Liquid pharmaceutical preparation of CNP compound

Abstract

A liquid pharmaceutical preparation comprising a CNP compound, a buffer, an isotonic agent, a preservative, and optionally an antioxidant, and a method for producing and using such a preparation.

Description

Technical Field

[0001] The present invention relates to a liquid pharmaceutical preparation containing a CNP compound, a buffer, an isotonic agent, a preservative, and optionally an antioxidant, and a method for producing and using the preparation.

Background Art

[0002] Achondroplasia (ACH) is a genetic disorder caused by an autosomal dominant mutation in the fibroblast growth factor receptor 3 (FGFR3) gene, which causes abnormal cartilage formation and leads to dwarfism. C-type natriuretic peptide (CNP) is a hormone that binds to peptide receptor B (NPR-B), activates it, and results in the inhibition of FGFR3 downstream signaling. This in turn induces cartilage growth and skeletal overgrowth, as observed in both mice and humans overexpressing CNP. The dwarfism of achondroplastic mice is normalized by overproduction of CNP in cartilage or continuous delivery of CNP by intravenous (iv) injection, suggesting that administration of CNP at supra-physiological levels is a strategy for the treatment of ACH.

[0003] Human prepro-CNP containing 126 amino acids is further cleaved by furin to yield CNP-53. CNP-53 has biological activity, but typically it is processed by an unknown mechanism to become the 22-amino acid form with biological activity in circulation, namely CNP-22. The biological activity of CNP is tightly regulated and its clearance from plasma is very rapid. Therefore, considering the short in vivo half-life of CNP (2 minutes after intravenous injection), its use as a therapeutic agent is considered to require continuous infusion and is thus difficult in the pediatric population.

[0004] Various approaches have been investigated to increase the in vivo half-life of CNP. For example, Lorget et al. (Am. J. Hum. Genet. 91, 1108-1114, 2012) disclosed a recombinant CNP-39 (also referred to as BMN111) consisting of the 37 C-terminal amino acids of human CNP-53, which is similar in pharmacological activity to human CNP, plus glycine and proline added to the N-terminus of the peptide. BMN111 is more resistant to NEP cleavage, but since it has a half-life of only 20 minutes, when administered once a day, it is associated with short-term exposure to effective drug levels.

Summary of the Invention

Problems to be Solved by the Invention

[0005] The expansion of the scope of in vivo half-life extension of CNP based on the attachment to a water-soluble carrier moiety such as PEG via a reversible prodrug linker has been investigated in WO2016 / 110577A1, WO2017 / 118693A1, WO2017 / 118698A1, WO2017 / 118700A1, WO2017 / 118703A1, WO2017 / 118704A1 and WO2017 / 118707A1. Pharmaceutical formulations of such reversible CNP conjugates, in which a carrier is attached to CNP via a reversible linkage, must provide sufficient CNP conjugate stability to avoid early CNP release during storage. If the reversible linkage between the carrier and CNP deteriorates during storage, the concentration of the drug that can exert its pharmacological effect increases, thereby potentially increasing the risk of overdose at the time of administration, which can lead to hypotension. Furthermore, the drug released during storage is rapidly cleared by the kidneys immediately after administration to the patient, thus shortening the time for the long-acting composition to provide a therapeutically necessary amount of the drug. This creates a risk that medical needs are not met.

[0006] Since the decomposition products or oligomers that may be generated during storage may impair the physiological activity of the CNP moiety, it is desirable to minimize their generation. Furthermore, the reversible linkage between the CNP moiety and the carrier moiety in the reversible CNP conjugate makes the storage of pharmaceutical formulations containing the CNP conjugate difficult.

[0007] WO2020 / 165081A1 shows a dry pharmaceutical formulation containing a reversible CNP conjugate that requires reconstitution with an aqueous solution to provide an injectable pharmaceutical.

[0008] Also, it is important to identify a suitable liquid pharmaceutical formulation of a reversible CNP conjugate containing a CNP compound, particularly a CNP covalently linked to a carrier moiety via a reversible linker, in which the peptide exhibits an acceptable impurity profile. Also, particularly in the case of a reversible CNP conjugate, it is important to identify a suitable liquid formulation that exhibits limited early CNP release even after long-term storage.

[0009] Therefore, in the art, there remains a need for an easy-to-use liquid formulation of a CNP compound, particularly a reversible CNP conjugate, that exhibits storage stability and provides ease of use.

[0010] Therefore, an object of the present invention is to at least partially overcome the above drawbacks.

Means for Solving the Problems

[0011] This object is achieved by a liquid pharmaceutical formulation comprising a CNP compound, a buffer, an isotonic agent, a preservative, and optionally an antioxidant.

Effects of the Invention

[0012] Surprisingly, it has been found that the liquid pharmaceutical formulation of the present invention allows for stable long-term storage. The liquid pharmaceutical formulation of the present invention does not require freezing, such as storage in a freezer, and exhibits a significant degree of peptide stability even after storage for several months.

Mode for Carrying Out the Invention

[0013] Within the scope of the meaning of the present invention, terms are used as follows.

[0014] As used herein in combination with a numerical value, the term "about" refers to, in addition to the numerical value itself, a range of plus / minus 20% or less of the numerical value, plus / minus 10% or less of the numerical value, in certain embodiments plus / minus 8% or less of the numerical value, in certain embodiments plus / minus 5% or less of the numerical value, and in certain embodiments plus / minus 2% or less of the numerical value. For example, the phrase "about 200" means a range of 200+ / -10%, i.e., a range of 180 to 220, in certain embodiments 200+ / -8%, i.e., a range of 184 to 216, in certain embodiments 200+ / -5%, i.e., a range of 190 to 210, and in certain embodiments 200+ / -2%, i.e., a range of 196 to 204. It is understood that the percentage indicated as "about 20%" does not mean a range of "20%+ / -10%", i.e., 10 to 30%, but rather "about 20%" means a range of plus / minus 10% of the numerical value 20, i.e., 18 to 22.

[0015] As used herein, the term "antibacterial agent" refers to chemical substances such as chemical substances that kill or inhibit the growth of microorganisms such as bacteria, fungi, yeasts, protozoa, molds, and / or destroy viruses.

[0016] As used herein, the term "adsorption inhibitor" primarily refers to an ionic or non-ionic surfactant, protein, or soluble polymer used to coat or competitively adsorb onto the inner surface of a container containing a formulation. The concentration and type of excipient selected are determined by the effect to be avoided, but typically a monolayer of surfactant is formed at the interface just above the critical micelle concentration (CMC).

[0017] As used herein, the term "buffer" or "buffering agent" refers to a compound that maintains the pH within a desired range. Physiologically tolerated buffering agents are, for example, sodium phosphate, histidine, bicarbonate, citrate, acetate, sulfate, nitrate, chloride, and pyruvate. Antacids such as Mg(OH)2 or ZnCO3 can also be used.

[0018] As used herein, the term "CNP" refers, in certain embodiments, to all CNP polypeptides derived from mammalian species, such as human and mammalian species, particularly human and murine species, as well as variants, analogs, orthologs, homologs, and derivatives thereof, and fragments thereof, which are characterized by the control of growth, proliferation, and differentiation of chondrocyte growth plate chondrocytes. The term "CNP" also includes all CNP variants, analogs, orthologs, homologs, derivatives, and fragments thereof. The CNP variants, analogs, orthologs, homologs, derivatives, and fragments thereof disclosed in WO2009 / 067639A2 and WO2010 / 135541A2 are incorporated herein by reference.

[0019] As used herein, the term "CNP polypeptide variant" refers to a polypeptide derived from the same species that is different from the reference CNP polypeptide. Generally, since the differences are limited, the amino acid sequences of the reference and the variant are very similar overall and identical in many regions. In certain embodiments, the CNP polypeptide variant is at least 70%, 80%, 90% or 95% identical to the reference CNP polypeptide. A polypeptide having a query amino acid sequence and, for example, at least 95% "identical" amino acid sequence means that the amino acid sequence of the target polypeptide is identical to the query sequence, except that it may contain 5 or fewer amino acid changes per 100 amino acids of the query amino acid sequence. Such changes to the reference sequence may occur at the amino-terminal (N-terminal) or carboxy-terminal (C-terminal) positions of the reference amino acid sequence, but may occur anywhere between such terminal positions, individually scattered among the residues within the reference sequence, or scattered as one or more adjacent groups within the reference sequence. The query sequence may be the complete amino acid sequence of the reference sequence or any of the specified fragments described herein. Such CNP polypeptide variants can be natural variants, such as natural allelic variants encoded by one of several alternative forms of CNP occupying a given locus on a chromosome or in an organism, or isoforms encoded by natural splice variants derived from a single primary transcript. Alternatively, the CNP polypeptide variant may be a variant that is not known to be natural and can be made by mutagenesis techniques known in the art. It is known in the art that one or more amino acids can be deleted from the N-terminus or C-terminus of a bioactive peptide or protein without substantially losing its physiological function. Such N-terminal and / or C-terminal deletions are also encompassed by the term CNP polypeptide variant.

[0020] It is also recognized by those skilled in the art that it is possible to modify some of the amino acid sequences of the CNP polypeptide without significantly affecting the structure or function of the peptide. Such variants contain deletions, insertions, inversions, repeats, and substitutions selected according to principles known in the art so as to have little effect on activity. For example, guidelines regarding methods for making phenotypically silent amino acid substitutions are provided in Bowie et al. (1990), Science 247:1306-1310, which is hereby incorporated by reference in its entirety, where the authors show that there are two main approaches for examining the tolerance of amino acid sequence changes.

[0021] As used herein, the term "CNP analog" refers to CNP from a different unrelated organism that performs the same function in each organism but does not originate from the ancestral structure commonly possessed by the ancestors of those organisms. Instead, the analog CNP evolved separately and then evolved to perform the same or similar functions. In other words, an analog CNP polypeptide is a polypeptide having quite different amino acid sequences that achieves the same biological activity, namely, the control of the growth, proliferation, and differentiation of chondrocytes in the growth plate of cartilage.

[0022] As used herein, the term "CNP ortholog" refers to CNP present in two different species, the sequences of which are related to each other through a common homologous CNP in an ancestral species but have evolved to be different from each other.

[0023] As used herein, the term "CNP homolog" refers to a CNP of a different organism that performs the same function in each organism and is derived from an ancestral structure that the ancestors of those organisms had in common. In other words, a homologous CNP polypeptide is a polypeptide having very similar amino acid sequences that achieve the same biological activity, namely, the control of the growth, proliferation, and differentiation of chondrocytes in the growth plate of cartilage. In certain embodiments, a CNP polypeptide homolog can be defined as a polypeptide that exhibits at least 40%, 50%, 60%, 70%, 80%, 90%, or 95% identity to a reference CNP polypeptide.

[0024] Accordingly, a CNP polypeptide can be, for example, (i) a CNP polypeptide in which at least one of the amino acid residues is substituted, in certain embodiments, by a conservative amino acid residue or a non-conservative amino acid residue, such that the substituted amino acid residue may or may not be encoded by the genetic code; and / or (ii) a CNP polypeptide in which at least one of the amino acid residues contains a substituent; and / or (iii) a CNP polypeptide in which the CNP polypeptide is fused to another compound, such as a compound that lengthens the half-life of the polypeptide (e.g., polyethylene glycol); and / or (iv) an additional amino acid is fused to the CNP polypeptide, such as an IgG Fc fusion region peptide, or a leader sequence, or a secretion sequence, or a sequence used for purification of the polypeptide of the above forms, or a preprotein sequence.

[0025] As used herein, the term "CNP polypeptide fragment" refers to any peptide that includes a contiguous range of a portion of the amino acid sequence of a CNP polypeptide.

[0026] More specifically, the CNP polypeptide fragment contains at least 6 consecutive amino acids of the CNP polypeptide, for example, at least 8, at least 10, or at least 17 consecutive amino acids. The CNP polypeptide fragment can further be described as a subclass of the CNP polypeptide containing at least 6 amino acids, where "at least 6" is defined as any integer between 6 and the integer representing the C-terminal amino acid of the CNP polypeptide. Further, the molecular species are included which are CNP polypeptide fragments of at least 6 amino acid lengths as described above and which are further specified with respect to the N-terminal and C-terminal positions. Also, the term "CNP polypeptide fragment" includes, as individual molecular species, all CNP polypeptide fragments of at least 6 amino acid lengths as described above that can be specifically specified by the N-terminal and C-terminal positions. That is, it is every combination of N-terminal and C-terminal positions that a fragment of at least 6 consecutive amino acid residues in length can occupy on any given amino acid sequence of the CNP polypeptide.

[0027] Since the term CNP includes the above variants, analogs, orthologs, homologs, derivatives and fragments of CNP, any reference to a specific position within the reference sequence includes, even if not explicitly mentioned, the equivalent positions in the variants, analogs, orthologs, homologs, derivatives and fragments of the CNP moiety.

[0028] Native CNP-22 (SEQ ID NO: 1) has the following sequence. GLSKGCFGLKLDRIGSMSGLGC Wherein the cysteines at positions 6 and 22 are linked via a disulfide bridge.

[0029] In certain embodiments, "CNP" refers to the following peptide sequence: SEQ ID NO: 2 (CNP-53): DLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 3 (G-CNP-53): GDLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 4 (M-CNP-53): MDLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 5 (P-CNP-53): PDLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 6 (CNP-53 M48N): DLRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSNSGLGC; SEQ ID NO: 7 (CNP-53 Δ15-31): DLRVDTKSRAAWARGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 8 (CNP-52): LRVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 9 (CNP-51): RVDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 10 (CNP-50): VDTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 11 (CNP-49): DTKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 12 (CNP-48): TKSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 13 (CNP-47): KSRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 14 (CNP-46): SRAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 15 (CNP-45): RAAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 16 (CNP-44): AAWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 17 (CNP-44 Δ14-22): AAWARLLQEHPNAGLSKGCFGLKLDRIGSMSGLGC; Accession No. 18 (CNP-44 Δ15-22): AAWARLLQEHPNARGLSKGCFGLKLDRIGSMSGLGC; Accession No. 19 (CNP-43): AWARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 20 (CNP-42): WARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 21 (CNP-41): ARLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 22 (CNP-40): RLLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 23 (CNP-39): LLQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession No. 24 (CNP-38): LQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC, wherein the cysteines at positions 22 and 38 are linked by a disulfide bridge; Accession number 25 (CNP-37): QEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 26 (CNP-37 Q1pQ, where pQ = pyroglutamic acid): pQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 27 (G-CNP-37): GQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 28 (P-CNP-37): PQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 29 (M-CNP-37): MQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 30 (PG-CNP-37): PGQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 31 (MG-CNP-37): MGQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 32 (CNP-37 M32N): QEHPNARKYKGANKKGLSKGCFGLKLDRIGSNSGLGC; Accession number 33 (G-CNP-37 M32N): GQEHPNARKYKGANKKGLSKGCFGLKLDRIGSNSGLGC; Accession number 34 (G-CNP-37 K14Q): GQEHPNARKYKGANQKGLSKGCFGLKLDRIGSMSGLGC; Accession number 35 (G-CNP-37 K14P): GQEHPNARKYKGANPKGLSKGCFGLKLDRIGSMSGLGC; Accession number 36 (G-CNP-37 K14Q, Δ15): GQEHPNARKYKGANQGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 37 (G-CNP-37 K14Q, K15Q): GQEHPNARKYKGANQQGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 38 (CNP-36): EHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 39 (CNP-35): HPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 40 (CNP-34): PNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 41 (CNP-33): NARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 42 (CNP-32): ARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 43 (CNP-31): RKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 44 (CNP-30): KYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 45 (CNP-29): YKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 46 (CNP-28): KGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 47 (GHKSEVAHRF-CNP-28): GHKSEVAHRFKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 48 (CNP-27): GANKKGLSKGCFGLKLDRIGSMSGLGC; Accession number 49 (CNP-27 K4Q, K5Q): GANQQGLSKGCFGLKLDRIGSMSGLGC; Accession number 50 (CNP-27 K4R, K5R): GANRRGLSKGCFGLKLDRIGSMSGLGC; Accession number 51 (CNP-27 K4P, K5R): GANPRGLSKGCFGLKLDRIGSMSGLGC; Accession number 52 (CNP-27 K4S, K5S): GANSSGLSKGCFGLKLDRIGSMSGLGC; Accession number 53 (CNP-27 K4P, K5R): GANGANPRGLSRGCFGLKLDRIGSMSGLGC; Accession number 54 (CNP-27 K4R, K5R, K9R): GANRRGLSRGCFGLKLDRIGSMSGLGC; Accession number 55 (CNP-27 K4R, K5R, K9R, M22N): GANRRGLSRGCFGLKLDRIGSNSGLGC; Accession number 56 (P-CNP-27 K4R, K5R, K9R): PGANRRGLSRGCFGLKLDRIGSMSGLGC; Accession number: 57 (M-CNP-27 K4R, K5R, K9R): MGANRRGLSRGCFGLKLDRIGSMSGLGC; Accession number 58 (HSA fragment-CNP-27): GHKSEVAHRFKGANKKGLSKGCFGLKLDRIGSMSGLG; Accession number 59 (HSA fragment-CNP-27 M22N): GHKSEVAHRFKGANKKGLSKGCFGLKLDRIGSNSGLGC; Accession number 60 (M-HSA fragment-CNP-27): MGHKSEVAHRFKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 61 (P-HSA fragment-CNP-27): PGHKSEVAHRFKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 62 (CNP-26): ANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 63 (CNP-25): NKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 64 (CNP-24): KKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 65 (CNP-23): KGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 66 (R-CNP-22): RGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 67 (ER-CNP-22): ERGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 68 (R-CNP-22 K4R): RGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 69 (ER-CNP-22 4KR): ERGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 70 (RR-CNP-22): RRGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 71 (HRGP fragment-CNP-22): GHHSHEQHPHGANQQGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 72 (HRGP fragment-CNP-22): GAHHPHEHDTHGANQQGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 73 (HRGP fragment-CNP-22): GHHSHEQHPHGANPRGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 74 (IgG1(F c ) fragment - CNP - 22): GQPREPQVYTLPPSGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 75 (HSA fragment - CNP - 22): GQHKDDNPNLPRGANPRGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 76 (HSA fragment - CNP - 22): GERAFKAWAVARLSQGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 77 (osteocrin NPR C inhibitory fragment - CNP22): FGIPMDRIGRNPRGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 78 (FGF2 heparin - binding region fragment - CNP22): GKRTGQYKLGSKTGPGPKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 79 (IgG1(F c ) fragment - CNP - 22 K4R): GQPREPQVYTGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 80 (HSA fragment - CNP - 22 K4R): GVPQVSTSTGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 81 (fibronectin fragment - CNP - 22 K4R): GQPSSSSQSTGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 82 (fibronectin fragment - CNP - 22 K4R): GQTHSSGTQSGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 83 (fibronectin fragment - CNP - 22 K4R): GSTGQWHSESGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 84 (Zinc Finger Fragment - CNP - 22 K4R): GSSSSSSSSSGANQQGLSRGCFGLKLDRIGSMSGLGC; SEQ ID NO: 85 (CNP - 21): LSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 86 (CNP - 20): SKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 87 (CNP - 19): KGCFGLKLDRIGSMSGLGC; SEQ ID NO: 88 (CNP - 18): GCFGLKLDRIGSMSGLGC; SEQ ID NO: 89 (CNP - 17): CFGLKLDRIGSMSGLGC; SEQ ID NO: 90 (BNP Fragment - CNP - 17 - BNP Fragment): SPKMVQGSGCFGLKLDRIGSMSGLGCKVLRRH; SEQ ID NO: 91 (CNP - 38 L1G): GQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 92 (Ac - CNP - 37; where Ac = acetyl): Ac - QEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC; SEQ ID NO: 93: QEHPNARX1YX2GANX3X4GLSX5GCFGLX6LDRIGSMSGLGC, where X1, X2, X3, X4, X5 and X6 are independently selected from the group consisting of K, R, P, S and Q, provided that at least one of X1, X2, X3, X4, X5 and X6 is selected from the group consisting of R, P, S and Q; in certain embodiments, X1, X2, X3, X4, X5 and X6 are selected from the group consisting of K and R, provided that at least one of X1, X2, X3, X4, X5 and X6 is R; SEQ ID NO: 94: QEHPNARKYKGANX1X2GLSX3GCFGLX4LDRIGSMSGLGC, Here, X1, X2, X3, and X4 are independently selected from the group consisting of K, R, P, S, and Q, provided that at least one of X1, X2, X3, and X4 is selected from the group consisting of R, P, S, and Q; in certain embodiments, X1, X2, X3, and X4 are selected from K and R, provided that at least one of X1, X2, X3, and X4 is R; SEQ ID NO: 95: QEHPNARKYKGANX1X2GLSKGCFGLKLDRIGSMSGLGC, Here, X1X2 is selected from the group consisting of KR, RK, KP, PK, SS, RS, SR, QK, QR, KQ, RQ, RR, and QQ.

[0030] It is understood that in SEQ ID NOs: 2 to 95, those corresponding to cysteines at positions 22 and 38 of SEQ ID NO: 24 are linked by disulfide bridges.

[0031] The term "CNP" also includes poly(amino acid) conjugates having the above sequences but having a backbone that includes both amide bonds and non-amide bonds such as ester bonds, such as depsipeptides. A depsipeptide is a chain of amino acid residues whose backbone contains both amide (peptide) and ester bonds. Thus, as used herein, the term "side chain" refers to the portion attached to the α-carbon of an amino acid moiety when the amino acid moiety is linked by an amine bond as in a polypeptide, or to any carbon atom-containing portion attached to the backbone of a poly(amino acid) conjugate, such as in the case of a depsipeptide. In certain embodiments, the term "CNP" refers to a polypeptide having a backbone formed by amide (peptide) bonds.

[0032] As used herein, the term "loop portion" refers to a continuous series of amino acid residues of CNP that are located between two cysteine residues forming an intramolecular disulfide bridge or between homologous amino acid residues linked via a chemical crosslinking agent. Preferably, the loop portion is between two cysteine residues forming an intramolecular disulfide bridge. These two cysteines correspond to the cysteines at positions 22 and 38 in the sequence of CNP-38 (SEQ ID NO: 24). Thus, when CNP has the sequence of CNP-38, amino acids 23 to 37 are located in the aforementioned loop portion.

[0033] Regardless of the length of the CNP portion, the sequence of the loop portion of wild-type CNP is FGLKLDRIGSMSGLG (SEQ ID NO: 96). In certain embodiments, the loop portion of CNP is FGLKLDRIGSNSGLG (SEQ ID NO: 97).

[0034] Since the term CNP includes the above-mentioned variants, analogs, orthologs, homologs, derivatives and fragments of CNP, the term "loop portion" also includes the corresponding variants, analogs, orthologs, homologs, derivatives and fragments of the sequence of SEQ ID NO: 96. Thus, any reference to a specific position within the reference sequence also includes the equivalent position in variants, analogs, orthologs, homologs, derivatives and fragments of the CNP portion, even if not explicitly mentioned.

[0035] As used herein, the term "CNP agonist" refers to any molecule that activates the natriuretic peptide receptor B (NPR-B). In certain embodiments, the CNP agonist has an EC50 that is up to 50-fold higher than the NPR-B activity of CNP-22 (SEQ ID NO: 1).

[0036] As used herein, the term "CNP conjugate" refers to any compound, crystal, or mixture that includes at least one CNP portion linked to at least one other moiety.

[0037] As used herein, the term "reversible CNP conjugate" refers to a CNP conjugate comprising at least one CNP moiety, wherein the at least one CNP moiety is covalently and reversibly attached to a carrier moiety and is released with a release half-life of at least 6 hours under physiological conditions.

[0038] As used herein, the term "stable CNP conjugate" refers to a CNP conjugate comprising at least one CNP moiety, wherein the at least one CNP moiety is covalently and stably attached to a carrier moiety, i.e., attached via a stable bond with a half-life of more than 6 months under physiological conditions.

[0039] When an amount in units of mg / mL follows the phrase "CNP conjugate, its CNP moiety" as used herein, the liquid formulation contains the CNP conjugate, meaning that for the corresponding amount, only the CNP moiety and not the complete CNP conjugate is considered. That is, parts of the CNP conjugate other than the CNP moiety, such as the carrier moiety, are not calculated. The amount of the CNP moiety in the CNP conjugate can be determined by quantitative amino acid analysis or by a known analytical method capable of quantifying an unknown sample by comparison with a CNP conjugate of known CNP moiety content.

[0040] As used herein, the term "cryoprotectant" refers to a compound added to a formulation to protect a CNP compound during the freezing process.

[0041] The term "C 1-4 alkyl" used alone or in combination herein means a straight-chain or branched alkyl moiety having 1 to 4 carbon atoms. When present at the end of a molecule, straight-chain or branched C 1-4 Examples of alkyl are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl and tert-butyl. When two parts of a molecule are joined by C 1-4 alkyl, such C 1-4Examples of alkyl groups are -CH2-, -CH2-CH2-, -CH(CH3)-, -CH2-CH2-CH2-, -CH(C2H5)-, -C(CH3)2-. C 1-4 Each hydrogen of the alkyl carbon may optionally be replaced by a substituent as defined above. Optionally, C 1-4 One or more moieties as defined below may be inserted into the alkyl.

[0042] The term "C 1-6 alkyl" as used herein alone or in combination means a straight or branched alkyl moiety having from 1 to 6 carbon atoms. When present at the end of a molecule, straight-chain and branched C 1-6 Examples of alkyl groups are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, 2-methylbutyl, 2,2-dimethylpropyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl, 2,3-dimethylbutyl, and 3,3-dimethylpropyl. When two moieties of a molecule are joined by a C 1-6 alkyl group, such C 1-6 Examples of alkyl groups are -CH2-, -CH2-CH2-, -CH(CH3)-, -CH2-CH2-CH2-, -CH(C2H5)-, and -C(CH3)2-. C 1-6 Each hydrogen atom of the carbon may optionally be replaced by a substituent as defined above. Optionally, C 1-6 One or more moieties as defined below may be inserted into the alkyl.

[0043] Thus, "C 1-10 alkyl", "C 1-20 alkyl", or "C 1-50 alkyl" each means an alkyl chain having from 1 to 10, 1 to 20, or 1 to 50 carbon atoms, respectively, and C 1-10 C 1-20 or C 1-50 Each hydrogen atom of the carbon may optionally be replaced by a substituent as defined above. Optionally, C 1-10or C 1-50 One or more moieties as defined below may be inserted into the alkyl.

[0044] The term "C 2-6 alkenyl", used herein alone or in combination, means a straight or branched hydrocarbon moiety having from 2 to 6 carbon atoms and containing at least one carbon-carbon double bond. When present at the end of a molecule, examples are -CH=CH2, -CH=CH-CH3, -CH2-CH=CH2, -CH=CH-CH2-CH3 and -CH=CH-CH=CH2. When two moieties of a molecule are joined by C 2-6 alkenyl, an example of such C 2-6 alkenyl is -CH=CH-. Each hydrogen atom of the C 2-6 alkenyl moiety may optionally be replaced by a substituent as defined above. Optionally, one or more moieties as defined below may be inserted into the C 2-6 alkenyl.

[0045] Thus, the term "C 2-10 alkenyl", "C 2-20 alkenyl" or "C 2-50 alkenyl", used herein alone or in combination, means a straight or branched hydrocarbon moiety having from 2 to 10, from 2 to 20, or from 2 to 50 carbon atoms and containing at least one carbon-carbon double bond. Each hydrogen atom of the C 2-10 alkenyl, C 2-20 alkenyl or C 2-50 alkenyl group may optionally be replaced by a substituent as defined above. Optionally, one or more moieties as defined below may be inserted into the C 2-10 alkenyl, C 2-20 alkenyl or C 2-50 alkenyl.

[0046] The term "C 2-6"Alkynyl" means a straight-chain or branched hydrocarbon moiety having from 2 to 6 carbon atoms and containing at least one carbon-carbon triple bond. When present at the end of a molecule, examples are -C≡CH, -CH2-C≡CH, CH2-CH2-C≡CH, and CH2-C≡C-CH3. When two moieties of a molecule are linked by an alkynyl group, an example is -C≡C-. C 2-6 Each hydrogen atom of the alkynyl group may optionally be replaced by a substituent as defined above. Optionally, one or more double bonds may be present. Optionally, C 2-6 One or more moieties as defined below may be interposed in the alkynyl.

[0047] Accordingly, the term "C 2-10 alkynyl", "C 2-20 alkynyl", or "C 2-50 alkynyl" as used herein, alone or in combination, each means a straight-chain or branched hydrocarbon moiety having from 2 to 10, 2 to 20, or 2 to 50 carbon atoms and containing at least one carbon-carbon triple bond. C 2-10 alkynyl, C 2-20 alkynyl, or C 2-50 alkynyl group may each hydrogen atom optionally be replaced by a substituent as defined above. Optionally, one or more double bonds may be present. Optionally, C 2-10 alkynyl, C 2-20 alkynyl, or C 2-50 alkynyl may one or more moieties as defined below be interposed.

[0048] As described above, C 1-4 alkyl, C 1-6 alkyl, C 1-10 alkyl, C 1-20 alkyl, C 1-50 alkyl, C 2-6 alkenyl, C 2-10 alkenyl, C 2-20 alkenyl, C 2-50 alkenyl, C 2-6 alkynyl, C 2-10 alkynyl, C2-20 Alkenyl or C 2-50 In alkynyl, one or more moieties may optionally intervene, and said one or more moieties, in certain embodiments, [Chemical formula] (wherein, The dashed line indicates the bond to the remainder of said moiety or reagent, -R and -R a are, independently of each other, selected from the group consisting of -H, methyl, ethyl, propyl, butyl, pentyl and hexyl) selected from the group consisting of.

[0049] As used herein, the term "C 3-10 cycloalkyl" means a cyclic alkyl chain having 3 to 10 carbon atoms, which may be saturated or unsaturated, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, cycloheptyl, cyclooctyl, cyclononyl or cyclodecyl. Each hydrogen atom of C 3-10 cycloalkyl carbon may be replaced by a substituent as defined above. The term "C 3-10 cycloalkyl" also includes bridged bicyclics such as norbornane or norbornene.

[0050] The term "8- to 30-membered carbopolycyclic" or "8- to 30-membered carbon polycyclic" means a polycyclic moiety having 8 to 30 ring atoms, wherein two adjacent rings share at least one ring atom, and which can contain up to the maximum number of double bonds (completely saturated, partially saturated or unsaturated, aromatic or non-aromatic rings). Preferably, 8- to 30-membered carbopolycyclic means a cyclic moiety of 2, 3, 4 or 5 rings, more preferably 2, 3 or 4 rings.

