A useful marker for reinforcement strategies for the treatment of osteoarthritis

Abstract

The present invention relates to pharmacogenetics, more specifically, to strategies involving biomarkers associated with the clinical response to a compound before or during treatment of cartilage disorders such as osteoarthritis. The present invention relates more particularly to combinations of JSW measurements and levels of specific proteins present in blood, serum, synovial fluid, or urine, which can be used for strategies before or during treatment, such as patient reinforcement in clinical trials, patient selection strategies, or for fitting patient treatment within a treatment framework for cartilage disorders, such as osteoarthritis.

Description

[0001] Field of the present invention

[0002] The present invention relates to pharmacogenetics, more specifically, to a strategy involving a compound and a marker associated with the clinical response to the compound before or during treatment of a cartilage disorder such as osteoarthritis. The invention is particularly related to a combination of joint space (JSW) measurements and levels of specific proteins present in blood, serum, synovial fluid, or urine, which may be used for strategies before or during treatment, such as patient reinforcement or patient selection strategies in clinical trials, or for fitting patient treatment within a treatment framework for a cartilage disorder, such as osteoarthritis.

[0003] The present invention further discloses diagnostic tools and kits based on a combination of JSW measurements and specific biomarker levels. Accordingly, the present invention can be used to predict the response to a compound for the treatment of cartilage disorders before or during treatment using said compound. This strategy can be used to select / identify subjects to be treated by intra-articular administration of an active compound for treating cartilage disorders, and in patient enhancement strategies. For example, the use of these strategies in diagnosis can lead to increased benefits and reduced risks for subjects.

[0004] Background of the present invention

[0005] Chondrosis is a broad term for diseases characterized by metabolic degeneration in connective tissue, manifested by pain, stiffness, and limited movement in the affected body part. These disorders may be caused by pathology or are the result of trauma or injury. Among these, chondrosis includes osteoarthritis (OA), cartilage damage (comprehensive sports injuries of cartilage and joints, and surgical injuries such as microfracture(s)). Mature cartilage, in particular, has limited self-repair ability due to the absence of blood vessels and the limited potential for proliferation of mature chondrocytes. Furthermore, cartilage does not nitrate easily and has low oxygen pressure. Replacing damaged cartilage caused by injury or disease, especially articular cartilage, is a major challenge for physicians, and available surgical treatments are not fully predictable and are considered effective only for a limited time. Therefore, most younger subjects are advised not to seek treatment or to postpone treatment for as long as possible. When treatment is required, standard surgical procedures are age-dependent and vary between total arthroplasty, cartilage fragment transplantation, or bone marrow stimulation techniques (e.g., microfractures). Microfractures is a common surgical procedure involving the infiltration of the subchondral bone to stimulate cartilage deposition by bone marrow stem cells. However, this technique has been found to fail to adequately repair cartilage defects, and the newly formed cartilage is primarily fibrocartilage, which leads to inadequate or altered function and biomechanics. In fact, fibrocartilage does not possess the same durability and may not attach precisely to the surrounding hyaline cartilage. For this reason, newly synthesized fibrocartilage can be more easily destroyed (estimated timeframe: 5–10 years).

[0006] For individuals with osteoarthritis, non-surgical treatments consist particularly of physical therapy, lifestyle modifications (e.g., increased physical activity), support devices, oral and injectable medications (e.g., non-steroidal anti-inflammatory drugs), gait aids, and medical symptom management. If these treatments fail, surgery, such as joint replacement, is the primary option for the individual. Tibial or femoral osteotomy (cutting the bone to rebalance joint wear) can reduce symptoms, help maintain an active lifestyle, and delay the need for total joint replacement. Total joint replacement can alleviate the symptoms of progressive osteoarthritis, but generally requires changes in the individual's lifestyle and / or activity level.

[0007] At this time, commercially available drug treatments are primarily for pain relief. There are no commercially available treatments to repair or delay cartilage damage (see [Lotz, 2010]).

[0008] Fibroblast growth factor 18 (FGF-18) is a member of the FGF protein family, which is closely related to FGF-8 and FGF-17. FGF-18 has been shown to be a proliferative agent for chondrocytes and osteoblasts (references [Ellsworth et al., 2002; Shimoaka et al., 2002; Gigout et al., 2017]). FGF-18 has been proposed for the treatment of cartilage disorders such as osteoarthritis and chondrosis, either alone (WO2008 / 023063) or in combination with hyaluronic acid (WO2004 / 032849).

[0009] Sprifermin, a truncated form of human FGF-18, is being studied in clinical trials for the treatment of both osteoarthritis and chondrosis (see, e.g., NCT01033994, NCT00911469, and NCT01066871). The current regimen for sprifermin is once a week for three weeks (one treatment cycle), and the drug is administered via intra-articular injection. This treatment cycle may be repeated. This regimen is described in WO2008 / 023063.

[0010] At this time, treatment of OA and cartilage damage using FGF-18 is provided to subjects during clinical trials without predictive information about the response (literature [Lohmander et al., 2014; Dahlberg et al., 2016]), that is, without knowledge of whether the treatment will be highly effective, moderately effective, or have little or no effect. Currently, a large number of treated subjects show a moderate / high response to Sprifermin treatment after at least one treatment cycle according to cartilage thickness and WOMAC scores measured by MRI technology, while some other subjects do not respond to the treatment (i.e., no or limited increase in cartilage thickness measured by MRI) or respond while presenting higher WOMAC scores compared to the control group.

[0011] WO2014 / 023703 describes a genetic marker (a combination of SNPs IL-1RN rs9005 and IL-1RN rs315952) associated with the quality of clinical response to FGF-18 treatment for cartilage disorders, e.g., OA, cartilage damage, or microfracture(s). These markers are useful, through pre-treatment genetic screening, for identifying a specific response to FGF-18 treatment, e.g., a subgroup of subjects more likely to exhibit a very good clinical response to FGF-18 treatment, or conversely, those for whom the therapy may fail.

[0012] Knowledge regarding the types of clinical responses in subjects to treatment can be used to optimize regimens, such as selecting or stratifying patients for clinical trials, choosing a regimen, selecting treatment using compounds as a first-line therapy, or fitting a dosing regimen. This information will be clinically useful in the management of clinical trials in subjects or in the medical management of cartilage disorders, such as OA and / or chondrosis. Furthermore, this predictive information can also be clinically useful in guiding decisions regarding dosing regimens. To provide a complete range of solutions for subjects to be treated or for physicians seeking the optimal treatment for their patients, it is necessary to identify additional markers that aid in predicting placebo responses, optimizing regimens, or selecting regimens that assist in clinical trial management.

[0013] Summary of the present invention

[0014] As described herein, metabolic biomarkers such as proC2 or CTX-II combined with the physical marker JSW (joint gap) may be used for the detection, stratification, diagnosis, and / or treatment of patients with osteoarthritis or cartilage disorders. For example, the expression level (or amount) of at least one of these biomarkers (or combinations thereof) combined with the physical marker JSW may be used to better classify patients or to detect patients to be included in or, conversely, excluded from a specific therapy.

[0015] The present invention relates to a method for eliminating the risk of a clinical trial or determining a placebo effect during a clinical trial (wherein the clinical trial relates to the treatment of cartilage disorders in subjects using an active compound) or treatment using an active compound, and

[0016] a) a step of measuring the joint space (JSW) at least one knee of the subject,

[0017] b) A step to predict the risk of the placebo effect from the results of step a).

[0018] It is a method that includes.

[0019] According to the above method, the presence of JSW higher than 3.5 ± 2SD mm predicts a placebo effect. Conversely, the presence of JSW lower than 3.5 ± 2SD mm predicts no or low placebo effect.

[0020] The present invention also relates to a method for eliminating the risk of clinical trials associated with the treatment of cartilage disorders in subjects with an active compound, and

[0021] a) a step of determining the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject;

[0022] b) a step of measuring the joint gap (JSW) at at least one knee of the subject,

[0023] c) predicting the susceptibility of the subject to treatment using the active compound from the results of steps a) and b).

[0024] It includes, where steps a) and b) are performed in any order.

[0025] According to the above method, the presence of JSW exceeding 3.5 ± 2SD mm with CTX-II exceeding 350 ± 2SD ng / mmol or ProC2 exceeding 4.2 ± 2SD ng / mL predicts low or no sensitivity to treatment with the active compound. Conversely, the presence of 1) JSW less than 3.5 ± 2SD mm with CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL, 2) JSW greater than 3.5 ± 2SD mm with CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL, or 3) JSW less than 3.5 ± 2SD mm with CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL predicts susceptibility to treatment with the active compound.

[0026] also,

[0027] a) determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from a subject, and measuring the joint gap (JSW) at least one knee of the subject, wherein the amount of at least one of these proteins and the gap of the JSW predict the risk of the subject being susceptible or insensitive to the treatment, and

[0028] b) A step of selecting a susceptible subject suitable for the above treatment or clinical trial

[0029] A method for selecting subjects having cartilage disorders to be included in or excluded from treatment or clinical trials using an active compound is described, based on the possibility of their susceptibility to treatment, including.

[0030] According to the above method, the presence of CTX-II in excess of 350 ± 2SD ng / mmol, or ProC2 in excess of 4.2 ± 2SD ng / mL, together with JSW in excess of 3.5 ± 2SD mm, preferably excludes the subject from treatment with the active compound. Conversely, the presence of 1) JSW of 3.5 ± 2SD mm or less with CTX-II of less than 350 ± 2SD ng / mmol and / or ProC2 of less than 4.2 ± 2SD ng / mL, 2) JSW of greater than 3.5 ± 2SD mm with CTX-II of less than 350 ± 2SD ng / mmol and / or ProC2 of less than 4.2 ± 2SD ng / mL, or 3) JSW of less than 3.5 ± 2SD mm with CTX-II of greater than 350 ± 2SD ng / mmol and / or ProC2 of greater than 4.2 ± 2SD ng / mL leads to inclusion of the said subject from treatment with the active compound.

[0031] In addition, a kit comprising means for carrying out the method according to the present invention and instructions for use is described.

[0032] Additionally, the active compound for use in treating a subject with a cartilage disorder is included, characterized in that the subject exhibits 1) CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL together with JSW less than 3.5 ± 2SD mm, 2) CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL together with JSW greater than 3.5 ± 2SD mm, or 3) CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL together with JSW less than 3.5 ± 2SD mm.

[0033] The present invention also relates to a method for treating a subject having cartilage disorders with an active compound, and

[0034] a) determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject, and measuring the joint gap (JSW) at at least one knee of the subject, wherein the amount and gap predict the risk of the subject being susceptible to treatment with the active compound

[0035] b) Step of selecting an object having the following:

[0036] i. CTX-II < 350 ± 2SD ng / mmol, and / or ProC2 < 4.2 ± 2SD ng / mL, together with JSW ≤ 3.5 ± 2SD mm,

[0037] ii. CTX-II < 350 ± 2SD ng / mmol and / or ProC2 < 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm or

[0038] iii. CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL with JSW less than 3.5 ± 2SD mm

[0039] c) A step of administering the active compound into the joint of the selected subject.

[0040] It is a method that includes.

