CNP compounds

Abstract

The problem that this invention aims to solve is to provide a CNP compound having improved pharmaceutical properties. [Solution] The present invention relates to a CNP compound comprising a CNP peptide and a modifying group, wherein the net charge (effective charge) of the compound at physiological pH is 0 or negative, the CNP peptide comprises an amino acid sequence according to formula I, and the modifying group comprises Chem.A, Chem.B, and Chem.C.

Description

[Technical Field]

[0001] This invention relates to novel C-type natriuretic peptide (CNP) compounds, pharmaceutical compositions containing these compounds, and these compounds for use as pharmaceuticals. [Background technology]

[0002] Natriuretic peptides are a family of three structurally related hormones that play specific roles within the cardiovascular system. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) are expressed in the heart and released in response to volume-induced contraction of the atria and ventricles, respectively. Their physiological effects include the regulation of intracardiac structure, blood pressure, and blood volume.

[0003] C-type natriuretic peptide (CNP) is highly expressed in endothelial cells, where it is constitutively released. Other cells in the cardiovascular system, including cardiomyocytes and fibroblasts, also produce CNP, but to a lesser degree. CNP has direct effects on inflammation, fibrosis, cardiac contractility, endothelial function, angiogenesis, and blood pressure.

[0004] Three receptors for natriuretic peptides are known. Natriuretic peptide receptor-1 (NPR1) is a fine-particle guanylyl cyclase that catalyzes the synthesis of cGMP upon binding by ANP or BNP. NPR1 is expressed in the kidneys, lungs, adipose tissue, adrenal glands, brain, heart, testes, and vascular smooth muscle tissue. NPR2, expressed in bone, brain, fibroblasts, heart, kidneys, liver, lungs, uterus, and vascular smooth muscle tissue, is homologous to NPR1 but is selectively activated by CNP. In contrast, NPR3 contains only a 37-residue intracellular domain and lacks guanylyl cyclase activity. It regulates local natriuretic peptide concentrations via receptor-mediated internalization and degradation, although evidence for the signaling function of NPR3 continues to grow. All three natriuretic peptides bind to NPR3 with high affinity, and this receptor is the most widely and abundantly expressed of the three receptors. Furthermore, the degree of conservation of all three receptors is very high, and consistent with CNP, NPR2 is the most highly conserved.

[0005] Clinical and preclinical data have demonstrated the crucial role of CNP and its two receptors in cardiorenal metabolic function, and growing data supports the therapeutic potential of targeting this system in a wide range of cardiovascular, renal, and metabolic diseases (Non-Patent Literature 1; Non-Patent Literature 2; Non-Patent Literature 3). Similarly, human genetics, as well as preclinical and clinical data, supports NPR2 and CNP (Non-Patent Literature 4; Non-Patent Literature 5) as bone growth regulators with therapeutic potential in a wide range of short stature indications, including FGFR3-related osteodysplasia (Non-Patent Literature 6; Non-Patent Literature 7, Non-Patent Literature 8) and RAS disease (Non-Patent Literature 9). Two CNP compounds are currently in clinical development for achondroplasia, a type of dwarfism (BMN111 (Vosoritide) and TransCon CNP). The Mayo Clinic previously studied a CNP compound designed to activate both NPR2 and NPR1 in heart failure (Non-Patent Literature 10).

[0006] CNP clearance in human plasma is very rapid, with a calculated half-life of several minutes. Given this short half-life, it would be beneficial to develop novel, long-acting CNP compounds that can be administered at lower frequencies while maintaining an acceptable clinical profile.

[0007] To provide a longer-lasting action profile, various different approaches have been used to modify the structure of CNP. Patent Document 1 discloses a fusion peptide of the CNP domain and the Fc domain. Patent Document 2 discloses a fatty acid-modified NPR1 agonist. [Prior art documents] [Patent Documents]

[0008] [Patent Document 1] WO2013 / 058833 [Patent Document 2] WO2018 / 175534 [Non-patent literature]

[0009] [Non-Patent Document 1] Int.J.Mol.Sci.2019,20,2281 [Non-Patent Document 2] Cardiovasc.Res.2022,118,2085-2102 [Non-Patent Document 3] Cardiovasc.Res.2022 Aug 25;cvac125 [Non-Patent Document 4] Lancet,396:684;J Pharmacol Exp Ther,370:459 [Non-Patent Document 5] Hisado-Oliva(2018)Genet Med,20:91 [Non-Patent Document 6] Sabir and Cole Orphanet Journal of Rare Diseases(2019)14:300

Non-Patent Document 7

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Summary of the Invention

[0010] The present invention provides a CNP compound having improved pharmaceutical properties.

[0011] In one aspect, the present invention is a CNP compound comprising a CNP peptide and a modifying group, wherein the net charge (effective charge) of the compound at physiological pH is 0 or negative, and the CNP peptide has the formula I: AA 28 , 26 , 24 , 29 , 27 , 25 , 23 -AA 02 -AA 03 -AA 04 -AA 05 -AA 06 -AA 07 ... -AA 08 -AA 09 -AA 10 -AA 11 -AA 12 -AA 13 -AA 14 -AA 15 -AA 16 -AA 17 -AA 18 -AA 19 -AA 20 -AA 21 -AA 22 -AA 23 -AA 24 -AA 25 -AA 26 -AA 27 -AA 28 -AA 29 ... -AA30 -AA 31 -AA 32 -AA 33 -AA 34 -AA 35 -AA 36 -AA 37 (In the formula, AA 01 However, it is either Gln or absent, AA 02 However, it is either Glu or absent, AA 03 However, His is either absent or not. AA 04 However, whether they are Pro or absent, AA 05 However, it is either Asn, Gln, or Glu, or it is absent. AA 06 However, Ala is either absent or not present. AA 07 However, it is either Arg, His, or Ala, or absent. AA 08 However, it is either Lys, Ser, or His, or absent. AA 09 However, it is either Tyr or Glu, or absent. AA 10 However, it is either Lys, Glu, Gln, or His, or it is absent. AA 11 However, it is Gly, AA 12 However, it is Ala, AA 13 However, it is Gln or Asn or Glu, AA 14 However, it is either Lys, His, or Glu. AA 15 However, it is Lys, Ser or Glu or Thr or His, AA 16 However, it is Gly, AA 17However, it is Leu or Gly or Ser or Val, AA 18 However, it is either Ser or His. AA 19 However, it is Gln or Ser or Lys or His, AA 20 However, it is Gly, AA 21 However, it is Cys, AA 22 However, it is Phe, AA 23 However, it is Gly, AA 24 However, it is Leu, AA 25 However, it is either Pro or Lys. AA 26 However, it is Leu, AA 27 However, it is either Asp or Glu. AA 28 However, it is Arg, AA 29 However, it is Ile, AA 30 However, it is Gly, AA 31 However, it is Ser, AA 32 However, it is Leu or Nle or Met, AA 33 However, it is Ser, AA 34 However, it is Gly, AA 35 However, it is Leu, AA 36 However, it is Gly, AA 37 However, it contains an amino acid sequence that follows Cys. The modifying groups include Chem.A, Chem.B, and Chem.C. Chem.A, [ka] (In the formula, p is an integer in the range of 14 to 20, * is selected from the group consisting of (where * represents the amide bond connecting Chem.A and Chem.B), Chem.B [ka] (In the formula, q is an integer in the range of 1 to 8, * represents the amide bond connecting Chem.A- and Chem.B-. ** is selected from the group consisting of (where ** represents an amide bond connecting Chem.B- and Chem.C-), Chem.C [ka] (In the formula, r is an integer in the range of 0 to 4, s is an integer in the range of 0 to 3. t is an integer in the range of 0 to 1, ** represents the amide bond connecting Chem.B- and Chem.C-, A CNP compound selected from the group consisting of *** (where *** represents the amide bond connecting Chem.C- and the N-terminal alpha-amine on the CNP peptide).

[0012] In one embodiment, the present invention provides a CNP compound, the CNP peptide having any one of the following amino acid sequences. GAQKKGSSQGCFGLPLDRIGSLSGLGC (Sequence ID 136), ARKYKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 77), YKGAQKKGGSQGCFGLPLDRIGSLSGLGC (Sequence ID 88) YKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 103), QEHPQARKYKGAQKKGLSSGCFGLPLDRIGSLSGLGC (Sequence ID 67), and QEHPQARKYKGAQKKGLSSGCFGLPLERIGSLSGLGC (Sequence ID 104)

[0013] In another embodiment, the present invention provides a pharmaceutical composition comprising a compound according to the present invention and one or more pharmaceutically acceptable excipients.

[0014] In one aspect, the present invention also provides the use of the compounds of the present invention as pharmaceuticals for treating or preventing cardiorenal metabolic diseases, including heart failure, and chondrodysplasia.

[0015] Due to its short half-life of approximately 2 minutes, wt CNP is unsuitable for pharmaceutical applications. In this invention, the half-life of CNP is extended by fatty acid acylation, which promotes albumin binding. However, the combination of a net positively charged CNP peptide (at physiological pH) and fatty acid albumin-binding modification (when the CNP compound is positively charged overall) exacerbates injection site reactions during subcutaneous injection and reduces bioavailability. This is surprising, as a net positively charged CNP peptide without fatty acid albumin binding does not exhibit an observable injection site reaction. To address these issues, amino acid substitutions are introduced into the CNP peptide, and negative charges are introduced into the modifying groups (including fatty acids). These modification combinations yield compounds with good subcutaneous bioavailability and no to mild subcutaneous injection site reactions, exhibiting an overall net negative charge at physiological pH.

[0016] Surprisingly, adding a negative charge also reduces the in vitro and in vivo potency of the CNP compound, in a manner in which a higher net negative charge generally correlates with lower in vivo potency. Only a small number of net negative charges are introduced to maintain sufficient potency of the CNP compound. However, a small number of net negative charges reduces the solubility and biophysical stability of the CNP compound when formulated for subcutaneous administration at a physiologically relevant pH. This disclosure provides a CNP compound that balances these parameters and, surprisingly, also possesses sufficient potency, as well as sufficient solubility and biophysical stability for liquid formulations.

[0017] Furthermore, wt CNP does not exhibit the desired chemical stability in liquid formulations necessary for convenient drug administration. In this invention, selected amino acid substitutions are introduced into the wt CNP sequence to improve chemical stability in the formulation.

[0018] This disclosure provides a CNP compound that has sufficient in vivo efficacy for therapeutic use, as well as high chemical and biophysical stability in liquid formulations, while also possessing an extended half-life and good tolerability for subcutaneous administration.

[0019] The present invention can also solve further problems that become apparent from the disclosure of exemplary embodiments. [Brief explanation of the drawing]

[0020] [Figure 1] Figure 1 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 0776, as described in Example 11. [Figure 2] Figure 2 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 0312, as described in Example 11. [Figure 3] Figure 3 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1225, as described in Example 11. [Figure 4] Figure 4 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1233, as described in Example 11. [Figure 5] Figure 5 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1227, as described in Example 11. [Figure 6] Figure 6 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1241, as described in Example 11. [Figure 7] Figure 7 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1351, as described in Example 11. [Figure 8]Figure 8 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1354, as described in Example 11. [Figure 9] Figure 9 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1356, as described in Example 11. [Figure 10] Figure 10 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1375, as described in Example 11. [Figure 11] Figure 11 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1376 described in Example 11. [Figure 12] Figure 12 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1377, as described in Example 11. [Figure 13] Figure 13 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1352, as described in Example 11. [Figure 14] Figure 14 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1379, as described in Example 11. [Figure 15] Figure 15 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1434, as described in Example 11. [Figure 16] Figure 16 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 9384, as described in Example 11. [Figure 17] Figure 17 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 9407, as described in Example 11. [Figure 18] Figure 18 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1420, as described in Example 11. [Figure 19] Figure 19 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1378, as described in Example 11. [Figure 20]Figure 20 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1381, as described in Example 11. [Figure 21] Figure 21 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1386, as described in Example 11. [Figure 22] Figure 22 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1389, as described in Example 11. [Figure 23] Figure 23 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1392, as described in Example 11. [Figure 24] Figure 24 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1426, as described in Example 11. [Figure 25] Figure 25 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 9435, as described in Example 11. [Figure 26] Figure 26 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 1235, as described in Example 11. [Figure 27] Figure 27 shows the pharmacodynamic response (cGMP) to rats after intravenous administration of compound ID 9482, as described in Example 11. [Figure 28] Figure 28 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 0312, as described in Example 12. [Figure 29] Figure 29 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 0776, as described in Example 12. [Figure 30] Figure 30 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9384, as described in Example 12. [Figure 31] Figure 31 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9407, as described in Example 12. [Figure 32]Figure 32 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9435, as described in Example 12. [Figure 33] Figure 33 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9480, as described in Example 12. [Figure 34] Figure 34 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9482, as described in Example 12. [Figure 35] Figure 35 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after intravenous administration of compound ID 9483, as described in Example 12. [Figure 36] Figure 36 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 0312, as described in Example 12. [Figure 37] Figure 37 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 0776, as described in Example 12. [Figure 38] Figure 38 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9384, as described in Example 12. [Figure 39] Figure 39 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9407, as described in Example 12. [Figure 40] Figure 40 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9435, as described in Example 12. [Figure 41] Figure 41 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9480, as described in Example 12. [Figure 42] Figure 42 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9482, as described in Example 12. [Figure 43]Figure 43 shows the pharmacodynamic response (cGMP) to Göttingen miniature pigs after subcutaneous administration of compound ID 9483, as described in Example 12. [Figure 44] Figure 44 shows the pharmacodynamic response (cGMP) to livestock LYD pigs after intravenous administration of compound ID 0312 described in Example 12. [Figure 45] Figure 45 shows the pharmacodynamic response (cGMP) after intravenous administration of compound ID 0776, described in Example 12, to domestic LYD pigs. [Figure 46] Figure 46 shows the pharmacodynamic response (cGMP) to livestock LYD pigs after intravenous administration of compound ID 1225 described in Example 12. [Figure 47] Figure 47 shows the pharmacodynamic response (cGMP) to livestock LYD pigs after intravenous administration of compound ID 1227 described in Example 12. [Figure 48] Figure 48 shows the pharmacodynamic response (cGMP) to livestock LYD pigs after intravenous administration of compound ID 1235 described in Example 12. [Modes for carrying out the invention]

[0021] The present invention provides CNP compounds having improved pharmaceutical properties. The present invention also provides pharmaceutical compositions, as well as the use of the compounds and compositions of the present invention as pharmaceuticals for treating diseases.