[0051] As used herein, the term "3- to 10-membered heterocyclyl" or "3- to 10-membered heterocyclic ring" has 3, 4, 5, 6, 7, 8, 9, or 10 ring atoms, with at least 1 and up to a maximum of 4 ring atoms replaced by heteroatoms selected from the group consisting of sulfur (-S(O)-, -S(O)2- included), oxygen, and nitrogen (=N(O)- included), and can contain up to the maximum number of double bonds (fully saturated, partially saturated, or unsaturated, aromatic or non-aromatic rings), and means a ring bonded to the remainder of the molecule by a carbon or nitrogen atom. Examples of 3- to 10-membered heterocyclic rings include, but are not limited to, aziridine, oxirane, thiirane, azirine, oxirene, thiirene, azetidine, oxetane, thietane, furan, thiophene, pyrrole, pyrroline, imidazole, imidazoline, pyrazole, pyrazoline, oxazole, oxazoline, isoxazole, isoxazoline, thiazole, thiazoline, isothiazole, isothiazoline, thiadiazole, thiadiazoline, tetrahydrofuran, tetrahydrothiophene, pyrrolidine, imidazolidine, pyrazolidine, oxazolidine, isoxazolidine, thiazolidine, isothiazolidine, thiadiazolidine, sulfolane, pyran, dihydropyran, tetrahydropyran, imidazolidine, pyridine, pyridazine, pyrazine, pyrimidine, piperazine, piperidine, morpholine, tetrazole, triazole, triazolidine, tetrazolidine, diazepane, azepine, and homopiperazine. Each hydrogen atom of a 3- to 10-membered heterocyclyl or 3- to 10-membered heterocyclic ring moiety may optionally be replaced by a substituent as defined below.

[0052] As used herein, the term "8- to 11-membered heterobicyclic" or "8- to 11-membered heterocyclic bicyclic" refers to a bicyclic heterocyclic moiety having 8 to 11 ring atoms and capable of containing up to the maximum number of double bonds (completely saturated, partially saturated, or unsaturated, aromatic or non-aromatic rings), wherein at least one ring atom is shared by both rings, and at least one, up to a maximum of 6 ring atoms, is replaced by a heteroatom selected from the group consisting of sulfur (-S(O)-, -S(O)2-), oxygen, and nitrogen (=N(O)-), and the ring is attached to the remainder of the molecule by a carbon or nitrogen atom. Examples of 8- to 11-membered heterocyclic bicyclics include indole, indoline, benzofuran, benzothiophene, benzoxazole, benzisoxazole, benzothiazole, benzisothiazole, benzimidazole, benzimidazoline, quinoline, quinazoline, dihydroquinazoline, quinoline, dihydroquinoline, tetrahydroquinoline, decahydroquinoline, isoquinoline, decahydroisoquinoline, tetrahydroisoquinoline, dihydroisoquinoline, benzazepine, purine, and pteridine. The term 8- to 11-membered heterocyclic bicyclic also includes bicyclic spiro structures such as 1,4-dioxa-8-azaspiro[4.5]decane or bridged heterocycles such as 8-aza-bicyclo[3.2.1]octane. Each hydrogen atom of an 8- to 11-membered heterobicyclic or 8- to 11-membered heterocyclic bicyclic carbon may optionally be replaced by a substituent as defined below.

[0053] Similarly, the term "8- to 30-membered hetero polycyclic" or "8- to 30-membered hetero polycycle" means a heterocyclic moiety having 8 to 30 ring atoms and capable of containing up to a maximum number of double bonds (fully saturated, partially saturated, or unsaturated, aromatic or non-aromatic rings), which is a polycycle of more than two rings, in certain embodiments a 3-, 4- or 5-ring heterocyclic moiety, wherein two adjacent rings share at least one ring atom, and at least one ring atom, up to a maximum of 10 ring atoms, is replaced by a heteroatom selected from the group consisting of sulfur (including -S(O)-, -S(O)2-), oxygen, and nitrogen (including =N(O)-), and said ring is attached to the remainder of the molecule by a carbon or nitrogen atom.

[0054] The following structure:

Chemical formula

Chemical formula

[0055] The following structure:

Chemical formula

Chemical formula

[0056] As used herein, the term "dry pharmaceutical formulation" means that a pharmaceutical formulation that can be stored in a dried form before administration, for example, for at least 1 week, for example, for at least 2 weeks, for example, for at least 1 month, for example, for at least 6 months, for example, for at least 12 months, for example, for at least 18 months, for example, for at least 24 months, for example, for at least 36 months, or for example, for at least 48 months, is provided in a dried form. Suitable drying methods are spray drying and freeze drying, i.e., lyophilization. Such dry pharmaceutical formulations containing CNP compounds have a residual moisture content of up to 5%, in certain embodiments less than 2%, and in certain embodiments less than 1% as measured by the Karl Fischer method. In certain embodiments, the dry pharmaceutical formulations of the invention are dried by lyophilization.

[0057] As used herein, the term "drug" refers to a substance used in the treatment, cure, prevention or diagnosis of a disease, or in other ways to promote physical or mental well-being. When a drug such as CNP is attached to another moiety, the resulting product moiety derived from that drug is referred to as a "drug moiety".

[0058] As used herein, the "EC 50 " for a CNP agonist or CNP conjugate refers to the concentration of the CNP agonist or CNP conjugate at which half of the maximal cGMP production is induced. The EC 50 of CNP agonists, CNP conjugates, and CNP-22 in terms of NPR-B activity is measured by culturing NIH-3T3 (mouse embryonic fibroblast cell line) cells expressing NPR-B on the cell surface and incubating the cells with a CNP agonist, a CNP conjugate, the corresponding released CNP, or CNP-22, respectively, and measuring the intracellular production of the second messenger cGMP in a standard cGMP assay. In particular, the assay is performed as follows: (1) Mouse NIH-3T3 cells expressing endogenous NPR-B are cultured in DMEM F-12 medium containing 5% FBS and 5 mM glutamine at 37 °C and 5% CO2; (2) For each assay, 50,000 cells are resuspended in Dulbecco's PBS containing IBMX and incubated at different concentrations with either a CNP agonist, a CNP conjugate, the corresponding released CNP or CNP-22; (3) After incubation at 37 °C and 5% CO2 for 30 minutes, the cells are lysed and the cGMP level is measured; (4) An EC 50 value is generated from the measured cGMP levels.

[0059] Preferably, the IBMX concentration in step (2) is 0.5 mM.

[0060] Step (3) can be carried out using any assay for measuring cGMP, which is a standard procedure well known to those skilled in the art. Preferably, step (3) is carried out using a cGMP TR-FRET assay, more preferably, the cGMP TR-FRET assay from Cisbio, catalog number 62GM2PEB. During such experiments, since the reversible CNP complex releases a certain amount of CNP that may affect the results, the measurement of the NPR-B activity of the CNP complex is preferably carried out in the form of a stable CNP complex that does not release CNP.

[0061] As used herein, the term "excipient" refers to a compound administered together with a CNP compound, such as a buffering agent, an isotonicity adjusting agent, a preservative, a stabilizer, an adsorption inhibitor, an antioxidant, or other adjuvants. However, in some cases, one excipient may have dual or triple functions. The term "excipient" can also refer to a diluent, an adjuvant or a carrier used together to administer a CNP compound. Such pharmaceutical additives can be sterile liquids, such as water and oils, and the oils include those of petroleum, animal, plant or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, etc., but are not limited thereto. When the pharmaceutical preparation is administered orally, water is a preferred excipient. When the pharmaceutical preparation is administered intravenously or subcutaneously, physiological saline and glucose aqueous solution are preferred excipients. In certain embodiments, as the liquid excipient for the injection solution, physiological saline, glucose aqueous solution and glycerol solution are used. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, mannitol, trehalose, gelatin, malt, rice, wheat flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, nonfat dry milk, glycerol, propylene, glycol, water, ethanol, etc. The pharmaceutical preparation can also contain a small amount of a wetting agent or an emulsifier, a pH buffering agent, such as acetate, succinate, Tris (tris (hydroxymethyl) aminomethane), carbonate, phosphate, HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), MES (2-(N-morpholino) ethanesulfonic acid), etc., or a surfactant, such as Tween (registered trademark), poloxamer, poloxamine, CHAPS, Igepal (registered trademark), etc., or an amino acid, such as glycine, lysine or histidine, etc. These pharmaceutical preparations can take forms such as solutions, suspensions, emulsions, tablets, pills, capsules, powders, sustained-release preparations, etc. The pharmaceutical preparation can be formulated as a suppository using conventional binders and additives, such as triglycerides, etc.Oral formulations can contain standard excipients, e.g., pharmaceutical mannitol, lactose, starch, magnesium stearate, sodium saccharin, cellulose, magnesium carbonate, etc. Such formulations will contain a therapeutically effective amount of a CNP compound together with a suitable amount of excipients to provide the form for proper administration to a patient. The formulation should be suitable for the method of administration.

[0062] As used herein, the term "formulation" or "drug formulation" refers to a formulation containing one or more CNP compounds and one or more excipients, and to a product that is directly or indirectly produced by the combination, complexation or aggregation of any two or more of the components of the composition, or by the dissociation of one or more of those components, or by one or more other types of reaction or interaction of those components. Thus, the pharmaceutical formulations of the present invention include formulations or compositions made by mixing one or more CNP compounds and pharmaceutically acceptable excipients, e.g., buffers and isotonic agents.

[0063] As used herein, the term "free form" of a medicament refers to the medicament in an unmodified, pharmacologically fully active form, e.g., after being released from a reversible CNP conjugate.

[0064] As used herein, the term "functional group" means an atomic group capable of reacting with other atomic groups. Functional groups include, but are not limited to, the following groups: carboxylic acid (-(C=O)OH), primary or secondary amine (-NH2, -NH-), maleimide, thiol (-SH), sulfonic acid (-(O=S=O)OH), carbonate, carbamate (-O(C=O)N<), hydroxyl (-OH), aldehyde (-(C=O)H), ketone (-(C=O)-), hydrazine (>N-N<), isocyanate, isothiocyanate, phosphoric acid (-O(P=O)OHOH), phosphonic acid (-O(P=O)OHH), haloacetyl, alkyl halide, acryloyl, aryl fluoride, hydroxylamine, disulfide, sulfonamide, sulfonic acid, vinyl sulfone, vinyl ketone, diazoalkane, oxirane, and aziridine, among others.

[0065] As used herein, the term "halogen" means fluoro, chloro, bromo, or iodo. Halogen is generally preferably fluoro or chloro.

[0066] As used herein, the term "interrupted" means that a moiety is inserted between two carbon atoms, or, in the case where the insertion is at one end of the moiety, between a carbon or heteroatom and a hydrogen atom, and in certain embodiments between a carbon and a hydrogen atom.

[0067] As used herein, the term "tonicity agent" refers to a compound that reduces pain, irritation, and tissue damage that can result from cell damage due to an osmotic pressure difference between an injected solution and plasma.

[0068] As used herein, the term "liquid pharmaceutical formulation" means that the pharmaceutical formulation is provided and can be stored in liquid form for a period of time prior to administration, such as at least 1 week, such as at least 2 weeks, such as at least 1 month, such as at least 6 months, such as at least 12 months, such as at least 18 months, such as at least 24 months, such as at least 36 months, or such as at least 48 months.

[0069] As used herein, the term "lyophilized formulation" means that a formulation containing a CNP compound is first frozen and then water is removed by reduced pressure. This term does not exclude additional drying steps performed in the manufacturing process prior to filling the formulation into the final container.

[0070] As used herein, the terms "lyophilization" or "freeze-drying" are used interchangeably and refer to a dehydration process characterized by freezing a formulation and then reducing the ambient pressure and optionally applying heat to directly sublimate the frozen water in the formulation from the solid phase to the gas phase. Typically, the displaced water is recovered by sublimation.

[0071] As used herein, the term "moiety" means a part of a molecule that lacks one or more atoms compared to the corresponding drug. For example, when a reagent of the formula "H-X-H" reacts with another reagent and becomes part of the reaction product, the corresponding part of the reaction product has the structure "H-X-" or "-X-", where each "-" represents a bond to another part. Thus, a pharmaceutical moiety such as a CNP moiety is released as a pharmaceutical such as CNP from a reversible CNP conjugate.

[0072] When the arrangement or chemical structure of an atomic group that is bonded to two parts or inserted into one part is provided, unless otherwise clearly stated, it is understood that the said arrangement or chemical structure may be bonded to the two parts in any orientation. For example, the moiety "-C(O)N(R 1 )-" may be "-C(O)N(R 1 )-" or "-N(R1 ) As "-C(O)-", it can be bonded to two moieties or can also insert into one moiety. Similarly, the moiety: [Chemical formula] is [Chemical formula] such that it can be bonded to two moieties or can insert into one moiety.

[0073] When the CNP moiety contains one or more acidic or basic groups, the liquid pharmaceutical formulation also includes their corresponding pharmaceutically or toxicologically acceptable salts, in particular their pharmaceutically available salts. Thus, a CNP moiety containing one or more acidic groups can be present and can be used, for example, as an alkali metal salt, an alkaline earth metal salt or an ammonium salt. More detailed examples of such salts include sodium salts, potassium salts, calcium salts, magnesium salts, or salts with ammonia or organic amines (e.g., ethylamine, ethanolamine, triethanolamine, etc.) or amino acids, as well as other salts or amines known to those skilled in the art. A CNP moiety containing one or more basic groups, i.e., groups that can be protonated, can also be present and can be used in the form of their addition salts with inorganic or organic acids. Examples of suitable acids include hydrogen chloride, hydrogen bromide, phosphoric acid, sulfuric acid, nitric acid, methanesulfonic acid, p-toluenesulfonic acid, naphthalenedisulfonic acid, oxalic acid, acetic acid, tartaric acid, lactic acid, salicylic acid, benzoic acid, formic acid, propionic acid, pivalic acid, diethylacetic acid, malonic acid, succinic acid, pimelic acid, fumaric acid, maleic acid, malic acid, sulfamic acid, phenylpropionic acid, gluconic acid, ascorbic acid, isonicotinic acid, citric acid, adipic acid, and other acids known to those skilled in the art. Further methods for converting basic groups to cations, such as alkylation of amine groups to provide suitable counterions for positively charged ammonium groups and their salts, are known to those skilled in the art. When the CNP moiety contains both acidic and basic groups simultaneously, the pharmaceutical formulations according to the invention also include inner salts or betaines (zwitterions) in addition to the salt forms mentioned. Each salt can be obtained, for example, by contacting these conjugates with an organic or inorganic acid or base in a solvent or dispersant, or by anion exchange or cation exchange with other salts, by conventional methods known to those skilled in the art. The formulations according to the invention are not directly suitable for use in pharmaceuticals due to low physiological compatibility, but include all salts of the CNP conjugate that can be used, for example, as intermediates in chemical reactions or in the preparation of pharmaceutically acceptable salts.

[0074] As used herein, the term "natural product" refers to an organic compound isolated and purified from a natural source produced by a primary or secondary metabolic pathway.

[0075] As used herein, the term "oligonucleotide" preferably refers to double-stranded or single-stranded RNA and DNA of 2 to 1000 nucleotides, and any modifications thereof. Modifications include, for example, modifications that provide additional charge, polarizability, hydrogen bonding, electrostatic interactions, and other chemical groups incorporated into the nucleic acid ligand base or the entire nucleic acid ligand. Such modifications include, for example, 2'-sugar modifications, 5-position pyrimidine modifications, 8-position purine modifications, modifications at exocyclic amines, substitution of 4-thiouridine, substitution of 5-bromo or 5-iodo-uracil, backbone modifications, methylation, combinations of abnormal base pairing such as isobases isocytidine and isoguanidine. Modifications also include 3' and 5' modifications such as capping and changes in stereochemistry. This term also includes aptamers.

[0076] As used herein, the term "antioxidant" or "oxidation protector" refers to a compound that inhibits the oxidation of other compounds such as peptides.

[0077] As used herein, the term "pH adjuster" refers to a compound used to adjust the pH of a solution.

[0078] As used herein, the term "pharmaceutically acceptable" means a substance that does not cause harm when administered to a patient, and preferably means that it has been approved for use in animals, preferably for use in humans, by a regulatory authority, such as the EMA (Europe) and / or the FDA (USA) and / or the regulatory authority of any other country.

[0079] As used herein, the term "physiological conditions" refers to aqueous buffer conditions of pH 7.4 and 37°C.

[0080] As used herein, the term "polypeptide" refers to a chain of 2 to 50 amino acid monomer moieties linked by peptide (amide) linkages. Only in the case of CNP, sequences having more than 50 amino acids are also referred to simply as "polypeptides".

[0081] As used herein, the term "preservative" refers to a chemical substance that has an antibacterial effect and / or prevents chemical decomposition.

[0082] As used herein, the term "protein" refers to a chain of amino acid monomer moieties having more than 50 amino acid monomer moieties linked by peptide linkages, preferably 12,000 or fewer amino acid monomers linked by peptide linkages, for example, 10,000 or fewer amino acid monomer moieties, 8,000 or fewer amino acid monomer moieties, 5,000 or fewer amino acid monomer moieties, or 2,000 or fewer amino acid monomer moieties.

[0083] As used herein, the term "polymer" refers to a molecule containing repeating structural units, i.e., monomers, linked linearly, cyclically, branched, cross-linked, or dendrimer-like or in combinations thereof by chemical bonds, which may be of synthetic origin, biological origin, or a combination of both. It is understood that a polymer may also contain one or more other chemical groups and / or moieties, such as one or more functional groups. In certain embodiments, a soluble polymer has a molecular weight of at least 0.5 kDa, for example, at least 1 kDa, at least 2 kDa, at least 3 kDa, or at least 5 kDa. When the polymer is soluble, in certain embodiments, it has a molecular weight of up to 1000 kDa, for example, up to 750 kDa, for example, up to 500 kDa, for example, up to 300 kDa, for example, up to 200 kDa, for example, up to 100 kDa.

[0084] It is understood that a protein or polypeptide is also a polymer in which the amino acids are repeating structural units, even though the side chains of each amino acid may be different.

[0085] As used herein, the terms "polymeric" or "polymeric moiety" mean a reagent or moiety that includes one or more polymers or polymeric moieties. The polymeric agent or moiety may optionally also include one or more other moieties, which, in certain embodiments, are selected from the group consisting of: · C 1-50 alkyl, C 2-50 alkenyl, C 2-50 alkynyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl, and tetralinyl, and · linkages selected from the group consisting of:

Chemical formula

[0086] One of ordinary skill in the art will understand that polymerization products obtained from polymerization reactions do not necessarily all have the same molecular weight, but rather exhibit a molecular weight distribution. As a result, the molecular weight ranges, molecular weights, monomer number ranges in the polymer, and monomer numbers in the polymer used herein refer to the number average molecular weight and the average number of monomers, i.e., the arithmetic mean of the molecular weights of the polymer or polymeric moiety, and the arithmetic mean of the number of monomers in the polymer or polymeric moiety.

[0087] Thus, in the case of a polymer moiety containing "x" monomer units, any integer applied to "x" corresponds to the arithmetic mean of the monomers. Any range of integers applied to "x" provides the range of integers in which the arithmetic mean of the monomers exists. The integer of "x" indicated as "about x" means that the arithmetic mean of the monomers is in the integer range of x+ / -10%, in certain embodiments in the integer range of x+ / -8%, in certain embodiments in the integer range of x+ / -5%, and in certain embodiments in the integer range of x+ / -2%.

[0088] As used herein, the term "PEG-based" with respect to a moiety or reagent means that the moiety or reagent contains PEG. In certain embodiments, the PEG-based moiety or reagent contains at least 10 wt% PEG, such as at least 20 wt% PEG, such as at least 30 wt% PEG, such as at least 40 wt% PEG, such as at least 50 wt%, such as at least 60 wt% PEG, such as at least 70 wt% PEG, such as at least 80 wt% PEG, such as at least 90 wt%, such as at least 95 wt% PEG. The remaining weight percentage of the PEG-based moiety or reagent is other moieties selected from the following moieties and linkages. ·C 1-50 alkyl, C 2-50 alkenyl, C 2-50 alkynyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl and tetralinyl, and ·linkages selected from the group consisting of the following:[[]]

Chemical formula

[0089] As used herein, the term "PEG-based containing at least X% PEG" with respect to a moiety or reagent means that the moiety or reagent contains at least X weight % of ethylene glycol units (-CH2CH2O-), and the ethylene glycol units can be arranged in an alternating block pattern or can be randomly distributed within the moiety or reagent. In certain embodiments, all of the ethylene glycol units of the moiety or reagent are present within one block, and the remaining weight percentage of the PEG-based moiety or reagent is other moieties selected from the following moieties and linkages in certain embodiments. ·C 1-50 alkyl, C 2-50 alkenyl, C 2-50 alkynyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, phenyl, naphthyl, indenyl, indanyl and tetralinyl, and ·linkages selected from the group consisting of:

Chemical formula

[0090] As used herein, the term "hyaluronic acid-based containing at least X% hyaluronic acid" is used as appropriate.

[0091] It is also recognized by those skilled in the art that the reversible CNP conjugate contained in the liquid preparation of the present invention is a prodrug. As used herein, the term "prodrug" refers to a pharmaceutical moiety, such as a CNP moiety, reversibly and covalently bound to a carrier moiety, such as -Z, via a reversible linker moiety. A prodrug releases the reversibly and covalently bound pharmaceutical moiety in the form of its corresponding pharmaceutical. That is, a prodrug is a reversible conjugate containing a pharmaceutical moiety, such as a CNP moiety, covalently and reversibly bound to a carrier moiety via a reversible linker moiety, and the covalent and reversible binding of the carrier moiety to the reversible linker moiety is direct or via a spacer. Such a prodrug or conjugate releases the previously bound pharmaceutical moiety in the form of a free drug.

[0092] As used herein, the term "random coil" refers to a peptide or protein that adopts / has / forms a three-dimensional structure substantially lacking a defined secondary and tertiary structure, as determined by circular dichroism spectroscopy performed in an aqueous buffer at pH 7.4 at room temperature. In certain embodiments, room temperature (ambient temperature) is about 20 °C, i.e., 18 °C to 22 °C, while in other certain embodiments, the environmental temperature is 20 °C.

[0093] As used herein, the term "reversible linkage" refers to a linkage that is cleavable in the absence of an enzyme under physiological conditions (aqueous buffer at pH 7.4, 37 °C), and the half-life ranges from 1 hour to 6 months, such as 1 hour to 4 months, such as 1 hour to 3 months, 1 hour to 2 months, or 1 hour to 1 month. Thus, a stable bond is a bond with a half-life of 6 months or more under physiological conditions (aqueous buffer at pH 7.4, 37 °C).

[0094] As used herein, the term "reagent" means a compound containing at least one functional group for reaction with another compound or a functional group of a pharmaceutical. It is understood that a pharmaceutical containing a functional group (e.g., a primary or secondary amine or a hydroxy functional group) is also a reagent.

[0095] As used herein, the term "reversible linker moiety" is a moiety that is covalently attached to a pharmaceutical moiety such as a CNP moiety by a reversible linkage and is also covalently attached to a carrier moiety such as -Z, where the covalent attachment to the carrier moiety is either direct or via a spacer moiety such as -L 2 -. In certain embodiments, the linkage between -Z and -L 2 - is a stable linkage.

[0096] As used herein, the term "reconstitution" means adding a liquid to a dry pharmaceutical formulation to return it to its original form, such as a solution formulation.

[0097] As used herein, the term "small molecule" or "small drug molecule" refers to a molecule or drug that is an organic compound having a molecular weight of less than 1000 Da, such as less than 900 Da or less than 800 Da.

[0098] As used herein, the term "sealing a container" means sealing the container so that it is airtight, preventing any gas exchange between the outside and the inside, and maintaining the sterility of the contents.

[0099] As used herein, the term "spacer" or "spacer moiety" refers to a moiety suitable for connecting two moieties. Suitable spacers can be selected from the group consisting of C 1-50 alkyl, C 2-50 alkenyl or C 2-50 alkynyl, and C 1-50 alkyl, C 2-50 alkenyl or C 2-50 alkynyl may be interrupted by one or more groups selected from -NH-, -N(C 1-4 alkyl)-, -O-, -S-, -C(O)-, -C(O)NH-, -C(O)N(C 1-4 alkyl)-, -O-C(O)-, -S(O)-, -S(O)2-, 4- to 7-membered heterocyclyl, phenyl and naphthyl.

[0100] As used herein, the term "substituted" means that one or more -H atoms of a molecule or moiety are replaced by different atoms or groups of atoms referred to as "substituents".

[0101] In certain embodiments, such one or more substituents are, independently of each other, halogen, -CN, -COOR x1 , -OR x1 , -C(O)R x1 , -C(O)N(R x1 R x1a ), -S(O)2N(R x1 R x1a ), -S(O)N(R x1 R x1a ), -S(O)2R x1 , -S(O)R x1 , -N(R x1 ), S(O)2N(R x1a R x1b ), -SR x1 , -N(R x1 R x1a ), -NO2, -OC(O)R x1 , -N(R x1 ), C(O)R x1a , -N(R x1 ), S(O)2R x1a , -N(R x1 ), S(O)R x1a , -N(R x1 ), C(O)OR x1a , -N(R x1 ), C(O)N(R x1a R x1b ), -OC(O)N(R x1 R x1a ), -T 0 , C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; -T 0 , C 1-50 alkyl, C 2-50 alkenyl and C 2-50 alkynyl may be substituted with one or more -R x2 that are the same or different, C 1-50Alkyl, C 2-50 Alkenyl and C 2-50 Alkynyl may be interrupted by one or more groups selected from the group consisting of -T 0 -, -C(O)O-, -O-, -C(O)-, -C(O)N(R x3 ), -, -S(O)2N(R x3 ), -, -S(O)N(R x3 ), -, -S(O)2-, -S(O)-, -N(R x3 )S(O)2N(R x3a ), -, -S-, -N(R x3 ), -, -OC(OR x3 )(R x3a ), -, -N(R x3 )C(O)N(R x3a ), - and -OC(O)N(R x3 ); -R x1 -, -R x1a -, -R x1b are, independently of one another, -H, -T 0 , C 1-50 Alkyl, C 2-50 Alkenyl, and C 2-50 Alkynyl selected from the group consisting of; -T 0 , C 1-50 Alkyl, C 2-50 Alkenyl, and C 2-50 Alkynyl may be substituted with one or more -R x2 which are the same or different, and C 1-50 Alkyl, C 2-50 Alkenyl, and C 2-50 Alkynyl is -T 0 -, -C(O)O-, -O-, -C(O)-, -C(O)N(R x3 ), -, -S(O)2N(R x3 ), -, -S(O)N(R x3 ); -S(O)2-, -S(O)-, -N(R x3 )S(O)2N(R x3a ), -, -S-, -N(R x3 ), -, -OC(OR x3 )(R x3a ), -, -N(R x3 )C(O)N(R x3a ), - and -OC(O)N(Rx3 )- may be interrupted by one or more groups selected from the group consisting of; each T 0 is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, and 8- to 11-membered heterobicyclic; each T 0 may be independently substituted with one or more -R x2 which are the same or different; each -R x2 is independently selected from the group consisting of halogen, -CN, oxo (=O), -COOR x4 , -OR x4 , -C(O)R x4 , -C(O)N(R x4 R x4a ), -S(O)2N(R x4 R x4a ), -S(O)N(R x4 R x4a ), -S(O)2R x4 , -S(O)R x4 , -N(R x4 )(O)2N(R x4a R x4b ), -SR x4 , -N(R x4 R x4a ), -NO2, -OC(O)R x4 , -N(R x4 )(O)R x4a , -N(R x4 )(O)2R x4a , -N(R x4 )(O)R x4a , -N(R x4 )(O)OR x4a , -N(R x4 )(O)N(R x4a R x4b ), -OC(O)N(R x4 R x4a ), and C 1-6 alkyl; C 1-6 alkyl may be optionally substituted with one or more halogen which are the same or different; each -R x3 , -R x3a , -Rx4 , -R x4a , -R x4b are each independently selected from the group consisting of -H and C 1-6 alkyl; C 1-6 alkyl may be substituted with one or more halogens, which may be the same or different.

[0102] In certain embodiments, said one or more substituents are each independently halogen, -CN, -COOR x1 , -OR x1 , -C(O)R x1 , -C(O)N(R x1 R x1a ), -S(O)2N(R x1 R x1a ), -S(O)N(R x1 R x1a ), -S(O)2R x1 , -S(O)R x1 , -N(R x1 ), S(O)2N(R x1a R x1b ), -SR x1 , -N(R x1 R x1a ), -NO2, -OC(O)R x1 , -N(R x1 ), C(O)R x1a , -N(R x1 ), S(O)2R x1a , -N(R x1 ), S(O)R x1a , -N(R x1 ), C(O)OR x1a , -N(R x1 ), C(O)N(R x1a R x1b ), -OC(O)N(R x1 R x1a ), -T 0 , C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl; -T 0 , C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl may be substituted with one or more -R, which may be the same or different. x2may be replaced by C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl may be interrupted by one or more groups selected from the group consisting of -T 0 -, -C(O)O-, -O-, -C(O)-, -C(O)N(R x3 ), -S(O)2N(R x3 ), -S(O)N(R x3 ), -S(O)2-, -S(O)-, -N(R x3 )S(O)2N(R x3a ), -S-, -N(R x3 ), -OC(OR x3 )(R x3a ), -N(R x3 )C(O)N(R x3a ), and -OC(O)N(R x3 ); each -R x1 , -R x1a , -R x1b , -R x3 , -R x3a is independently selected from the group consisting of -H, halogen, C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl; each T 0 is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, and 8- to 11-membered heterobicyclyl; each T 0 may be independently substituted by one or more -R x2 which are the same or different; each -R x2 is independently halogen, -CN, oxo(=O), -COOR x4 , -OR x4 , -C(O)R x4 , -C(O)N(R x4 R x4a ), -S(O)2N(R x4 R x4a ), -S(O)N(R x4 R x4a ), -S(O)2Rx4 、 -S(O)R x4 、 -N(R x4 )S(O)2N(R x4a R x4b )、 -SR x4 、 -N(R x4 R x4a )、 -NO2、 -OC(O)R x4 、 -N(R x4 )C(O)R x4a 、 -N(R x4 )S(O)2R x4a 、 -N(R x4 )S(O)R x4a 、 -N(R x4 )C(O)OR x4a 、 -N(R x4 )C(O)N(R x4a R x4b )、 -OC(O)N(R x4 R x4a )、 and C 1-6 selected from the group consisting of alkyl; C 1-6 alkyl may be substituted with one or more halogens that are the same or different; each -R x4 、 -R x4a 、 -R x4b is independently selected from the group consisting of -H, halogen, C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl.