[0041] In general, in a specific embodiment of the present invention, i.e., in any method or use described herein, the subject has a cartilage disorder selected from the group consisting of osteoarthritis, cartilage damage, fractures affecting articular cartilage, or surgical procedures affecting articular cartilage (e.g., microfractures), and the active compound may be selected from the group consisting of, for example, FGF-18 compounds, BMP-2, BMP-7, GDF-5, FGFβ, FGF-9, SOX-9 enhancers, TGFβ, Wnt inhibitors, anti-MMP13 inhibitors, anti-ADAMTS5 inhibitors, calcitonin, and any variants or fusion proteins thereof.

[0042] In any method or use described herein, it should be understood that, before determining the amount of at least one of the proteins, it is necessary to obtain a sample (or test sample) of the subject, for example, through the collection of blood, serum, synovial fluid, or urine. Additionally, it should also be understood that any method or use described herein is performed in vitro rather than on an animal or human body.

[0043] definition

[0044] - As used herein, the terms “FGF-18 compound” or “FGF-18” are intended to be proteins that retain one or more biological activities of wild-type human FGF-18 protein (e.g., increased osteoblast activity, see WO98 / 1664, or increased chondrogenesis, see WO2008 / 023063). FGF-18 may be natural (Sequence 3), its mature form (corresponding to the amino acid sequence of residues 28 (Glu) to 207 (Ala) of Sequence 3), or its truncated form, e.g., sprifermin (as presented in Sequence 4; having amino acid residues 2 to 170 of Sequence 4 corresponding to amino acid residues 28 to 196 of Sequence 3). The term “FGF-18 compound” also includes variants or mutants of the natural, mature, or truncated forms of FGF-18, as well as fusion proteins comprising (biologically) active FGF-18 residues coupled to heterologous proteins or chemical compounds (e.g., disclosed in the patent family EP17192467.3). In such fusion proteins, the FGF-18 residues may be the natural, mature, or truncated forms of the FGF-18 protein or its variants or mutants.

[0045] - The term "BMP-2" as used herein refers to a protein that promotes bone and cartilage formation by inducing matrix synthesis and promoting cartilage repair, as well as playing an important role in the differentiation of osteoprogenitor cells into osteoblasts (Reference [Deng et al., 2018]). The full-length natural form of human BMP-2 is presented in Sequence 6. One of the recombinant forms of the BMP-2 protein is known as divotermine alpha. The term "BMP-2" also includes its variants or fusion proteins containing BMP-2 residues.

[0046] - The term "BMP-7" as used herein refers to a protein known for its osteogenic properties, suggested to have strong anabolic effects on cartilage by stimulating the synthesis of cartilage matrix components and increasing the synthesis of proteoglycans and collagen (Reference [Deng et al., 2018]). The full-length natural form of human BMP7 is presented in Sequence 7. One of the recombinant forms of the BMP-2 protein is known as eptortermin alpha. The term also includes its variants or fusion proteins containing BMP-7 residues.

[0047] - The term "GDF-5," also known as LAP-4 or radotermin as used herein, is a protein that has a stimulating effect on the synthesis of the matrix in human articular chondrocytes cultured in vitro from both healthy subjects and OA patients (Reference [Parrish et al., 2017]). The full-length natural form of human GDF-5 is presented in Sequence 8. This term also includes its variants or fusion proteins containing GDF-5 residues.

[0048] - The terms "FGFβ" or "FGF-2" as used herein refer to proteins known for cartilage repair. They have also been shown to stimulate the proliferation of chondrocytes in immature rabbits (Reference [Ameye and Young, 2006]). The full-length natural form of human FGF-2 is presented in Sequence 9. One of the recombinant forms of the FGFβ protein is known as trapermin. This term also includes its variants or fusion proteins containing FGFβ residues.

[0049] - The term "FGF-9" as used herein is a protein known to delay articular chondrosis degradation in OA subjects while having a somewhat negative effect on osteophyte formation (reference [Zhou et al., 2016]). The full-length natural form of human FGF-9 is presented in Sequence 10. This term also includes its variants or fusion proteins containing FGF-9 residues.

[0050] - The term "TGF-β" as used herein refers to TGF-β, a protein belonging to the TGF-β family that plays an important role in cartilage maintenance. TGF-β has been suggested as a cartilage enhancer (reference [Wang 2014]). This term also includes its variants or fusion proteins containing TGF-β residues.

[0051] - As used herein, the term "SOX-9" enhancer is intended to refer to a compound that promotes the production of SOX9. In fact, SOX9 is a transcription factor suggested to be essential for the formation of the extracellular matrix (ECM) of chondrocytes.

[0052] - The term "calcitonin" as used herein is a 32-amino acid peptide (sequence 5) of the salmon calcitonin type, which has been proven to have protective activity against both bone and cartilage, more specifically.

[0053] - As used herein, the term "Wnt inhibitor" is intended to refer to a compound that interferes with the WNT pathway.

[0054] - As used herein, the term "anti-MMP13 inhibitor" is intended to refer to a compound that inhibits the activity of matrix metalloproteinase 13 (MMP13). MMP13 is one of the major collagen type II degrading enzymes.

[0055] - As used herein, the term "anti-ADAMTS4 or 5 inhibitor" is intended to refer to a compound that inhibits the enzymatic activity of disintegrin and metalloproteinase (ADAMTS4 or ADAMTS5) having a thrombospondin motif 4 or 5.

[0056] - The terms "biomarker" or "protein marker" are used interchangeably. In the context of the present invention, these are peptides or proteins. "Prognostic biomarkers" provide information about the subject's condition, including but not limited to disease evolution, disease severity, or disease outcome, regardless of any therapy. "Predictive biomarkers" provide information about the effect of the therapy received, including but not limited to efficacy and safety outcomes. The definitions of prognosis and prediction are not mutually exclusive, and thus a biomarker may be both prognostic and predictive. The amount of a biomarker or the expression level of a biomarker is expressed herein in nmol, μmol, mmol, ng, μg, mg, or g for a given protein. The amount or level may be expressed as an absolute value (e.g., 10 ng or 2 μg) or as a concentration (e.g., 10 ng / mL, 2 μg / mL, 10 ng / mmol, or 2 μg / mmol).

[0057] - The term "metabolic biomarker" refers to biomarkers, e.g., CTX-II, ProC2, PIIANP, C2M, ARGS, and AGNx1, to a lesser extent. More specifically, the term "metabolic biomarker" refers to biomarkers of cartilage metabolism, namely collagen and agrecan turnover, such as the degradation and / or synthesis of cartilage matrix components. Different collagen and agrecan biomarkers have been described (reference [Karsdal et al., 2016]).

[0058] - The term "CTX-II" or "CTXII" refers to the C-terminal telopeptide of type II collagen. See Sequence 1. It is a biomarker of type II collagen degradation, which is part of osteoarthritis (e.g., see reference [Duclos et al., 2010]).

[0059] - The term ProC2 refers to a collagen type II neo-epitope generated during the synthesis of collagen type II, and the sequence QDVRQP is recognized as an epitope in the ProC2 assay: ProC2 is a biomarker for the assay of chondrogenesis (type II collagenogenesis). Refer to Sequence 2. ProC2 is associated with the ability for repair and may be a valuable tool for identifying patients with chondrogenesis-induced diseases. ProC2 has been tested as a biomarker in preclinical studies of clinical samples (e.g., see literature [Gudmann et al., 2016; Munk et al., 2016; Gudmann et al., 2014]).

[0060] - The term PIIANP refers to a type II collagen propeptide: type IIA procollagen contains an N-terminal 69-amino acid, cysteine-rich globular domain coded by exon 2 of the collagen type II gene. Type IIA procollagen has been found to be synthesized by osteoarthritis chondrocytes in affected cartilage and may serve as a specific arthritis biomarker reflecting attempts by chondrocytes to repair affected cartilage (Reference [Valdes et al., 2014]).

[0061] - The term C2M refers to a serological type II collagen-degrading neoepitope associated with chondrolysis. C2M is an interhelical fragment of type II collagen produced by MMPs. This biomarker has been described as being associated with drug response, pain measurements, and radiographic severity (Reference [valdes et al. 2014]).

[0062] - The term ARGS refers to neoepitopes generated during agrecan degradation. The assay detects agrecan degradation products in serum and synovial fluid. ARGS levels are associated with progression in chondrosis (references [Struglics et al., 2011; Struglics et al., 2015]).

[0063] - The term AGNx1 relates to a assay for detecting another neoepitope generated during agrecan decomposition.

[0064] - In relation to biomarkers, the terms "quantity" or "expression level" may be used interchangeably.

[0065] The term "physical marker" refers to the joint gap (JSW) and its evaluation in the context of the present invention.

[0066] - The term "SD" refers to the standard deviation and relates to the normal deviation of any test / system.

[0067] - As used herein, the term "chondral disorder" encompasses disorders resulting from damage caused by injury, such as traumatic injury, chondropathy, or arthritis. Examples of chondral disorders that may be treated by the administration of the compounds described herein include, but are not limited to, arthritis, e.g., osteoarthritis, cartilage damage, fractures affecting articular cartilage, or surgical procedures affecting articular cartilage (e.g., microfractures). Degenerative diseases / disorders of cartilage or joints, e.g., chondrocalcinosis, polychondritis, recurrent polychondritis, ankylosing spondylitis, or osteomyelitis, are also included in the above term. The International Cartilage Repair Society proposed an arthroscopic grading system to assess the severity of cartilage defects: Grade 0: (Normal) healthy cartilage, Grade 1: Cartilage has soft spots or vesicles, Grade 2: Visible secondary tears in the cartilage, Grade 3: The lesion has a deep fissure (exceeding 50% of the cartilage layer), Grade 4: The cartilage tear exposes the underlying (subchondral) bone (see, e.g., page 13 of http: / / www.cartilage.org / _files / contentmanagement / ICRS_evaluation.pdf).

[0068] - The term "osteoarthritis" is used to refer to the most common form of arthritis. The term "osteoarthritis" includes both primary and secondary osteoarthritis (see, for example, the literature [The Merck Manual, 17th edition, page 449]). The most common method of classifying / grading osteoarthritis is the use of the Kellgren-Lawrence radiographic grading scale (see table below). Osteoarthritis can be caused by the destruction of cartilage. The cartilage bit may be destroyed, causing pain and swelling in the joints between the bones. Over time, the cartilage may be completely worn away, and the bones will rub together. Osteoarthritis can affect any joint, but it typically concerns the hands and weight-bearing joints, such as the hips, knees, feet, and spine. In a preferred example, osteoarthritis may be knee osteoarthritis or hip osteoarthritis. Osteoarthritis is one of the desirable cartilage disorders that can be treated by administering the compound according to the present invention.

[0069] The Kellgren-Laurence Radiographic Rating Scale (KL) for osteoarthritis is described as follows:

[0070]

[0071] Grades 1 and 2 can be considered less severe forms of the disease, while grades 3 and 4 can be considered more severe forms of the disease.

[0072] - As used herein, the term "chondral damage" refers to cartilage disorder or damage caused particularly by trauma. Chondral damage may occur particularly after traumatic mechanical destruction, especially after additional accidents or surgery (e.g., microfracture surgery). The term "chondral damage" also includes cartilage or osteochondral fractures, damage to the meniscus, and the term microfracture. Additionally, within these definitions, sports-related damage or sports-related wear of joint tissues is considered.

[0073] - The "WOMAC total score" or "WOMAC score" ("WOMAC" for the "Western Ontario and McMaster University Osteoarthritis Index") measures pain (WOMAC pain score), function (WOMAC function score), and stiffness (WOMAC stiffness score). When applied to the assessment of pain and functional impairment associated with cartilage damage, it consists of a questionnaire containing 24 items divided into three subscales (5 items for pain, 2 items for stiffness, and 17 items for physical function) (see references [Bellamy et al., 1988; Wolfe, 1999]). This is a widely known device, particularly widely used in the assessment of OA severity.