[0022] C-type natriuretic peptide C-type natriuretic peptide (CNP) (GenBank accession number NP 077720) is a small single-chain peptide in the family of peptides (ANP, BNP, CNP) that have a 17-amino acid disulfide ring structure and play important roles in several biological processes. CNP interacts with natriuretic peptide receptor 2 (NPR2, NPR-B, GC-B) to stimulate the production of cyclic guanosine monophosphate (cGMP). CNP is more broadly expressed, including in the central nervous system, reproductive tract, bone, and vascular endothelium. Clearance of CNP in human plasma is rapid, with a half-life of several minutes.

[0023] Natural CNP genes and polypeptides have been previously described. U.S. Patent No. 5,352,770 discloses CNP-22 isolated and purified from porcine brain, which has the same sequence as human CNP (human wild-type CNP-22: GLSKGCFGLKLDRIGSMSGLGC (SEQ ID NO: 01)). U.S. Patent No. 6,034,231 discloses the human gene and polypeptide of prepro-CNP (126 amino acids), as well as the human CNP-53 gene and peptide. Human wild-type CNP-37 has the sequence QEHPNARKYKGANKKGLSKGCFGLKLDRIGSMSGLGC (SEQ ID NO: 02).

[0024] CNP compound In one embodiment, the present invention provides a CNP compound comprising a CNP peptide and a modifying group, wherein the net charge of the compound at physiological pH is 0 or negative.

[0025] The CNP compounds described herein have been manipulated to achieve an overall neutral or net negative charge. This was achieved by introducing amino acid substitutions to the CNP peptide and introducing negative charges to the modifying groups. Maintaining the biological activity of these CNP compounds against the NPR2 receptor while reducing their charge is not straightforward.

[0026] As used herein, the term "net charge" refers to the sum of all negative and positive charges on the CNP compound at a given pH, and the charge is defined solely by the acid dissociation constant of each ionizable group in the CNP compound.

[0027] In one embodiment, the net charge of the CNP compound at physiological pH is negative, as determined as described in Example 4.

[0028] In one embodiment, the net charge of the CNP compound at physiological pH is 0 to -4, as determined as described in Example 4.

[0029] As used herein, the term "physiological pH" refers to a pH in the range of 7.35 to 7.45, more typically, an average of 7.4.

[0030] In one embodiment, the CNP compound described herein has an intramolecular disulfide bridge (a disulfide bond between two cysteine ​​residues) that gives rise to a cyclic structure.

[0031] CNP peptide The term "CNP peptide," as used herein, refers to a peptide of 37 to 22 amino acids in length containing the CNP-22 amino acid sequence (SEQ ID NO: 01), and its amino acid sequence containing one or more amino acid modifications (e.g., one or more amino acid substitutions, additions, and / or deletions). The term "CNP peptide," as used herein, also encompasses CNP variants.

[0032] The term "CNP variant," as used herein, refers to a CNP peptide that is an amino acid variant of the CNP-37 sequence (SEQ ID NO: 02). In other words, a CNP variant is a CNP peptide that contains one or more amino acid modifications, i.e., at least one amino acid is altered compared to SEQ ID NO: 02. These modifications may independently represent one or more amino acid substitutions, additions, and / or deletions.

[0033] CNP variants may be described by referring to the number of altered amino acid residues (i.e., the corresponding positions in the CNP-37 sequence (SEQ ID NO: 02)) and the changes (e.g., the identity of amino acid residues in the variant at a given position). That is, a numerical reference to a particular amino acid residue in a CNP peptide refers to the CNP-37 sequence (SEQ ID NO: 2) unless otherwise specified (i.e., residue 1 is glutamine (Q1) and residue 37 is cysteine ​​(C37)). The following are non-restrictive examples of preferred variant nomenclature: des1~4, 5Q, 13Q, 32Nle. A CNP variant specifies a CNP peptide sequence having the following amino acid changes compared to CNP-37: deletion of amino acid residues at positions 1-4, as well as substitution of asparagine (N) at position 5 with glutamine (Q), substitution of asparagine (N) at position 13 with glutamine (Q), and substitution of methionine (M) at position 32 with norleucine.

[0034] The term “amino acid” refers to any amino acid that is either naturally occurring (including the 20 standard amino acids encoded by the standard human genetic code) or not naturally occurring. Amino acid residues may be identified by their full name, their one-letter code, and / or their three-letter code. These three methods are entirely equivalent. The term “uncoded amino acid” refers to all amino acids that are not among the 20 standard amino acids encoded by the standard human genetic code. Uncoded amino acids may be naturally occurring or purely synthesized. Non-limiting examples of uncoded amino acids are the D isomers of coded amino acids and glycine residues that have a side chain bonded to a nitrogen atom rather than an alpha-carbon atom. D isomers of coded amino acids may be referred to as (i) “D” followed by the full name of the amino acid, a one-letter code, or a three-letter code, or (ii) the lowercase one-letter code of the amino acid. Further abbreviations for uncoded amino acids as used in this application are listed below. [Table 1]

[0035] In one aspect of the present invention, the CNP peptide has the formula I: AA 01 -AA 02 -AA 03 -AA 04 -AA 05 -AA 06 -AA 07 -AA 08 -AA 09 -AA 10 -AA 11 -AA 12 -AA 13 -AA 14 -AA 15 -AA 16 -AA 17 -AA 18 -AA 19 -AA 20 -AA 21 -AA 22 -AA 23 -AA 24 -AA 25 -AA 26 -AA 27 -AA 28 -AA 29 -AA 30 -AA 31 -AA 32 -AA 33 -AA 34 -AA 35 -AA 36 -AA 37 (wherein, AA 01 is Gln or absent, AA 02 is Glu or absent,<000057​​​​​​​​​​​​​​​​AA 07 However, it is either Arg, His, or Ala, or absent. AA 08 However, it is either Lys, Ser, or His, or absent. AA 09 However, it is either Tyr or Glu, or absent. AA 10 However, it is either Lys, Glu, Gln, or His, or it is absent. AA 11 However, it is Gly, AA 12 However, it is Ala, AA 13 However, it is Gln or Asn or Glu, AA 14 However, it is either Lys, His, or Glu. AA 15 However, it is Lys, Ser or Glu or Thr or His, AA 16 However, it is Gly, AA 17 However, it is Leu or Gly or Ser or Val, AA 18 However, it is either Ser or His, AA 19 However, it is Gln or Ser or Lys or His, AA 20 However, it is Gly, AA 21 However, it is Cys, AA 22 However, it is Phe, AA 23 However, it is Gly, AA 24 However, it is Leu, AA 25 However, it is either Pro or Lys. AA 26 However, it is Leu, AA 27 However, it is either Asp or Glu, AA 28 However, it is Arg, AA 29 However, it is Ile, AA 30 However, it is Gly, AA 31 However, it is Ser, AA 32 However, it is Leu or Nle or Met, AA 33 However, it is Ser, AA 34 However, it is Gly, AA 35 However, it is Leu, AA 36 However, it is Gly, AA 37 It contains an amino acid sequence that follows the Cys gene.

[0036] In certain embodiments of the present invention, amino acid substitutions are introduced into the CNP peptide to reduce its positive charge. For example, to remove the positive charge, Lys25(AA 25 ) is replaced with Pro, and Lys19 (AA19) is replaced with Gln or Ser.

[0037] In a particular embodiment of the present invention, the AA of the CNP peptide 13 is Gln, AA 19 is Gln or Ser, AA 25 It is Pro, AA 32 It is Leu.

[0038] In a particular embodiment of the present invention, AA5 of the CNP peptide is Gln, and AA 13 is Gln, AA 19 is Gln or Ser, AA 25 It is Pro, AA 32 It is Leu.

[0039] In a particular embodiment of the present invention, the CNP peptide has the following amino acid sequence: GAQKKGSSQGCFGLPLDRIGSLSGLGC (Sequence ID 136), ARKYKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 77), YKGAQKKGGSQGCFGLPLDRIGSLSGLGC (Sequence ID 88) YKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 103), QEHPQARKYKGAQKKGLSSGCFGLPLDRIGSLSGLGC (Sequence ID 67), and It has one of the following: QEHPQARKYKGAQKKGLSSGCFGLPLERIGSLSGLGC (Sequence ID 104).

[0040] Modifying group In one aspect of the present invention, the CNP compound includes a modifying group covalently bonded to an amino acid residue of the CNP peptide.

[0041] In one aspect of the present invention, the modifying group can form a non-covalent association with albumin, thereby promoting the circulation of CNP compounds in the bloodstream. This has the effect of extending the plasma exposure of CNP compounds and prolonging their plasma half-lives compared to the plasma half-lives of CNP-22 (SEQ ID NO: 01) and CNP-37 (SEQ ID NO: 02). For this reason, modifying groups that have the effect of prolonging and extending the duration of action of CNP compounds are sometimes also called extension groups.

[0042] In some embodiments of the present invention, the modifying group is covalently bonded to the N-terminal amino acid of the CNP peptide.

[0043] In some embodiments of the present invention, the modifying group is covalently bonded to the N-terminal alpha-amine of the CNP peptide.

[0044] In one aspect of the present invention, the modifying group comprises Chem.A, Chem.B, and Chem.C, where Chem.A is [ka] (In the formula, p is an integer in the range of 14 to 20, * is selected from the group consisting of (where * represents the amide bond connecting Chem.A and Chem.B), Chem.B [ka] (In the formula, q is an integer in the range of 1 to 8, * represents the amide bond connecting Chem.A- and Chem.B-. ** is selected from the group consisting of (where ** represents an amide bond connecting Chem.B- and Chem.C-), Chem.C [ka] (In the formula, r is an integer in the range of 0 to 4, s is an integer in the range of 0 to 3. t is an integer in the range of 0 to 1, u is an integer in the range of 0 to 3. ** represents the amide bond connecting Chem.B- and Chem.C-, *** is selected from the group consisting of (where *** represents the amide bond connecting Chem.C- and the N-terminal alpha-amine on the CNP peptide).

[0045] The following are abbreviations used to modify group elements in this application. [Table 2]

[0046] In some embodiments of the present invention, the modifying group is, in non-limiting examples, Chem.E, Chem.F, Chem.G, Chem.H, Chem.I, and Chem.J [ka] [ka] (In the formula, the dotted line defines the bond to the CNP peptide via an amide bond) is selected from the following.

[0047] The modifying groups of the present invention may exist in different stereoisomeric forms, having the same molecular formula and arrangement of bonded atoms, but differing only in the three-dimensional orientation of those atoms in space. The stereoisomers of the illustrated modifying groups of the present invention are shown in the experimental section, by name and structure, using standard nomenclature. Unless otherwise specified, the present invention relates to all stereoisomeric forms of the claimed modifying groups.

[0048] Functional characteristics In some aspects of the present invention, the CNP compound has desirable biophysical properties. The CNP compound exhibits an extended in vivo half-life. Furthermore, the CNP compound of the present invention is biologically active. In addition, the compound of the present invention has desirable stability. Furthermore, the compound of the present invention has desirable solubility. Furthermore, the compound of the present invention can be administered subcutaneously without inducing undesirable levels of local tissue reactions such as necrosis.

[0049] biological activity In one aspect of the present invention, the compound is biologically active in vitro, and this biological activity may be determined as the ability to activate the NPR2 receptor in vitro in cell lines expressing the NPR2 receptor, and this ability may be determined as the ability to increase the production and concentration of cGMP, which represents downstream receptor activation of NPR2 in cells. The biological activity in vitro may be determined, for example, as described in Example 2.

[0050] In one aspect of the present invention, the compound is biologically active in vivo, and this biological activity may be determined as the ability to increase the concentration of cGMP in plasma after intravenous or subcutaneous administration in rats, Göttingen miniature pigs, or domestic LYD pigs.

[0051] Rats are an example of a suitable animal model, and activity may be determined in vivo in such rats, as described in Example 11.

[0052] Göttingen miniature pigs or domestic LYD pigs are other examples of suitable animal models, and activity may be determined in vivo in such pigs as described in Example 12.

[0053] Plasma half-life In one aspect of the present invention, the CNP compound exhibits extended plasma exposure and a prolonged in vivo plasma half-life compared to natural CNP, which can be determined in suitable in vivo pharmacokinetic studies. The prolonged plasma exposure may be determined as the plasma half-life (T1 / 2) after intravenous or subcutaneous administration to animals such as rats, Göttingen miniature pigs, or domestic LYD pigs.

[0054] In some embodiments of the present invention, the plasma half-life of the CNP compound after intravenous administration to rats is determined as described in Example 11 to be at least 4 hours, more preferably at least 8 hours, or most preferably at least 10 hours.

[0055] In some embodiments, the in vivo plasma half-life of the CNP compound after intravenous or subcutaneous administration to Göttingen miniature pigs is determined as described in Example 12 to be at least 10 hours, more preferably at least 40 hours, or most preferably at least 60 hours.

[0056] Bioavailability In one aspect of the present invention, the compound has suitable bioavailability after subcutaneous injection. The bioavailability (F%) may be determined in any suitable animal model as is known in the art.

[0057] Göttingen miniature pigs or domestic LYD pigs are examples of suitable animal models, and bioavailability may be determined in vivo in such Göttingen miniature pigs or domestic LYD pigs, as described in Example 12.

[0058] In some embodiments of the present invention, the CNP compound has a bioavailability of at least 40%, 50%, 60%, or 70% after subcutaneous administration to Göttingen miniature pigs.

[0059] stability In one aspect of the present invention, the compound has suitable physical and chemical stability. A lack of physical or chemical stability can lead to structural changes in the compound, resulting in the formation of chemical degradation products that may have reduced biological activity, decreased solubility, and / or increased immunogenicity compared to the intact compound.

[0060] Physical stability can be evaluated, for example, by measuring the tendency for fibril formation using the ThT assay described in Example 6.