[0103] In certain embodiments, said one or more substituents are, independently of each other, halogen, -CN, -COOR x1 、 -OR x1 、 -C(O)R x1 、 -C(O)N(R x1 R x1a )、 -S(O)2N(R x1 R x1a )、 -S(O)N(R x1 R x1a )、 -S(O)2R x1 、 -S(O)R x1 、 -N(R x1 )S(O)2N(R x1a R x1b )、 -SR x1 、 -N(Rx1 R x1a )、 -NO2, -OC(O)R x1 、 -N(R x1 )C(O)R x1a 、 -N(R x1 )S(O)2R x1a 、 -N(R x1 )S(O)R x1a 、 -N(R x1 )C(O)OR x1a 、 -N(R x1 )C(O)N(R x1a R x1b )、 -OC(O)N(R x1 R x1a )、 -T 0 、 C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl selected from the group consisting of; -T 0 、 C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl may be substituted with one or more -R x2 that are the same or different, and C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl may be interrupted by one or more groups selected from the group consisting of -T 0 -, -C(O)O-, -O-, -C(O)-, -C(O)N(R x3 ), -S(O)2N(R x3 ), -S(O)N(R x3 ), -S(O)2-, -S(O)-, -N(R x3 )S(O)2N(R x3a ), -S-, -N(R x3 ), -OC(OR x3 )(R x3a ), -N(R x3 )C(O)N(R x3a ), and -OC(O)N(R x3 ); each -R x1 、 -R x1a 、 -R x1b 、 -R x2 、 -R x3 、 -R x3ais independently selected from the group consisting of -H, halogen, C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl; each T 0 is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, and 8- to 11-membered heterobicyclic; each T 0 is independently optionally substituted with one or more -R x2 .

[0104] In certain embodiments, up to 6 H atoms of the optionally substituted molecule are independently replaced by substituents, for example 5 H atoms are independently replaced by substituents, 4 H atoms are independently replaced by substituents, 3 H atoms are independently replaced by substituents, 2 H atoms are independently replaced by substituents, or 1 H atom is independently replaced by a substituent.

[0105] As used herein, the terms "stable" and "stability" with respect to a pharmaceutical formulation containing a reversible CNP conjugate mean that after a storage period, e.g., after 1 month, 2 months, 4 months, 6 months, 8 months, 12 months, 18 months, 24 months, 36 months, 48 months, 60 months, and particularly after a specified storage period, the pharmaceutical formulation contains less than 5% of the CNP in free form and less than 20%, e.g., less than 10%, or e.g., less than 5% of impurities, and impurities resulting from the oxidation of methionine. Such storage is preferably done at 2 - 8°C, i.e., by storing the formulation in a liquid state. The impurities can be quantified by RP - HPLC or SEC based on the individual peak area relative to the total peak area of all CNP conjugate - related peaks in the chromatogram. The impurities in the CNP moiety of the CNP conjugate can be determined by proteolytic digestion or peptide mapping and quantified based on the individual peak area relative to the peak area of the corresponding unmodified proteolytic peptide. Thus, the terms "stable" and "stability" with respect to a pharmaceutical formulation containing a CNP agonist or a stable CNP conjugate mean that after a storage period of 1 month, 2 months, 4 months, 6 months, 8 months, 12 months, 18 months, 24 months, 36 months, 48 months, 60 months, etc., and after a specific storage period, the pharmaceutical formulation contains less than 20%, e.g., less than 10% or e.g., less than 5% of impurities such as impurities resulting from the oxidation of methionine. The impurities can be quantified by RP - HPLC or SEC based on the respective peak area relative to the total peak area of all CNP agonist or stable CNP conjugate - related peaks, particularly based on the CNP conjugate - related peaks within the chromatogram. The impurities within the CNP agonist are determined by proteolytic digestion or peptide mapping and can be quantified based on the respective peak area relative to the peak area of the corresponding unmodified proteolytic peptide.

[0106] As used herein, the term "stabilizer" refers to a compound used to stabilize a CNP compound. Stabilization can be achieved by enhancing the peptide stabilizing power or by direct binding of an excipient to the CNP compound.

[0107] As used herein, the term "surfactant" refers to a wetting agent that reduces the surface tension of a liquid.

[0108] As used herein, the term "therapeutically effective amount" means an amount sufficient to cure, alleviate or partially prevent the clinical symptoms of a given disease and its complications. The amount effective for each purpose is determined not only by the severity of the disease or injury, but also by the body weight and general condition of the subject. As will be appreciated, determining the appropriate dosage can be done using routine experimentation by constructing a matrix of numerical values and testing various points within that matrix, all of which are within the ordinary skill of a skilled physician. Within the scope of the present invention, a therapeutically effective amount relates to a dosage targeted to achieve a therapeutic effect over a long period of time, for at least 1 day, such as 2 days, such as 3 days, such as 4 days, such as 5 days, such as 6 days, such as 1 week or such as 2 weeks.

[0109] As used herein, the term "non-tracer linker" means a reversible linker that releases the drug in its free form upon cleavage.

[0110] As used herein, the term "unit dose" means the amount of a medicament administered to a patient in a single dose.

[0111] As used herein, in relation to a polymer moiety, the term "water-soluble" means that when such a carrier moiety, such as such a polymer moiety, is part of a CNP conjugate, at least 1 g of a CNP conjugate containing such a water-soluble carrier moiety can be dissolved in 1 liter of water at 20 °C to form a homogeneous solution.

[0112] Generally, the terms "comprise" or "comprising" include "consist of", "consisting of", or "consisting essentially of".

[0113] When the present disclosure refers to a range of values of a parameter, it should be understood that the disclosure also discloses the integers within that range and the sub-ranges formed between the integers within that range.

[0114] In certain embodiments, the CNP compound is a CNP conjugate.

[0115] In certain embodiments, the CNP compound is a reversible CNP conjugate, the CNP conjugate including at least one CNP moiety covalently and reversibly bound to a carrier moiety, such that at least one CNP moiety is released with a release half-life of at least 6 hours under physiological conditions.

[0116] In certain embodiments, the CNP compound is a stable CNP conjugate, the CNP conjugate including at least one CNP moiety covalently and stably bound to a carrier moiety via a stable linkage having a half-life of greater than 6 months under physiological conditions.

[0117] In certain embodiments, the CNP compound is a stable CNP conjugate, the CNP conjugate having a CNP moiety covalently and stably bound to a fatty acid derivative.

[0118] In certain embodiments, the CNP portion has a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, SEQ ID NO: 26, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 31, SEQ ID NO: 32, SEQ ID NO: 33, SEQ ID NO: 34, SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 37, SEQ ID NO: 38, SEQ ID NO: 39, SEQ ID NO: 40, SEQ ID NO: 41, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46, SEQ ID NO: 47, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 50, SEQ ID NO: 51, SEQ ID NO: 52, SEQ ID NO: 53, SEQ ID NO: 54, SEQ ID NO: 55, SEQ ID NO: 56, SEQ ID NO: 57, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94 and SEQ ID NO: 95.

[0119] In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 20. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 21. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 22. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 23. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 24. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 25. In certain embodiments, the CNP moiety has the sequence of SEQ ID NO: 30.

[0120] In certain embodiments, the ring moiety of the CNP moiety has SEQ ID NO: 96 or SEQ ID NO: 97. In certain embodiments, the ring moiety of the CNP moiety has SEQ ID NO: 96. In certain embodiments, the ring moiety of the CNP moiety has SEQ ID NO: 97.

[0121] In certain embodiments, the carrier moiety is a polymeric moiety defined by variable -Z, which is described in more detail elsewhere in this specification.

[0122] To maintain a particular pH or pH range, the liquid pharmaceutical formulation includes a buffer. The buffer maintains the pH of the liquid pharmaceutical formulation within a desired range.

[0123] The buffer can be selected from the group consisting of acetic acid, succinic acid, citric acid, lactic acid, glutamic acid, fumaric acid, aspartic acid, glutaric acid, phosphoric acid, histidine, gluconic acid, tartaric acid, malic acid, tris (tris (hydroxymethyl) aminomethane), and mixtures thereof. It will be apparent to those skilled in the art that the corresponding conjugate bases or salts of buffers such as acetates, succinates, citrates, lactates, glutamates, fumarates, aspartates, glutarates, phosphates, gluconates, tartrates, malates, and mixtures thereof are also included.

[0124] In certain embodiments, the buffer is selected from the group consisting of acetic acid, succinic acid, citric acid, lactic acid, glutamic acid, fumaric acid, aspartic acid, glutaric acid, phosphoric acid, histidine, gluconic acid, tartaric acid, and malic acid.

[0125] In certain embodiments, the buffer is acetic acid. In certain embodiments, the buffer is succinic acid. In certain embodiments, the buffer is citric acid. In certain embodiments, the buffer is lactic acid. In certain embodiments, the buffer is glutamic acid. In certain embodiments, the buffer is fumaric acid. In certain embodiments, the buffer is aspartic acid. In certain embodiments, the buffer is glutaric acid. In certain embodiments, the buffer is phosphoric acid. In certain embodiments, the buffer is histidine. In certain embodiments, the buffer is gluconic acid. In certain embodiments, the buffer is tartaric acid. In certain embodiments, the buffer is malic acid.

[0126] Acetic acid has been found to be a particularly useful buffer because no acetylated adduct impurities are formed that are detectable by RP-HPLC and / or RP-HPLC-MS for use under long-term storage.

[0127] In certain embodiments, the buffer is not citric acid.

[0128] In certain embodiments, the concentration of the buffer is in the range of 5 to 50 mM. In certain embodiments, the concentration of the buffer is in the range of 10 to 50 mM. In certain embodiments, the concentration of the buffer is about 10 mM.

[0129] In certain embodiments, under basic conditions, the reversible linkage within the reversible CNP conjugate may not be stable, and thus the pH of the liquid pharmaceutical formulation is 6 or less.

[0130] In certain embodiments, the pH of the liquid pharmaceutical formulation is from about pH 3.5 to about pH 5.5. In certain embodiments, the pH of the liquid pharmaceutical formulation is from about pH 3.5 to about pH 5.0. In certain embodiments, the pH of the liquid pharmaceutical formulation is from about pH 4.5 to about pH 5.0. In certain embodiments, the pH of the liquid pharmaceutical formulation is from about pH 4.0 to about pH 4.5. In certain embodiments, the pH of the liquid pharmaceutical formulation is about 4.5. In certain embodiments, the pH of the liquid pharmaceutical formulation is about 4.0. In certain embodiments, the pH of the liquid pharmaceutical formulation is 4.5. In certain embodiments, the pH of the liquid pharmaceutical formulation is 4.0.

[0131] The liquid pharmaceutical formulation according to the present invention contains an isotonic agent. The isotonic agent can be selected from the group consisting of mannitol, trehalose, sucrose, raffinose, gelatin, lactose, dibasic calcium phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, dextrose, dextran, Ficoll®, propylene glycol, glycerol, and mixtures thereof.

[0132] In certain embodiments, the isotonic agent is selected from the group consisting of mannitol, trehalose, sucrose, raffinose, gelatin, lactose, dibasic calcium phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, dextrose, dextran, propylene glycol, glycerol, and mixtures thereof.

[0133] In certain embodiments, the isotonic agent is selected from the group consisting of mannitol, trehalose, sucrose, raffinose, gelatin, lactose, dibasic calcium phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, dextrose, dextran, glycerol, and propylene glycol.

[0134] In certain embodiments, the isotonic agent is mannitol. In certain embodiments, the isotonic agent is trehalose, such as trehalose dihydrate. In certain embodiments, the isotonic agent is sucrose. In certain embodiments, the isotonic agent is raffinose. In certain embodiments, the isotonic agent is gelatin. In certain embodiments, the isotonic agent is lactose. In certain embodiments, the isotonic agent is dibasic calcium phosphate. In certain embodiments, the isotonic agent is sorbitol. In certain embodiments, the isotonic agent is xylitol. In certain embodiments, the isotonic agent is glycine. In certain embodiments, the isotonic agent is histidine. In certain embodiments, the isotonic agent is ethanol. In certain embodiments, the isotonic agent is hydroxyethyl starch. In certain embodiments, the isotonic agent is potassium chloride. In certain embodiments, the isotonic agent is sodium chloride. In certain embodiments, the isotonic agent is dextrose. In certain embodiments, the isotonic agent is dextran. In certain embodiments, the isotonic agent is Ficoll®. In certain embodiments, the isotonic agent is propylene glycol. In certain embodiments, the isotonic agent is glycerol.

[0135] In certain embodiments, the isotonic agent is selected from the group consisting of mannitol and trehalose.

[0136] In certain embodiments, the isotonic agent is a mixture of mannitol and trehalose.

[0137] As defined herein, the term "trehalose" is intended to encompass all salts and hydrated forms of trehalose, such as trehalose anhydrate or trehalose dihydrate. In certain embodiments, the term "trehalose" refers to trehalose anhydrate. In certain embodiments, the term "trehalose" refers to trehalose dihydrate.

[0138] As defined herein, the term "mannitol" is intended to encompass both D-mannitol and L-mannitol, and mixtures thereof. In certain embodiments, the term "mannitol" refers to L-mannitol. In certain embodiments, the term "mannitol" refers to D-mannitol. In certain embodiments, the term "mannitol" refers to a mixture of L-mannitol and D-mannitol.

[0139] In certain embodiments, the concentration of the isotonic agent ranges from 10 to 200 mg / mL. In certain embodiments, the concentration of the isotonic agent is about 100 mg / mL. In certain embodiments, the concentration of the isotonic agent is about 84 mg / mL. In certain embodiments, the concentration of the isotonic agent is about 80 mg / mL. In certain embodiments, the concentration of the isotonic agent is about 50 mg / mL. In certain embodiments, the concentration of the isotonic agent is about 41 mg / mL.

[0140] In some embodiments, the liquid formulation is substantially isotonic, meaning an osmotic pressure of about 250 - 350 mOsm / kg water. The liquid formulation can also be hypertonic (>350 mOsm / kg water) or hypotonic (<250 mOsm / kg water).

[0141] In certain embodiments, reducing sugars should be avoided because they may react with the CNP moiety. Thus, in certain embodiments, the pharmaceutical formulation does not contain reducing sugars.

[0142] The liquid pharmaceutical preparation according to the present invention contains a preservative. The preservative can be selected from the group consisting of m-cresol, benzyl alcohol, benzoic acid, phenol, methyl paraben, ethyl paraben, propyl paraben, butyl paraben, potassium sorbate, chlorobutanol, benzyl alcohol, phenylmercuric nitrate, thimerosal, sorbic acid, potassium sorbate, chlorocresol, benzalkonium chloride, 2-ethoxyethanol, chlorhexidine, chlorobutanol, phenylethyl alcohol, phenylmercuric acetate, and mixtures thereof.

[0143] In certain embodiments, the preservative is m-cresol. In certain embodiments, the preservative is benzyl alcohol. In certain embodiments, the preservative is benzoic acid. In certain embodiments, the preservative is phenol. In certain embodiments, the preservative is methyl paraben. In certain embodiments, the preservative is ethyl paraben. In certain embodiments, the preservative is propyl paraben. In certain embodiments, the preservative is butyl paraben. In certain embodiments, the preservative is potassium sorbate. In certain embodiments, the preservative is chlorocresol. In certain embodiments, the preservative is benzyl alcohol. In certain embodiments, the preservative is phenylmercuric nitrate. In certain embodiments, the preservative is thimerosal. In certain embodiments, the preservative is sorbic acid. In certain embodiments, the preservative is potassium sorbate. In certain embodiments, the preservative is chlorocresol. In certain embodiments, the preservative is benzalkonium chloride. In certain embodiments, the preservative is 2-ethoxyethanol. In certain embodiments, the preservative is chlorhexidine. In certain embodiments, the preservative is chlorobutanol. In certain embodiments, the preservative is phenylethyl alcohol. In certain embodiments, the preservative is phenylmercuric acetate.

[0144] In certain embodiments, the concentration of the preservative is in the range of 1 to 6 mg / mL. In certain embodiments, the concentration of the preservative is in the range of 3 to 6 mg / mL. In certain embodiments, the concentration of the preservative is in the range of 2 to 3 mg / mL. In certain embodiments, the concentration of the preservative is about 3 mg / mL. In certain embodiments, the concentration of the preservative is 3 mg / mL.

[0145] The liquid pharmaceutical preparation according to the present invention may further contain a pH adjuster. In certain embodiments, the pH adjuster is a base. Examples of bases can be selected from the group consisting of sodium hydroxide, tris (tris (hydroxymethyl) aminomethane), potassium hydroxide, lysine, and mixtures thereof.

[0146] In certain embodiments, the pH adjuster is a base selected from the group consisting of sodium hydroxide, sodium acetate, tris (tris (hydroxymethyl) aminomethane), potassium hydroxide, and lysine.

[0147] In certain embodiments, the pH adjuster is selected from the group consisting of sodium hydroxide and tris (tris (hydroxymethyl) aminomethane).

[0148] In certain embodiments, the pH adjuster is sodium hydroxide. In certain embodiments, the pH adjuster is sodium acetate. In certain embodiments, the pH adjuster is Tris. In certain embodiments, the pH adjuster is potassium hydroxide. In certain embodiments, the pH adjuster is lysine.

[0149] In certain embodiments, the pH adjuster is an acid. Examples of acids can be selected from the group consisting of hydrochloric acid, phosphoric acid, carbonic acid, nitric acid, and mixtures thereof.

[0150] In certain embodiments, the pH adjuster is an acid selected from the group consisting of hydrochloric acid, phosphoric acid, carbonic acid, and nitric acid.

[0151] In certain embodiments, the pH regulator is hydrochloric acid. In certain embodiments, the pH regulator is phosphoric acid. In certain embodiments, the pH regulator is carbonic acid. In certain embodiments, the pH regulator is nitric acid.

[0152] In certain embodiments, the pH regulator is a mixture of at least one base and at least one acid. In certain embodiments, the pH regulator is a mixture of one base and one acid. In certain embodiments, the pH regulator is a mixture of sodium hydroxide and hydrochloric acid.

[0153] In certain embodiments, the concentration of the pH regulator or mixture of pH regulators is in the range of 0.01 - 5 mg / mL. In certain embodiments, the concentration of the pH regulator or mixture of pH regulators is in the range of 0.04 - 2.5 mg / mL. In certain embodiments, the concentration of the pH regulator or mixture of pH regulators is in the range of 0.08 - 1.25 mg / mL. In certain embodiments, the concentration of the pH regulator or mixture of pH regulators is about 0.4 mg / mL. In the case of a mixture of pH regulators, it is understood that the concentration provided refers to the total concentration of all pH regulators.

[0154] The liquid pharmaceutical formulation according to the present invention optionally contains an antioxidant. Examples of antioxidants can be selected from the group consisting of methionine, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, propyl gallate, ascorbic acid, sodium bisulfite, ethylenediaminetetraacetic acid (EDTA), cysteine, glutathione, monothioglycerol, poly(ethyleneimine), vitamin E, ectoine, molybdine, and mixtures thereof.

[0155] In certain embodiments, the antioxidant is selected from the group consisting of methionine, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, propyl gallate, ascorbic acid, sodium bisulfite, ethylenediaminetetraacetic acid (EDTA), cysteine, glutathione, monothioglycerol, poly(ethyleneimine), vitamin E, ectoine, and molybdine.

[0156] Methionine has been observed to be particularly useful in preventing residues within the CNP compound, such as methionine residues, from being oxidized to methionine sulfoxide or methionine sulfone.

[0157] In certain embodiments, the antioxidant is methionine. In certain embodiments, the antioxidant is ascorbic acid. In certain embodiments, the antioxidant is butylated hydroxytoluene. In certain embodiments, the antioxidant is butylated hydroxyanisole. In certain embodiments, the antioxidant is tocopherol. In certain embodiments, the antioxidant is propyl gallate. In certain embodiments, the antioxidant is sodium bisulfite. In certain embodiments, the antioxidant is monothioglycerol. In certain embodiments, the antioxidant is EDTA. In certain embodiments, the antioxidant is cysteine. In certain embodiments, the antioxidant is glutathione. In certain embodiments, the antioxidant is poly(ethyleneimine). In certain embodiments, the antioxidant is vitamin E. In certain embodiments, the antioxidant is ectoine. In certain embodiments, the antioxidant is molybdenum.

[0158] As defined herein, the term "methionine" is intended to include both D-methionine and L-methionine, and mixtures thereof. In certain embodiments, the term "methionine" refers to L-methionine. In certain embodiments, the term "methionine" refers to D-methionine. In certain embodiments, the term "methionine" refers to a mixture of D-methionine or L-methionine.

[0159] As defined herein, the term "EDTA" encompasses all EDTA forms known in the art, such as EDTA salts, such as EDTA metal salts, such as EDTA disodium salt, EDTA dipotassium salt, calcium EDTA salt, magnesium EDTA disodium salt, or mixtures thereof. In certain embodiments, EDTA refers to the EDTA disodium salt. In certain embodiments, the term "EDTA" refers to the calcium EDTA salt. In certain embodiments, the term "EDTA" refers to EDTA anhydride.

[0160] In certain embodiments, the liquid pharmaceutical formulation does not contain sodium bisulfite. In certain embodiments, the liquid pharmaceutical formulation does not contain cysteine. In certain embodiments, the liquid pharmaceutical formulation does not contain glutathione. In certain embodiments, the liquid pharmaceutical formulation does not contain monothioglycerol. In certain embodiments, the liquid pharmaceutical formulation does not contain ascorbic acid.

[0161] In certain embodiments, the concentration of the antioxidant is in the range of about 0.1 mg / mL to about 1.5 mg / mL. In certain embodiments, the concentration of the antioxidant is in the range of about 0.5 mg / mL to about 1.0 mg / mL.

[0162] In certain embodiments, the molar ratio of the antioxidant to CNP or a CNP agonist is from about 0.1:1 to about 100:1. In certain embodiments, the molar ratio of the antioxidant to CNP or a CNP agonist is from about 0.1:1 to about 70:1. In certain embodiments, the molar ratio of the antioxidant to CNP or a CNP agonist is from about 0.1:1 to about 15:1. In certain embodiments, the molar ratio of the antioxidant to CNP or a CNP agonist is from about 1:1 to about 10:1. In certain embodiments, the molar ratio of the antioxidant to CNP or a CNP agonist is from about 3:1 to about 7:1.

[0163] In certain embodiments, the liquid pharmaceutical formulation of the present invention does not contain an antioxidant.

[0164] The liquid pharmaceutical preparation of the present invention may contain a detergent. Examples of detergents include octyl sucrose, Triton X-100, Triton X-114, polysorbate-20, polysorbate-80, NP-40, CA-630, Brij-35, Brij-58, n-dodecyl-β-maltoside, octyl-β-glucoside, octylthioglucoside, sodium dodecyl sulfate, CHAPS, Pluronic® F-127, and Pluronic® F-68, and can be selected from the group consisting of them.

[0165] In certain embodiments, the liquid pharmaceutical preparation of the present invention does not contain a detergent.

[0166] Preparations intended for administration to humans are preferably manufactured according to Good Manufacturing Practice (GMP) approved or approvable by the FDA or regulatory authorities in countries other than the United States, such as the European Medicines Agency, with respect to the preparation of pharmaceuticals for human administration. Typically, the preparation is sterile and is sterilized by sterile filtration using, for example, a 0.2 μm or 0.22 μm filter.

[0167] In certain embodiments, the liquid pharmaceutical preparation of the present invention contains a CNP agonist at a concentration of about 0.1 to about 20 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 0.5 to about 15 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 0.75 to about 10 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 1 to about 8 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 2 to about 6 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 10 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 8 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 6 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 4 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 2 mg / mL. In certain embodiments, the concentration of the CNP agonist is about 1 mg / mL.

[0168] In certain embodiments, the liquid pharmaceutical preparation of the present invention contains a CNP agonist at a concentration of 0.1 to 20 mg / mL. In certain embodiments, the concentration of the CNP agonist is 0.5 to 15 mg / mL. In certain embodiments, the concentration of the CNP agonist is 0.75 to 10 mg / mL. In certain embodiments, the concentration of the CNP agonist is 1 to 8 mg / mL. In certain embodiments, the concentration of the CNP agonist is 2 to 6 mg / mL. In certain embodiments, the concentration of the CNP agonist is 10 mg / mL. In certain embodiments, the concentration of the CNP agonist is 8 mg / mL. In certain embodiments, the concentration of the CNP agonist is 6 mg / mL. In certain embodiments, the concentration of the CNP agonist is 4 mg / mL. In certain embodiments, the concentration of the CNP agonist is 2 mg / mL. In certain embodiments, the concentration of the CNP agonist is 1 mg / mL.

[0169] In certain embodiments, the liquid pharmaceutical formulation of the present invention comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 0.1 to about 20 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 0.5 to about 15 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 0.75 to about 10 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 1 to about 8 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 2 to about 6 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 10 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 8 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 6 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 4 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 2 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of about 1 mg / mL. It is understood that the concentrations provided above refer to the amount of the CNP moiety and not to the entire CNP conjugate.

[0170] In certain embodiments, the liquid pharmaceutical formulation of the present invention comprises a CNP conjugate in which the CNP moiety is present at a concentration of 0.1 to 20 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 0.5 to 15 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 0.75 to 10 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 1 to 8 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 2 to 6 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 10 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 8 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 6 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 4 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 2 mg / mL. In certain embodiments, the liquid pharmaceutical formulation comprises a CNP conjugate in which the CNP moiety is present at a concentration of 1 mg / mL. It is understood that the concentrations provided above refer to the amount of the CNP moiety and not to the entire CNP conjugate.

[0171] In certain embodiments, the reversible CNP conjugate is of the following formula (Ia) or (Ib).

Chemical formula

[0172] -D in formula (Ia) or (Ib) is -L 1 - is covalently and reversibly bonded thereto.

[0173] In certain embodiments, the liquid pharmaceutical formulation of the present invention contains a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, D-mannitol: 10 to 200 mg / mL, m-cresol: 1 to 10 mg / mL and.

[0174] In certain embodiments, the liquid pharmaceutical formulation of the present invention contains a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, trehalose dihydrate: 10 to 200 mg / mL, m-cresol: 1 to 10 mg / mL and.

[0175] In certain embodiments, -Z in formula (Ia) or (Ib) is a polymer moiety. In certain embodiments, -Z in formula (Ia) or (Ib) is a water-soluble polymer moiety.

[0176] In certain embodiments, x in formula (Ia) is an integer selected from the group consisting of 1, 2, 3, 4, 6, and 8. In certain embodiments, x in formula (Ia) is an integer selected from the group consisting of 1, 2, 4, and 6. In certain embodiments, x in formula (Ia) is an integer selected from the group consisting of 1, 4, and 6, and in certain embodiments, x in formula (Ia) is 1.

[0177] In certain embodiments, y of formula (Ib) is an integer selected from the group consisting of 3, 4, and 5. In certain embodiments, y of formula (Ib) is an integer selected from the group consisting of 4 and 5. In certain embodiments, y of formula (Ib) is an integer selected from the group consisting of 2 and 3. In certain embodiments, y of formula (Ib) is 2. In certain embodiments, y of formula (Ib) is 3. In certain embodiments, y of formula (Ib) is 4. In certain embodiments, y of formula (Ib) is 5.

[0178] In certain embodiments, the reversible CNP conjugate is of formula (Ia) where x = 1.

[0179] In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, or SEQ ID NO: 25.

[0180] In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 20. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 21. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 22. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 23. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 24. In certain embodiments, -D of formula (Ia) or (Ib) has the sequence of SEQ ID NO: 25.

[0181] Sub - L of formula (Ia) or (Ib) 1- is attached to the functional group of the side chain of the amino acid residue of -D, attached to the N-terminal amine functional group or the C-terminal carboxyl functional group of -D, or attached to the nitrogen atom in the backbone polypeptide chain of -D. The attachment to either the N-terminus or the C-terminus can be direct through the corresponding amine or carboxyl functional group, or can be indirect through a spacer moiety that is first attached to the amine or carboxyl functional group to which -L 1 - is attached.

[0182] The moiety -L of formula (Ia) or (Ib) 1 - is a pharmaceutical, i.e., a reversible prodrug linker from which D-H is released in its free or unmodified form, i.e., -L 1 - is a linker that leaves no trace. Suitable reversible linkers, such as those disclosed in WO2005 / 099768A2, WO2006 / 136586A2, WO2011 / 089216A1, and WO2013 / 024053A1, which are incorporated herein by reference, are known in the art.

[0183] In certain embodiments, -L 1 - is a reversible linker as described in WO2011 / 012722A1, WO2011 / 089214A1, WO2011 / 089215A1, WO2013 / 024052A1, and WO2013 / 160340A1, which are incorporated herein by reference.

[0184] The moiety -L 1 - can be attached to -D through any type of linker, provided that it is reversible. In certain embodiments, -L 1 - is attached to -D through a linker selected from the group consisting of amide, ester, carbamate, acetal, aminals, imine, oxime, hydrazone, disulfide, and acylguanidine. In certain embodiments, -L 1- is linked to -D via a linking moiety selected from the group consisting of amide, ester, carbamate and acylguanidine. These linking moieties may not be reversible by themselves, -L 1 It is understood that the adjacent groups contained in -L may be such that they render the linking moiety reversible.

[0185] In certain embodiments, moiety -L 1 - is linked to -D via an amide linking moiety.

[0186] Moiety -L 1 - is disclosed in WO2009 / 095479A2. Thus, in certain embodiments, moiety -L 1 - has the formula (II):

Chemical formula

[0187] In certain embodiments, -L of formula (II) 1 - is a single moiety -L 2 - is substituted with -Z.

[0188] In certain embodiments, -L of formula (II) 1 - is further unsubstituted.

[0189] -R of formula (II) 3 / -R 3a When they combine with the nitrogen atom to which they are attached to form a 3- to 10-membered heterocyclic ring, it is understood that only such 3- to 10-membered heterocyclic rings in which the atom directly attached to the nitrogen is an sp 3 hybridized carbon atom can be formed. In other words, -R 3 / -R 3a such 3- to 10-membered heterocyclic rings formed by -R

Chemical formula

[0190] It is also understood that the 3- to 10-membered heterocyclic ring may be further substituted.