[0074] - To evaluate cartilage repair, cartilage volume measurements were performed via magnetic resonance imaging (MRI), which include the lateral volume of cartilage (also referred to as LFTC), the medial volume of cartilage (also referred to as MFTC), the total volume of cartilage (also referred to as LFTC+ MFTC), and the new total average cartilage thickness.

[0075] - The term "thin" cartilage refers to cartilage having a JSW of 3.5 ± 2 SD mm or less.

[0076] - The term "thick" cartilage refers to cartilage having a JSW greater than 3.5 ± 2 SD mm.

[0077] - The term "baseline" means before treatment (i.e., at the time of study participation). It refers to clinical variables, specifically, at the time of study participation (i.e., before treatment with the active compound or placebo), equal to but not limited to the cartilage volume and WOMAC total score of one given subject.

[0078] - The terms "subject" or "patient" refer to both human and non-human animals. The term non-human includes mammals, such as rodents (including mice), rabbits, cats, dogs, horses, cattle, sheep, or primates.

[0079] - "Sensitive" refers to subjects who exhibit a response to the treatment of cartilage disorders using the active compound. Preferably, sensitive subjects (or subjects who exhibit / are likely to exhibit sensitivity / response to treatment) exhibit a particularly higher increase in total cartilage thickness and / or cartilage volume than placebo-treated subjects; that is, they exhibit cartilage repair. Additionally, sensitive subjects exhibit at least a similar improvement in the WOMAC total score compared to placebo. The terms "sensitive" (including both "highly sensitive" and "moderately sensitive") and "insensitive" (including "lowly sensitive") refer to different groups of subjects based specifically on the increase in cartilage volume after treatment with the active compound. Highly sensitive subjects exhibit a high response (i.e., high cartilage repair) to treatment with the active compound, moderately sensitive subjects exhibit an intermediate response (i.e., moderate cartilage repair) to treatment with the active compound, and insensitive subjects exhibit no response or a low response to treatment with the active compound. Highly susceptible subjects preferably have an improvement in WOMAC total scores similar to placebo. Conversely, non-responders have a significantly smaller improvement in WOMAC total scores than placebo.

[0080] More specifically, the terms “intermediate-sensitive,” “highly sensitive,” “unsensitive,” “intermediate responder,” “highly responder,” and “unresponsive” (including low responder) relate to different subject groups based on the increase in cartilage volume and improvement in the WOMAC total score following treatment with the active compound.

[0081] The proposed criteria for evaluating susceptibility / response may be selected from one or more of the following (but not limited to):

[0082] 1. Benign cartilage increase compared to baseline,

[0083] 2. Significantly higher change in cartilage increase than the change in placebo (tested with a linear model adjusted for BMI, KL grade, sex, and age, e.g., alpha = 5%),

[0084] 3. Improvement in WOMAC score compared to baseline, i.e., a decrease (e.g., a decrease of more than 5 points),

[0085] 4. Changes in WOMAC scores were not significantly higher than changes in placebo (e.g., as tested with a linear model adjusted for BMI, KL grade, sex, and age and alpha = 5%).

[0086] - "Response" or "susceptibility" to treatment with an active compound for cartilage disorders is understood as at least one year or even better two years after the first injection, and must be measured by 1) an increase in cartilage volume measured, e.g., by MRI or X-ray, 2) a decrease in the WOMAC total score, or 3) a change in the WOMAC total score not significantly higher than that from placebo (see also definition of "susceptibility").

[0087] - As used herein, the term "placebo effect" should be understood as a change compared to baseline (e.g., dilution of inflammatory components in synovial fluid) caused by the placebo itself, due to contextual influence (e.g., procedure of intra-articular injection by a physician) and patient expectations. The term "low placebo effect" refers to a response size equivalent to the response size at baseline within the standard deviation of the evaluation method.

[0088] - As used herein, the term “storage device” is intended to include any suitable computing or processing device or other device configured or adapted for storing data or information. Examples of electronic devices suitable for use in the present invention include standalone computing devices, local area networks (LANs), wide area networks (WANs), data telecommunications networks including the Internet, intranets, and extranets, and local and distributed computer processing systems. Storage devices include, but are not limited to, magnetic storage media such as floppy disks, hard disk storage media, and magnetic tapes; optical storage media such as CD-ROMs and DVDs; electronic storage media such as RAM, ROM, EPROM, EEPROM, etc.; general hard disks; and hybrids of these categories, such as magnetic / optical storage media.

[0089] - As used herein, the term “stored” refers to a process of encoding information for a storage device. A person skilled in the art may readily adopt any currently known method for recording information on a known medium to produce a product containing expression level information.

[0090] Detailed description of the invention

[0091] It is necessary to predict not only 1) the placebo response in subjects with cartilage disorders, but also 2) the clinical efficacy (particularly regarding cartilage repair) of active compound therapy for the treatment of subjects with cartilage disorders, such as osteoarthritis, cartilage damage, fractures affecting articular cartilage, or surgical procedures affecting articular cartilage (e.g., microfractures). To optimize the management of clinical trials or the treatment of said subjects, particularly regarding cartilage repair, it is important to identify markers that can be used as predictors of 1) the placebo response and / or 2) the response of a given subject to active compound therapy. These predictive biomarkers can be used to identify 1) a group of subjects at risk of exhibiting a placebo effect and / or 2) a high-risk group that is less sensitive to treatment, conversely sensitive (e.g., moderate or good sensitivity), or even highly sensitive. For example, if it is known that one subject with osteoarthritis is at high risk of an intermediate response (or intermediate sensitivity) to treatment, the physician may decide to adjust the dosage regimen, for example, to increase the dose of the active compound administered to said subject and / or to administer the drug over a longer period. This predictive information may be clinically useful in guiding medical decisions, particularly regarding the dosage regimen or the length of treatment to be applied to one patient, or, if necessary (for example, when treatment with the active compound is predicted to be less effective), the timing of joint replacement surgery.

[0092] The remarkable findings of the present invention are based on different studies aimed at identifying potential markers associated with the administration of different active compounds. The biomarkers used in these studies consisted of numerous protein biomarkers. The physical marker was the JSW measurement. The association between protein markers and / or physical markers and clinical response variables was evaluated. The rationale for this type of analysis was to identify combinations of markers capable of predicting 1) placebo response and / or 2) clinical outcomes (particularly regarding cartilage repair) for subjects to be treated with active compounds, such as FGF-18 compounds, BMP-2, BMP-7, GDF-5, FGFβ, FGF-9, SOX-9 enhancers, TGFβ, Wnt inhibitors, anti-MMP13 inhibitors, anti-ADAMTS4 or 5 inhibitors, calcitonin, and any of its variants or fusion proteins. In particular, surprisingly, it was found that 1) JSW measurements can be used to predict the placebo effect, and 2) protein biomarkers can be used in conjunction with JSW measurements to stratify and target specific subject populations.

[0093] Different markers have demonstrated changes in response to active compound therapy or presented changes in outcomes (i.e., cartilage thickness or volume, WOMAC score) by applying criteria for high and low values. Examples include C1M, C3M, and hs-CRP. Surprisingly, the inventors have discovered associations with specific metabolic proteins in combination with physical markers and outcomes (e.g., cartilage repair). The proteins CTX-II and ProC2 are of particular interest. While only CTX-II and / or ProC2 are specifically described herein, it should be noted that other metabolic biomarkers, such as PIIANP, C2M, ARGS, or AGNx1, may be used. Based on the teachings of the present invention, it would be routine for a person skilled in the art to discover thresholds for each of these biomarkers. These proteins are described in the literature as potentially associated with osteoarthritis. For example, CTX-II and ProC2 can be considered as metabolic biomarkers (Bay-Jensen, 2016).

[0094] Surprisingly, the inventors found that for a given subject (having cartilage impairment) exhibiting a JSW higher than 3.5 ± 2 SD mm, a placebo response was high regardless of the biomarker levels: among these subjects, there was a strong placebo effect that likely biased the results of clinical trials and subsequently biased treatment by masking the actual lack of effect of the given active compound. Also surprisingly, it was found that when the amount of metabolic biomarkers, such as CTX-II and / or ProC2, is reduced and the JSW is below a certain threshold, this combination is associated with a superior response to treatment with active compounds, such as FGF-18 compounds, BMP-2, BMP-7, GDF-5, FGFβ, FGF-9, SOX-9 enhancers, TGFβ, Wnt inhibitors, anti-MMP13 inhibitors, anti-ADAMTS4 or 5 inhibitors, calcitonin, and any of its variants or fusion proteins, in subjects suffering from cartilage impairment. These subjects are referred to as highly sensitive (or highly responsive). Conversely, surprisingly, the inventors have also revealed that when the amounts of CTX-II and / or ProC2 increase and JSW exceeds a certain limit, this is associated with the absence or low response to treatment with the active compound (i.e., low sensitivity or insensitivity to treatment with said active compound) in subjects suffering from cartilage disorders. These subjects are referred to as insensitive (or insensitive). Separately, 1) when the amounts of metabolic biomarkers, such as CTX-II and / or ProC2, decrease and JSW exceeds a certain limit, or 2) when the amounts of metabolic biomarkers, such as CTX-II and / or ProC2, increase and JSW is below a certain limit, this may be associated with an intermediate response, i.e., the subject will be of intermediate sensitivity (or intermediate response).

[0095] Accordingly, it is a finding of the present invention that metabolic biomarkers, such as CTX-II and / or ProC2, combined with the physical marker JSW, can be used as predictive markers of a subject's responsiveness to active compound treatments, such as FGF-18 compounds, BMP-2, BMP-7, GDF-5, FGFβ, FGF-9, SOX-9 enhancers, TGFβ, Wnt inhibitors, anti-MMP13 inhibitors, anti-ADAMTS4 or 5 inhibitors, calcitonin and any of its variants or fusion proteins. Preferably, the subject has a cartilage disorder, such as osteoarthritis, cartilage damage, a fracture affecting articular cartilage, or a surgical procedure affecting articular cartilage (e.g., microfracture). In certain embodiments, if the amount of CTX-II is greater than 350 ± 2SD ng / mmol (greater than 240-260% of the normal mean) and / or the amount of ProC2 is greater than 4.2 ± 2SD ng / mL (greater than 120-280% of the normal mean) and JSW is greater than 3.5 ± 2SD mm, the subject will be predicted to be non-sensitive to treatment with the active compound. Conversely, if the amount of CTX-II is less than 350 ± 2SD ng / mmol and / or the amount of ProC2 is less than 4.2 ± 2SD ng / mL and JSW is 3.5 ± 2SD mm or less, the subject will be predicted to be highly sensitive to treatment with the active compound. 1) if the amount of CTX-II is less than 350 ± 2SD ng / mmol and / or the amount of ProC2 is less than 4.2 ± 2SD ng / mL and JSW is greater than 3.5 ± 2SD mm, or 2) if the amount of CTX-II is greater than 350 ± 2SD ng / mmol and / or the amount of ProC2 is greater than 4.2 ± 2SD ng / mL and JSW is less than 3.5 ± 2SD mm, the subject will be predicted to be moderately sensitive to treatment with the active compound.