[0061] Chemical stability can be evaluated, for example, by measuring the amount of chemical decomposition products (isomers, isoAsp, and hydrolysis products, etc.) at various time points after exposure to different conditions, such as increased temperature, as described in Example 8.

[0062] Stability can also be evaluated by examining the molecular ability to form covalent dimers and polymers called HMWPs, as described in Example 7, for example.

[0063] solubility According to the functional aspects of the present invention, the CNP compound has the desired solubility. Low solubility of CNP significantly impairs the properties and therapeutic applications of its pharmaceutical formulations; therefore, developing CNP compounds with high solubility in relevant formulations will improve therapeutic usefulness.

[0064] As described herein, the solubility is measured as described in Example 5. In certain embodiments, the CNP compound of the present invention has a solubility of at least 2000 μM, 3000 μM, or 4000 μM at either pH 4 or pH 6.5.

[0065] Pharmaceutical composition In one embodiment, the present invention relates to a pharmaceutical composition comprising a compound according to the present invention and a pharmaceutically acceptable excipient. The composition is suitable for parenteral administration.

[0066] In one embodiment, the compound is present in the formulation at a concentration of approximately 0.1 mg / ml to approximately 50 mg / ml. In another embodiment, the compound is present in the formulation at a concentration of approximately 10 mg / ml to approximately 30 mg / ml. In another embodiment, the formulation has a pH of 3.0 to 8.0. In yet another embodiment, the formulation has a pH of 3.5 to 5.5. In a further embodiment, the formulation has a pH of 6.0 to 7.0. Pharmaceutical compositions containing compounds according to the present invention can be prepared by conventional techniques, for example, Remington's Pharmaceutical Sciences, 1985 or Remington: The Science and Practice of Pharmacy, 1985. th It may be prepared as described in edition, 1995.

[0067] This formulation may further contain a buffer system, preservatives, isotonic agents, chelating agents, stabilizers, and / or surfactants. The use of such excipients in pharmaceutical compositions is known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy, 19. th See edition 1995.

[0068] In one embodiment, the pharmaceutical formulation is a liquid formulation, i.e., a formulation containing water. Such formulations are typically solutions or suspensions. In a further embodiment of the present invention, the pharmaceutical formulation is an aqueous solution. The term “aqueous formulation” is defined as a formulation containing at least 50% w / w water. Similarly, the term “aqueous solution” is defined as a solution containing at least 50% w / w water, and the term “aqueous suspension” is defined as a suspension containing at least 50% w / w water.

[0069] In another embodiment, the pharmaceutical formulation is a lyophilized formulation to which a physician or patient adds a solvent and / or diluent before use. In another embodiment, the pharmaceutical formulation is a dry (e.g., lyophilized or spray-dried) formulation ready for use without any prior dissolution. "Dry form" refers to a liquid pharmaceutical composition or formulation that has been freeze-dried (i.e., lyophilized, see Williams and Polli (1984) J. Parenteral Sci. Technol. 38:48-59) or spray-dried (see Masters (1991) in Spray-Drying Handbook (5)). th It is intended to be dried by either Longman Scientific and Technical, Essez, UK (ed.), pp. 491-676, Broadhead et al. (1992) Drug Devel. Ind. Pharm. 18:1169-1206, and Mumenthaler et al. (1994) Pharm. Res. 11:12-20, or by air drying (see Carpenter and Crowe (1988) Cryobiology 25:459-470, and Roser (1991) Biopharm. 4:47-53).

[0070] In further embodiments of the present invention, the buffer is selected from the group consisting of acetate, carbonate, citrate, glycylglycine, histidine, glycine, lysine, arginine, dihydrogen phosphate, hydrogen phosphate, phosphate, and tris(hydroxymethyl)aminomethane, bicine, tricine, malic acid, lactic acid, succinate, maleic acid, fumaric acid, tartaric acid, and aspartic acid, or mixtures thereof. Each of these specific buffers constitutes an alternative embodiment of the present invention.

[0071] In another embodiment of the present invention, the formulation further comprises a pharmaceutically acceptable preservative. In a further embodiment of the present invention, the formulation further comprises an isotonic agent, such as propylene glycol, mannitol, or glycerol. In a further embodiment of the present invention, the formulation further comprises a chelating agent.

[0072] In another embodiment of the present invention, the formulation further comprises a stabilizer. The use of stabilizers in pharmaceutical compositions is known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy, 19 th See edition 1995.

[0073] The pharmaceutical composition of the present invention may further contain an amount of amino acid bases sufficient to reduce the aggregation of peptides during storage of the composition.

[0074] "Amino acid base" means an amino acid, or a combination of amino acids in which any given amino acid exists either in the form of its free base or a salt thereof. When a combination of amino acids is used, all amino acids may exist in the form of their free base, all may exist in the form of their salts, or some may exist in the form of their free bases and others in the form of salts. In one embodiment, the amino acids used in the preparation of the compositions of the present invention are those having charged side chains, such as arginine, lysine, aspartic acid, and glutamic acid. Any stereoisomer (i.e., L, D, or a combination thereof) of a particular amino acid (e.g., methionine, histidine, imidazole, arginine, lysine, isoleucine, aspartic acid, tryptophan, threonine, and mixtures thereof), or combinations of these stereoisomers, may be present in the pharmaceutical compositions of the present invention, insofar as the particular amino acid exists either in the form of its free base or a salt thereof. In one embodiment, the L-stereoisomer is used. The compositions of the present invention may also be formulated using derivatives of these amino acids. Suitable arginine derivatives include, for example, aminoguanidine, ornithine, and N-monoethyl L-arginine; suitable methionine derivatives include ethionine and butionine; and suitable cysteine ​​derivatives include S-methyl-L-cysteine. As with other amino acids, amino acid derivatives are incorporated into the composition either in the form of their free bases or their salts. In another embodiment of the present invention, an amino acid or an amino acid derivative is used in a concentration sufficient to prevent or delay protein aggregation.

[0075] In another embodiment of the present invention, the formulation further comprises a surfactant. In another embodiment of the present invention, the formulation further comprises a protease inhibitor. The use of a protease inhibitor is particularly useful in pharmaceutical compositions comprising a protease enzyme precursor to inhibit autocatalysis.

[0076] Other components may be present in the pharmaceutical formulation of the present invention. Such additional components may include wetting agents, emulsifiers, antioxidants, fillers, tonicity modifiers, chelating agents, metal ions, oily excipients, proteins (e.g., human serum albumin, gelatin, or proteins) and zwitterions (e.g., amino acids such as betaine, taurine, arginine, glycine, lysine, or histidine). Such additional components should, of course, not adversely affect the overall stability of the pharmaceutical formulation of the present invention.

[0077] Pharmaceutical compositions according to the present invention may be administered to patients requiring such treatment at several sites, including local sites (e.g., skin and mucous membrane sites), sites that bypass absorption (e.g., intra-arterial, intra-venous, intracardiac administration), and sites involved in absorption (e.g., intracutaneous, subcutaneous, intramuscular, or intraabdominal administration).

[0078] The administration of the pharmaceutical composition according to the present invention can be obtained to a patient requiring such treatment via several routes of administration, e.g., by tongue, sublingual, buccal, oral, oral, gastrointestinal, nasal, lung, e.g., bronchi and alveoli or a combination thereof, epidermal, dermal, transdermal, transvaginal, rectal, oral cavity, e.g., conjunctiva, and parenterally.

[0079] The compositions of the present invention can be administered in several drug forms, for example, as solutions, suspensions, emulsions, microemulsions, multilayer emulsions, foams, ointments, pastes, plasters, ointments, tablets, coated tablets, rinses, capsules, such as hard gelatin capsules or soft gelatin capsules, suppositories, rectal capsules, drops, gels, sprays, powders, aerosols, inhalants, eye drops, eye ointments, eye washes, pessaries, vaginal rings, vaginal ointments, injections, insight conversion solutions, such as insight gelation, insight coagulation, insight precipitation, or insight crystallization, infusions, and implants.

[0080] The compositions of the present invention may be further formulated or coupled to drug carriers, drug delivery systems, and advanced drug delivery systems, for example, via covalent, hydrophobic, or electrostatic interactions, to further enhance the stability of the compound, increase bioavailability, increase solubility, mitigate adverse effects, achieve chronotherapy known to those skilled in the art, and improve patient medication adherence, or any combination thereof.

[0081] The compositions of the present invention are useful for formulations of solids, semi-solids, powders, and solutions for pulmonary administration of the CNP compounds of the present invention, using, for example, metered-dose inhalers, dry powder inhalers, and nebulizers, all of which are known to those skilled in the art.

[0082] The compositions of the present invention are useful for formulations of controlled, sustained, extended, delayed, and sustained-release drug delivery systems.

[0083] Parenteral administration may be carried out by subcutaneous, intramuscular, intraperitoneal, or intravenous injection using a syringe, optionally a pen-like syringe. Alternatively, parenteral administration may be carried out using an infusion pump. A further option is a composition that may be a solution or suspension for administration of the CNP compound of the present invention in the form of a nasal spray or lung spray. As a further option, the pharmaceutical composition containing the peptide of the present invention may also be adapted for transdermal administration, for example, by needleless injection, or by patch, optionally an iontophoresis patch, or by transmucosal administration, for example, oral administration.

[0084] The CNP compounds of the present invention can be administered via the pulmonary route in a vehicle as a solution, suspension, or dry powder using any known type of apparatus suitable for pulmonary drug delivery. Examples, but not limited to, of these include three common types of aerosol generation for pulmonary drug delivery, which include jet or ultrasonic nebulizers, metered-dose inhalers, or dry powder inhalers (see Yu J, Chien YW. Pulmonary drug delivery: Physiologic and mechanistic aspects. Crit Rev Ther Drug Carr Sys 14(4)(1997) 395-453).

[0085] In one embodiment of the present invention, a pharmaceutical formulation containing the compound of the present invention is stable for use for more than 6 weeks and for storage for more than 3 years. In another embodiment of the present invention, a pharmaceutical formulation containing the compound of the present invention is stable for use for more than 2 weeks and for storage for more than 2 years.

[0086] In one embodiment, a process for preparing a pharmaceutical composition comprising a compound according to the present invention comprises mixing the compound according to the present invention with at least one pharmaceutically acceptable excipient.

[0087] Indications The present invention also relates to compounds for use as pharmaceuticals.

[0088] As used herein, the term “treatment” refers to any medical treatment for a human subject who requires it. The timing and purpose of such treatment may vary from person to person, depending on the subject’s health condition. Such treatment may be preventive, palliative, symptomatic, and / or curative.

[0089] In one embodiment, the compounds of the present invention may be used in the treatment of cardiorenal metabolic diseases. Cardiorenal metabolic diseases include, but are not limited to, hypertension, arteriosclerosis, atherosclerosis, restenosis, acute myocardial infarction, pulmonary hypertension, acute decompensated heart failure, congestive heart failure, cardiac edema, renal edema, hepatic edema*, acute renal failure, chronic renal failure, insulin resistance, and type 2 diabetes.

[0090] In one embodiment, the compound of the present invention is used to treat congestive heart failure.

[0091] In one embodiment, the compound of the present invention is used to treat acute decompensated heart failure.

[0092] In one embodiment, the compounds of the present invention may be used to treat growth disorders, including short stature, which may be associated with FGFR3-related osteodysplasia and related comorbidities.

[0093] In some embodiments, the compounds of the present invention may be used to treat diseases selected from the group consisting of chondrodysplasia, hypochondrodysplasia, lethal dysplasia type 1 and type 2, SHOX deficiency, Noonan syndrome, Costello, LEOPARD syndrome, idiopathic short stature, autosomal dominant short stature, growth hormone deficiency, hypophosphatemic rickets, CNP deficiency, aggrecan deficiency, heterozygous NPR2 mutation, osteoarthritis, craniosynostosis (e.g., Muwenke syndrome, Crouzon syndrome, Apert syndrome, Jackson-Weiss syndrome, Pfeiffer syndrome, or Crouzon syndrome with acanthosis nigricans), Lacrimo-Auriculo-Dento-Digital syndrome (LADD), syringodysplasia, and SADDAN (severe chondrodysplasia with delayed developmental acanthosis nigricans).

[0094] In some embodiments, the compounds of the present invention may be used to treat diseases selected from the group consisting of osteogenesis imperfecta, achondroplasia, chondrodysplasia punctata, homozygous chondrodysplasia punctata, rhomboid chondrodysplasia punctata, congenital spondyloepiphyseal dysplasia, congenital short femur, Langer's intermediate limb dysplasia, neurofibromatosis, Regius syndrome, and neurofibromatosis type 1.

[0095] In some embodiments, the compounds of the present invention may be used to treat phenotypes associated with growth disorders, including short stature, which may be associated with FGFR3-related osteodysplasia, selected from the group consisting of growth retardation, cranial deformity, corrective defects, cervical spinal cord compression, spinal cord stenosis, pain associated with skeletal dysplasia, hydrocephalus, hearing loss due to chronic otitis, cardiovascular disease, neurological disorders, and obesity.

[0096] Unless otherwise indicated herein, terms presented in the singular form may encompass multiple contexts.

[0097] List of embodiments The present invention can be further described by the following non-limiting embodiments.

