasts, with A = induction group (without CBF) and B = induction group with CBF. A marked increase in NFKB gene expression was observed in periodontal ligament fibroblasts cultured under uremic conditions compared to the control. This increase could be significantly reduced (p < 0.05) by CBF treatment. In periodontal ligament fibroblasts cultured under uremic conditions and treated with CBF, NFKB gene expression was significantly reduced compared to the induction group without CBF.

[0127] Quantification of protein levels

[0128] To quantify the effect of CBF on NF-κB at the protein level under uremic conditions, percutaneous diuretic liver failure cells (PDLFs) were stimulated with uremic toxins, with and without CBF, for 24 h. The cells were then lysed, and the same concentration of protein was isolated from each sample by 12% SDS-C gel electrophoresis. The proteins were subsequently transferred to a nitrocellulose membrane, blocked with 5% milk powder for 1 h at room temperature, and detected using specific antibodies. Anti-NF-κB antibody (Cell Signaling, 8242S), anti-β-actin antibody (MP Biomedicals, 691001), HRP-conjugated antibody (Cell Signaling, 7074S), and IgG HRP antibody (Cell Signaling, 7076S) were used for detection. Primary antibodies were incubated overnight in 5% milk powder at 4 °C. Secondary antibodies were incubated in 5% milk powder at room temperature for 1 h. Protein bands were visualized by chemiluminescence.NFKB levels were determined in relation to β-actin levels. Results were normalized to a control (control medium only, no CBF).

[0129] Figure 3B shows the protein levels of NFKB in human periodontal ligament fibroblasts, with A = induction group (without CBF) and B = induction group with CBF. At the protein level, it was also shown that CBF treatment leads to lower levels of NFKB in human periodontal ligament fibroblasts.

Claims

Claims 1. Peptide containing or consisting of an amino acid sequence selected from the group consisting of SEQ ID NO: 1 , SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 1 1 , SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, for use in a procedure for the treatment and / or prevention of periodontal degeneration, where the degeneration of the periodontal apparatus includes degeneration of the periodontal ligament and / or the cementum.

2. Peptide for use according to claim 1, wherein the peptide contains or consists of an amino acid sequence according to SEQ ID NO: 15, or which contains or consists of an amino acid sequence which differs from the amino acid sequence according to SEQ ID NO: 15 by a maximum of 2 amino acids, preferably by a maximum of 1 amino acid.

3. Peptide for use according to claim 1, wherein the peptide consists of 4 to 10 amino acids, preferably 4 to 9 amino acids, more preferably 4 to 8 amino acids, particularly preferably 4 to 7 amino acids, more preferably 4 to 6 amino acids, and even more preferably 4 to 5 amino acids.

4. Peptide for use according to claim 1 or 3, wherein the peptide consists of an amino acid sequence selected from the group consisting of SEQ ID NOs: 3 to 12.

5. Pharmaceutical composition comprising a peptide as defined in any one of claims 1 to 4, for use in a method for treating and / or preventing periodontal degeneration, wherein periodontal degeneration includes periodontal ligament and / or cementum degeneration.

6. Peptide for use according to any one of claims 1 to 4 or composition for use according to claim 5, wherein the degeneration of the periodontal apparatus refers to degeneration of the periodontal ligament and / or the cementum.

7. Peptide for use according to any one of claims 1 to 4 or composition for use according to claim 5 or 6, wherein the degeneration of the periodontal apparatus is caused by or associated with reduced renal function, preferably by chronic renal insufficiency.

8. Peptide for use according to any one of claims 1 to 4 or composition for use according to claim 5 or 6, wherein the degeneration of the periodontal apparatus is caused by or associated with nicotine exposure, preferably chronic nicotine exposure.

9. Peptide for use according to any one of claims 1 to 4 or composition for use according to claim 5 or 6, wherein the degeneration of the periodontal apparatus is triggered by heart failure.

Images