[0096] Accordingly, the present invention relates to a method for eliminating the risk of a clinical trial or determining a placebo effect during a clinical trial (wherein the clinical trial relates to the treatment of cartilage disorders in subjects using an active compound) or treatment using an active compound, and

[0097] a) a step of measuring the joint space (JSW) at least one knee of one subject, and

[0098] b) A step of predicting the risk of a placebo effect on the subject from the result of step a).

[0099] It is a method that includes.

[0100] According to the above method, the presence of JSW greater than 3.5 ± 2 SD mm predicts a placebo effect. Conversely, the presence of JSW less than or equal to 3.5 ± 2 SD mm predicts no or low placebo effect. From the above prediction:

[0101] - For clinical trials, clinicians can easily classify subjects based on their likelihood of exhibiting or not exhibiting a placebo effect.

[0102] - For the treatment of cartilage disorders, a physician can easily determine whether the subject is likely to be sensitive to a given active compound treatment (including both moderate-sensitive and high-sensitive).

[0103] Alternatively, the present invention relates to a method for eliminating the risk of a clinical trial or determining a placebo effect in a subject during a clinical trial (wherein the clinical trial relates to the treatment of cartilage disorders in a subject using an active compound) or treatment using an active compound, and

[0104] a) a step of determining the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject;

[0105] b) a step of measuring the joint space (JSW) at least one knee of the subject, and

[0106] c) a step of predicting the risk of placebo effect from the results of steps a) and b),

[0107] It is a method that includes, wherein steps a) and b) are performed in any order.

[0108] Before determining the amount of at least one of the biomarkers, it is necessary to obtain a sample of the subject (or biological sample or test sample) by, for example, collecting blood, serum, synovial fluid, or urine. Accordingly, the present invention relates to a method for determining a placebo effect in a subject during a clinical trial (wherein the clinical trial relates to the treatment of cartilage disorders in a subject using an active compound) or treatment using an active compound, and

[0109] a) obtaining a sample from the patient and determining the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from the sample;

[0110] b) a step of measuring the joint space (JSW) at least one knee of the subject, and

[0111] c) a step of predicting the risk of a placebo effect on the subject from the results of steps a) and b),

[0112] It is a method that includes, wherein steps a) and b) are performed in any order.

[0113] According to the above method, the presence of CTX-II greater than 350 ± 2SD ng / mmol (240-260% greater than the normal mean) and / or ProC2 greater than 4.2 ± 2SD ng / mL (120-280% greater than the normal mean) with JSW greater than 3.5 ± 2SD mm predicts a placebo effect. Similarly, 1) CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm, or 2) CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL with JSW less than 3.5 ± 2SD mm predicts a placebo effect. Conversely, the presence of CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL, along with JSW less than 3.5 ± 2SD mm, is predicted to result in no or low placebo effect.

[0114] From the above prediction:

[0115] - For clinical trials, clinicians can easily classify subjects based on their likelihood of exhibiting or not exhibiting a placebo effect.

[0116] - For the treatment of cartilage disorders, a physician can easily determine whether the subject is likely to be sensitive to a given active compound treatment (including both moderate-sensitive and high-sensitive).

[0117] The present invention also relates to a method for eliminating the risk of clinical trials associated with the treatment of cartilage disorders in subjects with active compounds, and

[0118] c) a step of determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject,

[0119] d) a step of measuring the joint space (JSW) at at least one knee of the subject, and

[0120] e) a step of predicting the response of the subject to treatment using the active compound from the results of steps a) and b),

[0121] It is a method that includes, wherein steps a) and b) are performed in any order.

[0122] Before determining the amount of at least one of the biomarkers, it is necessary to obtain a sample of the subject (or biological sample or test sample) by, for example, collecting blood, serum, synovial fluid, or urine. Accordingly, the present invention relates to a method for eliminating the risk of clinical trials associated with the treatment of cartilage disorders in a subject with an active compound, and

[0123] a) obtaining a sample from the patient and determining the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from the sample,

[0124] b) a step of measuring the joint space (JSW) at least one knee of the subject, and

[0125] c) a step of predicting the response of the subject to treatment using the active compound from the results of steps a) and b),

[0126] It is a method that includes, wherein steps a) and b) are performed in any order.

[0127] According to the above method, the presence of CTX-II greater than 350 ± 2SD ng / mmol (greater than 240-260% of the normal mean) and / or ProC2 greater than 4.2 ± 2SD ng / mL (greater than 120-280% of the normal mean) along with JSW greater than 3.5 ± 2SD mm predicts no or low response to treatment with the active compound (i.e., insensitivity or low sensitivity). Therefore, the subject will be predicted to be insensitive (or non-responder). Conversely, the presence of CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL along with JSW less than 3.5 ± 2SD mm predicts a good response to treatment with the active compound (i.e., good sensitivity). Therefore, the subject is predicted to be highly sensitive to treatment with the active compound (i.e., will be a highly responder). The presence of 1) CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL and JSW greater than 3.5 ± 2SD mm, or 2) CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and JSW less than 3.5 ± 2SD mm predicts an intermediate response (i.e., intermediate sensitivity) to treatment with the active compound. Therefore, the subject is predicted to be moderately sensitive to treatment with the active compound (i.e., will be a moderate responder). From the above prediction, the physician can easily select only subjects predicted to be sensitive to treatment with a given active compound, including both moderate sensitivity and excellent sensitivity.

[0128] The present application also includes a method for selecting a subject having a cartilage disorder based on the possibility of his susceptibility to said treatment in order to include or exclude him / her in a treatment or clinical trial using an active compound, and

[0129] a) determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject, and measuring the joint gap (JSW) at at least one knee of the subject, wherein the amount of at least one of these proteins and the gap of the JSW predict the risk of the subject being susceptible or insensitive to the treatment, and

[0130] b) selecting an intermediate-sensitive and / or good-sensitive subject as suitable for the treatment or clinical trial

[0131] Includes

[0132] Before determining the amount of at least one of the biomarkers, it is necessary to obtain a sample of the subject (or biological sample or test sample) by, for example, collecting blood, serum, synovial fluid, or urine. Accordingly, the present invention relates to a method for selecting a subject having a cartilage disorder to be included in or excluded from treatment or clinical trials using an active compound based on the potential for its susceptibility to treatment, and

[0133] a) obtaining a sample from the patient, determining from the sample the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2, and measuring the joint gap (JSW) at at least one knee of the subject, wherein the amount of at least one of these proteins and the gap of the JSW predict the risk of the subject being susceptible or insensitive to the treatment, and

[0134] b) selecting an intermediate-sensitive and / or good-sensitive subject as suitable for the treatment or clinical trial

[0135] Includes

[0136] According to the above method, subjects exhibiting CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL, along with JSW greater than 3.5 ± 2SD mm, will preferably be excluded from treatment with the active compound. Conversely, subjects exhibiting CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL, along with JSW less than 3.5 ± 2SD mm, will preferably be included for treatment with the active compound (i.e., preferably selected). Subjects exhibiting 1) CTX-II of less than 350 ± 2SD ng / mmol and / or ProC2 of less than 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm, or 2) subjects exhibiting CTX-II of greater than 350 ± 2SD ng / mmol and / or ProC2 of greater than 4.2 ± 2SD ng / mL with JSW less than 3.5 ± 2SD mm will also be included in an alternative administration regimen for treatment with the active compound, possibly in which the dose of the active compound is increased or / or the active compound is administered for a longer treatment period.

[0137] Alternatively, a method for treating a subject with cartilage disorders with an active compound is described herein, and

[0138] a) determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject, and measuring the joint gap (JSW) at at least one knee of the subject, wherein the amount and width predict the risk of the subject being excellently susceptible to treatment with the active compound,

[0139] b)

[0140] i. CTX-II < 350 ± 2SD ng / mmol, and / or ProC2 < 4.2 ± 2SD ng / mL and JSW ≤ 3.5 ± 2SD mm,

[0141] ii. CTX-II < 350 ± 2SD ng / mmol and / or ProC2 < 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm, or

[0142] iii. A step of selecting a subject having CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL, along with JSW less than 3.5 ± 2SD mm, and

[0143] c) a step of administering the active compound to the selected target.

[0144] Includes

[0145] Before determining the amount of at least one of the biomarkers, it is necessary to obtain a sample of the subject (or biological sample or test sample) by, for example, collecting blood, serum, synovial fluid, or urine. Accordingly, the present invention relates to a method for treating a subject having a cartilage disorder with an active compound, and

[0146] a) obtaining a sample from the patient, measuring the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from the sample, and measuring the joint gap (JSW) in one or more knees of the subject, wherein the amount and width predict the risk of the subject being excellently susceptible to treatment with the active compound,

[0147] b)

[0148] i. CTX-II < 350 ± 2SD ng / mmol, and / or ProC2 < 4.2 ± 2SD ng / mL and JSW ≤ 3.5 ± 2SD mm,

[0149] ii. CTX-II < 350 ± 2SD ng / mmol and / or ProC2 < 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm, or

[0150] iii. A step of selecting a subject having CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL, along with JSW less than 3.5 ± 2SD mm, and

[0151] c) a step of administering the active compound to the selected target.

[0152] Includes

[0153] Intermediately susceptible subjects (1) having CTX-II of less than 350 ± 2SD ng / mmol and / or ProC2 of less than 4.2 ± 2SD ng / mL with JSW greater than 3.5 ± 2SD mm or 2) having either CTX-II of greater than 350 ± 2SD ng / mmol and / or ProC2 of greater than 4.2 ± 2SD ng / mL with JSW of less than 3.5 ± 2SD mm) can be treated with the same dosing regimen as good-susceptible subjects (having CTX-II of less than 350 ± 2SD ng / mmol and / or ProC2 of less than 4.2 ± 2SD ng / mL and JSW of less than 3.5 ± 2SD mm), but these subjects may have better benefits with an alternative dosing regimen.

[0154] Accordingly, a method for treating a subject with cartilage disorders with an active compound is also described herein, and

[0155] a) determining the amount of at least one biomarker selected from the group consisting of CTX-II and / or ProC2 from a sample from the subject, and measuring the joint gap (JSW) at at least one knee of the subject, wherein the amount and width predict the risk of the subject having intermediate sensitivity to treatment with the active compound,

[0156] b)

[0157] i. CTX-II < 350 ± 2SD ng / mmol, and / or ProC2 < 4.2 ± 2SD ng / mL and JSW < 3.5 ± 2SD mm or

[0158] ii. A step of selecting a subject having CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and JSW less than 3.5 ± 2SD mm, and

[0159] c) a step of administering the active compound to the selected subject according to an alternative administration regimen in which the dose of the active compound is increased and / or the active compound is administered for a longer treatment period than the usual administration regimen for the active compound.

[0160] Includes

[0161] Before determining the amount of at least one of the biomarkers, it is necessary to obtain a sample of the subject (or biological sample or test sample) by, for example, collecting blood, serum, synovial fluid, or urine. Accordingly, the present invention relates to a method for treating a subject having a cartilage disorder with an active compound, and

[0162] a) obtaining a sample from the patient, measuring the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2 from the sample, and measuring the joint gap (JSW) in one or more knees of the subject, wherein the amount and width predict the risk of the subject being of intermediate sensitivity to treatment with the active compound,

[0163] b)

[0164] i. CTX-II < 350 ± 2SD ng / mmol, and / or ProC2 < 4.2 ± 2SD ng / mL and JSW < 3.5 ± 2SD mm, or

[0165] ii. A step of selecting a subject having CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and JSW less than 3.5 ± 2SD mm, and

[0166] c) a step of administering the active compound to the selected subject according to an alternative administration regimen in which the dose of the active compound is increased and / or the active compound is administered for a longer treatment period than the usual administration regimen for the active compound.