[0098] Embodiment 1: A CNP compound comprising a CNP peptide and a modifying group, wherein the net charge of the compound at physiological pH is 0 or negative, and the CNP peptide is of formula I: AA 01 -AA 02 -AA 03 -AA 04 -AA 05 -AA 06 -AA 07 -AA 08 -AA 09 -AA 10 -AA 11 -AA 12 -AA 13 -AA 14 -AA 15 -AA 16 -AA 17 -AA 18 -AA 19 -AA 20 -AA 21 -AA 22 -AA23 -AA 24 -AA 25 -AA 26 -AA 27 -AA 28 -AA 29 -AA 30 -AA 31 -AA 32 -AA 33 -AA 34 -AA 35 -AA 36 -AA 37 (In the formula, AA 01 However, it is either Gln or absent, AA 02 However, it is either Glu or absent, AA 03 However, His is either absent or not. AA 04 However, whether they are Pro or absent, AA 05 However, it is either Asn, Gln, or Glu, or it is absent. AA 06 However, Ala is either absent or not present. AA 07 However, it is either Arg, His, or Ala, or absent. AA 08 However, it is either Lys, Ser, or His, or absent. AA 09 However, it is either Tyr or Glu, or absent. AA 10 However, it is either Lys, Glu, Gln, or His, or it is absent. AA 11 However, it is Gly, AA 12 However, it is Ala, AA 13 However, it is Gln or Asn or Glu, AA 14 However, it is either Lys, His, or Glu. AA 15However, it is Lys, Ser or Glu or Thr or His, AA 16 However, it is Gly, AA 17 However, it is Leu or Gly or Ser or Val, AA 18 However, it is either Ser or His, AA 19 However, it is Gln or Ser or Lys or His, AA 20 However, it is Gly, AA 21 However, it is Cys, AA 22 However, it is Phe, AA 23 However, it is Gly, AA 24 However, it is Leu, AA 25 However, it is either Pro or Lys. AA 26 However, it is Leu, AA 27 However, it is either Asp or Glu, AA 28 However, it is Arg, AA 29 However, it is Ile, AA 30 However, it is Gly, AA 31 However, it is Ser, AA 32 However, it is Leu or Nle or Met, AA 33 However, it is Ser, AA 34 However, it is Gly, AA 35 However, it is Leu, AA 36 However, it is Gly, AA 37 However, it contains an amino acid sequence that follows Cys. The modifying groups include Chem.A, Chem.B, and Chem.C. Chem.A [Chemical formula] (where p is an integer in the range of 14 - 20, * represents an amide bond connecting Chem.A and Chem.B) and is selected from the group consisting of Chem.B is Chem.B1 and Chem.B2 [Chemical formula] (where q is an integer in the range of 1 - 8, * represents an amide bond connecting Chem.A- and Chem.B-, ** represents an amide bond connecting Chem.B- and Chem.C-) and is selected from the group consisting of Chem.C is [Chemical formula] (where r is an integer in the range of 0 - 4, s is an integer in the range of 0 - 3, t is an integer in the range of 0 - 1, ** represents an amide bond connecting Chem.B- and Chem.C-, *** represents an amide bond connecting Chem.C- and the N-terminal alpha-amine on the CNP peptide) and is selected from the group consisting of, a CNP compound. Embodiment 2: AA of the CNP peptide 13 is Gln, AA 19 is Gln or Ser, AA 25 is Pro, AA 32 is Leu, the CNP compound according to Embodiment 1. Embodiment 3: AA5 of the CNP peptide is Gln, AA 13 is Gln, AA 19 is Gln or Ser, AA 25 is Pro, AA 32 is Leu, the CNP compound according to Embodiment 1. Embodiment 4: The CNP compound according to Embodiment 1, wherein the CNP peptide comprises one of the following amino acid sequences. [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5] Embodiment 5: The CNP compound according to Embodiment 1, wherein the CNP peptide has one of the following amino acid sequences. [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6] Embodiment 6: The CNP peptide has the following amino acid sequence: GAQKKGSSQGCFGLPLDRIGSLSGLGC (Sequence ID 136), ARKYKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 77), YKGAQKKGGSQGCFGLPLDRIGSLSGLGC (Sequence ID 88) YKGAQKKGLSQGCFGLPLDRIGSLSGLGC (Sequence ID 103), QEHPQARKYKGAQKKGLSSGCFGLPLDRIGSLSGLGC (SEQ ID NO: 67), and the CNP compound according to Embodiment 1 having any one of QEHPQARKYKGAQKKGLSSGCFGLPLERIGSLSGLGC (SEQ ID NO: 104). Embodiment 7: The CNP peptide has the following amino acid sequence: the CNP compound according to Embodiment 1 having QEHPQARKYKGAQKKGLSSGCFGLPLDRIGSLSGLGC (SEQ ID NO: 67). Embodiment 8: The CNP compound according to any one of the preceding embodiments, wherein the modifying group is covalently bonded to the N-terminal alpha-amine of the CNP peptide. [[ID=IO]] Embodiment 9: The CNP compound according to any one of the preceding embodiments, wherein Chem. A of the modifying group has 16 p's. Embodiment 10: Chem. A of the modifying group is Chem. A2

Chemical formula

Chemical formula

Chemical formula

[0099] material and method List of Abbreviations 2-ClTrt 2-chlorotrityl resin Aldrithiol-4 4,4'-Dipyridyl disulfide Area under the AUC curve BEH Ethylene Crosslinked Hybrid Boc tert-butyloxycarbonyl C18d Octadecanoic Acid C20d Eicosanedioic Acid CAD charged aerosol detector cGMP Cyclic Guanosine Monophosphate CL Clearance CSH Charged Surface Hybrid DCM Dichloromethane DgGlu D-configured gamma-glutamoyl DIC N,N-diisopropylcarbodiimide DMEM Dulbecco's modified Eagle medium DMF Dimethylformamide DMSO (Dimethyl Sulfoxide) DTT (Dithiothreitol) EDTA (Ethylenediaminetetraacetic acid) ESI Electrospray Ionization F% Bioavailability Fd Faraday Fmoc Fluorenylmethyloxycarbonyl FPLC High-Speed ​​Protein Liquid Chromatography gGlu gamma-glutamoyl HEPES 2-[4-(2-hydroxyethyl)piperazine-1-yl]ethane-1-sulfonic acid HFIP Hexafluoro-2-propanol HMWP (High Molecular Weight Protein) HSA Human Serum Albumin iv intravenous IPA Isopropanol LCMS (Liquid Chromatography Mass Spectrometry) LLOQ Lower limit of quantification LYD Landrace, Yorkshire, and Duroc MeCN acetonitrile MMPX 8-Methoxymethyl-3-Isobutyl 1-Methylxanthine MS mass spectrometry No NAD anomalies detected. NCA Non-Compartmental Analysis No nd detections found. Nle (X) Norleucine NMP N-methyl-2-pyrrolidone OEG Oligoethylene Glycol Oxyma Pure Cyanohydroxyiminoethyl Acetate Pbf 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl PBS (phosphate-buffered saline) PD Pharmacodynamics PDA Photodiode Array Detection PK (Pharmacokinetics) QC quality control RP inverse phase RPM (Revolutions per minute) sc subcutaneous SEC size exclusion chromatography SIL stable isotope labeling SPPS Solid-Phase Peptide Synthesis T1 / 2 Half-life tBu tert-butyl Tetrazole-C18 17-(2H-tetrazole-5-yl)heptadecanoic acid TF time-of-flight type TFA (Trifluoroacetic Acid) Thioflavin T TIPS Triisopropylsilane Tris 2-amino-2-(hydroxymethyl)propane-1,3-diol Trt Trichl TUV Adjustable UV UPLC Ultra-High-Speed ​​Liquid Chromatography UV ultraviolet light

[0100] General preparation method Method A - Solid-phase peptide synthesis The peptides can be synthesized by solid-phase peptide synthesis known in the art, and the method used to synthesize exemplary compounds of the present invention is described below.

[0101] The compounds were synthesized using the SymphonyX solid-phase peptide synthesizer (GYROS Protein Technologies AB, Tucson, AZ).

[0102] Typically, the synthesis was carried out using 450 μmol of resin, although synthesis using 150 or 300 μmol of resin was also performed. In a typical synthesis, the resin was washed with DMF. After washing with DMF, the resin was incubated with Fmoc-protected amino acids (5 equivalents, 0.3 M in Oxyma Pure in DMF, 7.5 mL), DIC (5 equivalents, 0.75 M in DMF, 3.0 mL), and colidine (5 equivalents, 0.75 M in DMF, 3.0 mL) for 30 minutes. Then, additional DIC (5 equivalents, 0.75 M in DMF, 3.0 mL) was added, and the resin was incubated for 90 minutes. After each coupling, the resin was capped by treating it with acetic anhydride (1 M, 7.5 mL) and colidine (0.75 M, 3.0 mL) in DMF for 20 minutes. Fatty acids were coupled for 6 hours using fatty acid mono-tert-butyl ester (5 equivalents, 0.3 M in 0.3 M Oxyma Pure, 7.5 mL), DIC (5 equivalents, 0.75 M in DMF, 3.0 mL), and colidine (5 equivalents, 0.75 M in DMF, 3.0 mL).

[0103] The Fmoc-protected amino acids used are recommended standards supplied from, for example, Anaspec, Bachem, Iris Biotech, or NovabioChem: Fmoc-Ala-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Asn(Trt)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Cys(Trt)-OH , Fmoc-Gln(Trt)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Gly-OH, Fmoc-His(Trt)-OH, Fmoc-lle-OH, Fmo c-Leu-OH, Fmoc-Lys(Boc)-OH, Fmoc-Met-OH, Fmoc-Phe-OH, Fmoc-Pro-OH, Fmoc-Ser(tBu)-OH, F They were moc-Thr(tBu)-OH, Fmoc-Trp(Boc)-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Val-OH, and Fmoc-Nle-OH. Unless otherwise specified, the naturally occurring L-form of the amino acid was used. If serine or threonine is present in the peptide, a pseudo-proline dipeptide may be used (see also WRSampson et al., J.Pept.Sci. 1999, 5, 403-409).

[0104] For peptides acylated at any position in SPPS, the following preferred protected structural blocks, such as but not limited to Fmoc-8-amino-3,6-dioxaoctanoic acid and Boc-Glu(Fmoc)-OH, were supplied, for example, from Anaspec, Bachem, Iris Biotech, or Novabiochem.

[0105] Octadecanedioic acid mono-tert-butyl ester and eicosanedioic acid mono-tert-butyl ester were prepared as known in the art, for example, as described in WO2010 / 102886(A1).

[0106] 17-(2H-tetrazole-5-yl)heptadecanoic acid (tetrazole-C18) was incorporated by coupling it with (S)-4-(17-(1H-tetrazole-5-yl)heptadecanamide)-5-(tert-butoxy)-5-oxopentanoic acid, a protected structural block synthesized according to the following procedure. 5-chloro-1-((dimethylamino)(dimethylimino)methyl)-1H-benzo[d][1,2,3]triazole 3-oxide tetrafluoroborate (TCTU, 5.87 g, 16.5 mmol) and N-methylmorpholine (NMM, 8.00 mL, 72.8 mmol) were sequentially added to a stirred suspension of 17-(1H-tetrazole-5-yl)heptadecanoic acid (5.10 g, 15.0 mmol) in dry DMF (70 mL). The mixture was stirred at room temperature for 30 hours. HL-Glu-OtBu (1-tert-butyl glutamate, 4.58 g, 22.5 mmol) was added all at once. The suspension turned a yellow solution in 5 minutes. After standing at room temperature for 4 hours, it was poured into an ice-cold, stirred solution of hydrochloric acid (35%, 20 mL) in water (1 L). The crude product was filtered off, washed with water (3 × 100 mL), briefly dried on glass sintered frit by suction, and redissolved in DMF (70 mL). This solution was poured into water (1 L), the resulting suspension was stirred for 5 minutes, the solid was filtered off, washed with water (3 × 100 mL), and dried in vacuum. The crude product was dissolved in ethyl acetate (400 mL), filtered through a silica gel (Silicagel 60, 0.040~0.063 mm) column (100 g), and the column was washed with ethyl acetate (600 mL). The ethyl acetate was removed in vacuum. The solid residue was boiled in 1,2-dichloroethane (200 mL) with vigorous stirring for 5 minutes, the emulsion was cooled to room temperature in 5 minutes, and crystallized overnight at the same temperature. The waxy crystals were filtered off, washed with 1,2-dichloromethane (50 mL) and n-heptane (100 mL), and air-dried. This purification process was repeated once more. Finally, the compound was dissolved in 1,4-dioxane (50 mL), freeze-dried to obtain (S)-4-(17-(1H-tetrazole-5-yl)heptadecanamide)-5-(tert-butoxy)-5-oxopentanoic acid as an off-white powder.

[0107] For peptides characterized by a C-terminal cysteine, the resin used was pre-filled H-Cys(Trt)-2-ClTrt (e.g., from Bachem or Novabiochem).

[0108] Method B - Peptide cleavage from resin. After synthesis, the resin was washed with DCM, and the peptide was cleaved by treatment with TFA / TIPS / water / DTT (e.g., in a ratio of 90:5:2.5:2.5 or 92.5:2.5:2.5) for 2-3 hours, followed by precipitation with diethyl ether. The precipitate was centrifuged and washed several times with diethyl ether.

[0109] Method: C-disulfide crosslinking Natural CNPs contain disulfide crosslinks, which, when reduced, eliminate their binding to the NPR2 receptor. Methods for disulfide crosslink formation for synthetic peptides are known in the art, and three methods used to prepare exemplary compounds of the present invention are described below.

[0110] Folding using Aldrichio-4 The crude peptide precipitate from 150 μmol synthesis was dissolved in 10 ml of DMSO, followed by the addition of water to make 700 mL, and then 100 mL of MeCN was added. Finally, 2 mL of 0.5 M NaOAc buffer pH 5.0 was added to the peptide solution. The pH was adjusted to 5-6.5 with 1 M NaOH until the solution was clear. The amount of peptide in the solution was quantified using CAD, and a fresh stock solution of aldrithiol-4 (2 mg / ml in MeOH) was prepared. Compared to the peptide content, 1.0 equivalent of aldrithiol-4 in MeOH was added over 5-10 minutes with vigorous stirring. The solution was stirred for 10 minutes, and then another 0.5 or 1.0 equivalent of aldrithiol-4 stock solution was added. Once completion was determined by LC-MS, the reaction was quenched with 2 mL of TFA. The peptide was purified by RP-HPLC as described below.

[0111] Folding using DMSO The crude peptide precipitate from the 450 μmol synthesis was dissolved in 1:1 MeCN / water (50 mL), added to 500 mM Tris-HCl pH 8.0 buffer (1 L), and diluted with water (1.4 L) and DMSO (600 mL). The solution was allowed to stand at room temperature for 16-24 hours with gentle stirring. The reaction was quenched by adding 5.0 equivalents of iodoacetamide (50 mM in water), and after 10 minutes, the pH was acidified to 2 with TFA and diluted to 5 L with water. The peptide was purified by RP-HPLC as described below.