[0191] -R of formula (II) 3 / -R 3a Exemplary embodiments of suitable 3- to 10-membered heterocyclic rings formed by -R of formula (II) and the nitrogen atom to which they are attached are as follows:

Chemical formula

[0192] Optionally, -L in formula (II) 1 - may be further substituted. Generally, any substituent may be used as long as it does not affect the cleavage principle, that is, the hydrogen marked with an asterisk in formula (II) is not replaced, and the part of formula (II)

Chemical formula

[0193] In certain embodiments, -R in formula (II) 1 or -R 1a is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 2 or -R 2a is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 3 or -R 3a is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 4 is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 5 or -R 5a is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 6 is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 7 or -R 7a is substituted with -L 2 -Z. In certain embodiments, -R in formula (II) 8or -R 8a is -L 2 is replaced by -Z. In certain embodiments, -R of formula (II) 9 or -R 9a is -L 2 is replaced by -Z.

[0194] In certain embodiments, -R of formula (II) 4 is -L 2 is replaced by -Z.

[0195] In certain embodiments, -X- of formula (II) is -C(R 4 R 4a )- or -N(R 4 )-. In certain embodiments, -X- of formula (II) is -C(R 4 R 4a )-.

[0196] In certain embodiments, X of formula (II) 1 is C.

[0197] In certain embodiments, =X of formula (II) 3 is =O.

[0198] In certain embodiments, -X of formula (II) 2 - is -C(R 8 R 8a )-.

[0199] In certain embodiments, -R of formula (II) 8 and -R 8a are independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, at least one of -R 8 and -R 8a of formula (II) is -H. In certain embodiments, both -R 8 and -R 8a of formula (II) are -H.

[0200] In certain embodiments, -R of formula (II) 1 and -R1a is independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, -R of formula (II) 1 and -R 1a at least one of which is -H. In certain embodiments, -R of formula (II) 1 and -R 1a both are -H.

[0201] In certain embodiments, -R of formula (II) 2 and -R 2a is independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, -R of formula (II) 2 and -R 2a at least one of which is -H. In certain embodiments, -R of formula (II) 2 and -R 2a both are H.

[0202] In certain embodiments, -R of formula (II) 3 and -R 3a is independently selected from the group consisting of -H, methyl, ethyl, propyl and butyl. In certain embodiments, -R of formula (II) 3 and -R 3a at least one of which is methyl. In certain embodiments, -R of formula (II) 3 and -R 3a both are -H. In certain embodiments, -R of formula (II) 3 and -R 3a both are methyl. In certain embodiments, -R of formula (II) 3 is -H and -R of formula (II) 3a is methyl.

[0203] In certain embodiments, -R of formula (II) 4 and -R 4a is independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, -R of formula (II) 4 and -R 4aAt least one of them is -H. In certain embodiments, -R of formula (II) 4 and -R 4a are both -H.

[0204] In certain embodiments, the moiety -L 1 - is of formula (IIa):

Chemical formula

[0205] In certain embodiments, -L of formula (IIa) 1 - is replaced by one moiety -L 2 -Z.

[0206] In certain embodiments, the moiety -L of formula (IIa) 1 - is not further substituted.

[0207] In certain embodiments, -R of formula (IIa) 1 and -R 1a are independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, -R of formula (IIa) 1 and -R1a At least one of them is -H. In certain embodiments, -R of formula (IIa) 1 and -R 1a are both -H. In certain embodiments, -R of formula (IIa) 4 and -R 4a are independently selected from the group consisting of -H, methyl, and ethyl. In certain embodiments, -R of formula (IIa) 4 and -R 4a At least one of them is -H. In certain embodiments, -R of formula (IIa) 4 and -R 4a are both -H.

[0208] In certain embodiments, -X 2 - of formula (IIa) is -C(R 8 R 8a ).

[0209] In certain embodiments, -R 8 and -R 8a of formula (IIa) are independently selected from the group consisting of -H, methyl, and ethyl. In certain embodiments,, -R 8 and -R 8a of formula (IIa) At least one of them is -H. In certain embodiments, -R 8 and -R 8a of formula (IIa) are both -H.

[0210] In certain embodiments, -R 2 and -R 2a of formula (IIa) are independently selected from the group consisting of -H, methyl, and ethyl. In certain embodiments, -R 2 and -R 2a of formula (IIa) At least one of them is -H. In certain embodiments, -R 2 and -R 2a of formula (IIa) are both H.

[0211] In certain embodiments, -R 3 and -R 3ais independently selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, -R of formula (IIa) 3 and -R 3a of at least one is methyl. In certain embodiments, -R of formula (IIa) 3 and -R 3a are both -H. In certain embodiments, -R of formula (IIa) 3 and -R 3a are both methyl. In certain embodiments, -R of formula (IIa) 3 is -H and -R of formula (IIa) 3a is methyl.

[0212] In certain embodiments, the moiety -L 1 - is of formula (IIb):

Chemical formula

[0213] In certain embodiments, -L of formula (IIb) 1 - is substituted with one moiety -L 2 -Z. In certain embodiments, the moiety -L of formula (IIb) 1 - is not further substituted.

[0214] In certain embodiments, -X of formula (IIb) 2 - is -C(R 8 R 8a ).

[0215] In certain embodiments, -R and -R of formula (IIb) 8 and -R 8a are independently selected from the group consisting of -H, methyl, and ethyl. In certain embodiments, at least one of -R and -R of formula (IIb) 8 and -R 8a is -H. In certain embodiments, both -R and -R of formula (IIb) 8 and -R 8a are -H.

[0216] In certain embodiments, -R and -R of formula (IIb) 2 and -R 2a are independently selected from the group consisting of -H, methyl, and ethyl. In certain embodiments, at least one of -R and -R of formula (IIb) 2 and -R 2a is -H. In certain embodiments, both -R and -R of formula (IIb) 2 and -R 2a are H.

[0217] In certain embodiments, -R and -R of formula (IIb) 3 and -R 3a are independently selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, at least one of -R and -R of formula (IIb) 3 and -R 3a is methyl. In certain embodiments, both -R and -R of formula (IIb) 3 and -R 3a are both -H. In certain embodiments, both -R and -R of formula (IIb) 3 and -R 3a are both methyl. In certain embodiments, -R of formula (IIb) 3 is -H and -R of formula (IIb) 3a is methyl.

[0218] In certain embodiments, sub-L 1 - is of formula (IIb’):

Chemical formula

[0219] In certain embodiments, sub-L of formula (IIb’) 1 - is not further substituted.

[0220] In certain embodiments, -X of formula (IIb’) 2 - is -C(R 8 R 8a )-.

[0221] In certain embodiments, -R of formula (IIb’) 8 and -R 8a are independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, at least one of -R of formula (IIb’) 8 and -R 8a is -H. In certain embodiments, both -R of formula (IIb’) 8 and -R 8a are -H.

[0222] In certain embodiments, -R of formula (IIb’) 2 and -R2a is independently selected from the group consisting of -H, methyl and ethyl. In certain embodiments, -R of formula (IIb’) 2 and -R 2a at least one of which is -H. In certain embodiments, -R of formula (IIb’) 2 and -R 2a both are H.

[0223] In certain embodiments, -R of formula (IIb’) 3 and -R 3a are independently selected from the group consisting of -H, methyl, ethyl, propyl and butyl. In certain embodiments, -R of formula (IIb’) 3 and -R 3a at least one of which is methyl. In certain embodiments, -R of formula (IIb’) 3 and -R 3a both are -H. In certain embodiments, -R of formula (IIb’) 3 and -R 3a both are methyl. In certain embodiments, -R of formula (IIb’) 3 is -H and -R of formula (IIb’) 3a is methyl.

[0224] In certain embodiments, moiety -L 1 - has the formula (IIc):

Chemical formula

[0225] In certain embodiments, -L of formula (IIc) 1 - is a single moiety -L 2 -Z.

[0226] In certain embodiments, the moiety -L of formula (IIc) 1 - is further unsubstituted.

[0227] In certain embodiments, the moiety -L 1 - is of formula (IIc-a):

Chemical formula

[0228] In certain embodiments, -L of formula (IIc-a) 1 - is a single moiety -L 2 -Z.

[0229] In certain embodiments, the moiety -L of formula (IIc-a) 1 - is further unsubstituted.

[0230] In certain embodiments, the moiety -L 1 - is of formula (IIc-b):

Chemical formula

[0231] In a particular embodiment, -L of formula (IIc-b) 1 - is one part -L 2 -Z is replaced.

[0232] In a particular embodiment, the part -L of formula (IIc-b) 1 - is not further replaced.

[0233] In a particular embodiment, the part -L 1 - is of formula (IIc-i), (IIc-ii), (IIc-iii), (IIc-iv) and (IIc-v):

Chemical formula

[0234] In a particular embodiment, the part -L of formula (IIc-i), (IIc-ii), (IIc-iii), (IIc-iv) and (IIc-v) 1 - is not further replaced.

[0235] In a particular embodiment, the part -L 1 - is of formula (IIc-ii): [Chemical formula] (In the formula, The dashed line without a mark indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond. The dashed line with an asterisk indicates the bond to -L 2 -Z) is.

[0236] In certain embodiments, -L in formula (IIc-ii) 1 - is replaced by one moiety -L 2 -Z.

[0237] In certain embodiments, the moiety -L 1 - is of formula (IIc-i’), (IIc-ii’), (IIc-iii’), (IIc-iv’) and (IIc-v’): [Chemical formula] TIFF2025522281000025.tif29160 (In the formula, The dashed line without a mark indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond. The dashed line with an asterisk indicates the bond to -L 2 -Z) is selected from the group consisting of, optionally, said -L 1 - is further substituted, provided that the hydrogen with an asterisk in formula (IIc-i’), (IIc-ii’), (IIc-iii’), (IIc-iv’) and (IIc-v’) is not replaced by a substituent.

[0238] In certain embodiments, the moiety -L in formula (IIc-i’), (IIc-ii’), (IIc-iii’), (IIc-iv’) and (IIc-v’) 1 - is not further substituted.

[0239] In certain embodiments, the moiety -L1 - is the formula (IIc-ii’):

Chem.

[0240] In certain embodiments, -L in formula (IIc-ii’) 1 - is replaced by one moiety -L 2 -Z.

[0241] In certain embodiments, the moiety -L 1 - is of formula (IIc-i’’), (IIc-ii’’), (IIc-iii’’) and (IIc-iv’’):

Chem.

[0242] In certain embodiments, the moiety -L in formula (IIc-i’’), (IIc-ii’’), (IIc-iii’’) and (IIc-iv’’) 1 - is not further substituted.

[0243] In certain embodiments, Sub-L 1 - is of formula (IIc-ii''):

Chemical formula

[0244] In certain embodiments, -L of formula (IIc-ii'') 1 - is replaced by one Sub-L 2 -Z.

[0245] For -L in formulas (II), (IIa), (IIb), (IIb'), (IIc), (IIc-a), (IIc-b), (IIc-i), (IIc-ii), (IIc-iii), (IIc-iv), (IIc-v), (IIc-i'), (IIc-ii'), (IIc-iii'), (IIc-iv'), (IIc-v'), (IIc-i''), (IIc-ii''), (IIc-iii'') and (IIc-iv''), 1 any optional further substituents of - are as described above in certain embodiments.

[0246] Another Sub-L 1 - is disclosed in WO 2016 / 020373 A1. Thus, in certain embodiments, Sub-L 1 - is of formula (III):

Chemical formula

[0247] The optional further substituents of -L 1 - of formula (III) are, in certain embodiments, as described above.

[0248] In certain embodiments, -L 1 - of formula (III) is substituted with one moiety -L 2 -Z.

[0249] In certain embodiments, -L of formula (III) 1 - is further unsubstituted.

[0250] -L 1 Further embodiments for -L are disclosed in European Patent No. 1536334 B1, WO2009 / 009712 A1, WO2008 / 034122 A1, WO2009 / 143412 A2, WO2011 / 082368 A2, and U.S. Patent No. 8,618,124 B2, and the said patent documents are hereby incorporated herein by reference in their entirety.

[0251] -L 1 Further embodiments for -L are disclosed in U.S. Patent No. 8,946,405 B2 and U.S. Patent No. 8,754,190 B2, and the said patent documents are hereby incorporated herein by reference in their entirety. Thus, the moiety -L 1 - is of formula (IV):

Chemical formula

[0252] -L of formula (IV) 1 Any optional further substituents of - are as described above in certain embodiments

[0253] In certain embodiments, -L of formula (IV) 1 - is one moiety -L 2 - is replaced by -Z. In certain embodiments, -L of formula (IV) 1 - is not further substituted

[0254] The terms used solely in connection with formula (IV) have the following meanings:

[0255] As used herein, the term "alkyl" includes straight-chain, branched or cyclic saturated hydrocarbon groups having from 1 to 8 carbon atoms, or in some embodiments from 1 to 6 or 1 to 4 carbon atoms

[0256] The term "alkoxy" includes alkyl groups bonded to oxygen, including methoxy, ethoxy, isopropoxy, cyclopropoxy, cyclobutoxy and the like

[0257] The term "alkenyl" includes non-aromatic unsaturated hydrocarbons having a carbon-carbon double bond

[0258] The term "alkynyl" includes non-aromatic unsaturated hydrocarbons having a carbon-carbon triple bond

[0259] The term "aryl" includes aromatic hydrocarbon groups having 6 to 18 carbon atoms, such as phenyl, naphthyl, and anthracenyl, and in certain embodiments, 6 to 10 carbon atoms. The term "heteroaryl" includes groups such as pyrrolyl, pyridyl, pyrimidinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, quinolyl, indolyl, indenyl, and the like, containing 3 to 15 carbon atoms and at least one N, O, or S atom, and in certain embodiments, containing 3 to 7 carbon atoms and at least one N, O, or S atom, and including an aromatic ring.

[0260] In certain embodiments, the alkenyl, alkynyl, aryl, or heteroaryl moiety may be coupled to the remainder of the molecule by an alkylene bond. Substituents in these situations are referred to as alkenylalkyl, alkynylalkyl, arylalkyl, or heteroarylalkyl, which indicates that the alkylene moiety is between the alkenyl, alkynyl, aryl, or heteroaryl moiety and the molecule to which the alkenyl, alkynyl, aryl, or heteroaryl is coupled.

[0261] The term "halogen" includes bromo, fluoro, chloro, and iodo.

[0262] The term "heterocyclic ring" refers to a 4- to 8-membered aromatic or non-aromatic ring containing 3 to 7 carbon atoms and at least one N, O, or S atom. Examples are piperidinyl, piperazinyl, tetrahydropyranyl, pyrrolidine, and tetrahydrofuranyl, as well as the exemplary groups provided above for the term "heteroaryl".

[0263] When the ring system is optionally substituted, suitable substituents are selected from the group consisting of alkyl, alkenyl, alkynyl or a further ring, each of which is optionally further substituted. Optional substituents for any group including the above are halo, nitro, cyano, -OR, -SR, -NR2, -OCOR, -NRCOR, -COOR, -CONR2, -SOR, -SO2R, -SONR2, -SO2NR2, where each R is independently alkyl, alkenyl, alkynyl, aryl or heteroaryl, or two R groups together with the atom to which they are attached form a ring.

[0264] -L 1 Further embodiments for - are disclosed in WO2013 / 036857A1, which patent document is hereby incorporated by reference in its entirety. Thus, in certain embodiments, the moiety -L 1 - is of formula (V):

Chemical formula

[0265] The optional further substituents of -L in formula (V) 1 are, in certain embodiments, as described above.

[0266] In certain embodiments, -L in formula (V) 1 - is substituted with one moiety -L 2 -Z. In certain embodiments, -L in formula (V) 1 - is not further substituted.

[0267] Terms used solely in connection with formula (V) have the following meanings:

[0268] "Alkyl", "alkenyl" and "alkynyl" include straight-chain, branched or cyclic hydrocarbon groups of 1 to 8, or 1 to 6, or 1 to 4 carbon atoms, where alkyl is a saturated hydrocarbon, alkenyl contains one or more carbon-carbon double bonds, and alkynyl contains one or more carbon-carbon triple bonds. Unless otherwise specified, these contain 1 to 6 C.

[0269] "Aryl" includes aromatic hydrocarbon groups having 6 to 18 carbons, such as phenyl, naphthyl, and anthracene, and in certain embodiments, 6 to 10 carbons. "Heteroaryl" includes groups such as pyrrolyl, pyridyl, pyrimidinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, quinolyl, indolyl, indenyl, and the like, containing 3 to 15 carbons and at least one N, O, or S atom, and in certain embodiments, containing 3 to 7 carbons and at least one N, O, or S atom and including an aromatic ring.

[0270] The term "substituted" means an alkyl, alkenyl, alkynyl, aryl, or heteroaryl group containing one or more substituents in place of one or more hydrogen atoms. Substituents generally include halogen (including F, Cl, Br, and I), lower alkyl (including straight-chain, branched, and cyclic), lower haloalkyl (including fluoroalkyl, chloroalkyl, bromoalkyl, and iodoalkyl), OH, lower alkoxy (including straight-chain, branched, and cyclic), SH, lower alkylthio (including straight-chain, branched, and cyclic), amino, alkylamino, dialkylamino, silyl (including alkylsilyl, alkoxysilyl, and arylsilyl), nitro, cyano, carbonyl, carboxylic acid, carboxylic acid ester, carboxylic acid amide, aminocarbonyl, aminoacyl, carbamate, urea, thiocarbamate, thiourea, ketone, sulfone, sulfonamide, aryl (including phenyl, naphthyl, and anthracenyl), heteroaryl (including 5-membered heteroaryl such as pyrrole, imidazole, furan, thiophene, oxazole, thiazole, isoxazole, isothiazole, thiadiazole, triazole, oxadiazole, and tetrazole, 6-membered heteroaryl such as pyridine, pyrimidine, and pyrazine, and fused heteroaryl such as benzofuran, benzothiophene, benzoxazole, benzimidazole, indole, benzothiazole, benzisoxazole, and benzisothiazole), and may be selected from these.

[0271] -L 1 - Further embodiments of -L are disclosed in WO2022 / 115563A1, which patent document is incorporated herein by reference in its entirety. Thus, in certain embodiments, sub-L 1 - is of formula (Va):

Chemical formula

[0272] In certain embodiments, -L1- is of formula (Va), the dashed line marked with an asterisk indicates the bond to -L2-Z, and the unmarked dashed line indicates the bond to -D, where -D is the CNP moiety of the following amino acid sequence: SEQ ID NO: 106: LQEHPQARKYKGAQKKGLSKGCFGLKLDRIGSMSGLGC, (In the formula, the cysteines at positions 22 and 38 are linked via a disulfide bridge, and the bond to -L1- occurs at either the N-terminus of the peptide or the ring.)

[0273] -L 1 - Further embodiments of -L are disclosed in U.S. Patent No. 7585837B2, which patent document is incorporated herein by reference in its entirety. Thus, in certain embodiments, sub-L 1 - is of formula (VI):

Chemical formula

[0274] In certain embodiments, -L 1 - of formula (VI) is substituted with one moiety -L 2 -Z.

[0275] Any optional further substituents of -L 1 - of formula (VI) are, in certain embodiments, as described above.

[0276] In certain embodiments, -L 1 - of formula (VI) is not further substituted.

[0277] The terms used solely in connection with formula (VI) have the following meanings:

[0278] The terms "alkyl", "alkoxy", "alkoxyalkyl", "aryl", "alkaryl" and "aralkyl" mean an alkyl radical having 1 to 8 carbon atoms, in certain embodiments 1 to 4 carbon atoms, such as methyl, ethyl, propyl, isopropyl and butyl, and an aryl radical having 6 to 10 carbon atoms, such as phenyl and naphthyl. The term "halogen" includes bromo, fluoro, chloro and iodo.

[0279] -L 1 Further embodiments regarding - are disclosed in WO2002 / 089789A1, which patent document is hereby incorporated by reference in its entirety. Thus, the moiety -L 1 - is of formula (VII):

Chemical formula

[0280] In certain embodiments, -L of formula (VII) 1 - is substituted with one moiety -L 2 -Z.

[0281] -L of formula (VII) 1 Any optional further substituents of - are, in certain embodiments, as described above.

[0282] In certain embodiments, -L of formula (VII) 1 - is not further substituted.

[0283] Terms used solely in connection with formula (VII) have the following meanings:

[0284] The term "alkyl" is understood to include, for example, linear, branched, substituted C 3-8 alkyl such as alkoxy, C 1-12 cycloalkyl or substituted cycloalkyl.

[0285] The term "substituted" is understood to include adding one or more atoms contained in a functional group or a compound and replacing one or more different atoms.

[0286] The substituted alkyl includes carboxyalkyl, aminoalkyl, dialkylamino, hydroxyalkyl and mercaptoalkyl, the substituted cycloalkyl includes moieties such as 4-chlorocyclohexyl, the aryl includes moieties such as naphthyl, the substituted aryl includes moieties such as 3-bromo-phenyl, the aralkyl includes moieties such as toluyl, the heteroalkyl includes moieties such as ethylthiophene, the substituted heteroalkyl includes moieties such as 3-methoxythiophene, the alkoxy includes moieties such as methoxy, and the phenoxy includes moieties such as 3-nitrophenoxy. Halo- is understood to include fluoro, chloro, iodo and bromo.

[0287] In certain embodiments, -L 1 - is a substructure of formula (VIII) [Chemical formula] (wherein The dashed line with an asterisk indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond, The unmarked dashed line indicates the bond to the remainder of -L 1 -) and said -L 1 - is substituted with -L 2 -Z, and optionally, said -L 1 - is further substituted, where -L 2 - is a single chemical bond or a spacer, -Z is a carrier moiety, preferably a water-soluble polymer moiety.

[0288] In certain embodiments, -L of formula (VIII) 1 - is substituted with one moiety -L 2 -Z.

[0289] Any optional further substituents of -L of formula (VIII) 1 - are as described above.

[0290] In certain embodiments, -L of formula (VIII) 1 - is further unsubstituted.

[0291] In certain embodiments, -L 1 - is a substructure of formula (IX)

Chemical formula

[0292] Any optional further substituents of -L of formula (IX) 1 - are as described above.

[0293] In certain embodiments, -L of formula (IX) 1 - is substituted with one moiety -L 2 -Z.

[0294] In certain embodiments, -L of formula (IX) 1 - is further unsubstituted.

[0295] Moiety -D may be linked to -L 1 - via any functional group of D-H, and via the amine functional group of D-H to -L 1-is linked to this. This may be an amine functional group provided by the N-terminal amine functional group or the lysine side chain, i.e., when CNP has the sequence of SEQ ID NO: 24, the amine functional groups provided by lysine at positions 9, 11, 15, 16, 20, and 26.

[0296] The binding of -L to the ring of the CNP moiety 1 -significantly reduces the affinity of the CNP conjugate for NPR-B compared to the binding at the N-terminus or to the acyclic portion of the CNP. This reduced affinity for NPR-B then reduces the risk of cardiovascular side effects such as hypotension.

[0297] Therefore, -L 1 -is, in certain embodiments, attached to the side chain of the amino acid residue of the ring portion of -D or to the backbone of the ring portion of -D. In certain embodiments, -L 1 -is covalently and reversibly attached to the side chain of the amino acid residue of the ring portion of -D. When -D is a CNP moiety having the sequence of SEQ ID NO: 24, -L 1 -is, in certain embodiments, attached to the amine functional group provided by lysine at position 26 of the corresponding drug D-H.

[0298] The moiety -L 2 -is a chemical bond or a spacer moiety. In certain embodiments, -L 2 -is a chemical bond. In certain embodiments, -L 2 -is a spacer moiety.

[0299] The moiety -L 2 -can be attached to -L by replacing any -H present, unless specifically excluded. 1 -that is present.

[0300] -L 2 -is other than a single chemical bond, -L 2 -is -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y1 )-, -S(O)2N(R y1 )-, -S(O)N(Ry1 )-, -S(O)2-, -S(O)-, -N(R y1 )S(O)2N(R y1a )-, -S-, -N(R y1 )-, -OC(OR y1 )(R y1a )-, -N(R y1 )C(O)N(R y1a )-, -OC(O)N(R y1 )-, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl selected from; -T-, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl may be substituted with one or more identical or different -R y2 and C 1-50 alkyl, C 2-50 alkenyl and C 2-50 alkynyl is -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y3 )-, -S(O)2N(R y3 )-, -S(O)N(R y3 )-, -S(O)2-, -S(O)-, -N(R y3 )S(O)2N(R y3a )-, -S-, -N(R y3 )-, -OC(OR y3 )(R y3a )-, -N(R y3 )C(O)N(R y3a )-, and -OC(O)N(R y3 )- and may optionally be interrupted by one or more groups selected from the group consisting of; -R y1 and -R y1a are each independently, -H, -T, C 1-50 alkyl, C 2-50 alkenyl and C 2-50 alkynyl selected from the group consisting of; -T-, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl may be substituted with one or more identical or different -R y2 and C1-50 Alkyl, C 2-50 Alkenyl, and C 2-50 Alkynyl may optionally be interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y4 ), -S(O)2N(R y4 ), -S(O)N(R y4 ), -S(O)2-, -S(O)-, -N(R y4 )S(O)2N(R y4a )-, -S-, -N(R y4 )-, -OC(OR y4 )(R y4a )-, -N(R y4 )C(O)N(R y4a ), -OC(O)N(R y4 ); Each T is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbocyclic polycycle, and 8- to 30-membered heterocyclic polycycle; each T may independently be substituted with one or more identical or different -R y2 ; Each -R y2 is independently halogen, -CN, oxo (=O), -COOR y5 , -OR y5 , -C(O)R y5 , -C(O)N(R y5 R y5a ), -S(O)2N(R y5 R y5a ), -S(O)N(R y5 R y5a ), -S(O)2R y5 , -S(O)R y5 , -N(R y5 )S(O)2N(R y5a R y5b ), -SR y5 , -N(R y5 R y5a ), -NO2, -OC(O)R y5 , -N(R y5 )C(O)R y5a , -N(Ry5 )S(O)2R y5a 、 -N(R y5 )S(O)R y5a 、 -N(R y5 )C(O)OR y5a 、 -N(R y5 )C(O)N(R y5a R y5b )、 -OC(O)N(R y5 R y5a )、 and C 1-6 selected from the group consisting of alkyl; C 1-6 alkyl may be substituted with one or more identical or different halogens; each -R y3 、 -R y3a 、 -R y4 、 -R y4a 、 -R y5 、 -R y5a and -R y5b is independently selected from the group consisting of -H and C 1-6 alkyl; C 1-6 alkyl may be substituted with one or more identical or different halogens.

[0301] -L 2 - when other than a single chemical bond, -L 2 - is -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y1 )-, -S(O)2N(R y1 )-, -S(O)N(R y1 )-, -S(O)2-, -S(O)-, -N(R y1 )S(O)2N(R y1a )-, -S-, -N(R y1 )-, -OC(OR y1 )(R y1a )-, -N(R y1 )C(O)N(R y1a )-, -OC(O)N(R y1 )-, C 1-20 alkyl, C 2-20 alkenyl, and C 2-20 alkynyl; -T-, C 1-20 alkyl, C 2-20 alkenyl, and C 2-20The alkynyl may be the same or different and is one or more -R y2 and may be substituted with, C 1-20 alkyl, C 2-20 alkenyl and C 2-20 The alkynyl may optionally be interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y3 ), -S(O)2N(R y3 ), -S(O)N(R y3 ), -S(O)2-, -S(O)-, -N(R y3 )S(O)2N(R y3a ), -S-, -N(R y3 ), -OC(OR y3 )(R y3a ), -N(R y3 )C(O)N(R y3a ), and -OC(O)N(R y3 ); -R y1 and -R y1a are each independently selected from the group consisting of -H, -T, C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl; -T, C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 The alkynyl may be the same or different and is one or more -R y2 and may be substituted with, C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 The alkynyl may optionally be interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y4 ), -S(O)2N(R y4 ), -S(O)N(R y4 ), -S(O)2-, -S(O)-, -N(R y4 )S(O)2N(R y4a ), -S-, -N(R y4 ), -OC(OR y4 )(R y4a ), -N(R y4 )C(O)N(R y4a ), and -OC(O)N(Ry4 )-selected from the group consisting of may optionally be interrupted; Each T is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbocyclic polycycle, and 8- to 30-membered heterocyclic polycycle; each T is independently substituted with one or more identical or different -R y2 and may be; -R y2 is halogen, -CN, oxo (=O), -COOR y5 , -OR y5 , -C(O)R y5 , -C(O)N(R y5 R y5a ), -S(O)2N(R y5 R y5a ), -S(O)N(R y5 R y5a ), -S(O)2R y5 , -S(O)R y5 , -N(R y5 ), -S(O)2N(R y5a R y5b ), -SR y5 , -N(R y5 R y5a ), -NO2, -OC(O)R y5 , -N(R y5 ), -C(O)R y5a , -N(R y5 ), -S(O)2R y5a , -N(R y5 ), -S(O)R y5a , -N(R y5 ), -C(O)OR y5a , -N(R y5 ), -C(O)N(R y5a R y5b ), -OC(O)N(R y5 R y5a ), and C 1-6 alkyl; C 1-6 alkyl may optionally be substituted with one or more identical or different halogens; each -R y3 , -R y3a , -Ry4 , -R y4a , -R y5 , -R y5a and -R y5b are each independently selected from the group consisting of -H and C 1-6 alkyl; C 1-6 alkyl may be substituted with one or more identical or different halogen atoms.

[0302] -L 2 when - is other than a single chemical bond, -L 2 - is selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y1 ), -S(O)2N(R y1 ), -S(O)N(R y1 ), -S(O)2-, -S(O)-, -N(R y1 )S(O)2N(R y1a ), -S-, -N(R y1 ), -OC(OR y1 )(R y1a ), -N(R y1 )C(O)N(R y1a ), -OC(O)N(R y1 ), C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; -T-, C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl may be substituted with one or more identical or different -R y2 ; C 1-50 alkyl, C 2-50 alkenyl and C 2-50 alkynyl are selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R y3 ), -S(O)2N(R y3 ), -S(O)N(R y3 ), -S(O)2-, -S(O)-, -N(R y3 )S(O)2N(R y3a ), -S-, -N(R y3 ), -OC(OR y3 )(R y3a ), -N(Ry3 )(C(O)N(R y3a ))-, and -OC(O)N(R y3 )- from the group consisting of may optionally be interrupted; -R y1 and -R y1a are independently, -H, -T, C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl selected from the group consisting of; each T is independently phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbocyclic polycycle, and 8- to 30-membered heterocyclic polycycle selected from the group consisting of; each -R y2 is selected from the group consisting of halogen and C 1-6 alkyl; each -R y3 , -R y3a , -R y4 , -R y4a , -R y5 , -R y5a and -R y5b are independently selected from the group consisting of -H and C 1-6 alkyl; C 1-6 alkyl may be substituted with one or more identical or different halogens.