[0167] Includes

[0168] The amount (or expression level) of one or more biomarkers in the samples may be compared, if necessary, to a reference amount from a reference sample (alternatively a reference expression level, baseline amount, or baseline expression level). The reference level may be obtained from a healthy subject or from the very same patient to be diagnosed or treated before or during the treatment. If necessary, the width of the subject's joint space may be compared to a reference joint space width (or baseline joint space width). The reference width may be obtained from a healthy subject or from the very same patient to be diagnosed or treated before or during the treatment.

[0169] The present invention further comprises an active compound for use in the treatment of subjects having cartilage disorders, wherein the subjects are characterized by having any one of 1) CTX-II less than 350 ng / mmol and / or ProC2 less than 4.2 ng / mL and JSW less than 3.5 ± 2SD mm, 2) CTX-II less than 350 ng / mmol and / or ProC2 less than 4.2 ng / mL and JSW greater than 3.5 ± 2SD mm, or 3) CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and JSW less than 3.5 ± 2SD mm. Intermediately susceptible subjects may be treated with the same dosing regimen as good-susceptible subjects, but these subjects may have a better benefit with an alternative dosing regimen.

[0170] Accordingly, the present invention also comprises an active compound for use in an alternative administration regimen (e.g., increased dose of the active compound and / or a longer duration of treatment) in the treatment of subjects having cartilage disorders, wherein the subjects are characterized by having either 1) CTX-II of less than 350 ng / mmol and / or ProC2 of less than 4.2 ng / mL and JSW of greater than 3.5 ± 2SD mm, or 2) CTX-II of greater than 350 ± 2SD ng / mmol and / or ProC2 of greater than 4.2 ± 2SD ng / mL and JSW of less than 3.5 ± 2SD mm. Accordingly, subjects who do not meet these criteria are preferably excluded from treatment with the active compound.

[0171] The present invention also relates to a assay for determining sensitivity to treatment with an active compound or for determining a therapeutic regimen with a given active compound, wherein the assay comprises: (a) applying a test sample from a human subject diagnosed with cartilage disorder to at least one assay for determining the amount of at least one of CTX-II and / or ProC2; (b) determining the amount of at least one of CTX-II and / or ProC2; and (c) determining the sensitivity or insensitivity of the subject to treatment with said active compound in terms of joint space width measurements from the result of step b). According to said assay, the presence of CTX-II greater than 350 ng / mmol and / or ProC2 greater than 4.2 ng / mL with a JSW greater than 3.5 ± 2 SD mm predicts insensitivity to treatment with the active compound. Conversely, the presence of CTX-II at less than 350 ng / mmol and / or ProC2 at less than 4.2 ng / mL, along with JSW at less than 3.5 ± 2SD mm, predicts excellent susceptibility. Alternatively, 1) JSW at greater than 3.5 ± 2SD mm, along with CTX-II at less than 350 ng / mmol and / or ProC2 at less than 4.2 ng / mL, or 2) CTX-II at greater than 350 ± 2SD ng / mmol and / or ProC2 at greater than 4.2 ± 2SD ng / mL and JSW at less than 3.5 ± 2SD mm, predicts moderate susceptibility. From the results of the above test, physicians can easily select only subjects predicted to be susceptible to treatment with the active compound, including those with both moderate and excellent susceptibility. Before determining the amount of at least one of the biomarkers in the test disclosed above, it is necessary to obtain a sample of the biomarker (or test) of the subject by, for example, collecting blood, serum, synovial fluid, or urine.

[0172] The present invention also relates to a test for selecting a therapeutic regimen for a human subject having a cartilage disorder, wherein the test comprises: (a) applying a test sample from a human subject diagnosed as having a cartilage disorder to at least one test for determining the amount of at least one of CTX-II and / or ProC2; (b) determining from step a) the likelihood that the subject is moderately sensitive or highly sensitive to treatment with an active compound by considering joint space width measurements; and (c) determining an appropriate therapeutic regimen for the subject from the result of step b). Once the assay is performed, based on the recognition that the above amount / width is associated with a superior response to the compound, when the subject has less than 350 ng / mmol CTX-II and / or less than 4.2 ng / mL ProC2 with a JSW of 3.5 ± 2SD mm or less, or based on the recognition that the above amount / width is associated with an intermediate response to the compound, when the subject has 1) less than 350 ng / mmol CTX-II and / or less than 4.2 ng / mL ProC2 with a JSW greater than 3.5 ± 2SD mm, or 2) greater than 350 ± 2SD ng / mmol CTX-II and / or greater than 4.2 ± 2SD ng / mL ProC2 and less than 3.5 ± 2SD mm JSW, the subject may be selected and treated for an appropriate dosing regimen comprising an effective amount of the active compound; Subjects are excluded from treatment with the active compound when they have CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL along with JSW greater than 3.5 ± 2SD mm based on the recognition that the above amounts are associated with an inappropriate response to treatment with the compound. Intermediately susceptible subjects may be treated with the same dosing regimen as well-sensitive subjects, but these subjects may have a better benefit from an alternative dosing regimen.Therefore, alternatively, the intermediate-sensitive subjects may be selected for and treated with an alternative administration regimen of the active compound (e.g., increased dose of the active compound and / or a longer duration of treatment).

[0173] A method for treating a human subject having a cartilage disorder is further disclosed, comprising the steps of: (a) testing a biological sample of a subject diagnosed as having a cartilage disorder for the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2; (b) a step of measuring the joint space width and (c) a step of administering a therapeutic regimen comprising a composition containing an effective amount of an active compound to a subject when the subject 1) has CTX-II less than 350 ± 2SD ng / mmol and / or ProC2 less than 4.2 ± 2SD ng / mL and a JSW of 3.5 ± 2SD mm or less, 2) has CTX-II less than 350 ng / mmol and / or ProC2 less than 4.2 ng / mL with a JSW greater than 3.5 ± 2SD mm, or 3) has CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and a JSW less than 3.5 ± 2SD mm. Intermediately susceptible subjects may be treated with the same regimen as good-susceptible subjects, but these subjects will have a better benefit from an alternative regimen. Accordingly, a method for treating a human subject having a cartilage disorder is alternatively disclosed, comprising the steps of: (a) testing a biological sample of a subject diagnosed as having a cartilage disorder for the amount of one or more biomarkers selected from the group consisting of CTX-II and / or ProC2; (b) measuring the joint space width; and (c) when the subject 1) has less than 350 ± 2SD ng / mmol of CTX-II and / or less than 4.2 ± 2SD ng / mL of ProC2 and JSW greater than 3.5 ± 2SD mm, or 2) more than 350 ± 2SD ng / mmol of CTX-II and / or more than 4.2 ± 2SD ng / mL of ProC2 and JSW greater than 3.In cases where the duration is less than 5 ± 2 SD mm, the method includes the step of administering an alternative therapeutic regimen to the subject that includes a composition containing an active compound (e.g., an increased dose of the active compound and / or a longer duration of treatment).

[0174] Overall, in the context of the present invention, CTX-II and / or ProC2 have been found to be useful predictive biomarkers of a subject's response to an active compound for the treatment of placebo response and cartilage disorders. Therefore, they are considered predictive biomarkers. Other metabolic biomarkers, such as C2M, ARGS, or AGNx1, may be used alternatively.

[0175] In the context of the present invention as a whole, preferred cartilage disorders are selected from the group consisting of osteoarthritis, cartilage damage, fractures affecting articular cartilage, or surgical procedures affecting articular cartilage, such as microfractures.

[0176] In the context of the present invention as a whole, for example, in the context of any one of the methods, uses, tests or kits according to the present invention, the preferred active compound may be selected from FGF-18 compounds (including terminally determined FGF-18 compounds, e.g., sprifermin), BMP-2, BMP-7, GDF-5, FGFβ, FGF-9, SOX-9 enhancers, TGFβ, Wnt inhibitors, anti-MMP13 inhibitors, anti-ADAMTS4 or 5 inhibitors, calcitonin and any variants or fusion proteins thereof.

[0177] As an example of an administration schedule, when the active compound according to the present invention as a whole is an FGF-18 compound, said FGF-18 compound is typically administered intra-articularly to a subject (preferably a highly sensitive subject) once a week for three weeks per treatment cycle at a dose of 100 mcg per injection. For these subjects predicted to be highly sensitive, the proposed administration regimen is intra-articular administration of an FGF-18 compound at a dose of 30 mcg per injection once a week for three weeks per treatment cycle. At this time, the preferred dose is 100 mcg per injection, and in the case of hypersensitivity, it may be reduced to 30 mcg per injection, but it should be understood that the present invention is not limited to the above dosage. Accordingly, the FGF-18 compound may be administered intra-articularly at a dose of 50 to 300 mcg per injection, preferably 60 to 250 mcg, or even preferably 100 to 200 mcg. For hypersensitive subjects, the above dose may be reduced, for example, to about half or about one-third. For example, if the normal dose is 50 mcg per injection, the reduced dose may be 16 to 25 mcg per injection. The FGF-18 compound is generally administered for at least one treatment cycle. Preferably, the cycle is repeated at least once after the start of the first treatment cycle, for example, for 6 months (or about 26 weeks). Up to four treatment cycles within two years have shown promising results (see Fig. 1). Clearly, if the subject is predicted to be moderately sensitive to the active compound, an alternative administration regimen may consist of additional treatment cycles.

[0178] As another example of a dosing schedule, where the active compound according to the present invention as a whole is a calcitonin compound, said calcitonin compound should be administered orally, typically at a dose of 0.8 mg twice daily, for up to 24 months or longer if necessary, or with any typical dosing regimen administered for said compound. If necessary, the oral dose and duration of treatment may be adapted to the subject's response or potential for response to said treatment for intermediate responder benefit from calcitonin treatment.

[0179] In another example of an administration schedule, where the active compound according to the present invention as a whole is a BMP-7 compound, said BMP-7 compound is typically administered intra-articularly at a dose of about 0.03 mcg / ml to 0.3 mcg / ml per injection, e.g. 0.3, 0.1, or 0.3 mcg / ml, possibly repeated or in any typical administration regimen for said compound. If necessary, the oral dose and duration of treatment may be adapted to the subject's response or potential for response to said treatment.

[0180] In the context of the present invention as a whole, the positive test or other determination of one or more of the biomarkers of the present invention may be performed before or during treatment. In practice, the administration regimen during treatment may also need to be adapted to the new marker situation.

[0181] In the context of the present invention as a whole, if a patient has a CTX-II range of 350 ± 2SD ng / mmol or a ProC2 range of 4.2 ± 2SD ng / mL, it is recommended to complete diagnostic or biomarker testing using another metabolic biomarker. For example, if the level of CTX-II is in the range of 350 ± 2SD ng / mmol, the level(s) of ProC2 may be considered. In the rare case where a patient exhibits a CTX-II range of 350 ± 2SD ng / mmol and a ProC2 range of 4.2 ± 2SD ng / mL, it will be recommended to complete diagnostic or biomarker testing using another type of biomarker, such as an inflammatory biomarker or an SNP biomarker, such as that disclosed in WO2014023703.

[0182] In the context of the present invention as a whole, the width of the joint space is preferably determined by X-ray radiography or MRI technology. Obviously, a person skilled in the art may use alternative technologies if preferred.