[0112] Folding using cysteine / cystine or cysteamine / cystamine redox buffer. The crude peptide precipitate from the 150 μmol synthesis was dissolved in DMSO (20 mL) and added to a buffer of 50 mM Tris-HCl, 3 mM cysteine, and 0.3 mM cystine pH 8.2 (800 mL), or a buffer of 50 mM Tris-HCl, 3 mM cysteamine, and 0.3 mM cystamine pH 8.2. The solution was allowed to stand at room temperature for 16–24 hours with gentle stirring. The reaction product was quenched by adding 3 mL of TFA, diluted in 100 mL of MeCN, and purified by RP-HPLC as described below.

[0113] Method D - Purification Methods for purifying peptides are known in the art, and two methods used to purify the exemplary compounds of the present invention are described below.

[0114] Reverse-phase preparative HPLC using an acidic eluent Crude folded peptides were purified by preparative reverse-phase HPLC on a Waters Delta Prep 4000 or Gilson Model 322 H2 system equipped with a C18 column, e.g., Waters Xbridge Prep C18 OBD, 5 μm, 250 × 50 mm or Phenomenex Gemini Axia NX C18, 5 μm, 250 × 30 mm. The peptides were then subjected to a linear gradient of eluent A (0.1% TFA in water) and B (0.1% TFA in acetonitrile), e.g., 27–42% of eluent B over 30 minutes at a constant flow rate of 25 mL / min.

number

number

[0115] Reverse-phase preparative HPLC using a nearly neutral eluent. If further purification was required, the lyophilized TFA peptide salt was purified using a similar preparative HPLC system in neutral or near-neutral pH eluents, e.g., Eluent A: 100 mM NaOAc pH 6.5, 5% MeCN, Eluent B: MeCN. The peptide was then eluted using a linear gradient of eluents A and B, e.g., eluent B at a constant flow rate of 60 mL / min over 30 minutes, eluting 25–55% of the peptide.

number

[0116] Method E - Resalting and Desalting In some cases, it was desirable to change the counterion to sodium to facilitate formulation at pH 6.5. Methods for exchanging peptide counterions are known in the art, The following describes methods used to modify the counterions of some of the exemplary compounds of the present invention.

[0117] Size removal and desalting The TFA peptide salt was solubilized at 400 mM NaOAc pH 6.5 and 20% MeCN at a concentration of 2-3 mg / mL, and counterions were exchanged. The dissolved peptide was then desalted using a HiPrep 26 / 10 desalting column in an Akta Purifer FPLC system. The column was pre-equilibrated with 20% MeCN, the peptide was injected, and eluted at a uniform concentration of 1.5 CV of 20% MeCN at 6 mL / min. The peptide was collected, quantified by CAD analysis, and lyophilized to obtain a colorless powder of the sodium peptide salt.

[0118] General methods for detection and characterization To verify their identity, measure their purity, and quantify the amount of synthetic peptides prepared, the peptides were characterized using methods known in the art, such as LC-MS and reversed-phase ULC. The methods used to characterize the exemplary compounds of the present invention are described below.

[0119] Method F - Analytical chromatography method The prepared CNP compounds were characterized by RP-UPLC and LCMS, and the amount prepared was quantified by RP-UPLC / CAD.

[0120] LCMS34 LC-MS was performed using a setup consisting of an ACQUITY UPLC system and a Xevo G2-XS QTOF mass spectrometer. The system was equipped with a Waters ACQUITY UPLC BEH C18 column, 1.7 μm, 2.1 mm × 50 mm, at a column temperature of 60°C. UV detection was performed at 214 nm. The MS ionization mode was ESI+.

[0121] Eluent A: 0.1% formic acid in water, Eluent B: 0.1% formic acid in acetonitrile. The analysis was performed by injecting an appropriate volume of sample (preferably 1-10 μL) into the column and eluting it over 4.0 minutes at a constant flow rate of 0.4 mL / min with a linear gradient of eluent B to 5-95% of eluent A and B.

[0122] RP-UPLC107: Alternatively, LC-MS was performed using a setup consisting of a Waters ACQUITY UPLC system and a Waters QDA mass detector. The system was equipped with a Waters ACQUITY UPLC BEH C18 column, 1.7 μm, 2.1 mm × 50 mm, at a column temperature of 40°C. UV detection was performed at 214 nm.

[0123] Eluent A: 0.05% TFA in water, Eluent B: 0.05% TFA in acetonitrile. The analysis was performed by injecting an appropriate volume of sample (preferably 1-10 μL) into the column and eluting it with a linear gradient of 5-60% eluent B to eluent A and B over 1.6 minutes at a constant flow rate of 0.9 mL / min.

[0124] RP-UPLC02: RP-UPLC02 was performed using a Waters ACQUITY UPLC system equipped with a TUV or PDA detector. The system was fitted with a Waters ACQUITY UPLC BEH C18 column, 1.7 μm, 2.1 mm × 150 mm, column temperature 40°C. UV detection was performed at 214 nm. Eluent A: 0.05% TFA in water, Eluent B: 0.05% TFA in acetonitrile.

[0125] The analysis was performed by injecting an appropriate volume of sample (preferably 1-10 μL) into the column and eluting it at a constant flow rate of 0.4 mL / min for 16.0 minutes with a linear gradient between eluent A and eluent B, representing 5-95% of the sample.

[0126] RP-UPLC61: RP-UPLC61 was performed using a Waters ACQUITY UPLC system equipped with a TUV or PDA detector. The system was fitted with a Waters ACQUITY UPLC BEH Shield C18 column, 1.7 μm, 2.1 mm × 150 mm, column temperature 60°C. UV detection was at 214 nm. Eluent A: 20 mM Na2SO4, 2 mM Na2HPO4, 2 mM NaH2PO4 in 9:1 water / MeCN at pH 7.2; Eluent B: 7:3 MeCN / water. The analysis was performed by injecting an appropriate volume of sample (preferably 1–10 μL) into the column and eluting it with a linear gradient of eluents A and B at a constant flow rate of 0.4 mL / min: 5–20% B over 3 minutes, then 20–80% B over 17 minutes, then 80–90% B over 1 minute.

[0127] CAD02 peptide quantification: CAD was performed using a Thermo Scientific Vanquish UPLC system equipped with UV-DAD and CAD (charged aerosol detector). The system was fitted with a Waters ACQUITY UPLC CSH C18 column, 1.7 μm, 2.1 mm × 50 mm, column temperature 40°C. UV detection was at 214 nm. Eluent A: 0.1% v / v TFA in water, Eluent B: 0.1% v / v TFA in MeCN. Analysis was performed by injecting an appropriate volume of sample into the column and eluting it over 4 minutes at a constant flow rate of 0.45 mL / min with a linear gradient of 0–95% of eluent B for eluents A and B. Calibration was performed using a standard curve generated by injecting an external standard of Insulin Aspart, 3.55 mg / mL.

[0128] Method: Preparation of CNP compound formulations for G-in vivo experiments Formulations for in vivo research were prepared according to the principles outlined below.

[0129] Procedure: The formulations to be used in the in vivo study were prepared by dissolving the lyophilized peptide in a formulation buffer having the same composition as the final formulation (the final formulation composition is specified in Examples 9 to 15). The following final formulation compositions were used: • pH 4.0: 5 mM sodium acetate, 250 mM glycerol, pH 4.0 • pH 4.0: 5 mM sodium acetate, 240 mM propylene glycol, pH 4.0 pH 4.0: 5 mM sodium acetate, 250 mM glycerol, 0.007% polysorbate 20, pH 4.0 • pH 6.0: 20 mM sodium phosphate, 223 mM propylene glycol, pH 6.0 pH 6.5: 8 mM sodium phosphate, 250 mM glycerol, pH 6.5 pH 7.4: 8 mM sodium phosphate, 250 mM glycerol, pH 7.4 pH 7.4: 8 mM sodium phosphate, 250 mM glycerol, 0.007% polysorbate 20, pH 7.4 pH 7.5: 8 mM sodium phosphate, 250 mM glycerol, pH 7.5 pH 5.5: 1.33 mM citrate monohydrate, 3.67 mM citrate trisodium, 52 mg / ml trehalose, 15 mg / ml D-mannose, 0.73 mg / ml L-methionine, 0.05 mg / ml polysorbate 80, pH 5.5

[0130] For practical reasons, the formulation procedure differed slightly for the pH 6.5 formulation compared to the formulations of other pH targets.

[0131] Formulation at pH 6.5: Lyophilized peptide was dissolved in pH 7.4 buffer (8 mM sodium phosphate, 250 mM glycerol, pH 7.4) to a nominal concentration 20-50% above the target peptide concentration, and the pH was adjusted to pH 6.5 using 0.2 N NaOH or 0.2 N HCl. The actual peptide concentration was determined using CAD (Method F, CAD02), and a volume of pH 6.5 buffer (8 mM sodium phosphate, 250 mM glycerol, pH 6.5) corresponding to the nominal concentration of the target peptide concentration was added, and the pH was adjusted to pH 6.5 using 0.2 N NaOH or 0.2 N HCl. The actual peptide concentration was determined using CAD (Method F, CAD02). The formulation was sterile filtered (0.22 μm), the final peptide concentration was determined using CAD (Method F, CAD02), and the formulation was filled into sterile containers.

[0132] Formulation at pH 4.0, pH 6.0, pH 7.4, and pH 7.5: The lyophilized peptide was dissolved in a formulation buffer having a composition corresponding to the final formulation composition (specified in Examples 9 to 15) to a nominal concentration 20-50% above the target peptide concentration, and the pH was adjusted to the target using 0.2N NaOH or 0.2N HCl. The actual peptide concentration was determined using CAD (Method F, CAD02), and the same volume of formulation buffer was added to reach the nominal concentration corresponding to the target peptide concentration, and the pH was adjusted to the target using 0.2N NaOH or 0.2N HCl. The actual peptide concentration was determined using CAD (Method F, CAD02). The formulation was sterile filtered (0.22 μm), the final peptide concentration was determined using CAD (Method F, CAD02), and the formulation was filled into a sterile container.

[0133] [Example 1] CNP compound synthesis CNP compounds were synthesized and characterized according to the general preparation methods described above. Exemplary CNP compounds and their components are summarized in Table 1. The protractor and linker elements in Table 1 together form modifying groups of the CNP compounds. [Table 6-1] [Table 6-2] [Table 6-3] [Table 6-4] [Table 6-5] [Table 6-6]

[0134] Chem.1; Compound ID 0065; Sequence ID 1 hCNP22 [ka] Molecular weight: 2197.6008.LCMS34: m / 3 calculated value: 733.5336; m / 3 measured value: 733.7000; m / 4 calculated value: 550.4002; m / 4 measured value: 550.2800

[0135] Chem.2; Compound ID 1510; Sequence ID 2 hCNP37 [ka] Molecular weight: 3948.5589.UPLC107UPLC107: m / 3 Calculated value: 1317.1863

[0136] Chem.3; Compound ID 0312; Sequence ID 3 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-4,5Q,13Q,32Nle]-hCNP37 [ka] Molecular weight: 4182.9477.LCMS34LCMS34: m / 3 calculated value: 1395.3159; m / 3 measured value: 1395.1200; m / 4 calculated value: 1046.7369; m / 4 measured value: 1046.600

[0137] Chem.4; Compound ID 0776; Sequence ID 4 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,27E,32L]-hCNP37 [ka] Molecular weight: 4068.8451.LCMS34LCMS34: m / 3 calculated value: 1357.2817; m / 3 measured value: 1357.2694; m / 4 calculated value: 1018.2113; m / 4 measured value: 1018.2089

[0138] Chem.5; Compound ID 1235; Sequence ID 5 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,32L]-hCNP37 [ka] Molecular weight: 3674.2904.UPLC107: Calculated value per m / 3: 1225.7635; Measured value per m / 3: 1225.5700

[0139] Chem.6; Compound ID 0262; Sequence ID 6 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-8,32Nle]-hCNP37 [ka] Molecular weight: 4040.6131.LCMS34LCMS34: m / 3 calculated value: 1347.8710; m / 3 measured value: 1347.7200; m / 4 calculated value: 1011.1533; m / 4 measured value: 1010.7900

[0140] Chem.7; Compound ID 0296; Sequence ID 7 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-4,15S,19S,32Nle]-hCNP37 [ka] Molecular weight: 4427.9616.LCMS34: m / 3 calculated value: 1476.9872; m / 3 measured value: 1476.9200; m / 4 calculated value: 1107.9904; m / 4 measured value: 1108.1700

[0141] Chem.8; Compound ID 0313; Sequence ID 8 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-10,32Nle]-hCNP37 [ka] Molecular weight: 3620.1536.LCMS34: m / 3 calculated value: 1207.7179; m / 3 measured value: 1207.6400; m / 4 calculated value: 906.0384; m / 4 measured value: 905.9900

[0142] Chem.9; Compound ID 0334; Sequence ID 9 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-10,13Q,32Nle]-hCNP37 [ka] Molecular weight: 3634.1801.LCMS34: m / 3 calculated value: 1212.3934; m / 3 measured value: 1212.3100; m / 4 calculated value: 909.5450; m / 4 measured value: 909.2400

[0143] Chem.10; Compound ID 1221; Sequence ID 10 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,32L]-hCNP37 [ka] Molecular weight: 3700.3243.UPLC107: Calculated value per m / 3: 1234.4414; Measured value per m / 3: 1234.2700

[0144] Chem.11; Compound ID 1222; Sequence ID 11 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10E,13Q,32L]-hCNP37 [ka] Molecular weight: 3666.2651.LCMS34: m / 3 calculated value: 1223.0884; m / 3 measured value: 1222.9000;

[0145] Chem.12; Compound ID 1223; Sequence ID 12 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3701.266.LCMS34: m / 3 calculated value: 1234.7553; m / 3 measured value: 1234.5800;

[0146] Chem.13; Compound ID 1224; Sequence ID 13 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19S,32L]-hCNP37 [ka] Molecular weight: 3659.2294.LCMS34: m / 3 calculated value: 1220.7431; m / 3 measured value: 1220.3212; m / 4 calculated value: 915.8074; m / 4 measured value: 915.7400

[0147] Chem.14; Compound ID 1225; Sequence ID 14 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3669.2672.UPLC107: Calculated value per m / 3: 1224.0891; Measured value per m / 3: 1223.9400