[0303] In certain embodiments, -L 2 - is a C 1-20 alkyl chain which may optionally be interrupted by one or more groups independently selected from -O-, -T- and -C(O)N(R y1 )); C 1-20 alkyl chain may be independently substituted with one or more groups selected from -OH, -T and -C(O)N(R y6 R y6a )); -R y1 , -R y6 , -R y6a are independently, H and C 1-4Selected from the group consisting of alkyl, and T is phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 Selected from the group consisting of cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbopolycyclic, and 8- to 30-membered heteropolycyclic.

[0304] In certain embodiments, -L 2 - has a molecular weight in the range of 14 g / mol to 750 g / mol (including both ends).

[0305] In certain embodiments, -L 2 - has a chain length of 1 to 20 atoms.

[0306] As used herein, the moiety -L 2 The term "chain length" with respect to -L 1 refers to the number of atoms of -L 2 present in the shortest linkage between -L and -Z.

[0307] In certain embodiments, -L 2 - is of the following formula (i).

Chemical formula

[0308] In certain embodiments, -R of formula (i) 1is selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, -R of formula (i) 1 is selected from the group consisting of -H, methyl, ethyl, and propyl. In certain embodiments, -R of formula (i) 1 is selected from the group consisting of -H and methyl. In certain embodiments, -R of formula (i) 1 is methyl.

[0309] In certain embodiments, n of formula (i) is selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. In certain embodiments, n of formula (i) is selected from the group consisting of 0, 1, 2, 3, 4, and 5. In certain embodiments, n of formula (i) is selected from the group consisting of 0, 1, 2, and 3. In certain embodiments, n of formula (i) is selected from the group consisting of 0 and 1. In certain embodiments, n of formula (i) is 0.

[0310] In certain embodiments, -L 2 - is

Chemical formula

[0311] In certain embodiments, -L 2 - is

Chemical formula

[0312] In certain embodiments, -L 2 - is

Chemical formula

[0313] In certain embodiments, -L 2 - is of the following formula (xvi).

Chemical formula

[0314] In certain embodiments, the moiety -L 1 -L 2 - is

Chemical formula

[0315] In certain embodiments, the moiety -L 1 -L 2 - is of the following formula (IId-ii).

Chemical formula

[0316] In certain embodiments, moiety -L 1 -L 2 - is of the following formula (IId-ii′). [Chemical formula] In the formula, The unmarked dashed line indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond; The dashed line marked with an asterisk indicates the bond to -Z.

[0317] In certain embodiments, moiety -L 1 -L 2 - is [Chemical formula] selected from the group consisting of The unmarked dashed line indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond; The dashed line marked with an asterisk indicates the bond to -Z.

[0318] In certain embodiments, moiety -L 1 -L 2 - is of the following formula (IId-iia). [Chemical formula] In the formula, The unmarked dashed line indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond; The dashed line marked with an asterisk indicates the bond to -Z.

[0319] In certain embodiments, sub-L 1 -L 2 - is of the following formula (IId-iia′).

Chemical formula

[0320] In certain embodiments, sub-L 1 -L 2 - is

Chemical formula

[0321] In certain embodiments, sub-L 1 -L 2 - is of the following formula (IId-iib).

Chemical formula

[0322] In certain embodiments, sub-L 1 -L 2 - is of the following formula (IId-iib′).

Chemical formula

[0323] In certain embodiments, -Z of formula (Ia) or (Ib) has a molecular weight in the range of 5 to 200 kDa. In certain embodiments, -Z of formula (Ia) or (Ib) has a molecular weight in the range of 8 to 100 kDa. In certain embodiments, -Z of formula (Ia) or (Ib) has a molecular weight in the range of 10 to 80 kDa. In certain embodiments, -Z of (Ia) or (Ib) has a molecular weight in the range of 12 to 60 kDa. In certain embodiments, -Z of (Ia) or (Ib) has a molecular weight in the range of 15 to 40 kDa. In certain embodiments, -Z of (Ia) or (Ib) has a molecular weight of about 20 kDa. In certain embodiments, -Z of (Ia) or (Ib) has a molecular weight of about 40 kDa.

[0324] The polymer moiety -Z of formula (Ia) or (Ib) contains a polymer. In certain embodiments, -Z of formula (Ia) or (Ib) is poly(2-methacryloyloxyethyl phosphorylcholines), poly(acrylic acids), poly(acrylates), poly(acrylamides), poly(alkyloxy) polymers, poly(amides), poly(amideamines), poly(amino acids), poly(anhydrides), poly(aspartamides), poly(butyric acids), poly(glycolic acids), polybutylene terephthalates, poly(caprolactones), poly(carbonates), poly(cyanoacrylates), poly(dimethylacrylamides), poly(esters), poly(ethylenes), poly(ethylene glycols), poly(ethylene oxides), poly(ethyl phosphates), poly(ethyl oxazolines), poly(glycolic acids), poly(hydroxyethyl acrylates), poly(hydroxyethyl-oxazolines), poly(hydroxy methacrylates), poly(hydroxypropyl methacrylamides), poly(hydroxypropyl methacrylates), poly(hydroxypropyl oxazolines), poly(iminocarbonates), poly(lactic acids), poly(lactic-co-glycolic acids), poly(methacrylamides), poly(methacrylates), poly(methyl oxazolines), poly(organic phosphazenes), poly(orthoesters), poly(oxazolines), poly(propylene glycols), poly(siloxanes), poly(urethanes), poly(vinyl alcohols), poly(vinyl amines), poly(vinyl methyl ethers), poly(vinyl pyrrolidones), silicones, celluloses, carboxymethyl celluloses, hydroxypropyl methyl celluloses, chitins, chitosans, dextrans, dextrins, gelatins, hyaluronic acids and derivatives, functionalized hyaluronic acids, mannans, pectins, rhamnogalacturonans, starches, hydroxyalkyl starches, hydroxyethyl starches and other hydrocarbon-based polymers, xylans, and copolymers thereof.

[0325] In certain embodiments, -Z in formula (Ia) or (Ib) comprises a protein. Preferred proteins are the carboxyl-terminal peptide of chorionic gonadotropin as described in US2012 / 0035101A1 (incorporated herein by reference); albumin; the XTEN™ sequence as described in WO2011123813A2 (incorporated herein by reference); the proline / alanine random coil sequence as described in WO2011 / 144756A1 (incorporated herein by reference); the proline / alanine / serine random coil sequences as described in WO2008 / 155134A1 and WO2013 / 024049A1 (incorporated herein by reference); and Fc fusion proteins, and are selected from the group consisting of.

[0326] In certain embodiments, -Z in formula (Ia) or (Ib) is polysarcosine. In certain embodiments, -Z in formula (Ia) or (Ib) comprises poly(N-methylglycine). In certain embodiments, -Z in formula (Ia) or (Ib) comprises a random coil protein moiety. In certain embodiments, -Z in formula (Ia) or (Ib) comprises one random coil protein moiety. In certain embodiments, -Z in formula (Ia) or (Ib) comprises two random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises three random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises four random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises five random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises six random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises seven random coil protein moieties. In certain embodiments, -Z in formula (Ia) or (Ib) comprises eight random coil protein moieties.

[0327] In certain embodiments, such a random coil protein moiety comprises at least 25 amino acid residues and at most 2000 amino acids. In certain embodiments, such a random coil protein moiety comprises at least 30 amino acid residues and at most 1500 amino acid residues. In certain embodiments, such a random coil protein moiety comprises at least 50 amino acid residues and at most 500 amino acid residues.

[0328] In certain embodiments, -Z of formula (Ia) or (Ib) comprises a fatty acid derivative. In certain embodiments, -Z of formula (Ia) or (Ib) is a fatty acid derivative. In certain embodiments, -Z of formula (Ia) is a fatty acid derivative and x is 1.

[0329] In certain embodiments, -Z of formula (Ia) or (Ib) is a fatty acid derivative disclosed in WO2006 / 097537A1 which is incorporated herein by reference.

[0330] In certain embodiments, -Z of formula (Ia) or (Ib) is a fatty acid derivative disclosed in WO2021 / 055497A1 which is incorporated herein by reference. Thus, in certain embodiments, -Z of formula (Ia) or (Ib) has the following structure (w):

Chemical formula

[0331] In certain embodiments, -Z is of formula (w) and -L 1 - is of formula (V).

[0332] In certain embodiments, -Z-L 2 -L 1 - is of formula (w-a):

Chemical formula

[0333] In certain embodiments, CNP has a sequence selected from the group consisting of: PGQEHPQARRYRGAQRRGLSRGCFGLKLDRIGSMSGLGC (SEQ ID NO: 98); PGQEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC (SEQ ID NO: 30); PGQEHPNARRYRGANRRGLSRGCFGLKLDRIGSMSGLGC (SEQ ID NO: 99); and PGQEHPQARKYKGAQKKGLSKGCFGLKLDRIGSMSGLGC (SEQ ID NO: 100).

[0334] In certain embodiments, CNP has a sequence selected from the group consisting of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100, -Z is of formula (w), -L 1 - is a reversible linker moiety. In certain embodiments, CNP has a sequence selected from the group consisting of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100, -Z is of formula (w), -L 1 - is of formula (V). In certain embodiments, CNP has a sequence selected from the group consisting of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100, -Z-L 2 -L 1 - is of formula (w-a). Said -L 1 - may be attached to said CNP via a lysine other than the lysine within the ring structure, or may be attached to the N-terminus. In certain embodiments, the CNP of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100 further comprises an acetyl group such as an acetyl group at the N-terminus of the peptide. In certain embodiments, the CNP of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100 further comprises a C-terminal -OH or -NH2 group. In certain embodiments, the CNP of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99 and SEQ ID NO: 100 and -L 1- is attached to a residue of the ring portion of CNP or a site other than the CNP portion.

[0335] In certain embodiments, -L 1 - is attached to a lysine residue such as the bold lysine residues of SEQ ID NO: 98, SEQ ID NO: 30, SEQ ID NO: 99, and SEQ ID NO: 100: TIFF2025522281000056.tif30162

[0336] In certain embodiments, CNP is selected from the group consisting of: TIFF2025522281000057.tif46163

[0337] In certain embodiments, -Z of formula (Ia) or (Ib) is a hyaluronic acid-based polymer. In certain embodiments, -Z of formula (Ia) or (Ib) is a polymer moiety disclosed in WO2013 / 024047A1 (incorporated herein by reference).

[0338] In certain embodiments, -Z of formula (Ia) or (Ib) is a polymer moiety disclosed in WO2013 / 024048A1 (incorporated herein by reference).

[0339] In certain embodiments, -Z of formula (Ia) or (Ib) is a PEG-based polymer. In certain embodiments, -Z is a branched or multi-branched PEG-based polymer.

[0340] In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer. In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer having 1, 2, 3, 4, 5 or 6 branch points. In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer having 1, 2 or 3 branch points. In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer having 1 branch point. In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer having 2 branch points. In certain embodiments, -Z of formula (Ia) or (Ib) is a branched polymer having 3 branch points.

[0341] In certain embodiments, the branch point is selected from the group consisting of -N<, -CH< and C<.

[0342] In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) is PEG-based.

[0343] In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight in the range of 5 kDa to 500 kDa (including both ends). In certain embodiments, in certain embodiments, it has a molecular weight in the range of 10 kDa to 250 kDa (including both ends). In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight in the range of 10 kDa to 150 kDa (including both ends). In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight in the range of 12 kDa to 100 kDa (including both ends). In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight in the range of 15 kDa to 80 kDa (including both ends). In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight in the range of 10 kDa to 80 kDa (including both ends). In certain embodiments, the molecular weight is about 10 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 20 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 30 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 40 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 50 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 60 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 70 kDa. In certain embodiments, the molecular weight of such a branched moiety -Z of formula (Ia) or (Ib) is about 80 kDa. In certain embodiments, such a branched moiety -Z of formula (Ia) or (Ib) has a molecular weight of about 40 kDa.

[0344] In certain embodiments, -Z comprises the following moiety. [Chemical formula]

[0345] In certain embodiments, -Z contains an amide bond.

[0346] In certain embodiments, -Z of formula (Ia) or (Ib) contains a moiety of the following formula (a).

Chemical formula

[0347] Optionally, the moiety of formula (a) is substituted with one or more substituents.

[0348] In certain embodiments, BP in formula (a) a is -N<. In certain embodiments, BP in formula (a) a is -CR<. In certain embodiments, -R is -H. Thus, in certain embodiments, a in formula (a) is 0. In certain embodiments, BP in formula (a) a is >C<.

[0349] In certain embodiments, -S a - in formula (a) is a chemical bond.

[0350] In certain embodiments, -S a - in formula (a) is selected from the group consisting of C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl, and C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl is -C(O)O-, -O-, -C(O)-, -C(O)N(R 4 ), -, -S(O)2N(R 4 ), -, -S(O)N(R 4 ), -, -S(O)2-, -S(O)-, -N(R 4 )S(O)2N(R 4a ), -, -S-, -N(R 4 ), -, -OC(OR 4)(R 4a )-, -N(R 4 )C(O)N(R 4a )-, and -OC(O)N(R 4 )- may optionally be interrupted by one or more chemical groups selected from the group consisting of; -R 4 and -R 4a are independently selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, the -S a - of formula (a) is selected from the group consisting of methyl, ethyl, propyl, butyl, which may optionally be interrupted by one or more chemical groups selected from the group consisting of -O-, -C(O)-, and -C(O)N(R 4 ).

[0351] In certain embodiments, - the -S a′ - of formula (a) is a chemical bond.

[0352] In certain embodiments, the -S a′ - of formula (a) is selected from the group consisting of C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl, and C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl may optionally be interrupted by one or more chemical groups selected from the group consisting of -C(O)O-, -O-, -C(O)-, -C(O)N(R 4 )-, -S(O)2N(R 4 )-, -S(O)N(R 4 )-, -S(O)2-, -S(O)-, -N(R 4 )S(O)2N(R 4a )-, -S-, -N(R 4 )-, -OC(OR 4 )(R 4a )-, -N(R 4 )C(O)N(R 4a )-, and -OC(O)N(R 4 )-; -R 4 and -R 4ais independently selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, -S a′ - of formula (a) is selected from the group consisting of methyl, ethyl, propyl, and butyl, and one or more chemical groups selected from the group consisting of -O-, -C(O)-, and -C(O)N(R 4 )- may optionally intervene.

[0353] In certain embodiments, -S a″ - of formula (a) is a chemical bond.

[0354] In certain embodiments, -S a″ - of formula (a) is selected from the group consisting of C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl, and C 1-10 alkyl, C 2-10 alkenyl, and C 2-10 alkynyl may optionally be interrupted by one or more chemical groups selected from the group consisting of -C(O)O-, -O-, -C(O)-, -C(O)N(R 4 ), -, -S(O)2N(R 4 ), -, -S(O)N(R 4 ), -, -S(O)2-, -S(O)-, -N(R 4 ), S(O)2N(R 4a ), -, -S-, -N(R 4 ), -, -OC(OR 4 )(R 4a ), -, -N(R 4 ), C(O)N(R 4a ), - and -OC(O)N(R 4 ); -R 4 and -R 4a are independently selected from the group consisting of -H, methyl, ethyl, propyl, and butyl. In certain embodiments, -S a″ - of formula (a) is selected from the group consisting of methyl, ethyl, propyl, and butyl, and one or more chemical groups selected from the group consisting of -O-, -C(O)-, and -C(O)N(R 4 )- may optionally intervene.

[0355] In certain embodiments, the -S of formula (a) a″′ - is a chemical bond.

[0356] In certain embodiments, the -S of formula (a) a″′ - is C 1-10 alkyl, C 2-10 alkenyl and C 2-10 alkynyl, selected from the group consisting of C 1-10 alkyl, C 2-10 alkenyl and C 2-10 alkynyl is -C(O)O-, -O-, -C(O)-, -C(O)N(R 4 ), -, -S(O)2N(R 4 ), -, -S(O)N(R 4 ), -, -S(O)2-, -S(O)-, -N(R 4 ), S(O)2N(R 4a ), -, -S-, -N(R 4 ), -, -OC(OR 4 )(R 4a ), -, -N(R 4 ), C(O)N(R 4a ), - and -OC(O)N(R 4 ), - from the group consisting of one or more chemical groups selected may optionally intervene; -R 4 and -R 4a are independently selected from the group consisting of -H, methyl, ethyl, propyl and butyl. In certain embodiments, the -S of formula (a) a″′ - is selected from the group consisting of methyl, ethyl, propyl, butyl, which may optionally be interrupted by one or more chemical groups selected from the group consisting of -O-, -C(O)- and -C(O)N(R 4 ).

[0357] In certain embodiments, the -P of formula (a) a′ , -P a″ and -P a″′independently includes a polymer selected from the group consisting of poly(2-methacryloyloxyethyl phosphorylcholines), poly(acrylic acids), poly(acrylates), poly(acrylamides), poly(alkyloxy)polymers, poly(amides), poly(amideamines), poly(amino acids), poly(anhydrides), poly(aspartamides), poly(butyric acids), poly(glycolic acids), polybutylene terephthalates, poly(caprolactones), poly(carbonates), poly(cyanoacrylates), poly(dimethylacrylamides), poly(esters), poly(ethylenes), poly(ethylene glycols), poly(ethylene oxides), poly(ethyl phosphates), poly(ethyloxazolines), poly(glycolic acids), poly(hydroxyethyl acrylates), poly(hydroxyethyl-oxazolines), poly(hydroxy methacrylates), poly(hydroxypropyl methacrylamides), poly(hydroxypropyl methacrylates), poly(hydroxypropyl oxazolines), poly(iminocarbonates), poly(lactic acids), poly(lactic-co-glycolic acids), poly(methacrylamides), poly(methacrylates), poly(methyloxazolines), poly(organic phosphazenes), poly(orthoesters), poly(oxazolines), poly(propylene glycols), poly(siloxanes), poly(urethanes), poly(vinyl alcohols), poly(vinyl amines), poly(vinyl methyl ethers), poly(vinyl pyrrolidones), silicones, celluloses, carboxymethyl celluloses, hydroxypropyl methyl celluloses, chitins, chitosans, dextrans, dextrins, gelatins, hyaluronic acids and derivatives, functionalized hyaluronic acids, mannans, pectins, rhamnogalacturonans, starches, hydroxyalkyl starches, hydroxyethyl starches and other hydrocarbon-based polymers, xylans, and copolymers thereof.

[0358] In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′Independently, it has a molecular weight in the range of 5 kDa to 50 kDa (including both ends), and in certain embodiments, in the range of 5 kDa to 40 kDa (including both ends), in certain embodiments, in the range of 7.5 kDa to 35 kDa (including both ends), in certain embodiments, in the range of 7.5 to 30 kDa (including both ends), in certain embodiments, in the range of 10 to 30 kDa (including both ends).

[0359] In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 5 kDa. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 7.5 kDa. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 10 kDa. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 12.5 kDa. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 15 kDa. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ has a molecular weight of about 20 kDa.

[0360] In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ independently contains a PEG-based moiety. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′Independently, it comprises a PEG-based moiety comprising at least 20% PEG, in certain embodiments at least 30%, in certain embodiments at least 40% PEG, in certain embodiments at least 50% PEG, in certain embodiments at least 60% PEG, in certain embodiments at least 70% PEG, in certain embodiments at least 80% PEG and in certain embodiments at least 90% PEG.

[0361] In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ Independently, it comprises a protein moiety, in certain embodiments a random coil protein moiety, in certain embodiments a random coil protein moiety selected from the group consisting of PA, PAS, PAG, PG and XTEN™ moieties.

[0362] In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ is a PA moiety. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ is a PAS moiety. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ is a PAG moiety. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ is a PG moiety. In certain embodiments, -P of formula (a) a′ , -P a″ and -P a″′ is an XTEN™ moiety.

[0363] In certain embodiments, -Z contains one moiety of formula (a). In certain embodiments, -Z contains two moieties of formula (a). In another embodiment, -Z contains three moieties of formula (a). In certain embodiments, -Z contains four moieties of formula (a). In certain embodiments, -Z contains five moieties of formula (a). In certain embodiments, -Z contains six moieties of formula (a).

[0364] In certain embodiments, -Z contains a moiety of the following formula (b). [Chemical formula] In the formula, the dashed line represents the bond to -L 2 - or to the remainder of -Z; b1 is selected from the group consisting of 0, 1, 2, 3, 4, 5, 6, 7, and 8; b2 is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, and 8; b3 is an integer in the range of 150 to 1000 (including both ends); in certain embodiments, in the range of 150 to 500 (including both ends); in certain embodiments, in the range of 200 to 460 (including both ends); b4 is an integer in the range of 150 to 1000 (including both ends); in certain embodiments, in the range of 150 to 500 (including both ends); in certain embodiments, in the range of 200 to 460 (including both ends).

[0365] Optionally, the moiety of formula (b) is substituted with one or more substituents.

[0366] In certain embodiments, b3 and b4 of formula (b) are the same integer. In certain embodiments, both b3 and b4 of formula (b) are integers in the range of 200 to 250, and in certain embodiments, b3 and b4 of formula (b) are about 225. In certain embodiments, both b3 and b4 of formula (b) are integers in the range of 400 to 500, and in certain embodiments, b3 and b4 of formula (b) are about 450.

[0367] In certain embodiments, b1 of formula (b) is selected from the group consisting of 0, 1, 2, 3, and 4. In certain embodiments, b1 of formula (b) is selected from the group consisting of 1, 2, and 3. In certain embodiments, b1 of formula (b) is 2.

[0368] In certain embodiments, b2 of formula (b) is selected from the group consisting of 1, 2, 3, 4, and 5. In certain embodiments, b2 of formula (b) is selected from the group consisting of 2, 3, and 4. In certain embodiments, b2 of formula (b) is 3.

[0369] In certain embodiments, b1 of formula (b) is 2, b2 of formula (b) is 3, and both b3 and b4 are about 450. In certain embodiments, b1 of formula (b) is 2, b2 of formula (b) is 3, and both b3 and b4 are about 225.

[0370] In certain embodiments, -Z contains one moiety of formula (b). In certain embodiments, -Z contains two moieties of formula (b). In certain embodiments, -Z contains three moieties of formula (b). In certain embodiments, -Z contains four moieties of formula (b). In certain embodiments, -Z contains five moieties of formula (b). In certain embodiments, -Z contains six moieties of formula (b).

[0371] In certain embodiments, -Z contains a moiety of formula (c).

Chemical formula

[0372] Optionally, the moiety of formula (c) is substituted with one or more substituents.

[0373] In certain embodiments, both c1 and c2 of formula (c) are the same integer.

[0374] In certain embodiments, c1 and c2 of formula (c) are in the range of 200 to 250 (including both ends), and in certain embodiments, are about 225. In certain embodiments, c1 and c2 of formula (c) are in the range of 400 to 500 (including both ends), and in certain embodiments, c1 and c2 of formula (c) are about 450.

[0375] In certain embodiments, sub-Z is a branched PEG-based polymer containing at least 10% PEG, has one branch point and two PEG-based polymer arms, and has a molecular weight of about 40 kDa. Accordingly, each of the two PEG-based polymer arms has a molecular weight of about 20 kDa. In certain embodiments, the branch point is -CH<.

[0376] In certain embodiments, -Z contains one moiety of formula (c). In certain embodiments, -Z contains two moieties of formula (c). In certain embodiments, -Z contains three moieties of formula (c). In certain embodiments, -Z contains four moieties of formula (c). In certain embodiments, -Z contains five moieties of formula (c). In certain embodiments, -Z contains six moieties of formula (c).

[0377] In certain embodiments, sub-Z is of the following formula (d). [Chemical formula] Wherein, The dashed line represents a bond to -L 2 -; -Z b - is selected from the group consisting of C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl; C 1-50 alkyl, C 2-50 alkenyl, and C 2-50The alkynyl may be substituted by one or more identical or different -R 1 and may be C 1-50 alkyl, C 2-50 alkenyl and C 2-50 The alkynyl may optionally be interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R 2 ), -S(O)2N(R 2 ), -S(O)N(R 2 ), -S(O)2-, -S(O)-, -N(R 2 )S(O)2N(R 2a ), -S-, -N(R 2 ), -OC(OR 2 )(R 2a ), -N(R 2 )C(O)N(R 2a ), and -OC(O)N(R 2 ); Each -T- is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbocyclic polycycle, and 8- to 30-membered heterocyclic polycycle; each -T- may optionally be substituted by one or more identical or different -R 1 ; Each -R 1 is independently halogen, -CN, oxo (=O), -COOR 3 , -OR 3 , -C(O)R 3 , -C(O)N(R 3 R 3a ), -S(O)2N(R 3 R 3a ), -S(O)N(R 3 R 3a ), -S(O)2R 3 , -S(O)R 3 , -N(R 3 )S(O)2N(R 3a R 3b ), -SR 3 , -N(R 3 R 3a ), -NO2, -OC(O)R 3, -N(R 3 )C(O)R 3a , -N(R 3 )S(O)2R 3a , -N(R 3 )S(O)R 3a , -N(R 3 )C(O)OR 3a , -N(R 3 )C(O)N(R 3a R 3b , -OC(O)N(R 3 R 3a ), and C 1-6 alkyl selected from the group consisting of; C 1-6 alkyl may be substituted with one or more identical or different halogens; each -R 2 , -R 2a , -R 3 , -R 3a and -R 3b is independently selected from the group consisting of -H and C 1-6 alkyl, and C 1-6 alkyl may be substituted with one or more identical or different halogens; -Z a is as follows: [Chemical formula] where; In the formula, BP a , -S a -, -S a′ -, -S a″ -, -S a″′ , -P a′ , -P a″ , -P a″′ and a are used as defined for formula (a).

[0378] Optionally, the moiety of formula (d) is substituted with one or more substituents.

[0379] In certain embodiments, the BP of formula (d) a , -S a -, -S a′ -, -S a″ -, -Sa″′ -, -P a′ -, -P a″ -, -P a″′ is as defined above for formula (a).

[0380] In certain embodiments, -Z of formula (d) a is that of formula (b). In certain embodiments, b1, b2, b3, and b4 are as described for formula (b).

[0381] In certain embodiments, the moiety -Z of formula (Ia) or (Ib) is that of the following formula (e).

Chemical formula

Chemical formula

[0382] Optionally, the moiety of formula (e) is substituted with one or more substituents.

[0383] In certain embodiments, b1, b2, b3, and b4 of formula (e) are as defined above for formula (b).

[0384] In certain embodiments, e in formula (e) is 1. In certain embodiments, e in formula (e) is 2. In certain embodiments, e in formula (e) is 3. In certain embodiments, e in formula (e) is 4. In certain embodiments, e in formula (e) is 5. In certain embodiments, e in formula (e) is 6. In certain embodiments, e in formula (e) is 7. In certain embodiments, e in formula (e) is 8. In certain embodiments, e in formula (e) is 9. In certain embodiments, e in formula (e) is 10. In certain embodiments, e in formula (e) is 11. In certain embodiments, e in formula (e) is 12. In certain embodiments, e in formula (e) is 13. In certain embodiments, e in formula (e) is 14. In certain embodiments, e in formula (e) is 15.

[0385] In certain embodiments, e in formula (e) is selected from the group consisting of 2, 3, 4, 5, 6, 7, 8, and 9. In certain embodiments, e in formula (e) is selected from 3, 4, 5, and 6. In certain embodiments, e in formula (e) is 5.

[0386] In certain embodiments, e in formula (e) is 5, b1 in formula (e) is 2, b2 in formula (e) is 3, and both b3 and b4 in formula (e) are about 450.

[0387] In certain embodiments, sub-Z of formula (Ia) or (Ib) is of the following formula (e-i) or (e-i′).

Chemical formula

Chemical formula

[0388] In certain embodiments, b1, b2, b3, and b4 of formulas (e-i) and (e-i') are as defined above for formula (b).

[0389] In certain embodiments, e of formulas (e-i) and (e-i') is as described for formula (e).

[0390] In certain embodiments, b1 of formulas (e-i) and (e-i') is 2, b2 of formulas (e-i) and (e-i') is 3, and both b3 and b4 of formulas (e-i) and (e-i') are about 450.

[0391] In certain embodiments, -Z of formula (Ia) or (Ib) is that of formula (e-i).

[0392] In certain embodiments, moiety -Z is a branched PEG-based polymer containing at least 10% PEG, has 3 branch points and 4 PEG-based polymer arms, and has a molecular weight of about 40 kDa. Accordingly, each of the 4 PEG-based polymer arms has a molecular weight of about 10 kDa. In certain embodiments, each of the 3 branch points is -CH<.

[0393] In certain embodiments, moiety -Z is that of the following formula (f). [Chemical formula] wherein the dashed line represents a bond to -L 2 ; BP f is a branch point selected from the group consisting of -N<, -CR<, and C<; -R is selected from the group consisting of -H and C 1-6 alkyl; f is 0 when BP f is -N< or -CR<, and f is BPf is 1 when it is >C<; -S f -, -S f′ -, -S f″ - and -S f″′ - is independently a chemical bond or independently C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl selected from the group consisting of; C 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkyl, alkenyl, and C alkynyl may be substituted with one or more identical or different -R 1 and C alkyl, alkenyl, and C alkynyl may be interrupted by one or more groups selected from the group consisting of -T-, -C(O)O-, -O-, -C(O)-, -C(O)N(R 1-50 alkyl, C 2-50 alkenyl, and C 2-50 alkynyl, -S(O)2N(R 2 ), -S(O)N(R 2 ), -S(O)2-, -S(O)-, -N(R 2 ), -S-, -N(R 2 ), -S(O)2N(R 2a ), -S-, -N(R 2 ), -OC(OR 2 )(R 2a ), -N(R 2 ), -N(R 2a ), and -OC(O)N(R 2 ); each -T- is independently selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C 3-10 cycloalkyl, 3- to 10-membered heterocyclyl, 8- to 11-membered heterobicyclic, 8- to 30-membered carbocyclic polycycle, and 8- to 30-membered heterocyclic polycycle; each -T- may be independently substituted with one or more identical or different -R 1 ; each R 1 is independently halogen, -CN, oxo (=O), -COOR 3 , -OR 3 , -C(O)R 3 , -C(O)N(R 3 R3a ), -S(O)2N(R 3 R 3a ), -S(O)N(R 3 R 3a ), -S(O)2R 3 , -S(O)R 3 , -N(R 3 )S(O)2N(R 3a R 3b ), -SR 3 , -N(R 3 R 3a ), -NO2, -OC(O)R 3 , -N(R 3 )C(O)R 3a , -N(R 3 )S(O)2R 3a , -N(R 3 )S(O)R 3a , -N(R 3 )C(O)OR 3a , -N(R 3 )C(O)N(R 3a R 3b ), -OC(O)N(R 3 R 3a ), and C 1-6 alkyl; 1-6 The alkyl may be optionally substituted with one or more identical or different halogens; Each-R 2 , -R 2a , -R 3 , -R 3a and -R 3b are independently -H and C 1-6 alkyl, C 1-6 The alkyl may be optionally substituted with one or more identical or different halogens; -Z a′ , -Z a″ and -Z a″′ independently, [ka] and During the ceremony, BP a , -S a -, -S a′ -, -S a″-, -S a″′ -, -P a′ , -P a″ , -P a″′ and a is used as defined for formula (a).