[0183] In another embodiment of the present invention, a system for obtaining data (and a computer-readable medium for inducing a computer system) is also provided. The data may be used to evaluate suitability for a clinical trial for treatment with an active compound in particular in a subject, to monitor the therapeutic efficacy of the active compound for a given subject, or simply to monitor disease progression. The system may be used during a clinical trial, when treatment with an active compound for treating cartilage disorders is being considered, or when treatment with said compound is already in progress.

[0184] Accordingly, in one embodiment of the present invention, a computer system for obtaining data along with data related to JSW for said subject from at least one test sample obtained from said subject having a cartilage disorder is included, and the system includes (a) one or more determination modules configured to receive data related to JSW from said patient along with one or more test samples from said subject and to perform one or more analyses on said one or more test samples to determine the amount of one or more biomarkers according to the present invention; (b) one or more storage devices configured to store data output from said determination modules; and (c) one or more display modules for displaying content based partially on the data output from said determination modules (wherein the content includes a signal indicating the presence of one or more of these conditions and optionally the absence of any one of these conditions).

[0185] A computer-readable medium may have computer-readable instructions written thereon to define a software module for implementing a method on a computer. In such cases, the computer-readable storage medium may include (a) a command for comparing data stored on a storage device with reference data to provide a comparison result (wherein the comparison is based on the amount of at least one of the biomarkers and the width of the joint space according to the present invention); and (b) a command for displaying content based partially on the data output from the determination module (wherein the content includes a signal indicating the presence of at least one of the conditions and optionally the absence of one or more of the conditions).

[0186] A computer-readable storage medium may be any available type of medium that can be accessed by a computer. A computer-readable storage medium includes volatile and non-volatile, removable and non-removable type media implemented by any method or technique for storing information such as computer-readable instructions, data structures, program modules or other data. A computer-readable storage medium includes, but is not limited to, RAM (Random Access Memory), ROM (Read-Only Memory), EPROM (Erasable Programmable Read-Only Memory), EEPROM (Electrically Erasable Programmable Read-Only Memory), flash memory or other memory technologies, CD-ROM (Compact Disc Read-Only Memory), DVD (Digital Multifunction Disc) or other optical storage media, magnetic cassettes, magnetic tapes, magnetic disk storage or other magnetic storage media, other types of volatile and non-volatile memory, and any other type of medium that can be accessed by a computer and may be used to store desired information and may include any suitable combination of the above.

[0187] Computer-readable data implemented on one or more computer-readable media may define instructions as part of one or more programs that instruct a computer to perform one or more of the functions described herein and / or various embodiments, variations and combinations thereof as a result of execution by a computer, for example. Such instructions may be written in a plurality of programming languages, for example, Java, J#, Visual Basic, C, C#, C++, Fortran, Pascal, Eipel, Basic, COBOL assembly language, etc., or any various combination thereof. The computer-readable media on which such instructions are implemented may exist on one or more components of any one of the systems, or the computer-readable storage media described herein may be distributed across one or more of these components.

[0188] A computer-readable medium may be transportable so that instructions stored therein can be loaded onto any computer resource to implement aspects of the invention discussed herein.

[0189] Information determined by the determination module can be read by a storage device. The storage device is adapted or configured to record expression level or protein level information. This information may be provided in a digital form that can be transmitted and read electronically, for example, over the Internet, on a diskette, via a USB (Universal serial bus), or through any other suitable communication mode.

[0190] In summary, in the context of the present invention, it should be understood that in any one of the methods, uses, tests, computer systems, or kits according to the present invention, it is necessary to obtain a sample of a subject (or biological sample or test sample) by, for example, blood, serum, synovial fluid, or urine collection before determining the amount of at least one of the biomarkers of the present invention (e.g., CTX-II and / or ProC2). This may also be obtained from cells, tissues, cartilage, or synovial fluid, without limitation.

[0191] An individual suffering from a cartilage disorder and to be tested, tested and / or treated according to any of the methods, uses, tests, kits and other computer systems described herein is a human subject who is a candidate for treatment using an active compound for treating said cartilage disorder. In a preferred embodiment, the individual is diagnosed with a cartilage disorder or exhibits symptoms of a cartilage disorder.

[0192] In a further embodiment, the present invention comprises a kit comprising means for carrying out the method described above and instructions for use. Preferably, the kit comprises means for detecting the presence of one or more of the biomarkers according to the present invention (e.g., CTX-II and / or ProC2) and quantifying the same. The kit may comprise means for detecting the presence of at least two of the biomarkers according to the present invention and quantifying the same. Results obtained from the kit should be combined with JSW measurement results before any conclusions are reached.

[0193] The method and kit according to the present invention are useful for clinical diagnostic purposes. However, the term “diagnosis” as used herein is not limited to clinical or medical uses, and the diagnostic method and kit of the present invention claimed herein are also useful during any research application and clinical trial in which it is desirable to test a subject for the presence or absence of any marker described herein.

[0194] In the context of the present invention, the presence and quantification of one or more of the biomarkers according to the present invention (e.g., CTX-II and / or ProC2) may be detected by any technique known to a person skilled in the art, including ELISA.

[0195] Other embodiments of the invention within the scope of the claims of this application will be apparent to those skilled in the art by considering the specification or practice of the invention disclosed herein. This specification, together with the embodiments, is to be regarded merely as illustrative, and the scope and spirit of the invention are intended to be defined by the claims following the embodiments.

[0196] Description of the drawing:

[0197] Fig. 1: Schematic diagram of the administration regimen used for the FGF-18 compound in the FORWARD study.

[0198] Fig. 2:Proposed reinforcement strategy from the FDA Draft Guidance "Reinforcement Strategy for Clinical Trials Supporting Approval of Human Drugs and Biological Products" (FDA, 2012).

[0199] Fig. 3: Placebo response in KL2 and KL3 patients highlighted by total WOMAC scores (i.e., placebo effect as a function of JSW). A) Change in total WOMAC from baseline for KL2 and KL3 groups as a function of elapsed time since the mean of the initial IA injection group (data are mean + / -SD). B) Total WOMAC scores for KL2 and KL3 groups as a function of elapsed time since the mean of the initial IA injection group (data are median).

[0200] Fig. 4: Placebo response as a function of biomarker levels highlighted by total WOMAC scores. A) Change in total WOMAC in the groups as a function of elapsed time based on CTX-II levels (Group 1: ≤ 350 ng / mmol and Group 2: > 350 ng / mmol) (data are mean + / -SD). B) Change in total WOMAC in the groups as a function of elapsed time based on ProC2 levels (Group 1: ≤ 4.2 ng / mmol and Group 2: > 4.2 ng / mmol) (data are mean + / -SD).

[0201] Fig. 5: Placebo response combining both JSW measures and CTX-II levels, highlighted by the group mean total WOMAC score as a function of elapsed time (months). Data were retrieved from the placebo arm of the FORWARD study.

[0202] Fig. 6:a) Difference in active arm to placebo as a function of patient KL grade, as evidenced by total WOMAC score, and as a function of elapsed time since the first IA injection. b) Difference in active arm to placebo as a function of patient CTX-II level, as evidenced by total WOMAC score, and as a function of elapsed time since the first IA injection. c) Difference in active arm to placebo as a function of patient KL grade, as evidenced by cartilage thickness level, and as a function of elapsed time since the first IA injection. d) Difference in active arm to placebo as a function of patient CTX-II level, as evidenced by cartilage thickness level, and as a function of elapsed time since the first IA injection. In Figure 6 ad, each group of treated patients (Groups 2 through 5) from the FORWARD study was compared to the placebo group (Group 1) (comparison based on difference from placebo). Data are group medians.

[0203] Fig. 7: Placebo response to a combination of both JSW (expressed as KL2 and KL3) and CTX-II levels, highlighted by changes in the group mean (SEM) WOMAC pain score as a function of elapsed time (months). The number of patients in each subgroup is as follows: n≒380 (KL2 with low CTX-II), n≒90 (KL2 with high CTX-II), n≒40 (KL3 with low CTX-II), and n≒20 (KL3 with high CTX-II).

[0204] Fig. 8: Proposed reinforcement strategy for any clinical trial in osteoarthritis based on the 2012 FDA Draft Guidance.

[0205] Explanation of the sequence:

[0206] Sequence 1: Amino acid sequence of biomarker CTX-II.

[0207] Sequence 2: Amino acid sequence of biomarker ProC2.

[0208] Sequence 3: Amino acid sequence of natural human FGF-18.

[0209] Sequence 4: Amino acid sequence of recombinant truncated FGF-18 (trFGF-18).

[0210] Sequence 5: Amino acid sequence of salmon calcitonin.

[0211] Sequence 6: Amino acid sequence of human BMP-2.

[0212] Sequence 7: Amino acid sequence of human BMP-7.

[0213] Sequence 8: Amino acid sequence of human GDF-5.

[0214] Sequence 9: Amino acid sequence of human FGFβ.

[0215] Sequence 10: Amino acid sequence of human FGF-9.

[0216] Examples

[0217] Statistical methods

[0218] Treatment effects for primary endpoints were evaluated via dose-range using repeated measures analysis of variance (ANOVA, PROC MIXED in SAS) for absolute change from baseline, including baseline values, treatment groups, time, country as a factor, and time-period treatment as an interaction. The primary efficacy analysis consisted of testing linear dose relationships and overall treatment effects at 2 years. The significance level was set at 5% two-sided for both trials. Pairwise comparisons (between Sprifermin and placebo, and between Sprifermin dose and treatment groups) were performed within the context of this modeling framework. For each pairwise comparison, the difference between the treatment and the corresponding 95% confidence interval (CI) and p-value is indicated. Treatment effects for consecutive secondary endpoints, such as MRI endpoints, WOMAC endpoints (total, pain, function, and stiffness scores), and X-ray endpoints at each time point and over time, were evaluated using the same ANOVA model used for the primary endpoints. Treatment effects on binary efficacy endpoints, such as the OMERACT-OARSI responder rate, were evaluated using logistic regression. Point estimates for each pairwise comparison, along with corresponding 95% CI and p-values, are provided.

[0219] Pain and Functional Assessment

[0220] WOMAC is a validated instrument used to evaluate symptomatic variation in clinical OA studies. This clinical score was developed in 1981 and has been considered a valid instrument by both clinical researchers and regulatory agencies. WOMAC is widely used and extensively validated in clinical studies of hip and knee OA.

[0221] Subjects had to answer all 24 questions (i.e., 5 for pain, 2 for stiffness, and 17 for physical function assessment) using an 11-box NRS assessment (categories 0 to 10) based on the past 48 hours for Example 1 or a 100 mm VAS (visual analog scale, scoring each question from 0 to 100) based on the past 24 hours for Example 2. Different forms of the questionnaire exist for the right and left knees: to reduce confusion of the WOMAC response due to symptoms in the contralateral knee, subjects used a WOMAC questionnaire specific to the target knee.

[0222] For the administration of the questionnaire, the guidelines for the WOMAC 3.1 index were followed for both Examples 1 and 2.

[0223] Another device for evaluating pain and function is KOOS (Knee Injury and Osteoarthritis Outcome Score, literature [Collins et al. 2016]).

[0224] X-ray evaluation of JSW

[0225] Changes in JSW measured by X-rays are recognized endpoints approved by the European Medicines Agency and the U.S. Food and Drug Administration for use in efficacy studies in OA. JSW was measured using standardized techniques. Additionally, KL grades were evaluated in Example 2 using X-rays.