[0148] Chem.15; Compound ID 1226; Sequence ID 15 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,14E,25P,32L]-hCNP37 [ka] Molecular weight: 3670.2089.UPLC107: Calculated value per m / 3: 1224.4030; Measured value per m / 3: 1224.2000

[0149] Chem.16; Compound ID 1227; Sequence ID 16 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3628.1723.LCMS34: m / 3 calculated value: 1210.3908; m / 3 measured value: 1210.4200; m / 4 calculated value: 908.0431; m / 4 measured value: 908.0900

[0150] Chem.17; Compound ID 1228; Sequence ID 17 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,14E,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3629.114.LCMS34: m / 3 calculated value: 1210.7047; m / 3 measured value: 1210.5790; m / 4 calculated value: 908.2785; m / 4 measured value: 908.1400

[0151] Chem.18; Compound ID 1229; Sequence ID 18 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3666.2651.UPLC107: Calculated value per m / 3: 1223.0884; Measured value per m / 3: 1222.9300

[0152] Chem.19; Compound ID 1230; Sequence ID 19 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,13Q,14E,19S,32L]-hCNP37 [ka] Molecular weight: 3625.1701.UPLC107: Calculated value per m / 3: 1209.3900; Measured value per m / 3: 1209.2800

[0153] Chem.20; Compound ID 1231; Sequence ID 20 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10Q,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3666.222.UPLC107: Calculated value per m / 3: 1223.0740; Measured value per m / 3: 1222.9700

[0154] Chem.21; Compound ID 1232; Sequence ID 21 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3700.3243.UPLC107: Calculated value per m / 3: 1234.4414; Measured value per m / 3: 1234.2100

[0155] Chem.22; Compound ID 1233; Sequence ID 22 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3627.2306.UPLC107: Calculated value per m / 3: 1210.0769; Measured value per m / 3: 1209.9600

[0156] Chem.23; Compound ID 1236; Sequence ID 23 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3675.2321.LCMS34: m / 3 calculated value: 1226.0774; m / 3 measured value: 1225.8900;

[0157] Chem.24; Compound ID 1237; Sequence ID 24 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14E,32L]-hCNP37 [ka] Molecular weight: 3709.2913.UPLC107: Calculated value per m / 3: 1237.4304; Measured value per m / 3: 1237.3400

[0158] Chem.25; Compound ID 1240; Sequence ID 25 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14E,25P,32L]-hCNP37 [ka] Molecular weight: 3678.2342.UPLC107: Calculated value per m / 3: 1227.0781; Measured value per m / 3: 1226.9500

[0159] Chem.26; Compound ID 1241; Sequence ID 26 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3636.1976.LCMS34: m / 3 calculated value: 1213.0659; m / 3 measured value: 1213.1011; m / 4 calculated value: 910.0494; m / 4 measured value: 910.0900

[0160] Chem.27; Compound ID 1242; Sequence ID 27 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14E,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3637.1393.UPLC107: Calculated value per m / 3: 1213.3798; Measured value per m / 3: 1213.2100

[0161] Chem.28; Compound ID 1274; Sequence ID 28 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13E,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3670.2089.LCMS34: m / 3 calculated value: 1224.4030; m / 3 measured value: 1224.3150; m / 4 calculated value: 918.5522; m / 4 measured value: 918.4840

[0162] Chem.29; Compound ID 1287; Sequence ID 29 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-9,10E,13Q,14E,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3465.9407.LCMS34: m / 3 calculated value: 1156.3136; m / 3 measured value: 1156.5100; m / 4 calculated value: 867.4852; m / 4 measured value: 867.3800

[0163] Chem.30; Compound ID 1288; Sequence ID 30 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-9,10E,13Q,15E,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3465.9407.LCMS34: m / 3 calculated value: 1156.3136; m / 3 measured value: 1155.8400; m / 4 calculated value: 867.4852; m / 4 measured value: 867.3800

[0164] Chem.31; Compound ID 1289; Sequence ID 31 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,15E,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3629.114.LCMS34: m / 3 calculated value: 1210.7047; m / 3 measured value: 1210.5500; m / 4 calculated value: 908.2785; m / 4 measured value: 908.1600

[0165] Chem.32; Compound ID 1290; Sequence ID 16 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3757.2862.LCMS34: m / 3 calculated value: 1253.4287; m / 3 measured value: 1253.5700; m / 4 calculated value: 940.3216; m / 4 measured value: 940.1700

[0166] Chem.33; Compound ID 1302; Sequence ID 32 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,15T,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3609.1292.LCMS34: m / 3 calculated value: 1204.0431; m / 3 measured value: 1203.7523; m / 4 calculated value: 903.2823; m / 4 measured value: 903.3355

[0167] Chem.34; Compound ID 1303; Sequence ID 33 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3595.1026.LCMS34: m / 3 calculated value: 1199.3675; m / 3 measured value: 1199.3210; m / 4 calculated value: 899.7757; m / 4 measured value: 899.7320

[0168] Chem.35; Compound ID 1304; Sequence ID 34 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14H,15T,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3618.0962.LCMS34: m / 3 calculated value: 1207.0321; m / 3 measured value: 1206.7366; m / 4 calculated value: 905.5241; m / 4 measured value: 905.5760

[0169] Chem.36; Compound ID 1305; Sequence ID 35 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,14H,15T,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3609.1292.LCMS34: m / 3 calculated value: 1204.0431; m / 3 measured value: 1204.0040; m / 4 calculated value: 903.2823; m / 4 measured value: 903.2470

[0170] Chem.37; Compound ID 1309; Sequence ID 36 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3643.2333.LCMS34: m / 3 calculated value: 1215.4111; m / 3 measured value: 1215.4546; m / 4 calculated value: 911.8083; m / 4 measured value: 911.8567

[0171] Chem.38; Compound ID 1310; Sequence ID 37 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3602.1383.LCMS34: m / 3 calculated value: 1201.7128; m / 3 measured value: 1201.7507; m / 4 calculated value: 901.5346; m / 4 measured value: 901.5770

[0172] Chem.39; Compound ID 1311; Sequence ID 38 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3588.1117.LCMS34: m / 3 calculated value: 1197.0372; m / 3 measured value: 1197.0725; m / 4 calculated value: 898.0279; m / 4 measured value: 898.0776

[0173] Chem.40; Compound ID 1312; Sequence ID 39 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3561.0433.LCMS34: m / 3 calculated value: 1188.0144; m / 3 measured value: 1187.9800; m / 4 calculated value: 891.2608; m / 4 measured value: 891.2320

[0174] Chem.41; Compound ID 1322; Sequence ID 40 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14H,15H,27E,32L]-hCNP37 [ka] Molecular weight: 3740.3102.LCMS34: m / 3 calculated value: 1247.7701; m / 3 measured value: 1247.6970; m / 4 calculated value: 936.0776; m / 4 measured value: 936.0280

[0175] Chem.42; Compound ID 1323; Sequence ID 40 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14H,15H,27E,32L]-hCNP37 [ka] Molecular weight: 3869.4242.LCMS34: m / 3 calculated value: 1290.8081; m / 3 measured value: 1290.7130; m / 4 calculated value: 968.3561; m / 4 measured value: 968.2840

[0176] Chem.43; Compound ID 1324; Sequence ID 41 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,14H,15H,19Q,27E,32L]-hCNP37 [ka] Molecular weight: 3860.4141.LCMS34: m / 3 calculated value: 1287.8047; m / 3 measured value: 1287.7150; m / 4 calculated value: 966.1035; m / 4 measured value: 966.0310

[0177] Chem.44; Compound ID 1338; Sequence ID 42 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19S,25P,27E,32L]-hCNP37 [ka] Molecular weight: 3642.1988.LCMS34: m / 3 calculated value: 1215.0663; m / 3 measured value: 1214.9827; m / 4 calculated value: 911.5497; m / 4 measured value: 911.2273

[0178] Chem.45; Compound ID 1339; Sequence ID 43 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-9,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3464.999.LCMS34: m / 3 calculated value: 1155.9997; m / 3 measured value: 1155.9600; m / 4 calculated value: 867.2498; m / 4 measured value: 866.9667

[0179] Chem.46; Compound ID 1340; Sequence ID 44 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,14H,15H,19Q,32L]-hCNP37 [ka] Molecular weight: 3717.2736.LCMS34: m / 3 calculated value: 1240.0912; m / 3 measured value: 1240.0320; m / 4 calculated value: 930.3184; m / 4 measured value: 930.2630

[0180] Chem.47; Compound ID 1341; Sequence ID 45 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,14H,15H,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3686.2165.LCMS34: m / 3 calculated value: 1229.7388; m / 3 measured value: 1229.6760; m / 4 calculated value: 922.5541; m / 4 measured value: 922.5020

[0181] Chem.48; Compound ID 1345; Sequence ID 46 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14H,15H,32L]-hCNP37 [ka] Molecular weight: 3726.2836.LCMS34: m / 3 calculated value: 1243.0945; m / 3 measured value: 1243.0310; m / 4 calculated value: 932.5709; m / 4 measured value: 932.5240

[0182] Chem.49; Compound ID 1346; Sequence ID 47 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,14H,15H,25P,32L]-hCNP37 [ka] Molecular weight: 3695.2265.LCMS34: m / 3 calculated value: 1232.7422; m / 3 measured value: 1232.6930; m / 4 calculated value: 924.8066; m / 4 measured value: 924.7340

[0183] Chem.50; Compound ID 1347; Sequence ID 48 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-10,13Q,19H,25P,32L]-hCNP37 [ka] Molecular weight: 3385.947.LCMS34: m / 3 calculated value: 1129.6490; m / 3 measured value: 1129.2775; m / 4 calculated value: 847.4868; m / 4 measured value: 847.2063

[0184] Chem.51; Compound ID 1348; Sequence ID 49 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-10,13Q,18H,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3385.947.LCMS34: m / 3 calculated value: 1129.6490; m / 3 measured value: 1129.2775; m / 4 calculated value: 847.4868; m / 4 measured value: 847.4684

[0185] Chem.52; Compound ID 1350; Sequence ID 50 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,14H,15T,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3973.532.LCMS34: m / 3 calculated value: 1325.5107; m / 3 measured value: 1325.3463; m / 4 calculated value: 994.3830; m / 4 measured value: 994.2843

[0186] Chem.53; Compound ID 1351; Sequence ID 51 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,14H,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3959.5054.LCMS34: m / 3 calculated value: 1320.8351; m / 3 measured value: 1320.6271; m / 4 calculated value: 990.8764; m / 4 measured value: 990.7383

[0187] Chem.54; Compound ID 1352; Sequence ID 33 [N-terminus([2-[2-[[2-[[2-[2-[[2-[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3740.259.LCMS34: m / 3 calculated value: 1247.7530; m / 3 measured value: 1247.3048; m / 4 calculated value: 936.0648; m / 4 measured value: 935.9911

[0188] Chem.55; Compound ID 1353; Sequence ID 36 [N-terminus([2-[2-[[2-[[2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3788.3897.LCMS34: m / 3 calculated value: 1263.7966; m / 3 measured value: 1263.6780; m / 4 calculated value: 948.0974; m / 4 measured value: 947.7735

[0189] Chem.56; Compound ID 1354; Sequence ID 52 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4079.6524.LCMS34: m / 3 calculated value: 1360.8841; m / 3 measured value: 1360.7530; m / 4 calculated value: 1020.9131; m / 4 measured value: 1020.8160

[0190] Chem.57; Compound ID 1355; Sequence ID 22 [N-terminus([2-[2-[[2-[[2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3772.387.LCMS34: m / 3 calculated value: 1258.4623; m / 3 measured value: 1258.3439; m / 4 calculated value: 944.0968; m / 4 measured value: 944.0223

[0191] Chem.58; Compound ID 1356; Sequence ID 22 [N-terminus([2-[2-[[2-[[2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3901.501.LCMS34: m / 3 calculated value: 1301.5003; m / 3 measured value: 1301.3470; m / 4 calculated value: 976.3753; m / 4 measured value: 976.2725

[0192] Chem.59; Compound ID 1357; Sequence ID 5 [N-terminus([2-[2-[[2-[[2-[2-[[2-[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,9E,10H,13Q,32L]-hCNP37 [ka] Molecular weight: 3948.5608.LCMS34: m / 3 calculated value: 1317.1869; m / 3 measured value: 1317.0300; m / 4 calculated value: 988.1402; m / 4 measured value: 988.0348

[0193] Chem.60; Compound ID 1359; Sequence ID 53 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,9E,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3916.4792.LCMS34: m / 3 calculated value: 1306.4931; m / 3 measured value: 1306.0170; m / 4 calculated value: 980.1198; m / 4 measured value: 979.7797

[0194] Chem.61; Compound ID 1360; Sequence ID 54 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,9E,10H,13Q,14H,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3925.4462.LCMS34: m / 3 calculated value: 1309.4821; m / 3 measured value: 1309.3323; m / 4 calculated value: 982.3616; m / 4 measured value: 982.2667

[0195] Chem.62; Compound ID 1375; Sequence ID 14 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3959.5801.LCMS34: m / 3 calculated value: 1320.8600; m / 3 measured value: 1320.7200; m / 4 calculated value: 990.8950; m / 4 measured value: 990.5400

[0196] Chem.63; Compound ID 1376; Sequence ID 14 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(19-carboxynonadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3697.3204.LCMS34: m / 3 calculated value: 1233.4401; m / 3 measured value: 1233.3500; m / 4 calculated value: 925.3301; m / 4 measured value: 925.2600

[0197] Chem.64; Compound ID 1377; Sequence ID 55 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,17G,25P,32L]-hCNP37 [ka] Molecular weight: 3613.1609.LCMS34: m / 3 calculated value: 1205.3870; m / 3 measured value: 1205.3100; m / 4 calculated value: 904.2902; m / 4 measured value: 903.9800

[0198] Chem.65; Compound ID 1378; Sequence ID 56 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,6G,7G,8G,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3799.3262.LCMS34: m / 3 calculated value: 1267.4421; m / 3 measured value: 1267.3470; m / 4 calculated value: 950.8316; m / 4 measured value: 950.7240

[0199] Chem.66; Compound ID 1379; Sequence ID 57 [N-terminus([2-[2-[[[2-[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,8S,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4562.14.LCMS34: m / 3 calculated value: 1521.7133; m / 3 measured value: 1521.3220; m / 4 calculated value: 1141.5350; m / 4 measured value: 1141.2370