[0394] Optionally, the moiety of formula (f) is substituted with one or more substituents.

[0395] In certain embodiments, BP of formula (f) a , -S a -, -S a′ -, -S a″ -, -S a″′ -, -P a′ , -P a″ and -P a″′ are as defined above for formula (a).

[0396] In certain embodiments,, BP of formula (f) f is -CR< and r is 0. In certain embodiments, -R is -H.

[0397] In certain embodiments, -S of formula (f) f - is a chemical bond.

[0398] In certain embodiments, -Z of formula (f) a′ , -Z a″ and -Z a″′ have the same structure. In certain embodiments, -Z of formula (f) a′ , -Z a″ and -Z a″′ are those of formula (b).

[0399] In certain embodiments, b1, b2, b3 and b4 are as described for formula (b).

[0400] In certain embodiments, -S of formula (f) f - is a chemical bond and BP of formula (f) a is -CR< and -R is -H. In certain embodiments, -S of formula (f) f- is a chemical bond, and BP of formula (f) a is -CR<, -R is -H, and -Z of formula (f) a′ , -Z a″ and -Z a″′ are those of formula (b).

[0401] In certain embodiments, -Z is of the following formula (g).

Chemical formula

Chemical formula

[0402] Optionally, the moiety of formula (g) is substituted with one or more substituents.

[0403] In certain embodiments, the BP of formula (g) a , -S a (-), -S a′ (-), -S a″ (-), -S a″′ (-), -P a′ , -P a″ and -P a″′ are as defined above for formula (a).

[0404] In certain embodiments, the -S of formula (g) g (-) is selected from the group consisting of C 1-6 alkyl, C 2-6 alkenyl and C 2-6 alkynyl, which may be substituted with one or more identical or different -R 1 groups, -R 1 is halogen, oxo (=O), -COOR 3 , -OR 3 , -C(O)R 3 , -C(O)N(R 3 R 3a ), -S(O)2N(R 3 R 3a ), -S(O)N(R3 R 3a ), -S(O)2R 3 , -S(O)R 3 , -N(R 3 )S(O)2N(R 3a R 3b ), -SR 3 , -N(R 3 R 3a ), -NO2, -OC(O)R 3 , -N(R 3 )C(O)R 3a , -N(R 3 )S(O)2R 3a , -N(R 3 )S(O)R 3a , -N(R 3 )C(O)OR 3a , -N(R 3 )C(O)N(R 3a R 3b ), -OC(O)N(R 3 R 3a ), and C 1-6 alkyl; 1-6 The alkyl may be optionally substituted with one or more identical or different halogens; -R 3 , -R 3a and -R 3b is independently selected from: —H, methyl, ethyl, propyl, and butyl.

[0405] In certain embodiments, the -S of formula (g) g - is C 1-6 is selected from alkyl.

[0406] In certain embodiments, the -S of formula (g) g′ - is C 1-6 Alkyl, C 2-6 Alkenyl and C 2-6 alkynyl, which may be one or more of the same or different -R 1 may be substituted with -R 1 is halogen, oxo (=O), -COOR 3 , -OR 3 , -C(O)R3 ,-C(O)N(R 3 R 3a ),-S(O)2N(R 3 R 3a ),-S(O)N(R 3 R 3a ),-S(O)2R 3 ,-S(O)R 3 ,-N(R 3 ),S(O)2N(R 3a R 3b ),-SR 3 ,-N(R 3 R 3a ),-NO2,-OC(O)R 3 ,-N(R 3 ),C(O)R 3a ,-N(R 3 ),S(O)2R 3a ,-N(R 3 ),S(O)R 3a ,-N(R 3 ),C(O)OR 3a ,-N(R 3 ),C(O)N(R 3a R 3b ),-OC(O)N(R 3 R 3a ),and C 1-6 is selected from the group consisting of alkyl; C 1-6 alkyl may be substituted with one or more identical or different halogens; -R 3 ,-R 3a and -R 3b are independently selected from -H, methyl, ethyl, propyl, and butyl.

[0407] In certain embodiments, the -S g′ - of formula (g) is selected from C 1-6 alkyl.

[0408] In certain embodiments, the -S g″ - of formula (g) is selected from the group consisting of C 1-6 alkyl, C 2-6 alkenyl, and C 2-6 alkynyl, which may be substituted with one or more identical or different -R 1may be substituted with -R 1 is halogen, oxo (=O), -COOR 3 , -OR 3 , -C(O)R 3 , -C(O)N(R 3 R 3a ), -S(O)2N(R 3 R 3a ), -S(O)N(R 3 R 3a ), -S(O)2R 3 , -S(O)R 3 , -N(R 3 )S(O)2N(R 3a R 3b ), -SR 3 , -N(R 3 R 3a ), -NO2, -OC(O)R 3 , -N(R 3 )C(O)R 3a , -N(R 3 )S(O)2R 3a , -N(R 3 )S(O)R 3a , -N(R 3 )C(O)OR 3a , -N(R 3 )C(O)N(R 3a R 3b ), -OC(O)N(R 3 R 3a ), and C 1-6 alkyl; 1-6 The alkyl may be optionally substituted with one or more identical or different halogens; -R 3 , -R 3a and -R 3b is independently selected from: —H, methyl, ethyl, propyl, and butyl.

[0409] In certain embodiments, the -S of formula (g) g″ - is C 1-6 is selected from alkyl.

[0410] In certain embodiments, -Z of formula (g) a and -Z a′have the same structure. In certain embodiments, -Z of formula (g) a and -Z a′ is as defined in formula (b).

[0411] In certain embodiments, -Z of formula (Ia) or (Ib) is of the following formula (g-i).

Chemical formula

Chemical formula

Chemical formula

[0412] Optionally, the moiety of formula (g-i) is substituted with one or more substituents.

[0413] In certain embodiments, -Y a1 - and -Y a1′ - are both

Chemical formula

[0414] In certain embodiments, BP a , -S a , -S a′ , -S a″ , -S a″′ , -P a′ , -P a″ and -P a″′is as defined above for formula (a).

[0415] In certain embodiments, -S of formula (g-i) g -, -S g′ - and -S g″ - are as defined for formula (g).

[0416] In certain embodiments, -Z of formula (g-i) a and -Z a′ have the same structure. In certain embodiments, -Z of formula (g-i) a and -Z a′ are those of formula (b). In certain embodiments, b1, b2, b3 and b4 are as described for formula (b).

[0417] In certain embodiments, -Z is of the following formula (h).

Chemical formula

Chemical formula

[0418] Optionally, the moiety of formula (h) is substituted with one or more substituents.

[0419] In certain embodiments, both c1 of formula (h) are the same. In certain embodiments, both c1 of formula (h) are about 225.

[0420] In certain embodiments, -Z of formula (Ia) or (Ib) is the following formula (h-a):

Chemical formula

Chemical formula

Chemical formula

[0421] Optionally, the moiety of formula (h-a) is substituted with one or more substituents.

[0422] In certain embodiments, each k of formula (h-a) is independently selected from the group consisting of 2, 3, 4, 5, 6 and 7. In certain embodiments, both k's of formula (h-a) are the same.

[0423] In certain embodiments, both c1's of formula (h-a) are the same.

[0424] In certain embodiments, both c1's of formula (h-a) are about 225.

[0425] In certain embodiments, -Y a1 - and -Y a1′ - are both

Chemical formula

[0426] In certain embodiments, sub-Z is of the following formula (h-i).

Chemical formula

Chemical formula

[0427] Optionally, the moiety of formula (h-i) is substituted with one or more substituents.

[0428] In certain embodiments, both c1s in formula (h-i) are the same. In certain embodiments, both c1s in formula (h-i) are about 225.

[0429] In certain embodiments, sub-Z of formula (Ia) or (Ib) is of the following formula (h-ia).

Chemical formula

Chemical formula

Chemical formula

[0430] In certain embodiments, each k of formula (h-ia) is independently selected from the group consisting of 2, 3, 4, 5, 6, and 7. In certain embodiments, both k's of formula (h-ia) are the same.

[0431] In certain embodiments, both c1's of formula (h-ia) are the same. In certain embodiments, both c1's of formula (h-ia) are about 225.

[0432] In certain embodiments, -Y a1 - and -Y a1′ - are both

Chemical Structure

[0433] In certain embodiments, -Z of formula (Ia) or (Ib) includes a moiety selected from the group consisting of the following.

Chemical Structure

Chemical formula

[0434] In certain embodiments, -Y of formulas (j-iv), (j-v) and (j-vi) d1 - and -Y d2 - and -Y of formula (j-vii) d1 -,-Y d2 -,-Y d3 - and -Y d4 - is

Chemical formula

[0435] In certain embodiments, -Y of formulas (j-iv), (j-v) and (j-vi) d1 - and -Y d2 - and -Y of formula (j-vii) d1 -,-Y d2 -,-Y d3 - and -Y d4 - is

Chemical formula

[0436] In certain embodiments, -X of formula (j-i) f1 ,-X f2 ,-X f3 ,-X f4 ,-X f5 ,-X f6 ,-X f7 and -X f8 is -H; -X of formula (j-i) d1 and -X d2 is -OH; -X of formula (j-i) e1 and -X e2is selected from the group consisting of -H and methyl; s1, s2, and s3 of formula (j-i) are selected from the group consisting of 2, 3, 4, 5, and 6.

[0437] In certain embodiments, -X of formula (j-i) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 and -X f8 is -H; -X of formula (j-i) d1 and -X d2 is -OH; -X of formula (j-i) e1 and -X e2 is -H; s1, s2, and s3 of formula (j-i) are 4.

[0438] In certain embodiments, -X of formula (j-ii) f1 , -X f2 , -X f3 and -X f4 is -H; -X of formula (j-ii) d1 , -X d2 , -X d3 and -X d4 is -OH; -X of formula (j-ii) e1 , -X e2 , -X e3 and -X e4 is selected from the group consisting of -H and methyl; s1, s2, s3, s4, and s5 of formula (j-ii) are selected from the group consisting of 1, 2, 3, 4, 5, and 6. In certain embodiments, -X of formula (j-ii) f1 , -X f2 , -X f3 and -X f4 is -H; -X of formula (j-ii) d1 , -X d2 , -X d3 and -X d4 is -OH; -Xe1, -Xe2, -Xe3, and -X of formula (j-ii) e4 is -H; s1 of formula (j-ii) is 4, and s2, s3, s4, and s5 of formula (j-ii) are 1.

[0439] In certain embodiments, -X in formula (j-iii) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 and -X f10 is -H; -X in formula (j-iii) d1 , -X d2 , -X d3 and -X d4 is -OH; -X in formula (j-iii) e1 , -X e2 , -X e3 and -X e4 is selected from the group consisting of -H and methyl; s1, s2 and s3 in formula (j-iii) are selected from the group consisting of 2, 3, 4, 5 and 6. In certain embodiments, -X in formula (j-iii) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 and -X f10 is -H; -X in formula (j-iii) d1 , -X d2 , -X d3 and -X d4 is -OH; -X in formula (j-iii) e1 , -X e2 , -X e3 and -X e4 is -H; s1, s2 and s3 in formula (j-iii) are 4.

[0440] In certain embodiments, -X in formula (j-iv) f1 , -X f2 , -X f3 , -X f4 , -X f5 and -X f6is -H; s1, s2, s3, s4, s5, s6 and s7 of formula (j-iv) are selected from the group consisting of 1, 2, 3, 4, 5, 6 and 7; -Y d1 - and -Y d2 - is

Chemical formula

[0441] In certain embodiments, -X of formula (j-iv) f1 , -X f2 , -X f3 , -X f4 , -X f5 and -X f6 is -H; s1 of formula (j-iv) is 3, s2 of formula (j-iv) is 5, s3 of formula (j-iv) is 2, s4 of formula (j-iv) is 4, s5 of formula (j-iv) is 5, s6 of formula (j-iv) is 2, s7 of formula (j-iv) is 4; -Y of formula (j-iv) d1 - and -Y d2 - is

Chemical formula

[0442] In certain embodiments, -X of formula (j-iv) f1 , -X f2 , -X f3 , -X f4 , -X f5 and -X f6 is -H; s1 of formula (j-iv) is 3, s2 of formula (j-iv) is 5, s3 of formula (j-iv) is 2, s4 of formula (j-iv) is 4, s5 of formula (j-iv) is 5, s6 of formula (j-iv) is 2, s7 of formula (j-iv) is 4; -Y of formula (j-iv) d1 - and -Y d2 - is

Chemical formula

[0443] In a particular embodiment, -X of formula (j-v) f1 、 -X f2 、 -X f3 and -X f4 are -H; s1, s2, s3, s4 and s5 of formula (j-v) are selected from the group consisting of 1, 2, 3, 4, 5, 6 and 7; -Y of formula (j-v) d1 - and -Y d2 - are

Chemical formula

[0444] In a particular embodiment, -X of formula (j-v) f1 、 -X f2 、 -X f3 and -X f4 are -H; s1 of formula (j-v) is 3, s2 of formula (j-v) is 2, s3 of formula (j-v) is 1, s4 of formula (j-v) is 2, s5 of formula (j-v) is 1; -Y of formula (j-v) d1 - and -Y d2 - are

Chemical formula

[0445] In a particular embodiment, -X of formula (j-v) f1 、 -X f2 、 -X f3 and -X f4 are -H; s1 of formula (j-v) is 3, s2 of formula (j-v) is 2, s3 of formula (j-v) is 1, s4 of formula (j-v) is 2, s5 of formula (j-v) is 1; -Y of formula (j-v) d1 - and -Y d2 - are [Chemistry] and; In the formula, the dashed lines marked with asterisks are each -Z d1 , -Z d2 , -Z d3 and -Z d4 and are facing towards -L 2 -.

[0446] In a specific embodiment, -X f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 , -X f10 in formula (j-vi) is -H; s1, s2, s3, s4, s5, s6, s7, s8 and s9 in formula (j-vi) are selected from the group consisting of 1, 2, 3, 4, 5, 6 and 7; -Y d1 - and -Y d2 - are [Chemistry] selected from the group consisting of.

[0447] In a specific embodiment, -X f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 , -X f10 in formula (j-vi) is -H; s1 in formula (j-vi) is 4, s2 in formula (j-vi) is 5, s3 in formula (j-vi) is 2, s4 in formula (j-vi) is 4, s5 in formula (j-vi) is 4, s6 in formula (j-vi) is 5, s7 in formula (j-vi) is 2, s8 in formula (j-vi) is 4, s9 in formula (j-vi) is 4; -Y d1- and -Y d2 - is [Chem.] as follows.

[0448] In certain embodiments, -X in formula (j-vi) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 and -X f10 is -H; s1 in formula (j-vi) is 4, s2 in formula (j-vi) is 5, s3 in formula (j-vi) is 2, s4 in formula (j-vi) is 4, s5 in formula (j-vi) is 4, s6 in formula (j-vi) is 5, s7 in formula (j-vi) is 2, s8 in formula (j-vi) is 4, s9 in formula (j-vi) is 4; -Y in formula (j-v) d1 - and -Y d2 - is [Chem.] as follows; wherein the dashed lines marked with an asterisk are each directed towards -Z d1 , -Z d2 , -Z d3 and -Z d4 , and the unmarked dashed lines are directed towards -L 2 .

[0449] In certain embodiments, -X in formula (j-vii) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 , -X f10 , -X f11 , -X f12 , -X f13 and -Xf14 is -H; s1, s2, s3, s4, s5, s6, s7, s8, s9, s10, s11, s12, s13, s14 and s15 in formula (j-vii) are selected from the group consisting of 1, 2, 3, 4, 5, 6 and 7; -Y in formula (j-vii) d1 -, -Y d2 -, -Y d3 - and -Y d4 - is

Chemical formula

[0450] In a specific embodiment, -X in formula (j-vii) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 , -X f10 , -X f11 , -X f12 , -X f13 and -X f14 is -H; is -H; s1 in formula (j-vii) is 4, s2 in formula (j-vii) is 4, s3 in formula (j-vii) is 5, s4 in formula (j-vii) is 2, s5 in formula (j-vii) is 4, s6 in formula (j-vii) is 5, s7 in formula (j-vii) is 2, s8 in formula (j-vii) is 4, s9 in formula (j-vii) is 4, s10 in formula (j-vii) is 5, s11 in formula (j-vii) is 2, s12 in formula (j-vii) is 4, s13 in formula (j-vii) is 5, s14 in formula (j-vii) is 2, s15 in formula (j-vii) is 4; -Y in formula (j-vii) d1 -, -Y d2 -, -Y d3 - and -Y d4 - is

Chemical formula

[0451] In certain embodiments, -X of formula (j-vii) f1 , -X f2 , -X f3 , -X f4 , -X f5 , -X f6 , -X f7 , -X f8 , -X f9 , -X f10 , -X f11 , -X f12 , -X f13 and -X f14 is -H; is -H; s1 of formula (j-vii) is 4, s2 of formula (j-vii) is 4, s3 of formula (j-vii) is 5, s4 of formula (j-vii) is 2, s5 of formula (j-vii) is 4, s6 of formula (j-vii) is 5, s7 of formula (j-vii) is 2, s8 of formula (j-vii) is 4, s9 of formula (j-vii) is 4, s10 of formula (j-vii) is 5, s11 of formula (j-vii) is 2, s12 of formula (j-vii) is 4, s13 of formula (j-vii) is 5, s14 of formula (j-vii) is 2, s15 of formula (j-vii) is 4; -Y of formula (j-vii) d1 -, -Y d2 -, -Y d3 -and -Y d4 -are [Chemical formula] ; The dashed lines marked with asterisks are each directed towards -Z d1 , -Z d2 , -Z d3 and -Z d4 , and the unmarked dashed line is directed towards -L 2 -.

[0452] In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4have the same structure. In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4 is the PA moiety. In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4 is the PAS moiety. In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4 is the PAG moiety. In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4 is the PG moiety. In certain embodiments, -Z of formulas (j-i), (j-ii), (j-iii), (j-iv), (j-v), (j-vi) and (j-vii) d1 , -Z d2 , -Z d3 and -Z d4 is the XTEN™ moiety.

[0453] In certain embodiments, the reversible CNP conjugate has formula (Iif):

Chemical Formula

Chemical Formula

Chem.

[0454] In certain embodiments, each c1 of formula (Iif) is about 225.

[0455] In certain embodiments, the reversible CNP conjugate has formula (Iif -i):

Chem.

Chem.

Chem.

[0456] In certain embodiments, each c1 of formula (Iif-i) is about 225.

[0457] In certain embodiments, the CNP conjugate has formula (Iif-ii):

Chem.

[0458] In certain embodiments, each c1 of formula (Iif-ii) is about 225.

[0459] In certain embodiments, -D of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety, i.e., the reversible conjugates of formulas (Iif), (Iif-i) and (Iif-ii) are CNP conjugates. In certain embodiments, -D of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 24, SEQ ID NO: 25 or SEQ ID NO: 30. In certain embodiments, -D of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 24. In certain embodiments, the -D moiety of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 20. In certain embodiments, the -D moiety of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 21. In certain embodiments, the -D moiety of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 22. In certain embodiments, the -D moiety of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 23. In certain embodiments, the -D moiety of formulas (Iif), (Iif-i) and (Iif-ii) is a CNP moiety having the sequence of SEQ ID NO: 30.

[0460] In certain embodiments, -D in formulas (Iif), (Iif-i) and (Iif-ii) is the CNP moiety linked to -L through the nitrogen of the N-terminal amine functionality of CNP 1 - is the CNP moiety that is attached to

[0461] In certain embodiments, -D in formulas (Iif), (Iif-i) and (Iif-ii) is the CNP moiety linked to -L through the nitrogen provided by the amine functionality of the lysine side chain of the CNP moiety 1 - is the CNP moiety that is attached to

[0462] In certain embodiments, when the CNP moiety is that of SEQ ID NO: 24, the lysine side chain is not part of the ring formed by the disulfide bridge between the cysteine residues at positions 22 and 38

[0463] Thus, in certain embodiments, when CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the reversible CNP conjugates of formulas (Iif), (Iif-i) and (Iif-ii) through the amine functionality provided by the side chain of the lysine at position 9 1 - is linked to

[0464] In certain embodiments, when CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the reversible CNP conjugates of formulas (Iif), (Iif-i) and (Iif-ii) through the amine functionality provided by the side chain of the lysine at position 11 1 - is linked to

[0465] In certain embodiments, when CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the reversible CNP conjugates of formulas (Iif), (Iif-i) and (Iif-ii) through the amine functionality provided by the side chain of the lysine at position 15 1 - is linked to

[0466] In certain embodiments, when the CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the reversible CNP conjugates of formulae (Iif), (Iif-i) and (Iif-ii) via an amine functionality provided by the side chain of lysine at position 16 1 -.

[0467] In certain embodiments, when the CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the reversible CNP conjugates of formulae (Iif), (Iif-i) and (Iif-ii) via an amine functionality provided by the side chain of lysine at position 20 1 -.

[0468] In certain embodiments, when the CNP moiety is that of SEQ ID NO: 24, said lysine side chain is part of a ring formed by a disulfide bridge between cysteine residues at positions 22 and 38.

[0469] Thus, in certain embodiments, when the CNP has the sequence of SEQ ID NO: 24, the CNP moiety is linked to -L in the CNP conjugates of formulae (Iif), (Iif-i) and (Iif-ii) via an amine functionality provided by the side chain of lysine at position 26 1 -.

[0470] In certain embodiments, the reversible CNP conjugate has formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 24 and is linked to -L via an amine functionality provided by the side chain of lysine at position 26 1 -.

[0471] In certain embodiments, the reversible CNP conjugate has formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 20 and is linked to -L via an amine functionality provided by the side chain of lysine at position 30 1 -.

[0472] In certain embodiments, the reversible CNP conjugate has the formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 20, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 30 1 - is attached thereto.

[0473] In certain embodiments, the reversible CNP conjugate has the formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 20, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 30 1 - is attached thereto.

[0474] In certain embodiments, the reversible CNP conjugate has the formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 21, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 29 1 - is attached thereto.

[0475] In certain embodiments, the reversible CNP conjugate has the formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 21, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 29 1 - is attached thereto.

[0476] In certain embodiments, the reversible CNP conjugate has the formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 21, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 29 1 - is attached thereto.

[0477] In certain embodiments, the reversible CNP conjugate has the formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 22, and is attached to -L via an amine functional group provided by the side chain of the lysine at position 28 1 - is attached thereto.

[0478] In certain embodiments, the reversible CNP conjugate has formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 22, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 28 1 - thereto.

[0479] In certain embodiments, the reversible CNP conjugate has formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 22, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 28 1 - thereto.

[0480] In certain embodiments, the reversible CNP conjugate has formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 23, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 27 1 - thereto.

[0481] In certain embodiments, the reversible CNP conjugate has formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 23, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 27 1 - thereto.

[0482] In certain embodiments, the reversible CNP conjugate has formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 23, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 27 1 - thereto.

[0483] In certain embodiments, the reversible CNP conjugate has formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 30, and is attached to -L via an amine functionality provided by the side chain of the lysine at position 27 1 - thereto.

[0484] In certain embodiments, the reversible CNP conjugate has the formula (Iif-i), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 30, and is linked to -L via the amine functionality provided by the side chain of the lysine at position 27 1 -.

[0485] In certain embodiments, the reversible CNP conjugate has the formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 30, and is linked to -L via the amine functionality provided by the side chain of the lysine at position 27 1 -.

[0486] In certain embodiments, the reversible CNP conjugate has the formula (Iif-ii), where c1 is about 225, the CNP moiety has the sequence of SEQ ID NO: 24, and is linked to -L via the amine functionality provided by the side chain of the lysine at position 26 1 -.

[0487] The positions of the cysteine and lysine described above may vary depending on the length of the CNP moiety, and those skilled in the art will have no difficulty in identifying the corresponding cysteine and lysine in longer or shorter versions of the CNP moiety. For example, it is understood that some lysines may not be present in shorter CNP moieties. It is further understood that, as a result of site-directed mutagenesis, there may be more lysine residues in the acyclic and / or cyclic portions of the CNP moiety.

[0488] In certain embodiments, the reversible CNP conjugate has the formula (Iif), where c1 is about 225, -D is a CNP moiety having the sequence of SEQ ID NO: 24, and is linked to -L via the amine functionality provided by the side chain of the lysine at position 26 1 -.

[0489] In certain embodiments, the reversible CNP conjugate of the invention has the formula (Iif’):

Chemical Formula

Chemical formula

Chemical formula

[0490] In certain embodiments, each c1 of formula (Iif') is about 225.

[0491] In certain embodiments, the reversible CNP conjugate has formula (Iif-i'):

Chemical formula

Chemical formula

Chemical formula

[0492] In certain embodiments, each c1 of formula (Iif-i') is about 225.

[0493] In certain embodiments, the reversible CNP conjugate has the formula (Iif-ii’): [Chemical formula] (wherein The unmarked dashed line indicates the bond to the nitrogen provided by the side chain of lysine at position 26 of the CNP moiety of SEQ ID NO: 24 by forming an amide bond, The dashed line marked with an asterisk represents the structure: [Chemical formula] represents a bond to -Z having the formula, wherein each Z a is [Chemical formula] and each c1 in this formula is an integer in the range of 200 to 250).

[0494] In certain embodiments, each c1 of formula (Iif-ii’) is about 225.

[0495] In certain embodiments, the reversible CNP conjugate is of the following formula (Iifa). [Chemical formula] wherein The unmarked dashed line indicates the bond to the nitrogen of the -D which is the CNP moiety by forming an amide bond; The dashed line marked with an asterisk represents the following structure: [Chemical formula] represents a bond to -Z having the formula, k is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12; each -Z a is [Chemistry] and each c1 is independently an integer in the range of 200 to 250.

[0496] In certain embodiments, k of formula (Iifa) is selected from the group consisting of 2, 3, 4, 5, 6, and 7.

[0497] In certain embodiments, each c1 of formula (Iifa) is about 225.

[0498] In certain embodiments, the reversible CNP conjugate is of the following formula (Iifa-i). [Chemistry] Wherein The unmarked dashed line indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond; The dashed line marked with an asterisk is the following structure: [Chemistry] indicates the bond to -Z having k is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12; each -Z a is [Chemistry] and each c1 is independently an integer in the range of 200 to 250.

[0499] In certain embodiments, k of formula (Iifa-i) is selected from the group consisting of 2, 3, 4, 5, 6, and 7.

[0500] In certain embodiments, each c1 of formula (Iifa-i) is about 225.

[0501] In certain embodiments, the reversible CNP conjugate has the formula (Iifa-ii) below.

Chemical formula

Chemical formula

Chemical formula

[0502] In certain embodiments, each c1 of formula (Iifa-ii) is about 225.

[0503] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i) and (Iifa-ii) has the sequence of SEQ ID NO: 25.

[0504] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i) and (Iifa-ii) has the sequence of SEQ ID NO: 20.

[0505] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i) and (Iifa-ii) has the sequence of SEQ ID NO: 21.

[0506] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i), and (Iifa-ii) has the sequence of SEQ ID NO: 22.

[0507] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i), and (Iifa-ii) has the sequence of SEQ ID NO: 23.

[0508] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i), and (Iifa-ii) has the sequence of SEQ ID NO: 30.

[0509] In certain embodiments, the CNP moiety of the reversible CNP conjugates of formulas (Iifa), (Iifa-i), and (Iifa-ii) has the sequence of SEQ ID NO: 24.

[0510] In one embodiment, the CNP moiety is attached to -L in the reversible CNP conjugates of formulas (Iifa), (Iifa-i), and (Iifa-ii) via the nitrogen of the N-terminal amine functionality of CNP. 1 - is attached.

[0511] In certain embodiments, the reversible CNP conjugate is of the following formula (Iifa′).

Chemical formula

Chemical formula

[0512] In certain embodiments, k of formula (Iifa') is selected from the group consisting of 2, 3, 4, 5, 6, and 7.

[0513] In certain embodiments, each c1 of formula (Iifa') is about 225.

[0514] In certain embodiments, the reversible CNP conjugate is of the following formula (Iifa-i'). [Chemical formula] wherein The unmarked dashed line indicates a bond to the nitrogen provided by the side chain of the lysine at position 26 of the CNP moiety of SEQ ID NO: 24 by forming an amide bond; The dashed line marked with an asterisk is the following structure: [Chemical formula] represents a bond to -Z having k is selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, and 12; each Z a is [Chemical formula] and each c1 is independently an integer in the range of 200 to 250.

[0515] In certain embodiments, k of formula (Iifa-i') is selected from the group consisting of 2, 3, 4, 5, 6, and 7.

[0516] In certain embodiments, each c1 of formula (Iifa-i') is about 225.

[0517] In certain embodiments, the reversible CNP conjugate is of the following formula (Iifa-ii′).

Chemical formula

Chemical formula

Chemical formula

[0518] In certain embodiments, k of formula (Iifa-ii′) is selected from the group consisting of 2, 3, 4, 5, 6 and 7.

[0519] In certain embodiments, each c1 of formula (Iifa-ii′) is about 225.

[0520] In certain embodiments, the CNP compound is a CNP agonist selected from the group consisting of small molecules, natural products, oligonucleotides, polypeptides, and proteins.

[0521] In certain embodiments, the CNP agonist includes small molecules. In certain embodiments, the CNP agonist is a small molecule.