[0226] qMRI evaluation

[0227] The primary endpoint for DBPC treatment stage was the change from baseline in cartilage thickness at the common femorotibial joint as evaluated by qMRI at 2 years in mITT. Cartilage thickness at the common femorotibial joint was calculated in two ways:

[0228] 1. Average cartilage thickness (total volume divided by total surface area),

[0229] 2. Total cartilage thickness (sum of cartilage thickness in the medial and lateral compartments).

[0230] The treatment effect on the primary endpoint was evaluated via dose-range using repeated measures analysis of variance (ANOVA) on the absolute change from baseline, including treatment group, time point, and (pooled) country as a fixed factor, and baseline value as a covariate and treatment up to time point as an interaction. Repeated measures over time were explained using "unstructured" covariance patterns.

[0231] Pairwise comparisons of the absolute change in cartilage thickness from baseline (treatment group with compounds versus placebo) were performed within the context of the modeling framework described above. For each pairwise comparison, the difference between the treatment and the corresponding 95% confidence interval (CI) and p-value is indicated. P-values ​​(corresponding to Type 3 trials of fixed effects) are reported for all combined time points (i.e., baseline value, treatment, time point, treatment-specific time point interaction, country) and for all covariates in the original "whole" model for all time points. Estimated coefficients, p-values, and 95% CIs are indicated for (i) dose-relation (linear trend) and (ii) each pairwise comparison between the dose level and placebo, both collectively and at each time point.

[0232] To evaluate the robustness of the primary outcome, the linear dose-relationship and overall treatment effect trials were repeated using the PP analysis set. For the mITT analysis set, non-parameter analysis was performed on the ordered data of cartilage thickness in the total femorotibial joint as an alternative method for the primary analysis. The data were ordered by the magnitude of the absolute change from baseline over 2 years during the DBPC treatment phase using rank transformation.

[0233] Biomarker measurement

[0234] Serological and urinary biochemical markers of synovial inflammation, as well as bone and joint tissue turnover, were evaluated. Potential biomarkers of cartilage metabolism included, but were not limited to, neo-epitopes of collagen type II propeptides (ProC2) and C-telopeptide cross-links of type II collagen (CTX-II). Blood and urine samples for the evaluation of systemic biomarkers were collected at the following times: Week 0 (before the first injection of Sprifermin), Week 26, Week 54, Week 80, and Week 104. Additionally, for the time of administration of the injection, samples were collected prior to the injection. Synovial fluid samples were collected at the aforementioned times. These samples were taken immediately before the injection as part of the injection procedure (i.art.) and using the same needle used for the injection. For urine collection, the second morning excretion sample was obtained.

[0235] The following evaluation was conducted as an exploratory endpoint:

[0236] - Changes from baseline in serum and urine biomarkers associated with the administration of the compound.

[0237] - Baseline protein markers associated with response to treatment or disease progression (response evaluated by MRI and / or questionnaires).

[0238] Example 1. Placebo response and clinical efficacy in subjects treated with an FGF-18 compound

[0239] The FGF-18 compound used as a treatment in this example is Sprifermin (as defined in the “Definitions” section). Two strengths of Sprifermin were supplied to the study: 30 µg and 100 µg. Sprifermin was supplied as a white, sterile, freeze-dried powder in 3-mL glass vials. Each vial contained 31.5 µg or 105 µg of Sprifermin active substance; these amounts contained an excess of 5% that allowed for the extraction of 30 µg or 100 µg of Sprifermin active substance, respectively, after reconstitution with 0.9% w / v sodium chloride injection solution (referred herein as “saline solution”). The excipients of the formulation were sodium phosphate buffer (pH 7.2), sodium hydroxide, O-phosphate, sucrose, and poloxamer 188.

[0240] For all treatment groups, the administered volume was 2 mL.

[0241] This study was based on the FORWARD study (refer to study EMR700692-006) and studied five groups of patients:

[0242] · Group 1 (4 cycles placebo; hereinafter referred to as placebo or PBO): 108 subjects.

[0243] · Group 2 (2 cycles of Sprifermin 30 µg / injection alternating with 2 cycles of placebo; hereinafter referred to as Sprifermin / placebo 30 µg): 110 subjects.

[0244] · Group 3 (4 cycles Sprifermin 30 µg / injection; hereinafter referred to as Sprifermin 30 µg): 111 subjects.

[0245] · Group 4 (2 cycles of Sprifermin 100 µg / injection alternating with 2 cycles of placebo; hereinafter referred to as Sprifermin / placebo 100 µg): 110 subjects.

[0246] · Group 5 (4th cycle Sprifermin 100 µg / injection; hereinafter referred to as Sprifermin 100 µg): 110 subjects.

[0247] According to the FORWARD study, patients received four cycles of treatment at six-month intervals (each consisting of three intra-articular injections administered once a week over three consecutive weeks) (see Fig. 1). All injections were intra-articular (performed intra-articularly).

[0248] The primary efficacy endpoint was the change from baseline in cartilage thickness at the common femorotibial joint as assessed by MRI at 2 years. Exploratory endpoints included baseline protein markers associated with response to treatment or disease progression (response assessed by MRI and / or questionnaires).

[0249] This study enrolled adult subjects of either sex with primary femorotibial OA with a Kellgren-Lawrence grade of 2 or 3 according to the American College of Rheumatology (ACR) clinical and radiological criteria and a minimum joint gap (JSW) of ≥ 2.5 mm in the medial compartment. The subject must have pain within the target knee on most days, or require symptomatic treatment for knee pain with paracetamol (acetaminophen), systemic non-steroidal anti-inflammatory drugs (NSAIDs) including COX inhibitors (coxib), or tramadol on most days of the past month, and have 1) a history of pain due to OA within the target knee for at least 6 months, and 2) a pain score for the target knee of 4 to 9 points in response to Question 1 of the Western Ontario and McMaster University Osteoarthritis (WOMAC) Pain Index ("How much pain did you have [within the target knee, over the past 48 hours] when walking on a flat surface?") at screening and baseline, after at least 5 half-live discontinuations of the analgesic drug(s) (acetaminophen, topical or oral systemic NSAIDs, coxib, opioids, and / or tramadol). Women of childbearing age must use a form of contraception with an annual failure rate of less than 1% throughout the study.

[0250] Major exclusion criteria included misalignment of more than 5 degrees in the femorotibial axis of the target knee, clinical signs of inflammation (i.e., redness) in the target knee, administration of corticosteroids or hyaluronic acid into either knee within 6 months prior to screening, any plans for knee surgery (affecting the target or contralateral knee) within the next 2 years, concomitant conditions or treatments considered incompatible with study participation, contraindications to MRI scanning (including incompatibility with the scanner or knee coil), pregnancy or breastfeeding, participation in another clinical study within the past 30 days, and the statutory or restricted statutory dose.

[0251] Written prior consent must be obtained before any research-related activity.

[0252] Placebo effect based on JSW alone: As shown in Figure 3 (both A and B), the placebo effect is significant in KL2 patients (with JSW > 3.5 mm) versus KL3 patients (with JSW ≤ 3.5 mm). As is obvious, this plays a significant role in the total WOMAC score, where a simple injection of placebo (PBO) (i.e., salt) is sufficient to dramatically reduce the WOMAC score in KL2 patients over a very long period, whereas the WOMAC score continues to increase in KL3 patients despite the injection.

[0253] Placebo effect based on CTX-II or ProC2 levels: Figure 4 highlights that patients with high metabolic biomarker levels, e.g., CTX-II greater than 350 ng / mmol (Figure 4A) or ProC2 greater than 4.2 ng / mL (Figure 4B), show a higher placebo effect with respect to the placebo injection.

[0254] Placebo effect based on combination JSW + CTX-II levels:As shown in Figure 5, different distributions of CTX-II levels at baseline affect outcome measurements such as MRI and WOMAC. In terms of results, thin cartilage (JSW ≤ 3.5 mm) and low levels of metabolic biomarkers (e.g., CTX-II ≤ 350 ng / mmol) appear to be a suitable study population for clinical trials to mitigate elevated placebo responses.

[0255] After treatment with FGF18: As is evident from Figures 6a and 6b, when the placebo effect is taken into account, the shape of the response in terms of total WOMAC score changes dramatically for both KL2 and KL3 patients treated with various doses of Sprifermin. The same is true when CTX-2 levels are taken into account.

[0256] As is evident from Figures 6c and 6d, when the placebo effect is taken into account, the response shape in terms of cartilage thickness also changes deeply for both KL2 and KL3 patients treated with various doses of Sprifermin. The same is true when CTX-2 levels are taken into account.

[0257] From this embodiment, it is evident that stratification can eliminate the risk of clinical trials being otherwise biased by the placebo effect of some subgroups of patients.

[0258] Example 2. Placebo response in subjects treated with a placebo from a study using Compound X

[0259] The study was a double-blind, randomized, placebo-controlled, multicenter clinical trial for the evaluation of Compound X. Approximately 600 patients were enrolled in the placebo arm of this study. Patients aged 50–80 years with painful OA in at least one knee, but otherwise in good overall health, were recruited. Compound X was an oral medication, and the placebo was supplied in the same package. To be included, patients had to meet the ACR criteria for OA diagnosis. Both knees were evaluated during the study, but the target knee was identified prior to randomization for the evaluation of the primary efficacy endpoint. The target knee had to be painful on most days of the previous month. Additionally, patients had to meet at least one of the following criteria: age over 50 years, experience of morning stiffness lasting less than 30 minutes, or knee spotting. The target knee had to have a JSW of ≥ 2.0 mm in the medial tibiofemoral joint and a KL score of 2 or 3 on X-ray measurement. In WOMAC, patients had scores of ≥ 150 mm for pain and / or ≥ 510 mm for function. If both knees met the criteria, the target knee was defined as the knee meeting a KL score of 2. If both knees had a KL score of 2, the knee with the highest pain VAS was selected (as long as it was less than 80 mm).

[0260] Exclusion criteria included diseases (other than OA) and medications affecting bone or cartilage metabolism. Pain-relieving medications were permitted if taken at least 30 minutes after the study drug. Intra-articular injections of corticosteroids or hyaluronic acid into the target knee were prohibited during the study and 3 months prior to randomization.

[0261] Randomization was stratified by the center, ensuring that treatment allocations were unbiased and hidden from patients and investigator staff. The study was conducted in accordance with the Declaration of Helsinki. The protocol was approved by an independent ethics committee or institutional review board (IEC / IRB). All subjects or their legal representatives provided their informed consent for participation.

[0262] Placebo effect based on combination KL + CTX-II levels: As shown in Figure 7, the different distribution of CTX-II levels in the KL2 and KL3 subgroups at baseline affects outcome measures such as WOMAC pain. In terms of outcomes, thin cartilage (JSW ≤ 3.5 mm) and low levels of metabolic biomarkers (e.g., CTX-II ≤ 350 ng / mmol) appear to be a suitable study population for clinical trials to alleviate elevated placebo responses.

[0263] Overall conclusion

[0264] The examples clearly highlighted that a balanced design of KL3 enhancement and CTX-II enables the detection of symptomatic benefit for at least 3 years (see Fig. 8). Therefore, thin cartilage (≤3.5 ± 2SD mm) and low CTX-II (≤350 ± 2SD ng / mmol) appear to be the optimal study population for clinical trials to mitigate elevated placebo responses. In particular, it is clearly possible to use Allcomer data, i.e., without patient loss: this is merely a matter of stratification and patient management. Similarly, based on the results in the examples section, ProC2 levels can be used instead of CTX-II levels. In this case, thin cartilage and low ProC2 (≤4.2 ± 2SD ng / ml) would be equally appropriate study populations for clinical trials to mitigate elevated placebo responses.