[0200] Chem.67; Compound ID 1380; Sequence ID 58 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5E,8S,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4563.1248.LCMS34: m / 3 calculated value: 1522.0416; m / 3 measured value: 1521.8490; m / 4 calculated value: 1141.7812; m / 4 measured value: 1141.6240

[0201] Chem.68; Compound ID 1381; Sequence ID 59 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,7H,10H,13Q,14H,15H,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4625.1195.LCMS34: m / 3 calculated value: 1542.7065; m / 3 measured value: 1542.3070; m / 4 calculated value: 1157.2799; m / 4 measured value: 1156.9700

[0202] Chem.69; Compound ID 1382; Sequence ID 60 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,7H,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4882.5268.LCMS34: m / 3 calculated value: 1628.5089; m / 3 measured value: 1628.2280; m / 4 calculated value: 1221.6317; m / 4 measured value: 1221.2810

[0203] Chem.70; Compound ID 1383; Sequence ID 61 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,7H,10H,13Q,15H,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4745.2665.LCMS34: m / 3 calculated value: 1582.7555; m / 3 measured value: 1582.5080; m / 4 calculated value: 1187.3166; m / 4 measured value: 1186.9970

[0204] Chem.71; Compound ID 9384; Sequence ID 62 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-6,7G,8Q,9A,10P,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3704.2302.LCMS34: m / 3 calculated value: 1235.7434; m / 3 measured value: 1235.5400; m / 4 calculated value: 927.0576; m / 4 measured value: 926.9010

[0205] Chem.72; Compound ID 1385; Sequence ID 63 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-5,10E,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 4121.7322.LCMS34: m / 3 calculated value: 1374.9107; m / 3 measured value: 1374.6120; m / 4 calculated value: 1031.4331; m / 4 measured value: 1031.2130

[0206] Chem.73; Compound ID 1386; Sequence ID 64 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-5,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4079.6955.LCMS34: m / 3 calculated value: 1360.8985; m / 3 measured value: 1360.7680; m / 4 calculated value: 1020.9239; m / 4 measured value: 1020.8160

[0207] Chem.74; Compound ID 1387; Sequence ID 65 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-5,6G,7G,8G,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3895.4136.LCMS34: m / 3 calculated value: 1299.4712; m / 3 measured value: 1299.3300; m / 4 calculated value: 974.8534; m / 4 measured value: 974.7500

[0208] Chem.75; Compound ID 1388; Sequence ID 66 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),des1-5,6G,7G,8G,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 3936.5086.LCMS34: m / 3 calculated value: 1313.1695; m / 3 measured value: 1313.0250; m / 4 calculated value: 985.1272; m / 4 measured value: 985.2870

[0209] Chem.76; Compound ID 1389; Sequence ID 67 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),5Q,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4699.3224.LCMS34: m / 3 calculated value: 1567.4408; m / 3 measured value: 1567.4835; m / 4 calculated value: 1175.8306; m / 4 measured value: 1176.1075

[0210] Chem.77; Compound ID 1390; Sequence ID 68 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),5Q,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4700.2641.LCMS34: m / 3 calculated value: 1567.7547; m / 3 measured value: 1567.5480; m / 4 calculated value: 1176.0660; m / 4 measured value: 1175.8970

[0211] Chem.78; Compound ID 1391; Sequence ID 55 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,17G,25P,32L]-hCNP37 [ka] Molecular weight: 3903.4738.LCMS34: m / 3 calculated value: 1302.1579; m / 3 measured value: 1302.0100; m / 4 calculated value: 976.8685; m / 4 measured value: 976.5200

[0212] Chem.79; Compound ID 1392; Sequence ID 16 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 3918.4851.LCMS34: m / 3 calculated value: 1307.1617; m / 3 measured value: 1307.3500; m / 4 calculated value: 980.6213; m / 4 measured value: 980.5300

[0213] Chem.80; Compound ID 1393; Sequence ID 69 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3959.5371.LCMS34: m / 3 calculated value: 1320.8457; m / 3 measured value: 1321.0300; m / 4 calculated value: 990.8843; m / 4 measured value: 990.7800

[0214] Chem.81; Compound ID 1394; Sequence ID 70 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,10E,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4644.2869.LCMS34: m / 3 calculated value: 1549.0956; m / 3 measured value: 1548.7900; m / 4 calculated value: 1162.0717; m / 4 measured value: 1161.8500

[0215] Chem.82; Compound ID 1395; Sequence ID 71 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,7A,10E,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4559.1791.LCMS34: m / 3 calculated value: 1520.7264; m / 3 measured value: 1520.4600; m / 4 calculated value: 1140.7948; m / 4 measured value: 1140.600

[0216] Chem.83; Compound ID 1396; Sequence ID 70 [N-terminus([2-[2-[[[2-[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,10E,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4773.4009.LCMS34: m / 3 calculated value: 1592.1336; m / 3 measured value: 1592.1600; m / 4 calculated value: 1194.3502; m / 4 measured value: 1194.3700

[0217] Chem.84; Compound ID 1398; Sequence ID 72 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,8H,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5013.5738.LCMS34: m / 3 calculated value: 1672.1913; m / 3 measured value: 1672.1528; m / 4 calculated value: 1254.3935; m / 4 measured value: 1254.1224

[0218] Chem.85; Compound ID 1399; Sequence ID 72 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(19-carboxynonadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,8H,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5041.627.LCMS34: m / 3 calculated value: 1681.5423; m / 3 measured value: 1681.5050; m / 4 calculated value: 1261.4068; m / 4 measured value: 1261.1331

[0219] Chem.86; Compound ID 1400; Sequence ID 73 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,7H,13Q,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4955.5344.LCMS34: m / 3 calculated value: 1652.8448; m / 3 measured value: 1652.8279; m / 4 calculated value: 1239.8836; m / 4 measured value: 1239.8716

[0220] Chem.87; Compound ID 1401; Sequence ID 74 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,32L]-hCNP37 [ka] Molecular weight: 5578.2432.LCMS34: m / 3 calculated value: 1860.4144; m / 3 measured value: 1860.2654; m / 4 calculated value: 1395.5608; m / 4 measured value: 1395.4623

[0221] Chem.88; Compound ID 1402; Sequence ID 75 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5288.9151.LCMS34: m / 3 calculated value: 1763.9717; m / 3 measured value: 1763.8729; m / 4 calculated value: 1323.2288; m / 4 measured value: 1323.1718

[0222] Chem.89; Compound ID 1403; Sequence ID 76 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5232.8088.LCMS34: m / 3 calculated value: 1745.2696; m / 3 measured value: 1744.8611; m / 4 calculated value: 1309.2022; m / 4 measured value: 1309.1547

[0223] Chem.90; Compound ID 1404; Sequence ID 75 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5579.228.LCMS34: m / 3 calculated value: 1860.7427; m / 3 measured value: 1860.6008; m / 4 calculated value: 1395.8070; m / 4 measured value: 1395.7069

[0224] Chem.91; Compound ID 1405; Sequence ID 75 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(19-carboxynonadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5316.9683.LCMS34: m / 3 calculated value: 1773.3228; m / 3 measured value: 1773.2192; m / 4 calculated value: 1330.2421; m / 4 measured value: 1330.1785

[0225] Chem.92; Compound ID 1406; Sequence ID 75 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5450.114.LCMS34: m / 3 calculated value: 1817.7047; m / 3 measured value: 1817.5808; m / 4 calculated value: 1363.5285; m / 4 measured value: 1363.4484

[0226] Chem.93; Compound ID 9407; Sequence ID 77 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4830.4871.LCMS34: m / 3 calculated value: 1611.1624; m / 3 measured value: 1610.8330; m / 4 calculated value: 1208.6218; m / 4 measured value: 1208.3823

[0227] Chem.94; Compound ID 1419; Sequence ID 51 [N-terminus([2-[2-[[[2-[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,14H,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4088.6194.LCMS34: m / 3 calculated value: 1363.8731; m / 3 measured value: 1363.6902; m / 4 calculated value: 1023.1549; m / 4 measured value: 1023.0371

[0228] Chem.95; Compound ID 1420; Sequence ID 78 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,14H,15T,17V,25P,32L]-hCNP37 [ka] Molecular weight: 4258.8284.LCMS34: m / 3 calculated value: 1420.6095; m / 3 measured value: 1420.3875; m / 4 calculated value: 1065.7071; m / 4 measured value: 1065.5448

[0229] Chem.96; Compound ID 1421; Sequence ID 79 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4199.7994.LCMS34: m / 3 calculated value: 1400.9331; m / 3 measured value: 1400.7268; m / 4 calculated value: 1050.9499; m / 4 measured value: 1050.8062

[0230] Chem.97; Compound ID 1422; Sequence ID 80 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4235.8348.LCMS34: m / 3 calculated value: 1412.9449; m / 3 measured value: 1412.7313; m / 4 calculated value: 1059.9587; m / 4 measured value: 1059.8075

[0231] Chem.98; Compound ID 1423; Sequence ID 81 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,15T,17V,25P,32L]-hCNP37 [ka] Molecular weight: 4249.8614.LCMS34: m / 3 calculated value: 1417.6205; m / 3 measured value: 1417.3965; m / 4 calculated value: 1063.4654; m / 4 measured value: 1063.3148

[0232] Chem.99; Compound ID 1424; Sequence ID 82 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,15T,17V,25P,32L]-hCNP37 [ka] Molecular weight: 4370.0084.LCMS34: m / 3 calculated value: 1457.6695; m / 3 measured value: 1457.4309; m / 4 calculated value: 1093.5021; m / 4 measured value: 1093.5833

[0233] Chem.100; Compound ID 1425; Sequence ID 83 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,15T,17V,25P,32L]-hCNP37 [ka] Molecular weight: 4055.6244.LCMS34: m / 3 calculated value: 1352.8748; m / 3 measured value: 1352.6941; m / 4 calculated value: 1014.9061; m / 4 measured value: 1014.5325

[0234] Chem.101; Compound ID 1426; Sequence ID 84 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[17-(1H-tetrazole-5-yl)heptadecanoylamino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,15S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4144.7323.LCMS34: m / 3 calculated value: 1382.5774; m / 3 measured value: 1382.3732; m / 4 calculated value: 1037.1831; m / 4 measured value: 1037.0341

[0235] Chem.102; Compound ID 1431; Sequence ID 85 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10E,13Q,17G,19H,25P,32L]-hCNP37 [ka] Molecular weight: 3912.4408.LCMS34: m / 3 calculated value: 1305.1469; m / 3 measured value: 1304.9080; m / 4 calculated value: 979.1102; m / 4 measured value: 978.9250

[0236] Chem.103; Compound ID 1432; Sequence ID 86 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10E,13Q,14H,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4396.9476.LCMS34: m / 3 calculated value: 1466.6492; m / 3 measured value: 1466.3500; m / 4 calculated value: 1100.2369; m / 4 measured value: 1100.0354

[0237] Chem.104; Compound ID 1434; Sequence ID 87 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4040.57.LCMS34: m / 3 calculated value: 1347.8567; m / 3 measured value: 1347.9714; m / 4 calculated value: 1011.1425; m / 4 measured value: 1011.2499

[0238] Chem.105; Compound ID 9435; Sequence ID 88 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4031.603.LCMS34: m / 3 calculated value: 1344.8677; m / 3 measured value: 1344.7600; m / 4 calculated value: 1008.9008; m / 4 measured value: 1008.8210

[0239] Chem.106; Compound ID 1436; Sequence ID 89 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,17V,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4073.6828.LCMS34: m / 3 calculated value: 1358.8943; m / 3 measured value: 1358.6617; m / 4 calculated value: 1019.4207; m / 4 measured value: 1019.2703

[0240] Chem.107; Compound ID 1437; Sequence ID 90 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,17V,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4082.6498.LCMS34: m / 3 calculated value: 1361.8833; m / 3 measured value: 1361.7580; m / 4 calculated value: 1021.6625; m / 4 measured value: 1021.5750

[0241] Chem.108; Compound ID 1448; Sequence ID 91 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),5Q,10E,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 4741.359.LCMS34: m / 3 calculated value: 1581.4530; m / 3 measured value: 1581.2060; m / 4 calculated value: 1186.3398; m / 4 measured value: 1186.0240

[0242] Chem.109; Compound ID 1449; Sequence ID 51 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,14H,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4217.7334.LCMS34: m / 3 calculated value: 1406.9111; m / 3 measured value: 9407.0500; m / 4 calculated value: 1055.4334; m / 4 measured value: 1055.300

[0243] Chem.110; Compound ID 1450; Sequence ID 52 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,15T,17V,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4208.7664.LCMS34: m / 3 calculated value: 1403.9221; m / 3 measured value: 1403.7330; m / 4 calculated value: 1053.1916; m / 4 measured value: 1053.0130

[0244] Chem.111; Compound ID 1451; Sequence ID 92 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,10H,13Q,17V,19S,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4249.8614.LCMS34: m / 3 calculated value: 1417.6205; m / 3 measured value: 1417.4290; m / 4 calculated value: 1063.4654; m / 4 measured value: 1063.3310

[0245] Chem.112; Compound ID 1452; Sequence ID 77 [N-terminus([2-[2-[[[2-[2-[[2-[[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4685.3307.LCMS34: m / 3 calculated value: 1562.7769; m / 3 measured value: 1562.3981; m / 4 calculated value: 1172.3327; m / 4 measured value: 1172.3179

[0246] Chem.113; Compound ID 1453; Sequence ID 77 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4540.1743.LCMS34: m / 3 calculated value: 1514.3914; m / 3 measured value: 1514.0494; m / 4 calculated value: 1136.0436; m / 4 measured value: 1135.8064

[0247] Chem.114; Compound ID 1454; Sequence ID 77 [N-terminus([2-[2-[[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4701.3732.LCMS34: m / 3 calculated value: 1568.1244; m / 3 measured value: 1567.7415; m / 4 calculated value: 1176.3433; m / 4 measured value: 1176.0765

[0248] Chem.115; Compound ID 1455; Sequence ID 77 [N-terminus([2-[2-[[[2-[2-[[2-[[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4556.2167.LCMS34: m / 3 calculated value: 1519.7389; m / 3 measured value: 1519.3893; m / 4 calculated value: 1140.0542; m / 4 measured value: 1139.8102