[0522] In certain embodiments, the CNP agonist includes natural products. In certain embodiments, the CNP agonist is a natural product.

[0523] In certain embodiments, the CNP agonist comprises an oligonucleotide. In certain embodiments, such oligonucleotides are selected from the group consisting of antisense oligonucleotides, aptamers, RNAi, and siRNA. In certain embodiments, the CNP agonist is an oligonucleotide selected from the group consisting of antisense oligonucleotides, aptamers, RNAi, and siRNA.

[0524] In certain embodiments, the CNP agonist comprises a protein. In certain embodiments, the CNP agonist is a protein.

[0525] In certain embodiments, the CNP agonist comprises a polypeptide. In certain embodiments, the CNP agonist is a polypeptide.

[0526] In certain embodiments, the CNP agonist comprises a CNP molecule or moiety. In certain embodiments, the CNP agonist is CNP.

[0527] In certain embodiments, the CNP agonist comprises a CNP molecule or moiety having the sequence of SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30.

[0528] In certain embodiments, the CNP agonist is a CNP having the sequence of SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. In certain embodiments, the CNP agonist is a CNP having the sequence of SEQ ID NO: 24. In certain embodiments, the CNP agonist is a CNP having the sequence of SEQ ID NO: 25. In certain embodiments, the CNP agonist is a CNP having the sequence of SEQ ID NO: 30.

[0529] In certain embodiments, the liquid pharmaceutical formulation of the present invention may contain one or more excipients such as, for example, stabilizers, adsorption inhibitors, cryoprotectants, and other auxiliary agents. It is understood that one excipient may have multiple functions such as dual, triple, etc.

[0530] In certain embodiments, the liquid pharmaceutical formulation of the present invention may include a stabilizer selected from the group consisting of stabilizers such as alanine; arginine; aspartic acid; glycine; histidine; lysine; proline; sugars such as glucose, sucrose, trehalose; polyols such as glycerol, mannitol, sorbitol; salts such as potassium phosphate, sodium sulfate; chelating agents such as EDTA, hexaphosphate; ligands such as divalent metal ions; other salts or organic molecules such as phenol derivatives; oligomers or polymers such as cyclodextrin, dextran, dendrimer, PEG, PVP, protamine, HAS.

[0531] In certain embodiments, the liquid pharmaceutical formulation of the present invention may include an adsorption inhibitor selected from the group consisting of ionic or non-ionic surfactants or other proteins or soluble polymers used to competitively coat or adsorb onto the inner surface of the formulation or the container of the formulation, such as poloxamer (Pluronic™ F-68), PEG dodecyl ether (Brij® 35), dextran, polyethylene glycol, PEG-polyhistidine, BSA, HAS, gelatin. The type and concentration of the excipient are selected according to the effect to be avoided, but usually, at a concentration higher than CMC, a monolayer of surfactant is formed at the interface.

[0532] In certain embodiments, the liquid pharmaceutical formulation of the present invention can include a cryoprotectant selected from the group consisting of cryoprotectants such as saccharides, polyhydric alcohols, surfactants, amino acids, non-aqueous solvents, and peptides. During lyophilization or spray drying, the cryoprotectant can counteract the destabilizing effects caused by hydrogen bond disruption and water removal. Trehalose is particularly efficient in reducing moisture-induced aggregation and also improves the thermal stability that may be caused by the exposure of the hydrophobic groups of the compound to water. Mannitol and sucrose can also be used alone as lyoprotectants / cryoprotectants or in combination with each other, and in that case, it is known that the higher the ratio of mannitol:sucrose, the higher the physical stability of the compound in the liquid pharmaceutical formulation. Mannitol can also be combined with trehalose. Trehalose can also be combined with sorbitol, or sorbitol can be used alone as a stabilizer. Starches or starch derivatives can also be used.

[0533] In certain embodiments, the liquid pharmaceutical formulation of the present invention can include additional excipients selected from the group consisting of wetting agents, viscosity modifiers, and antibiotics.

[0534] The applicant has found that within the liquid pharmaceutical formulation of the present invention, the reversible linkage between CNP and the carrier is stable, eliminating the need for lyophilization and regeneration from the lyophilized product. However, if desired, the liquid pharmaceutical formulation of the present invention can be dried by lyophilization or the like to form a dried product such as a lyophilized pharmaceutical formulation.

[0535] In certain embodiments, the liquid pharmaceutical formulation of the present invention includes a CNP conjugate, acetic acid, mannitol, m-cresol, and methionine.

[0536] In certain embodiments, the liquid pharmaceutical formulation of the present invention includes a CNP conjugate, acetic acid, trehalose, m-cresol, and methionine.

[0537] Since there is a large difference in the weight range of patients, for example, from 3 to 80 kg, in certain embodiments, it is beneficial to have liquid pharmaceutical formulations with different strengths of CNP conjugate, that is, different concentrations, such as high-strength, medium-strength, and low-strength liquid pharmaceutical formulations. Thus, in one embodiment, the liquid pharmaceutical formulation is provided at a plurality of concentrations, such as two different concentrations, three different concentrations, four different concentrations, five different concentrations, six different concentrations, seven different concentrations, eight different concentrations, nine different concentrations, ten different concentrations, eleven different concentrations, twelve different concentrations, and so on.

[0538] In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for at least 6 months.

[0539] In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for at least 12 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for at least 24 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for at least 36 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for at least 48 months.

[0540] In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for 6 - 48 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation that is stored at 2 - 8°C for 6 - 36 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for 6 - 24 months. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 2 - 8°C for 6 - 12 months.

[0541] In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 25°C for 1 month. In certain embodiments, the present invention provides a liquid pharmaceutical formulation for storage at 30°C for 1 month.

[0542] The above liquid pharmaceutical preparation is a liquid pharmaceutical preparation that is stable for at least 6 months, for example, at least 7 months, for example, at least 8 months, for example, at least 9 months, for example, at least 10 months, for example, at least 11 months, for example, at least 12 months. In certain embodiments, the liquid pharmaceutical preparation is stable for at least 14 months, for example, at least 16 months, for example, at least 18 months, for example, at least 20 months, for example, at least 22 months, for example, at least 24 months, for example, at least 36 months, for example, at least 48 months.

[0543] In certain embodiments, the liquid pharmaceutical preparation is stable for at least 36 months when stored at 2°C to 8°C. In certain embodiments, the liquid pharmaceutical preparation is stable for at least 36 months when stored at 2°C. In certain embodiments, the liquid pharmaceutical preparation is stable for at least 6 months when stored at 5°C. In certain embodiments, the liquid pharmaceutical preparation is stable for at least 2 weeks when stored at 30°C.

[0544] In certain embodiments, the above liquid pharmaceutical preparation can be stored at a temperature of -80°C to 30°C, for example, -20°C to 25°C, for example, -15°C to 25°C, for example, -10°C to 25°C, for example, -5°C to 25°C, for example, 0°C to 25°C, for example, 2°C to 8°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 2°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 4°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 5°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 8°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 10°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 16°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 20°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 25°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 30°C. In certain embodiments, the liquid pharmaceutical preparation is stored at 40°C.

[0545] The liquid pharmaceutical preparation of the present invention contains at least about 80%, such as at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the peptide in its natural form (CNP or CNP moiety) after storage for at least 3 months, 6 months, 12 months, or at least 24 months. The storage is carried out at 5°C, 8°C, 15°C, 20°C, or 25°C, and the purity can be determined by any method known to those skilled in the art, such as SE-HPLC or RP-HPLC.

[0546] In certain embodiments, the liquid pharmaceutical preparation contains about 90%, 95%, 96%, 97%, or 99% of the peptide in its natural form (CNP or CNP moiety) after storage at 5°C for 3 or 6 months, as measured by, for example, SE-HPLC or RP-HPLC.

[0547] In certain embodiments, the liquid pharmaceutical preparation contains about 80%, such as at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the peptide in its natural form (CNP or CNP moiety) after storage at 5°C for 24 months, as measured by, for example, SE-HPLC or RP-HPLC.

[0548] In certain embodiments, the liquid pharmaceutical preparation contains about 80%, such as at least 85%, 90%, or 95% of the peptide in its natural form (CNP or CNP moiety) after storage at 5°C for 6 months, as measured by, for example, SE-HPLC or RP-HPLC.

[0549] The present invention (i) mixing a CNP compound with at least a buffer, an isotonic agent, a preservative, and optionally an antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture from step (ii) or (iii) corresponding to the desired number of administrations to a container; (v) sealing the container A method for manufacturing a liquid pharmaceutical preparation according to the present invention, comprising The order of step (ii) and step (iii) includes a method that may optionally be reversed.

[0550] In certain embodiments, step (ii) and step (iii) are not reversed.

[0551] In certain embodiments, the CNP compound in step (i) is mixed with a buffer, an isotonic agent, a preservative, and optionally an antioxidant.

[0552] In certain embodiments, the method for manufacturing a liquid pharmaceutical preparation according to the present invention (i) mixing a CNP compound with at least acetic acid, mannitol, m-cresol, and optionally an antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture from step (ii) or (iii) corresponding to the desired number of administrations to a container; (v) sealing the container comprises The order of step (ii) and step (iii) may optionally be reversed.

[0553] In certain embodiments, step (ii) and step (iii) are not reversed.

[0554] In certain embodiments, the CNP compound in step (i) is mixed with acetic acid, mannitol, m-cresol, and optionally an antioxidant.

[0555] In certain embodiments, the method for manufacturing a liquid pharmaceutical preparation according to the present invention (i) mixing a CNP conjugate with at least acetic acid, mannitol, m-cresol, and optionally methionine to obtain a CNP conjugate in which the CNP moiety is present at 0.1 - 20 mg / mL, acetic acid: 0.3 - 3 mg / mL, D-mannitol: 10 - 200 mg / mL, m-Cresol: 1 to 10 mg / mL obtaining a formulation containing; (ii) adjusting the pH of the mixture from step (i) to a pH in the range of pH 3.5 to pH 5.5 using NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture from step (ii) or (iii) corresponding to the desired number of administrations to a container; (v) sealing the container including, the order of step (ii) and step (iii) may optionally be reversed.

[0556] In certain embodiments, the method for producing a liquid pharmaceutical formulation according to the present invention is (i) mixing a CNP conjugate with at least acetic acid, trehalose, m-cresol, and optionally methionine to obtain a formulation containing a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, trehalose dihydrate: 10 to 200 mg / mL, m-cresol: 1 to 10 mg / mL obtaining a formulation containing; (ii) adjusting the pH of the mixture from step (i) to a pH in the range of pH 3.5 to pH 5.5 using NaOH and HCl; (iii) optionally, filtering the mixture from step (ii); (iv) transferring an amount of the mixture from step (ii) or (iii) corresponding to the desired number of administrations to a container; (v) sealing the container including, the order of step (ii) and step (iii) may optionally be reversed.

[0557] In certain embodiments, steps (ii) and (iii) are not reversed.

[0558] Another aspect of the present invention relates to a container for a liquid pharmaceutical formulation of the present invention. The container can be suitable for both storage and dispensing of the formulation. In certain embodiments, the container contains the liquid pharmaceutical formulation of the present invention. When using such a container, the kit may further include a dispenser, such as a syringe or a container with volume markings.

[0559] In certain embodiments, the container is made of glass. In certain embodiments, the inner surface of the container is coated with an inert material. In certain embodiments, the inert material is silicon.

[0560] In certain embodiments, the container may be selected from the group consisting of vials, syringes (such as dual-chamber syringes), ampoules, and cartridges (such as dual-chamber cartridges).

[0561] In certain embodiments, the cartridge is used with a pen-type injector, such as an auto-injector.

[0562] In certain embodiments, the liquid pharmaceutical formulation of the present invention is provided as a single dose, which means that the container containing the liquid pharmaceutical formulation contains one therapeutic dose. When the liquid pharmaceutical formulation is provided as a single dose, it is obvious to those skilled in the art that there is no preservative in the formulation.

[0563] In certain embodiments, the liquid pharmaceutical formulation contains multiple doses, which means that the container containing the liquid pharmaceutical formulation contains multiple therapeutic doses.

[0564] In certain embodiments, the multiple-dose liquid pharmaceutical formulation contains at least 2 doses, such as at least 4 doses, such as at least 6 doses, such as at least 8 doses, such as at least 10 doses, such as at least 12 doses of a CNP agonist or CNP, and in certain embodiments, contains at least 14 doses.

[0565] In certain embodiments, the multi-dose liquid pharmaceutical formulation comprises at least 2, 4, 6, 8, 10, 12, or 14 doses of a CNP agonist or CNP.

[0566] Accordingly, in another aspect of the invention, the liquid pharmaceutical formulation is provided as a multi-dose formulation.

[0567] Single-dose formulations containing a CNP compound, particularly a CNP conjugate, have a volume of 4 mL or less, such as from about 0.03 to about 1.1 mL. In certain embodiments, the volume is about 0.03 mL. In certain embodiments, the volume is about 0.05 mL. In certain embodiments, the volume is about 0.1 mL. In certain embodiments, the volume is about 0.3 mL. In certain embodiments, the volume is about 0.5 mL. In certain embodiments, the volume is about 0.8 mL. In certain embodiments, the volume is about 1 mL. In certain embodiments, the volume is about 1.5 mL. In certain embodiments, the volume is about 2 mL. In certain embodiments, the volume is about 3 mL. In certain embodiments, the volume is about 4 mL.

[0568] In certain embodiments, the CNP compound, particularly the CNP conjugate, is included in the formulation in an amount sufficient to provide a therapeutically effective amount of CNP over at least 3 days, such as at least 4 days, such as at least 5 days, such as at least 6 days, for a single administration.

[0569] In certain embodiments, the CNP compound, particularly the CNP conjugate, is included in the formulation in an amount sufficient to provide a therapeutically effective amount of CNP over a period of one week.

[0570] The liquid pharmaceutical preparation of the present invention can be administered, for example, via topical, enteral or parenteral administration, and by external application, injection or infusion, such as intra-articular, periaricular, intradermal, subcutaneous, intramuscular, intravenous, intraosseous, intraperitoneal, intrathecal, intracapsular, intraorbital, intravitreal, intratympanic, intravesical, intracardiac, transtracheal, subepidermal, subcapsular, subdural, intraspinal, intraventricular, intrasternal injection, infusion, intranasal, oral, transpulmonary and transdermal administration, direct delivery to the brain via an implantable device (e.g., an Ommaya reservoir) capable of delivering the present invention to brain tissue or cerebrospinal fluid, direct intraventricular injection or infusion, injection or infusion into the brain or brain-related regions, injection into the subchoroidal space, retrobulbar injection and eye drops, preferably by the method of subcutaneous injection.

[0571] In certain embodiments, the liquid pharmaceutical preparation of the present invention is administered by subcutaneous injection.

[0572] In certain embodiments, the liquid pharmaceutical preparation of the present invention is administered by subcutaneous injection using a syringe and needle, or a pen-type syringe such as an autoinjector.

[0573] In certain embodiments, the liquid pharmaceutical preparation of the present invention is administered by subcutaneous injection using a syringe and needle.

[0574] In certain embodiments, the liquid pharmaceutical preparation of the present invention is administered by subcutaneous injection using a pen-type syringe.

[0575] In certain embodiments, the liquid pharmaceutical preparation of the present invention is administered by subcutaneous injection using an autoinjector.

[0576] The period between two consecutive subcutaneous administrations, i.e., the dosing interval, is in certain embodiments at least 12 hours, 24 hours, 36 hours, 48 hours, 60 hours, 72 hours, 84 hours, 96 hours, 108 hours, 120 hours, 132 hours, 144 hours, 156 hours, once a week, once every two weeks, once every three weeks, or once every four weeks.

[0577] In certain embodiments, the period between two consecutive subcutaneous administrations is 12 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 24 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 48 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 72 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 96 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 120 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is 144 hours. In certain embodiments, the period between two consecutive subcutaneous administrations is one week.

[0578] Another aspect of the invention is the liquid pharmaceutical formulation of the invention for use as a medicament.

[0579] In another aspect, the invention relates to the liquid pharmaceutical formulation of the invention for use in the treatment, suppression, delay or prevention of one or more diseases that can be treated, suppressed, delayed or prevented by CNP agonists, particularly CNP.

[0580] In certain embodiments, the invention relates to the liquid pharmaceutical formulation of the invention for use in the treatment of one or more diseases that can be treated with CNP agonists, particularly CNP.

[0581] A further aspect of the invention is a method of treating, suppressing, delaying or preventing in a patient one or more diseases treatable by CNP agonists, particularly CNP, the method comprising administering to the patient a therapeutically effective amount of the liquid pharmaceutical formulation of the invention.

[0582] In certain embodiments, the patient is an adult. In certain embodiments, the patient is a pediatric patient.

[0583] The liquid pharmaceutical preparation of the present invention can be administered for at least 6 months, 1 year, 5 years, 10 years, until the patient reaches 18 years of age, until the epiphyseal line of the patient closes, or indefinitely. In certain embodiments, the liquid pharmaceutical preparation can be administered until the patient reaches 18 years of age. In certain embodiments, the liquid pharmaceutical preparation can be administered until the epiphyseal line of the patient closes.

[0584] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented with a CNP agonist, particularly CNP, are bone-related diseases such as skeletal dysplasia; cancer; autoimmune diseases; fibrotic diseases; inflammatory diseases; central nervous system diseases such as neurodegenerative diseases; infectious diseases; lung diseases; heart and vascular diseases; metabolic diseases; and ophthalmic diseases, selected from the group consisting of.

[0585] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented by CNP-acting agents, particularly CNP, are achondroplasia, hypochondroplasia, short stature, dwarfism, osteochondrodysplasia, lethal osteogenesis imperfecta, osteogenesis imperfecta, achondrogenesis, stippled chondrodysplasia, homozygous achondroplasia, campomelic dysplasia, congenital lethal hypophosphatasia, perinatal lethal osteogenesis imperfecta, short rib polydactyly syndrome, rhizomelic chondrodysplasia punctata, Jansen metaphyseal chondrodysplasia, congenital spondyloepiphyseal dysplasia, osteogenesis imperfecta, torsional osteodysplasia, congenital femoral shortening, Langer type intermediate limb dysplasia, Nievergelt type intermediate limb dysplasia, Robinow syndrome, Reinhardt syndrome, acroosteolysis, distal osteolysis, Knist bone dysplasia, fibrochondrogenesis, Roberts syndrome, distal intermediate limb dysplasia, micromelia, Morquio syndrome, Knist syndrome, dysplastic osteodysplasia, spondyloepiphyseal metaphyseal dysplasia, neurofibromatosis, Leopard syndrome, Noonan syndrome, hereditary gingival fibromatosis, neurofibromatosis type 1, Legius syndrome, heart-face-skin syndrome, Costello syndrome, SHOX deficiency, idiopathic short stature, growth hormone deficiency, osteoarthritis, cleidocranial dysplasia, craniosynostosis (e.g., Muenke syndrome, Crouzon syndrome, Apert syndrome, Jackson-Weiss syndrome, Pfeiffer syndrome, or Crouzonodermoskeletal syndrome), dactyly, brachydactyly, camptodactyly, polydactyly, syndactyly, segmental dysplasia, enchondroma, fibrous dysplasia, hereditary multiple exostoses, hypophosphatemic rickets, Jaffe-Lichtenstein syndrome, Marfan syndrome, McCune-Albright syndrome, osteopetrosis, osteopathia striata, hemorrhagic shock, hypertension, restenosis, arteriosclerosis, acute decompensated heart failure, congestive heart failure, cardiac edema, renal edema, hepatic edema, acute renal failure, chronic renal failure, glaucoma, ocular hypertension, multiple myeloma, myeloproliferative syndrome, leukemia, plasmacytic leukemia, lymphoma, glioblastoma, prostate cancer, bladder cancer, breast cancer, growth delay, cranial deformity, orthodontic defects, cervical spinal cord compression, spinal stenosis, hydrocephalus, hearing loss due to chronic otitis, cardiovascular disease, neurological disease and obesity, selected from the group consisting of.

[0586] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented by CNP are achondroplasia, such as homozygous achondroplasia, hypochondroplasia, short stature, dwarfism, osteochondrodysplasia, lethal osteogenesis imperfecta, osteogenesis imperfecta, chondrogenesis imperfecta, stippled chondrodysplasia, campomelic dysplasia, congenital lethal hypophosphatasia, perinatal lethal osteogenesis imperfecta, short rib polydactyly syndrome, rhizomelic chondrodysplasia punctata (proximal segment shortening type of chondrodysplasia punctata), Jansen metaphyseal dysplasia, congenital spondyloepiphyseal dysplasia, osteogenesis imperfecta, torsional osteodysplasia, congenital femoral shortening, Langer mesomelic dysplasia, Nievergelt mesomelic dysplasia, Robinow syndrome, Reinhardt syndrome, acroosteolysis, peripheral osteolysis, Kniest dysplasia, fibrocartilaginous dysplasia, Roberts syndrome, distal mesomelic dysplasia, micromelia, Morquio syndrome, Kniest syndrome, dysplastic osteodysplasia, spondyloepiphyseal metaphyseal dysplasia, neurofibromatosis, Legius syndrome, Leopard syndrome, Noonan syndrome, hereditary gingival fibromatosis, neurofibromatosis type 1, Legius syndrome, heart-face-skin syndrome, Costello syndrome, SHOX deficiency, idiopathic short stature, growth hormone deficiency, osteoarthritis, cleidocranial dysplasia, craniosynostosis (e.g., Muenke syndrome, Crouzon syndrome, Apert syndrome, Jackson-Weiss syndrome, Pfeiffer syndrome, or Crouzonodermoskeletal syndrome), dactyly, brachydactyly, clinodactyly, polydactyly, syndactyly, segmental dysplasia, enchondroma, fibrous dysplasia, hereditary multiple exostoses, hypophosphatemic rickets, Jaffe-Lichtenstein syndrome, Marfan syndrome, McCune-Albright syndrome, osteopetrosis, osteopathia striata, hemorrhagic shock, hypertension, restenosis, arteriosclerosis, acute decompensated heart failure, congestive heart failure, cardiac edema, renal edema, hepatic edema, acute renal failure, chronic renal failure, glaucoma, ocular hypertension, multiple myeloma, myeloproliferative syndrome, leukemia, plasmacytic leukemia, lymphoma, glioblastoma, prostate cancer, bladder cancer, breast cancer, growth delay, craniofacial deformity, orthodonticdefects), cervical spinal cord compression, spinal stenosis, hydrocephalus, hearing loss due to chronic otitis media, obesity, disorders involving abnormal RAS-mitogen-activated protein kinase signaling, pulmonary hypertension, vascular disorders, endothelial dysfunction, cirrhosis, hepatic ascites, hepatic fibrosis, hepatorenal syndrome, asthma, pulmonary fibrosis, chronic kidney disease, cardiorenal syndrome, dyspnea, and lysosomal storage diseases, such as mucopolysaccharidosis, selected from the group consisting of.

[0587] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented by a CNP-acting agent, particularly CNP, are arrhythmias, such as cardiac arrhythmias or sinus arrhythmias; atrial fibrillation; atrial flutter; bradycardia; Brugada syndrome; premature contractions; cardiac oscillation; heart block; QT prolongation syndrome; parasystole; early excitation syndrome; tachycardia; ventricular fibrillation; ventricular flutter; cardiac conduction disorders; low cardiac output; cardiac hypertrophy; cardiomyopathy, such as alcoholic, dilated, hypertrophic, restrictive, diabetic or Chagas cardiomyopathy; arrhythmogenic right ventricular dysplasia; endocardial fibroelastosis; endomyocardial fibrosis; glycogenosis type IIb; Kearns-Sayre syndrome; myocardial reperfusion injury; myocarditis; sarcoglycan abnormality; endocarditis, such as bacterial or non-infectious endocarditis; cardiac arrest; sudden cardiac death; out-of-hospital cardiac arrest; cardiorenal syndrome; paroxysmal dyspnea; cardiac edema, heart failure, such as diastolic or systolic heart failure; valvular heart disease; aortic valve insufficiency; aortic valve stenosis; cardiac valve prolapse; mitral valve insufficiency; mitral valve stenosis; pulmonary atresia; pulmonary valve insufficiency; pulmonary valve stenosis; tricuspid atresia; tricuspid valve insufficiency; tricuspid valve stenosis; myocardial ischemia; acute coronary syndrome; angina pectoris; coronary artery disease; Cornu syndrome; myocardial infarction; pulmonary heart disease; ventricular dysfunction, such as left ventricular or right ventricular dysfunction; ventricular outflow tract obstruction; aortic valve stenosis, pulmonary valve stenosis; hypertension; atherosclerosis; restenosis; severe lower limb ischemia; peripheral arterial disease; ischemia, such as ischemia-reperfusion injury or ischemic injury; abnormal fluid retention in cardiac edema and myocardial edema, one or more cardiovascular diseases selected from the group consisting of.

[0588] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented by a CNP agonist, particularly CNP, are ischemic heart diseases such as myocardial infarction; congestive heart failure; arrhythmia and atherosclerosis, and are selected from the group consisting thereof.

[0589] In certain embodiments, the one or more diseases that can be treated, suppressed, delayed or prevented by a CNP agonist, particularly CNP, are cerebral ischemia such as ischemic hypoxia; cerebral infarction; transient ischemic attack; vertebrobasilar insufficiency; cerebrovascular disorder; stroke; intracranial hemorrhage; corneal neovascularization; corneal transplantation; graft-versus-host disease; graft rejection; glaucoma such as closed angle, neovascularization, open angle or low-tension glaucoma; ischemic optic neuropathy; central serous chorioretinopathy; retinopathy such as diabetic or hypertensive retinopathy; retinal degeneration; macular degeneration; geographic atrophy; macular edema; Stargardt disease; vitreomacular dystrophy; exudative macular degeneration; retinal detachment; retinal detachment; retinal perforation; retinal hemorrhage; retinal neovascularization; retinal vein occlusion; retinal artery occlusion; retinopathy of prematurity; and one or more central nervous system diseases selected from the group consisting of proliferative vitreoretinopathy.

[0590] In certain embodiments, the one or more diseases are selected from the group consisting of hypophosphatasia, chondrodysplasia, Menkes syndrome, hypertension, osteogenesis imperfecta and achondroplasia.

[0591] In certain embodiments, the one or more diseases that can be treated by a CNP agonist, particularly CNP, are hypophosphatasia. In certain embodiments, the one or more diseases that can be treated by CNP are chondrodysplasia. In certain embodiments, the one or more diseases that can be treated by CNP are Menkes syndrome. In certain embodiments, the one or more diseases that can be treated by CNP are hypertension. In certain embodiments, the one or more diseases that can be treated by CNP are osteogenesis imperfecta. In certain embodiments, the one or more diseases that can be treated by CNP are achondroplasia.

[0592] The present invention is further illustrated by the following non-limiting items. 1. A liquid pharmaceutical preparation comprising a CNP compound, a buffer, an isotonic agent, a preservative, and optionally an antioxidant. 2. The liquid pharmaceutical preparation according to item 1, for storage at 2-8°C for at least 6 months. 3. The liquid pharmaceutical preparation according to item 1 or 2, for storage at 2-8°C for 6-48 months. 4. The liquid pharmaceutical preparation according to any one of items 1-3, for storage at 2-8°C for 6-36 months. 5. The liquid pharmaceutical preparation according to any one of items 1-4, for storage at 5°C for 6-36 months. 6. The liquid pharmaceutical preparation according to any one of items 1-5, wherein the CNP compound is a CNP conjugate. 7. The liquid pharmaceutical preparation according to any one of items 1-6, wherein the buffer is selected from the group consisting of acetic acid, succinic acid, citric acid, lactic acid, glutamic acid, fumaric acid, aspartic acid, glutaric acid, phosphoric acid, histidine, gluconic acid, tartaric acid, malic acid, and tris (tris (hydroxymethyl) aminomethane). 8. The liquid pharmaceutical preparation according to any one of items 1-7, wherein the buffer is acetic acid. 9. The liquid pharmaceutical preparation according to any one of items 1-8, wherein the isotonic agent is selected from the group consisting of mannitol, trehalose, sucrose, raffinose, gelatin, lactose, dibasic calcium phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, dextrose, dextran, propylene glycol, and glycerol. 10. The liquid pharmaceutical preparation according to any one of items 1-9, wherein the isotonic agent is mannitol or trehalose. 11. The liquid pharmaceutical preparation according to any one of items 1 to 10, wherein the preservative is selected from the group consisting of m-cresol, benzyl alcohol, benzoic acid, phenol, methyl paraben, ethyl paraben, propyl paraben, butyl paraben, potassium sorbate, chlorobutanol, benzyl alcohol, phenylmercuric nitrate, thimerosal, sorbic acid, potassium sorbate, chlorocresol, benzalkonium chloride, 2-ethoxyethanol, chlorhexidine, chlorobutanol, phenylethyl alcohol, and phenylmercuric acetate. 12. The liquid pharmaceutical preparation according to any one of items 1 to 11, wherein the preservative is m-cresol or phenol. 13. The liquid pharmaceutical preparation according to any one of items 1 to 12, wherein the pharmaceutical preparation contains an antioxidant selected from the group consisting of methionine, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, propyl gallate, ascorbic acid, sodium bisulfite, ethylenediaminetetraacetic acid (EDTA), cysteine, glutathione, monothioglycerol, poly(ethyleneimine), vitamin E, ectoine, and morin. 14. The liquid pharmaceutical preparation according to any one of items 1 to 13, wherein the pH of the liquid preparation is about pH 3.5 to about pH 5.5. 15. The liquid pharmaceutical preparation according to any one of items 1 to 14, wherein the pH of the liquid preparation is about 4.5. 16. The liquid pharmaceutical preparation according to any one of items 1 to 15, wherein the liquid preparation further contains a pH adjuster. 17. The liquid pharmaceutical preparation according to item 16, wherein the pH adjuster is selected from the group consisting of sodium hydroxide, sodium acetate, tris(tris(hydroxymethyl)aminomethane), potassium hydroxide, lysine, and mixtures thereof. 18. The liquid pharmaceutical preparation according to any one of items 1 to 17, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, an isotonic agent, a preservative, and optionally an antioxidant. 19. The liquid pharmaceutical preparation according to any one of items 1 to 18, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, trehalose dihydrate, a preservative, and optionally an antioxidant. 20. The liquid pharmaceutical preparation according to any one of items 1 to 19, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, trehalose dihydrate, m-cresol, and optionally an antioxidant. 21. The liquid pharmaceutical preparation according to any one of items 1 to 20, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, trehalose dihydrate, m-cresol, and methionine. 22. The liquid pharmaceutical preparation according to any one of items 1 to 17, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, mannitol, a preservative, and optionally an antioxidant. 23. The liquid pharmaceutical preparation according to any one of items 1 to 17 or 22, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, mannitol, m-cresol, and optionally an antioxidant. 24. The liquid pharmaceutical preparation according to any one of items 1 to 17, 22, or 23, wherein the liquid pharmaceutical preparation contains a CNP compound, acetic acid, mannitol, m-cresol, and methionine. 25. The liquid pharmaceutical preparation according to any one of items 1 to 24, wherein the CNP compound is a reversible CNP conjugate containing a CNP moiety covalently and reversibly bound to a carrier moiety. 26. The liquid pharmaceutical preparation according to item 25, wherein the CNP moiety has the sequence of SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. 27. The liquid pharmaceutical preparation according to item 25 or 26, wherein the CNP moiety has the sequence of SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25, or SEQ ID NO: 30. 28. The liquid pharmaceutical preparation according to any one of items 25 to 27, wherein the CNP moiety has the sequence of SEQ ID NO: 24. 29. The liquid pharmaceutical preparation according to any one of items 25 to 28, wherein the carrier moiety is a polymer moiety. 30. The liquid pharmaceutical preparation according to any one of items 1 to 29, wherein the CNP compound is a reversible CNP conjugate of the following formula (Ia). [Chemical formula] [In the formula, -D is a CNP moiety; -L 1 - is a reversible linker moiety; -L 2 - is a single chemical bond or a spacer moiety; -Z is a carrier moiety; x is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, and 16.] 31. The liquid pharmaceutical preparation according to item 30, wherein x in formula (Ia) is 1. 32. The liquid pharmaceutical preparation according to item 30 or 31, wherein -Z is a branched polymer moiety. 33. The liquid pharmaceutical preparation according to any one of items 30 to 32, wherein -Z is a branched PEG-based polymer moiety. 34. The liquid pharmaceutical preparation, a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, D-mannitol: 10 to 200 mg / mL, m-cresol: 1 to 10 mg / mL comprising the same, according to any one of items 6 to 18 or 22 to 33. 35. The liquid pharmaceutical preparation, a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, trehalose dihydrate: 10 to 200 mg / mL m-cresol: 1 to 10 mg / mL comprising the same, according to any one of items 6 to 21 or 25 to 33. 36. The said moiety -L 1- is the one of the following formula (II), the liquid pharmaceutical preparation according to any one of items 30 to 35.