[0265] Thanks to these findings regarding this proposed stratification and the two very different clinical trials involving two very different active compounds administered via different routes (one injected intra-articularly and the other administered orally), it is possible to reduce placebo response, avoid disparities between treatment groups, and eliminate the risk of clinical trials for any drug targeting OA due to the possibility of selecting a suitable patient population with structural and symptomatic benefits.

[0266] Refer to the references

[0267] 1) WO2008 / 023063

[0268] 2) WO2004 / 032849

[0269] 3) WO2014 / 023703

[0270] 4) Http: / / www.cartilage.org / _files / contentmanagement / ICRS_evaluation.pdf

[0271] 5) Literature [Lotz, 2010, Arthritis research therapy, 12: 211]

[0272] 6) Literature [Ellsworth et al., 2002, Osteoarthritis and Cartilage, 10: 308-320].

[0273] 7) References [Shimoaka et al., 2002, J. Bio. Chem. 277(9): 7493-7500]

[0274] 8) Literature [Gigout et al., 2017, Osteoarthritis and Cartilage, published online the 18.08.2018(https: / / doi.org / 10.1016 / j.joca.2017.08.004)]

[0275] 9) Literature [Lohmander et al., 2014, Arthritis Rheumatol., Jul; 66(7): 1820-31]

[0276] 10) References [Dahlberg et al., 2016, Clin Exp Rheumatol., May-Jun; 34(3): 445-50]

[0277] 11) Literature [Deng et al. 2018, Osteoarthritis and Cartilage, in press: 1-9].

[0278] 12) Parrish et al., 2017, Osteoarthritis Cartilage., Apr; 25(4): 554-560].

[0279] 13) Reference [Ameye and Young 2006. Curr Opin Rheumatol 18: 537-547].

[0280] 14) Literature [Zhou et al., 2016, Osteoarthritis Cartilage., Dec; 24(12): 2181-2192]

[0281] 15) Literature [Wang et al.2014. Birth defects Res C embryo today 102: 37-51]

[0282] 16) References [Karsdal et al., 2016, Osteoarthritis and Cartilage, 24(12): 2013-2021]

[0283] 17) Duclos et al., 2010, Osteoarthritis Cartilage. 2010 Nov; 18(11): 1467-76].

[0284] 18) References [Gudmann et al., 2016, Arthritis Res Ther., 18(1): 141].

[0285] 19) References [Munk et al., 2016, Rheumatol Int. 36(4): 541-9].

[0286] 20) Reference [Gudmann et al., 2014, Int J Mol Sci., 15(10): 18789-18803].

[0287] 21) Literature [Valdes et al., 2014, Osteoarthritis Cartilage., May; 22(5): 683-9]

[0288] 22) Struglics et al., 2011, Osteoarthritis Cartilage., Aug; 19(8): 1047-57]

[0289] 23) Struglics et al., 2015, Arthritis Rheumatol., Jul; 67(7): 1816-25]

[0290] 24) Literature [The Merck Manual, 17th edition, page 449]

[0291] 25) Literature [Bellamy et al., 1988, J. Rheumatology, 15: 1833-1840].

[0292] 26) Literature [Wolfe, 1999, Rheumatology, 38: 355-361]

[0293] 27) References [BAY-Jensen et al., 2016, Osteoarthritis Cartilage]. 24(1): 9-20]

[0294] 28) Literature [Collins et al. 2016]. Osteoarthritis Cartilage 24: 1317]

[0295] abbreviation

[0296] BMI = Body Mass Index; CI = Confidence Interval, DBPC = Double-blind placebo-controlled

[0297] CTX-II = C-telopeptide crosslinking of type II collagen,

[0298] DBPC = Double-blind placebo control group

[0299] ICOAP = Measurement of intermittent and constant osteoarthritis pain

[0300] ITT = Intention to Treat; JSW = Joint Space

[0301] KOOS Symptom Index = Knee Injury and Osteoarthritis Outcome Score Symptom Index Subscale

[0302] KOOS QOL = Quality of Life Subscale for Knee Injury and Osteoarthritis Outcomes

[0303] LOCF = the last observation performed prior to,

[0304] LFTC = Lateral femorotibial compartment

[0305] MFTC = Medial Femorotibial Compartment

[0306] mITT = Modified Treatment Intent

[0307] MOS SF-36 = Medical Outcomes Study Short Form-36 Overall Health Survey

[0308] MRI = Magnetic Resonance Imaging

[0309] NRS Pain Score = Numerical Rating Scale Pain Score

[0310] OA = Osteoarthritis

[0311] PGA = Patient's overall evaluation

[0312] PGIC = Impression of the patient's overall change

[0313] PROC2 = Neo-epitope of collagen type II propeptide

[0314] WOMAC = Western Ontario and McMaster University Osteoarthritis Index; W = State

Claims

Claim 1 A composition comprising an FGF-18 compound for use in a method for treating a subject having a cartilage disorder, wherein the subject’s sensitivity to the FGF-18 compound is determined by detecting the amount of one or both of CTX-II (C-terminal telopeptide of type II collagen) and ProC2 (neo-epitope of collagen type II) in a biological sample from the subject and measuring the joint gap (JSW) in at least one knee of the subject, wherein the subject has excellent sensitivity to treatment if the biological sample from the subject contains less than 350 ± 2SD ng / mmol of CTX-II and / or less than 4.2 ± 2SD ng / mL of ProC2 and less than 3.5 ± 2SD mm of JSW, or if the biological sample from the subject contains (i) less than 350 ± 2SD ng / mmol of CTX-II and / or less than 4.2 ± 2SD ng / mL of ProC2 and greater than 3.5 ± 2SD mm of A composition having intermediate sensitivity comprising JSW, or (ii) CTX-II greater than 350 ± 2SD ng / mmol and / or ProC2 greater than 4.2 ± 2SD ng / mL and JSW less than 3.5 ± 2SD mm, wherein the treatment method comprises administering the FGF-18 compound into the joint of the subject, wherein 30 to 100 μg of the FGF-18 compound per dose is administered into the joint of the subject having excellent sensitivity, and 100 μg of the FGF-18 compound per dose is administered into the joint of the subject having intermediate sensitivity. Claim 2 A composition according to claim 1, wherein cartilage disorder is selected from the group consisting of osteoarthritis, cartilage damage, fractures affecting articular cartilage or surgical procedures affecting articular cartilage, and microfractures. Claim 3 A composition according to claim 1, wherein the FGF-18 compound comprises 28 to 196 amino acid residues of sequence 3 fused to a heterologous protein or chemical compound. Claim 4 A composition according to claim 1, wherein the FGF-18 compound is sprifermin or sprifermin fused to a heterologous protein or chemical compound. Claim 5 A composition according to claim 1, wherein the FGF-18 compound is administered in a treatment cycle once a week for three weeks. Claim 6 A composition in which the treatment cycle is repeated, as described in paragraph 5. Claim 7 A composition comprising an FGF-18 compound for use in a method for treating a subject having a cartilage disorder, wherein the method for treating comprises the following steps: a) determining the amount of at least one biomarker selected from the group consisting of CTX-II (C-terminal telopeptide of type II collagen) and ProC2 (neo-epitope of collagen type II) from a sample from a subject, and measuring the joint gap (JSW) in at least one knee of said subject; b) i. having excellent sensitivity to treatment if the biological sample from the subject contains less than 350 ± 2 SD ng / mmol of CTX-II, and / or less than 4.2 ± 2 SD ng / mL of ProC2, and 3.5 ± 2 SD mm or less of JSW, or ii. iii. selecting a subject having moderate sensitivity to treatment when a biological sample from the subject contains less than 350 ± 2SD ng / mmol of CTX-II and / or less than 4.2 ± 2SD ng / mL of ProC2 and greater than 3.5 ± 2SD mm of JSW, or iii. selecting a subject having moderate sensitivity to treatment when a biological sample from the subject contains more than 350 ± 2SD ng / mmol of CTX-II and / or greater than 4.2 ± 2SD ng / mL of ProC2 and less than 3.5 ± 2SD mm of JSW; and c) administering the FGF-18 compound into the joint of the selected subject, wherein 30 to 100 μg of the FGF-18 compound per dose is administered into the joint of a subject having excellent sensitivity, and 100 μg of the FGF-18 compound per dose is administered into the joint of a subject having moderate sensitivity. Claim 8 A composition according to claim 7, wherein cartilage disorder is selected from the group consisting of osteoarthritis, cartilage damage, fractures affecting articular cartilage or surgical procedures affecting articular cartilage, and microfractures. Claim 9 A composition according to claim 7, wherein the FGF-18 compound comprises amino acid residues 28 to 196 of sequence 3 fused to a heterologous protein or chemical compound. Claim 10 A composition according to claim 7, wherein the FGF-18 compound is sprifermin or sprifermin fused to a heterogeneous protein or chemical compound. Claim 11 A composition according to claim 7, wherein the FGF-18 compound is administered in a treatment cycle once a week for three weeks. Claim 12 A composition in which the treatment cycle is repeated, as described in Clause 11. Claim 13 A kit comprising a assay means and instructions for use to predict sensitivity to FGF-18 compound treatment in subjects having cartilage disorders, wherein the kit comprises: a) a step of determining, from a sample from a subject, the amount of at least one biomarker selected from the group consisting of CTX-II (C-terminal telopeptide of type II collagen) and ProC2 (neo-epitope of collagen type II); b) a step of measuring the joint gap (JSW) in at least one knee of the subject; and c) from the results of steps a) and b), i. excellent sensitivity to treatment if the biological sample from the subject contains less than 350 ± 2 SD ng / mmol of CTX-II, and / or less than 4.2 ± 2 SD ng / mL of ProC2, and 3.5 ± 2 SD mm or less of JSW, ii. A kit used according to a method comprising the step of predicting: iii. intermediate susceptibility to treatment when a biological sample from a subject contains less than 350 ± 2SD ng / mmol CTX-II and / or less than 4.2 ± 2SD ng / mL ProC2 and greater than 3.5 ± 2SD mm JSW; iv. low susceptibility or insensitivity to treatment when a biological sample from a subject contains greater than 350 ± 2SD ng / mmol CTX-II and / or greater than 4.2 ± 2SD ng / mL ProC2 and less than 3.5 ± 2SD mm JSW; or iv. low susceptibility or insensitivity to treatment when a biological sample from a subject contains greater than 350 ± 2SD ng / mmol CTX-II or greater than 4.2 ± 2SD ng / mL ProC2 and greater than 3.5 ± 2SD mm JSW, wherein steps a) and b) are performed in either order. Claim 14 A kit in paragraph 13 in which cartilage disorders are selected from the group consisting of osteoarthritis, cartilage damage, fractures affecting articular cartilage or surgical procedures affecting articular cartilage, and microfractures. Claim 15 A kit according to claim 13, wherein the FGF-18 compound comprises amino acid residues 28 to 196 of sequence 3 fused to a heterologous protein or chemical compound. Claim 16 In paragraph 13, the kit is a sprifermin in which the FGF-18 compound is fused to sprifermin or a heterologous protein or chemical compound.

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