Chemical formula

Chemical formula

Chemical formula

Examples

[0593] Materials and Methods Peptides: CNP-38 TFA salt, CNP-27 TFA salt, CNP-53 acetate, PG-CNP-37 TFA salt were obtained from Caslo, Lyngby, Denmark. The peptide mapping reference peptide was obtained from Wuxi, China. Reagents: TFA (for sequencing, Thermo), water (for ULC / MS, Biosolve and hypotonic, sterilized, pyrogen-free, Carl Roth), acetonitrile (for ULC / MS, Biosolve), poloxamer 188 (for cell culture, Applichem), Tween-20 (Pharm. Eur, Carl Roth), thermolysin (Promega), NaH2PO4·H2O (ACS>98%, Sigma), 4M NaOH (Carl Roth), 1M NaOH (Reag. Ph. Eur., Carl Roth), 1M HCl (Reag. Ph. Eur., Carl Roth), NaCl (Pharm. Eur. >99%, Carl Roth), phosphoric acid (for ACS, >85%, Merck), CaCl2·2 hydrate (for ACS>99%), trehalose·2 hydrate (for USP, Pfanstiehl), succinic acid (for analysis, ACS, Applichem), acetic acid (ROTIPURAN 100%, p,a,, Carl Roth and Ph Eur, BP,, JP, USP, ACS, Merck), metacresol (for parenteral use according to Ph. Eur. / USP, Hedinger), phenol (puriss. p,a,, ACS reagent, reag. Ph. Eur., VWR), mannitol (Ph Eur, BP, USP, JP, Merck and Ph. Eur., USP, BP., Carl Roth), citric acid (ACS reagent, >99.5%, Sigma), citric acid·1 hydrate (Ph Eur, BP, ChP, JP, USP, E330, FCC), sodium citrate·2 hydrate (≥99%, Ph. Eur., Carl Roth), EDTA·2 sodium salt·2 hydrate (suitable for electrophoresis and molecular biology, 99.0 - 101% [titration], Sigma), methionine (Ph. Eur., USP, AppliChem), tris (certified by BioPerformance, conforming to EP and USP test specifications, suitable for cell culture, >99.9% [titration], Sigma), polysorbate 80 (tested according to Ph. Eur., Sigma Aldrich).

[0594] Preparation of CNP-38 formulation (F1 - F23) The necessary excipients were placed in a volumetric flask and filled with water to 90% of the desired volume to prepare the stock solution of excipients. The pH value of the stock solution was adjusted using 1M Tris solution or 4M NaOH solution as necessary and adjusted to the desired volume with water. The stock solution was filtered through a 0.2 μm PVDF filter. Phenol and m-cresol were added to the formulation as necessary. Next, the required amount of CNP-38 was dissolved in the required amount of the desired stock solution of excipients, the pH was checked, and adjusted with 1.0M Tris or 0.1M NaOH as necessary. The formulation was filtered through a 0.22 μm PVDF filter and aliquoted for incubation.

[0595] Preparation of CNP formulation (F33-45) The necessary excipients were placed in a volumetric flask and filled with water to 90% of the desired volume to prepare the stock solution of all excipients except phenol and m-cresol. The pH of the stock solution was adjusted using 1M NaOH solution as necessary and adjusted to the desired volume with water. The stock solution was filtered through a 0.2 μm PVDF filter. Stock solutions of phenol (1.75%) and m-cresol (1.05%) were prepared by adding preservatives to each stock solution of excipients after filtration. Next, the required amount of CNP was dissolved in the desired stock solution of excipients, the pH was checked, and adjusted with 1M NaOH as necessary. The formulation was filtered through a 0.22 μm PVDF filter and each preservative stock solution was added. The formulation was aliquoted for incubation.

[0596] Preparation of Compound (1) formulation (F24-F32) The solution of compound (1) was buffer-exchanged using diafiltration with the desired buffer solution and then concentrated by ultrafiltration. Diafiltration and ultrafiltration were carried out by TFF using an Akta Flux S TFF system and a Pellicon XL 50 cassette (Biomax 10 kDa). After ultrafiltration, the concentration was checked and the solution was diluted back to a concentration of 3.6 or 6.0 mg CNP-38 equivalent / mL. Next, the stock solution was aliquoted, methionine was added if necessary, the pH was checked and adjusted if necessary. Finally, m-cresol was added to the sample, which was then sterile filtered using a 0.22 μm PVDF filter and aliquoted for incubation.

[0597] Preparation of compound (1) formulations F46 - F50: The solution of compound (1) was buffer-exchanged by diafiltration into the designated buffer solution and then concentrated by ultrafiltration. Diafiltration and ultrafiltration were carried out using an ultrafiltration filter (Amicon® Ultra-15, Ultracel®-10K). After ultrafiltration, the concentration was checked and the solution was diluted back to a concentration of 2.8 mg CNP-38 equivalent / mL. Next, the pH was checked and adjusted with 1M HCl if necessary. The formulation was filtered using a 0.22 μm PVDF filter, and the stock preservative solution was added, thereby diluting the sample to 2.2 mg CNP-38 equivalent / mL. The formulation was aliquoted for incubation.

[0598] pH: The pH value of the formulation was measured at room temperature using a calibrated pH meter with a normal ionic strength electrode.

[0599] Analytical method RP-HPLC-MS for Measuring the Purity of CNP-38: Mobile phase A was composed of 0.05% aqueous TFA (by volume), and mobile phase B was composed of 0.04% TFA in acetonitrile (by volume). A Waters Acquity CSH C18, 130 Å, 1.7 μm, 2.1×100 mm column was used. The flow rate was set at 0.3 mL / min, detection was performed at a wavelength of 215 nm, and the operating temperature of the column was 60 °C (±1 °C). For LCMS, the HPLC system was connected to an LTQ Orbitrap Discovery XL instrument, and ESI-MS was run in the positive mode. A stepwise elution gradient was used, as shown in Table 21.

[0600] TIFF2025522281000150.tif71161

[0601] Preparative RP-HPLC for Isolating the CNP-38 Adduct. Mobile phase A was composed of water / acetonitrile / TFA (95 / 5 / 0.05, by volume), and mobile phase B was composed of water / acetonitrile / TFA (75 / 25 / 0.05, by volume). A Waters Acquity CSH C18, 130 Å, 10 μm, 10×150 mm column (+10×10 mm pre-column) was used. The flow rate was set at 10 mL / min, detection was performed at a wavelength of 215 nm, and the column operating temperature was at room temperature. The elution gradient is shown in Table 22.

[0602] TIFF2025522281000151.tif52161

[0603] RP-HPLC for determining the content and purity of compound (1) and detecting free CNP-38: Mobile phase A was an aqueous solution of 0.05% TFA (volume ratio), and mobile phase B was 0.04% TFA in acetonitrile (volume ratio). A Waters Acquity CSH C18, 130 Å, 1.7 μm, 2.1×100 mm column was used. The flow rate was set at 0.3 mL / min, detection was performed at a wavelength of 215 nm, the column operating temperature was 60 °C (±1 °C), and the sample injection volume was approximately 3 μg CNP-38 equivalent (20 μL of a 0.135 mg / mL sample). First, a stepwise eluent gradient operation was used from 5% B to 54% B over 25.5 minutes, and then from 54% B to 59% B over 10.0 minutes. The sample was diluted to 0.15 mg CNP-38 equivalent / mL with a dilution buffer (10 mM succinic acid, 84 g / L trehalose dihydrate, adjusted to pH 5 with 1 M Tris), and further diluted 9:1 (weight ratio) with a dilution buffer containing 0.5% Tween-20 (w / v). The content was determined by comparing the peak areas with a reference solution of known content. Free CNP-38 was identified by a reference peptide and quantified relative to the total peak area.

[0604] SE-HPLC for determining the purity of (1): The mobile phase was composed of 15 mM sodium phosphate pH 7.40, 135 mM sodium chloride, 0.2% poloxamer 188 aqueous solution (for example, 2.07 g of NaH2PO4·H2O, 7.89 g of NaCl and 2.00 g of poloxamer 188 in 1 L of HPLC water; the pH was adjusted to 7.40 using 4 M NaOH aqueous solution). A GE Superdex 200 Increase 10 / 300GL column was used. The constant composition flow rate was set at 0.75 mL / min, detection was performed at a wavelength of 215 nm, the column operating temperature was room temperature, and the sample injection volume was 17 μg CNP-38 equivalent (50 μL of 0.34 mg / mL sample). The sample was diluted to 0.34 mg / mL with a dilution buffer (10 mM succinic acid, 84 g / L trehalose dihydrate, adjusted to pH 5 with 1 M Tris) and a dilution buffer containing 0.1% poloxamer 188 (w / v) to give a final concentration of 0.05% poloxamer 188 (example: 20 μL of 3.6 mg CNP-38 equivalent / mL solution was diluted to 0.68 mg / mL with 86 μL of dilution buffer, and 50 μL of this solution was further diluted with 50 μL of dilution buffer containing 0.1% poloxamer 188).

[0605] Peptide mapping of (1) to evaluate the conversion of aspartic acid (28) to isoaspartic acid (28) (Isoasp) and the oxidation of methionine (33) to methionine sulfoxide (33) (Met(O)): The thermolysin digestion of (1) was carried out at a thermolysin / CNP-38 ratio = 1:20 (weight ratio). The pH was adjusted to pH 7.5 with a proteolysis buffer concentrate. After incubation at 37 °C for 7 h, 5% (v / v) phosphoric acid was added to stop the digestion. The resulting peptide mixture was separated by RP-HPLC on a Waters Acquity UPLC HSS T3, 100 Å, 1.8 μm, 2.1 × 150 mm column using 0.10% (v / v) aqueous TFA as mobile phase A and 0.09% (v / v) TFA in acetonitrile as mobile phase B, with detection at 210 nm. The flow rate was set at 0.28 L / min, the column operating temperature was 45 °C (±1 °C), and the sample injection volume was 19 μg of CNP-38 equivalent (e.g., 20 μL of 0.94 mg / mL sample). An eluent gradient from 0% B to 3% B over 31.0 min was used. Thermolysin fragments containing aspartic acid (28) (LisoDR) and methionine (33) (IGSM(O)SG) were identified by reference peptides and quantified based on their respective peak areas relative to the peak areas of the corresponding unmodified fragments (LDR; IGS+MSG).

[0606] Peptide mapping of CNP-38 and variants by LCMS. For (1), peptide mapping and RP-HPLC were performed. In the case of LCMS, the HPLC system was combined with an LTQ Orbitrap Discovery XL instrument, and ESI-MS was carried out in the positive mode.

[0607] The IEX-HPLC mobile phase A was composed of 20 mM sodium phosphate buffer pH 6.5 containing 10% (v / v) ethanol, and the mobile phase B was composed of 20 mM sodium phosphate buffer pH 6.5, 1 M sodium chloride containing 10% (v / v) ethanol. A Thermo Scientific ProPac Elite WCX 5 μm, 4 mm×250 mm column was used. The flow rate was set at 0.4 mL / min, the detection was carried out at a wavelength of 220 nm, the column operating temperature was 40 °C (±0.8 °C), and the sample injection volume was about 5.3 μg CNP-38 equivalent (7 μL of 0.75 mg / mL sample). The gradient is described in Table 23. The sample was diluted to 0.75 mg / mL with water.

[0608] TIFF2025522281000152.tif57161

[0609] RP-HPLC / MS for determining the content and purity of CNP(F33-45): The mobile phase A was an aqueous solution of 0.05% TFA (v / v), and the mobile phase B was an aqueous solution of 0.04% TFA (v / v) in acetonitrile. A Waters Acquity CSH C18, 130 Å, 1.7 μm, 2.1×100 mm column was used. The flow rate was set at 0.3 mL / min, the detection was carried out at a wavelength of 215 nm, the column operating temperature was 65 °C, and the sample injection volume was about 3 μg CNP-38 equivalent (20 μL of 0.135 mg / mL sample). The sample was diluted to 0.135 mg / mL with a dilution buffer (10 mM succinic acid, 84 g / L trehalose dihydrate, adjusted to pH 5 with 1 M Tris). The gradient is shown in Table 24. In the case of LCMS, the HPLC system was combined with an LTQ Orbitrap Discovery XL instrument, and ESI-MS was performed in the positive mode. The changes in the content of T1M, T3M, and T6M were determined by comparing the peak areas with the T0 sample. In the case of purity determination, all peaks higher than the LOQ (0.10%) and absent in the blank were integrated.

[0610] TIFF2025522281000153.tif57161

[0611] Example 1: Synthesis of Compound (1)

Chemical formula

[0612] Example 2: Stability of CNP-38 at Different pH Values in Succinate Buffer The effect of the pH value of a liquid pharmaceutical formulation on the stability of CNP-38 was evaluated. In this regard, six different formulations (F1, F2, F3, F4, F5, and F6) containing 2.8 mg / mL CNP-38 were prepared (Table 1).

[0613]

Table 1

[0614] The formulations were incubated at 37 °C for 7 days, and then the overall purity and the presence of succinate adducts were evaluated by using RP-HPLC and RP-HPLC-MS (MW + 100 Da of CNP-38). The results show that after incubation, the purity of formulation F3 at pH 4.5 was the highest (the least decomposition), and succinate adducts were detected in F1 to F5 but not in F6 (see Table 2).

[0615]

Table 2

[0616] Example 3: Identification of the Main CNP-38 Succinate Adduct Using the incubated CNP-38 from Example 2, the major succinic acid isomers were isolated by RP-HPLC. In parallel, succinylated CNP-38 was newly generated by incubating CNP-38 with succinic anhydride. To 10 mg of CNP-38 in 5 mL of PBS, 2.5 equivalents of succinic anhydride in 0.6 mL of PBS buffer were added, the pH of the mixture was adjusted to about pH 6.5 with 4N NaOH, and the mixture was incubated for 30 minutes. The addition and pH adjustment were repeated three times, and the major isomers were isolated by RP-HPLC and lyophilized. Peptide mapping and LCMS analysis revealed that succinylation was located at the N-terminus of the peptides of both compounds.

[0617] Example 4: Stability of CNP-38 at various pH values in acetate buffer The effect of the pH value of the liquid formulation on the stability of CNP-38 was evaluated. In this regard, five different formulations (F7, F8, F9, F10, F11) containing CNP-38 (1.8 mg / mL) in acetate buffer and one formulation (F12) in succinate buffer were prepared (Table 3).

[0618]

Table 3

[0619] The formulations were incubated at 37 °C, and samples were withdrawn and analyzed after 8 days and 28 days. RP-HPLC and RP-HPLC-MS were used to evaluate the overall purity of the samples and the pr...

Claims

1. A liquid pharmaceutical preparation comprising a CNP compound, a buffer, an isotonic agent, a preservative, and optionally an antioxidant.

2. The liquid pharmaceutical preparation according to claim 1, for storage at 2 to 8 °C for at least 6 months.

3. The liquid pharmaceutical preparation according to claim 1 or 2, for storage at 2 to 8 °C for 6 to 48 months.

4. The liquid pharmaceutical preparation according to any one of claims 1 to 3, for storage at 2 to 8 °C for 6 to 36 months.

5. The liquid pharmaceutical preparation according to any one of claims 1 to 4, for storage at 5 °C for 6 to 36 months.

6. The liquid pharmaceutical preparation according to any one of claims 1 to 5, wherein the CNP compound is a CNP conjugate.

7. The liquid pharmaceutical preparation according to any one of claims 1 to 6, wherein the buffer is selected from the group consisting of acetic acid, succinic acid, citric acid, lactic acid, glutamic acid, fumaric acid, aspartic acid, glutaric acid, phosphoric acid, histidine, gluconic acid, tartaric acid, malic acid, and tris (tris (hydroxymethyl) aminomethane).

8. The liquid pharmaceutical preparation according to any one of claims 1 to 7, wherein the buffer is acetic acid.

9. The liquid pharmaceutical preparation according to any one of claims 1 to 8, wherein the isotonic agent is selected from the group consisting of mannitol, trehalose, sucrose, raffinose, gelatin, lactose, dibasic calcium phosphate, sorbitol, xylitol, glycine, histidine, ethanol, hydroxyethyl starch, potassium chloride, sodium chloride, dextrose, dextran, propylene glycol, and glycerol.

10. The liquid pharmaceutical preparation according to any one of claims 1 to 9, wherein the isotonic agent is mannitol or trehalose.

11. The liquid pharmaceutical preparation according to any one of claims 1 to 10, wherein the isotonic agent is mannitol.

12. The liquid pharmaceutical preparation according to any one of claims 1 to 10, wherein the isotonic agent is trehalose.

13. The liquid pharmaceutical preparation according to any one of claims 1 to 12, wherein the preservative is selected from the group consisting of m-cresol, benzyl alcohol, benzoic acid, phenol, methyl paraben, ethyl paraben, propyl paraben, butyl paraben, potassium sorbate, chlorobutanol, benzyl alcohol, phenylmercuric nitrate, thimerosal, sorbic acid, potassium sorbate, chlorocresol, benzalkonium chloride, 2-ethoxyethanol, chlorhexidine, chlorobutanol, phenylethyl alcohol, and phenylmercuric acetate.

14. The liquid pharmaceutical preparation according to any one of claims 1 to 13, wherein the preservative is m-cresol or phenol.

15. The liquid pharmaceutical preparation according to any one of claims 1 to 14, wherein the preservative is m-cresol.

16. The liquid pharmaceutical preparation according to any one of claims 1 to 14, wherein the preservative is phenol.

17. The liquid pharmaceutical preparation according to any one of claims 1 to 16, wherein the pharmaceutical preparation contains an antioxidant selected from the group consisting of methionine, butylated hydroxytoluene, butylated hydroxyanisole, tocopherol, propyl gallate, ascorbic acid, sodium bisulfite, ethylenediaminetetraacetic acid (EDTA), cysteine, glutathione, monothioglycerol, poly(ethyleneimine), vitamin E, ectoine, and morin.

18. The liquid pharmaceutical preparation according to any one of claims 1 to 17, wherein the pH of the liquid preparation is from about pH 3.5 to about pH 5.

5.

19. The liquid pharmaceutical preparation according to any one of claims 1 to 18, wherein the pH of the liquid preparation is about 4.

5.

20. The liquid pharmaceutical preparation according to any one of claims 1 to 19, wherein the liquid preparation further contains a pH adjuster.

21. The liquid pharmaceutical preparation according to claim 20, wherein the pH adjuster is selected from the group consisting of sodium hydroxide, sodium acetate, tris(tris(hydroxymethyl)aminomethane), potassium hydroxide, lysine, and mixtures thereof.

22. The liquid pharmaceutical preparation according to any one of claims 1 to 21, wherein the CNP compound is a reversible CNP conjugate comprising a CNP moiety covalently and reversibly bound to a carrier moiety.

23. The liquid pharmaceutical preparation according to claim 22, wherein the carrier moiety is a polymer moiety.

24. The liquid pharmaceutical preparation according to any one of claims 1 to 21, wherein the CNP compound is a CNP-acting agent.

25. The liquid pharmaceutical preparation according to any one of claims 1 to 23, wherein the CNP compound is a reversible CNP conjugate of the following formula (Ia). 【Chemical 1】 [Wherein, -D is a CNP moiety; -L １ - is a reversible linker moiety; -L ２ - is a single chemical bond or a spacer moiety; -Z is a carrier moiety; x is an integer selected from the group consisting of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, and 16. ]]

26. The liquid pharmaceutical preparation according to claim 25, wherein x in formula (Ia) is 1.

27. The liquid pharmaceutical preparation according to claim 25 or 26, wherein -Z is a branched polymer moiety.

28. The liquid pharmaceutical preparation according to any one of claims 25 to 27, wherein -Z is a branched PEG-based polymer moiety.

29. The liquid pharmaceutical preparation is a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, D-mannitol: 10 to 200 mg / mL, m-cresol: 1 to 10 mg / mL The liquid pharmaceutical preparation according to any one of claims 6 to 11, 13 to 15, 17 to 23, or 25 to 28, comprising.

30. The liquid pharmaceutical preparation is a CNP conjugate in which the CNP moiety is present at 0.1 to 20 mg / mL, acetic acid: 0.3 to 3 mg / mL, trehalose dihydrate: 10 to 200 mg / mL m-cresol: 1 to 10 mg / mL The liquid pharmaceutical preparation according to any one of claims 6 to 10, 12 to 15, 17 to 23, or 25 to 28, comprising.

31. The partial-L １ - is the one of the following formula (II), the liquid pharmaceutical preparation according to any one of claims 25 to 30. [Chemical Formula 2] [Wherein, The dotted line indicates the bond to the nitrogen of -D, which is the CNP moiety, by forming an amide bond -X- is -C(R ４ R ４ａ ), -N(R ４ ), -O-, -C(R ４ R ４ａ ), -C(R ５ R ５ａ ), -C(R ５ R ５ａ ), -C(R ４ R ４ａ ), -C(R ４ R ４ａ ), -N(R ６ ), -N(R ６ ), -C(R ４ R ４ａ ), -C(R ４ R ４ａ ), -O-, -O-C(R ４ R ４ａ ), or -C(R ７ R ７ａ ); X １ is C or S(O); -X ２ - is -C(R ８ R ８ａ ) - or -C(R ８ R ８ａ )-C(R ９ R ９ａ ) - and; = X ３ is = O, = S or = N-CN; -R １ 、 -R １ａ 、 -R ２ 、 -R ２ａ 、 -R ４ 、 -R ４ａ 、 -R ５ 、 -R ５ａ 、 -R ６ 、 -R ８ 、 -R ８ａ 、 -R ９ 、 -R ９ａ is independently selected from the group consisting of -H; and C １－６ alkyl; -R ３ 、 -R ３ａ are each independently selected from the group consisting of -H and C １－６ alkyl, provided that when -R ３ , -R ３ａ or both are other than -H, they are linked to the N atom to which they are attached via an sp ３ hybridized carbon atom; -R ７ is -N(R １０ R １０ａ ) or -NR １０ -(C=O)-R １１ ; and -R ７ａ 、-R １０ 、-R １０ａ 、-R １１ are, independently of one another, -H; or C １－６ alkyl; Optionally, pair - R １ａ / - R ４ａ , - R １ａ / - R ５ａ , - R １ａ / - R ７ａ , - R ４ａ / - R ５ａ , - R ８ａ / - R ９ａ One or more of which form chemical bonds; Optionally, pair - R １ / - R １ａ , - R ２ / - R ２ａ , - R ４ / - R ４ａ , - R ５ / - R ５ａ , - R ８ / - R ８ａ , - R ９ / - R ９ａ One or more of them combine with the atoms to which they are attached and form a C ３－１０ cycloalkyl; or a 3 - to 10 - membered heterocyclic ring; Optionally, pair - R １ / - R ４ , - R １ / - R ５ , - R １ / - R ６ , - R １ / - R ７ａ , - R ４ / - R ５ , - R ４ / - R ６ , - R ８ / - R ９ , - R ２ / - R ３ One or more of them are integrated with the atoms to which they are attached to form ring A; Optionally, -R ３ / -R ３ａ is combined with the nitrogen atom to which it is attached to form a 3- to 10-membered heterocyclic ring; A is selected from the group consisting of phenyl, naphthyl, indenyl, indanyl, tetralinyl, C ３－１０ cycloalkyl, a 3- to 10-membered heterocyclic ring, and an 8- to 11-membered heterocyclic ring; -L １ - is -L ２ - is replaced by -Z, -L １ - may be further replaced, provided that the hydrogen marked with an asterisk in formula (II) is -L ２ - is not replaced by -Z or a substituent.]

32. Said partial-L １ -L ２ - is as represented by the following formula (II d-ii), the liquid pharmaceutical preparation according to any one of claims 25 to 31. 【Chemical Formula 3】 [Wherein, The dotted line without a mark indicates the bond of the amino acid side chain of the ring portion of -D to the nitrogen by forming an amide bond; the dotted line marked with an asterisk indicates the bond to -Z; -D has the sequence of SEQ ID NO: 24, and -L １ - is bonded to lysine at position 26; -Z is of the following formula (h): [Chemical 4] The dotted line indicates the connection to -L ２ -; each - Z ｃ is the following part: 【Chemical Formula 5】 and; each c1 is independently an integer in the range of about 200 to 250. ]]

33. A method for producing the liquid pharmaceutical preparation according to any one of claims 1 to 32, comprising (i) mixing the CNP compound according to any one of claims 1 to 32 with at least a buffer, an isotonic agent, a preservative, and optionally an antioxidant; (ii) adjusting the pH of the mixture of step (i); (iii) optionally filtering the mixture from step (ii); (iv) transferring an amount of the mixture from step (ii) or (iii) corresponding to the desired number of administrations to a container; (v) sealing the container including; The order of step (ii) and step (iii) may optionally be reversed.

34. The method according to claim 33, wherein the buffer is acetic acid.

35. The method according to claim 33 or 34, wherein the isotonic agent is selected from the group consisting of mannitol and trehalose.

36. The method according to any one of claims 33 to 35, wherein the isotonic agent is mannitol.

37. The method according to any one of claims 33 to 35, wherein the isotonic agent is trehalose.

38. A container containing the liquid pharmaceutical preparation according to any one of claims 1 to 37.

39. The container according to claim 38, wherein the container is selected from the group consisting of vials, syringes, ampoules, and cartridges.

40. The liquid pharmaceutical preparation according to any one of claims 1 to 32 for use in the treatment of one or more diseases treatable by CNP.

41. The diseases treatable by CNP are achondroplasia, hypochondroplasia, short stature, dwarfism, osteochondrodysplasia, lethal osteogenesis imperfecta, osteogenesis imperfecta, achondrogenesis, stippled chondrodysplasia, homozygous achondroplasia, campomelic dysplasia, congenital lethal hypophosphatasia, perinatal lethal osteogenesis imperfecta, short rib polydactyly syndrome, rhizomelic stippled chondrodysplasia, Jansen metaphyseal dysplasia, congenital spondyloepiphyseal dysplasia, osteogenesis imperfecta, torsional osteodysplasia, congenital femoral shortening, Langer mesomelic dysplasia, Nievergelt mesomelic dysplasia, Robinow syndrome, Reinhardt syndrome, acroosteolysis, distal osteolysis, Knist osteodysplasia, fibrochondrogenesis, Roberts syndrome, distal mesomelic dysplasia, micromelia, Morquio syndrome, Knist syndrome, dysplastic osteodysplasia, spondyloepimetaphyseal dysplasia, neurofibromatosis, Legius syndrome, Leopard syndrome, Noonan syndrome, hereditary gingival fibromatosis, neurofibromatosis type 1, cardiofaciocutaneous syndrome, Costello syndrome, SHOX deficiency, idiopathic short stature, growth hormone deficiency, osteoarthritis, cleidocranial dysplasia, craniosynostosis, dactyly, brachydactyly, camptodactyly, polydactyly, syndactyly, segmental dysplasia, enchondroma, fibrous dysplasia, hereditary multiple exostoses, hypophosphatemic rickets, Jaffe-Lichtenstein syndrome, Marfan syndrome, McCune-Albright syndrome, osteopetrosis, and osteopathia striata. The liquid pharmaceutical preparation for use according to claim 40, selected from the group consisting of.

42. A liquid pharmaceutical preparation for use according to claim 40, wherein the disease treatable by CNP is selected from the group consisting of achondroplasia, hypochondroplasia, short stature, Noonan syndrome, and SHOX deficiency.

43. A liquid pharmaceutical preparation for use according to claim 40, wherein the disease is achondroplasia.

44. A method of treating, reducing the risk of, or delaying a disease treatable by CNP in a human patient, the method comprising administering to the patient a therapeutically effective amount of the liquid pharmaceutical preparation according to any one of claims 1 to 32.

45. The method according to claim 44, wherein the disease is selected from the group consisting of achondroplasia, hypochondroplasia, short stature, Noonan syndrome, and SHOX deficiency.

46. The method according to claim 44 or 45, wherein the disease is achondroplasia.

47. A liquid pharmaceutical preparation according to any one of claims 1 to 32 for use in the manufacture of a medicament for the treatment of achondroplasia.

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