[0249] Chem.116; Compound ID 1456; Sequence ID 77 [N-terminus([2-[2-[[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4411.0603.LCMS34: m / 3 calculated value: 1471.3534; m / 3 measured value: 1471.0409; m / 4 calculated value: 1103.7651; m / 4 measured value: 1103.5541

[0250] Chem.117; Compound ID 1457; Sequence ID 93 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),-2G,-1Q,2P,3G,4Q,5A,6P,7G,8Q,9A,10P,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4410.9775.LCMS34: m / 3 calculated value: 1471.3258; m / 3 measured value: 1471.0251; m / 4 calculated value: 1103.7444; m / 4 measured value: 1103.7853

[0251] Chem.118; Compound ID 1458; Sequence ID 94 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-2,3G,4Q,5A,6P,7G,8Q,9A,10P,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4057.6038.LCMS34: m / 3 calculated value: 1353.5346; m / 3 measured value: 1353.3113; m / 4 calculated value: 1015.4010; m / 4 measured value: 1015.2496

[0252] Chem.119; Compound ID 1459; Sequence ID 95 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1,2G,3Q,4A,5P,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4738.3536.LCMS34: m / 3 calculated value: 1580.4512; m / 3 measured value: 1580.0658; m / 4 calculated value: 1185.5884; m / 4 measured value: 1185.5701

[0253] Chem.120; Compound ID 1460; Sequence ID 96 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 4669.3313.LCMS34: m / 3 calculated value: 1557.4438; m / 3 measured value: 1557.4031; m / 4 calculated value: 1168.3328; m / 4 measured value: 1168.0756

[0254] Chem.121; Compound ID 1461; Sequence ID 97 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,10H,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4096.6763.LCMS34: m / 3 calculated value: 1366.5588; m / 3 measured value: 1366.3214; m / 4 calculated value: 1025.1691; m / 4 measured value: 1025.0148

[0255] Chem.122; Compound ID 1462; Sequence ID 96 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 4830.5302.LCMS34: m / 3 calculated value: 1611.1767; m / 3 measured value: 1611.1900; m / 4 calculated value: 1208.6326; m / 4 measured value: 1208.6500

[0256] Chem.123; Compound ID 1463; Sequence ID 98 [N-terminus([2-[2-[[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,32L]-hCNP37 [ka] Molecular weight: 4861.5442.LCMS34: m / 3 calculated value: 1621.5147; m / 3 measured value: 1621.1760; m / 4 calculated value: 1216.3861; m / 4 measured value: 1216.1420

[0257] Chem.124; Compound ID 1464; Sequence ID 99 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4816.4606.LCMS34: m / 3 calculated value: 1606.4869; m / 3 measured value: 1606.1700; m / 4 calculated value: 1205.1152; m / 4 measured value: 1204.8820

[0258] Chem.125; Compound ID 1465; Sequence ID 100 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P]-hCNP37 [ka] Molecular weight: 4848.5256.LCMS34: m / 3 calculated value: 1617.1752; m / 3 measured value: 1616.8130; m / 4 calculated value: 1213.1314; m / 4 measured value: 1212.8620

[0259] Chem.126; Compound ID 1470; Sequence ID 96 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 4540.2173.LCMS34: m / 3 calculated value: 1514.4058; m / 3 measured value: 1514.1500; m / 4 calculated value: 1136.0543; m / 4 measured value: 1135.8500

[0260] Chem.127; Compound ID 1471; Sequence ID 75 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5159.8011.LCMS34: m / 3 calculated value: 1720.9337; m / 3 measured value: 1720.8900; m / 4 calculated value: 1290.9503; m / 4 measured value: 1290.9200

[0261] Chem.128; Compound ID 1472; Sequence ID 75 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 5030.6872.LCMS34: m / 3 calculated value: 1677.8957; m / 3 measured value: 1677.8700; m / 4 calculated value: 1258.6718; m / 4 measured value: 1258.6600

[0262] Chem.129; Compound ID 1473; Sequence ID 67 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4989.6352.LCMS34: m / 3 calculated value: 1664.2117; m / 3 measured value: 1664.2000; m / 4 calculated value: 1248.4088; m / 4 measured value: 1248.400

[0263] Chem.130; Compound ID 1474; Sequence ID 101 [N-terminus([2-[2-[[2-[2-[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),5Q,13Q,25P,32L]-hCNP37 [ka] Molecular weight: 5159.8442.LCMS34: m / 3 calculated value: 1720.9481; m / 3 measured value: 1720.8900; m / 4 calculated value: 1290.9611; m / 4 measured value: 1290.9200

[0264] Chem.131; Compound ID 1475; Sequence ID 77 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4830.4871.LCMS34: m / 3 calculated value: 1611.1624; m / 3 measured value: 1610.7510; m / 4 calculated value: 1208.6218; m / 4 measured value: 1208.5814

[0265] Chem.132; Compound ID 1476; Sequence ID 77 [N-terminus([2-[2-[[2-[[2-[2-[[(2S)-4-carboxy-2-[[2-[[(2S)-4-carboxy-2-[[2-[[(2S)-4-carboxy-2-[[2-[[(2S)-4-carboxy-2-[[2-[[(2S)-4-carboxy-2-(17-carboxyheptadecanoylamino)butanoyl]aminoacetyl]amino]butanoyl]amino]acetyl]amino]butanoyl]amino]acetyl]amino]butanoyl]amino]acetyl]amino]butanoyl]amino]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4768.3795.LCMS34: m / 3 calculated value: 1590.4598; m / 3 measured value: 1590.4137; m / 4 calculated value: 1193.0949; m / 4 measured value: 1193.0793

[0266] Chem.133; Compound ID 1477; Sequence ID 102 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,13Q,19Q,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4844.5137.LCMS34: m / 3 calculated value: 1615.8379; m / 3 measured value: 1615.4194; m / 4 calculated value: 1212.1284; m / 4 measured value: 1212.0844

[0267] Chem.134; Compound ID 1478; Sequence ID 103 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4087.7093.LCMS34: m / 3 calculated value: 1363.5698; m / 3 measured value: 1363.3427; m / 4 calculated value: 1022.9273; m / 4 measured value: 1022.7753

[0268] Chem.135; Compound ID 9480; Sequence ID 103 [N-terminus([2-[2-[[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4216.8233.LCMS34: m / 3 calculated value: 1406.6078; m / 3 measured value: 1406.6766; m / 4 calculated value: 1055.2058; m / 4 measured value: 1055.0295

[0269] Chem.136; Compound ID 1481; Sequence ID 103 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4216.8233.LCMS34: m / 3 calculated value: 1406.6078; m / 3 measured value: 1406.3542; m / 4 calculated value: 1055.2058; m / 4 measured value: 1055.0295

[0270] Chem.137; Compound ID 9482; Sequence ID 67 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),5Q,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4828.4363.LCMS34: m / 3 calculated value: 1610.4788; m / 3 measured value: 1610.4060; m / 4 calculated value: 1208.1091; m / 4 measured value: 1207.8275

[0271] Chem.138; Compound ID 9483; Sequence ID 104 [N-terminus([(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]butanoyl]amino]butanoyl]),5Q,13Q,19S,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4713.3489.LCMS34: m / 3 calculated value: 1572.1163; m / 3 measured value: 1572.1500; m / 4 calculated value: 1179.3372; m / 4 measured value: 1179.1200

[0272] Chem.139; Compound ID 1484; Sequence ID 105 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-6,7G,8Q,9A,10P,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3730.3106.LCMS34: m / 3 calculated value: 1244.4369; m / 3 measured value: 1244.7042; m / 4 calculated value: 933.5777; m / 4 measured value: 933.5259

[0273] Chem.140; Compound ID 1486; Sequence ID 106 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-6,7G,8Q,9A,10P,13Q,17S,19Q,25P,27E,32L]-hCNP37 [ka] Molecular weight: 3718.2568.LCMS34: m / 3 calculated value: 1240.4189; m / 3 measured value: 1240.3220; m / 4 calculated value: 930.5642; m / 4 measured value: 930.4920

[0274] Chem.141; Compound ID 1487; Sequence ID 107 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),-2G,-1Q,2P,3G,4Q,5A,6P,7G,8Q,9A,10P,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4437.0578.LCMS34: m / 3 calculated value: 9480.0193; m / 3 measured value: 1479.8291; m / 4 calculated value: 1110.2645; m / 4 measured value: 1110.3766

[0275] Chem.142; Compound ID 1488; Sequence ID 107 [N-terminus([(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]),-2G,-1Q,2P,3G,4Q,5A,6P,7G,8Q,9A,10P,13Q,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4146.7449.LCMS34: m / 3 calculated value: 1383.2483; m / 3 measured value: 1383.4402; m / 4 calculated value: 1037.6862; m / 4 measured value: 1037.5803

[0276] Chem.143; Compound ID 1489; Sequence ID 108 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,17G,19Q,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4045.6296.LCMS34: m / 3 calculated value: 1349.5432; m / 3 measured value: 1349.3710; m / 4 calculated value: 1012.4074; m / 4 measured value: 1012.2860

[0277] Chem.144; Compound ID 1493; Sequence ID 109 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,19Q,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4101.7359.LCMS34: m / 3 calculated value: 1368.2453; m / 3 measured value: 1368.0680; m / 4 calculated value: 1026.4340; m / 4 measured value: 1026.3130

[0278] Chem.145; Compound ID 1511; Sequence ID 62 [N-terminus([2-[2-[2-[[2-[2-[2-[[(2S)-4-carboxy-2-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-6,7G,8Q,9A,10P,13Q,17S,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 3704.2302.LCMS34: m / 3 calculated value: 1235.7434; m / 3 measured value: 1235.6700; m / 4 calculated value: 927.0576; m / 4 measured value: 927.0010

[0279] Chem.146; Compound ID 1512; Sequence ID 88 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(2S)-4-carboxy-2-[[(2S)-4-carboxy-2-(17-carboxyheptadecanoylamino)butanoyl]amino]butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-8,13Q,17G,19Q,25P,32L]-hCNP37 [ka] Molecular weight: 4031.603.LCMS34: m / 3 calculated value: 1344.8677; m / 3 measured value: 1344.7600; m / 4 calculated value: 1008.9008; m / 4 measured value: 1008.8080

[0280] Chem.147; Compound ID 1513; Sequence ID 110 [N-terminus (17-carboxyheptadecanoyl),-5E,-4E,-3E,-2E,-1E,5Q,13Q,19S,25P,32L]-hCNP37 [ka] Molecular weight: 4828.4363.LCMS34: m / 3 calculated value: 1610.4788; m / 3 measured value: 1610.2260; m / 4 calculated value: 1208.1091; m / 4 measured value: 1207.9160

[0281] Chem.148; Compound ID 1514; Sequence ID 111 [N-terminus (17-carboxyheptadecanoyl),-4E,-3E,-2E,-1E,5Q,13Q,19S,25P,27E,32L]-hCNP37 [ka] Molecular weight: 4713.3489.LCMS34: m / 3 calculated value: 1572.1163; m / 3 measured value: 1571.8940; m / 4 calculated value: 1179.3372; m / 4 measured value: 1179.1550

[0282] Chem.149; Compound ID 1265; Sequence ID 233 [N-terminus ([2-[2-[[2-[[2-[2-[[(4R)-4-carboxy-4-(17-carboxyheptadecanoylamino) Butanoyl[amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),des1-5,6a,7r,8k,9y,10k,12a,13Q,14k,15k,17l,18S,19k,21c,22f,24l,25k,26l,27e,28r,29i,31s,32l,33s,35l,37c]-hCNP37 [ka] Molecular weight: 4068.8451.LCMS34: m / 4 calculated value: 1018.2113; m / 4 measured value: 1018.1958

[0283] Chem.150; Compound ID 0106; Sequence ID 234 [-2P,-1G]-hCNP37 [ka] Molecular weight: 4102.7254.LCMS34: m / 3 calculated value: 1368.5751; m / 3 measured value: 1368.362; m / 4 calculated value: 1026.6814; m / 4 measured value: 1026.529

[0284] Chem.151; Compound ID 0089; Sequence ID 1 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl])]-hCNP22 [ka] Molecular weight: 2913.4725.LCMS34: m / 3 calculated value: 972.1575; m / 3 measured value: 971.844; m / 4 calculated value: 729.3681; m / 4 measured value: 729.131

[0285] Chem.152; Compound ID 0230; Sequence ID 235 [N-terminus([2-[2-[[2-[[2-[2-[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]),-5G,-4Q,-3A,-2P,-1G,2A,3P,4G,5Q,7P,8G,9Q,10A,11P,12G,13Q,14A,15P]-hCNP37 [ka] Molecular weight: 4680.3406.LCMS34: m / 3 calculated value: 1561.11; m / 3 measured value: 1560.773; m / 4 calculated value: 1171.0852; m / 4 measured value: 1170.834

[0286] Chem.153; Compound ID 0231; Sequence ID 1 [N-terminus([2-[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[(4S)-4-carboxy-4-(17-carboxyheptadecanoylamino)butanoyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl]amino]ethoxy]ethoxy]acetyl])-hCNP22 [ka] Molecular weight: 3784.4112.LCMS34: m / 3 calculated value: 1262.4704; m / 3 measured value: 1262.303; m / 4 calculated value: 947.1028; m / 4 measured value: 946.995

[0287] [Example 2] In vitro hNPR2 activity assay To evaluate the in vitro activity of CNP22(0065;Chem.1) and CNP compounds of the present invention, they were tested for their ability to induce the production of cyclic guanosine monophosphate (cGMP) in reporter cells expressing human natriuretic peptide receptor 2 (hNPR2).

[0288] Cell Culture: To produce a reporter cell line, HEK293 (ATCC® CRL-1573®) cells were stably transfected with the pGloSensor-42F reporter plasmid (GloSensor cGMP reporter-luciferase, Promega) and an expression plasmid encoding the hNPR2 receptor. Single-cell clones were isolated from the reporter cell line. When reporter cells are exposed to an hNPR2-activating compound, hNPR2 triggers the production of cGMP. GloSensor luciferase produced from the pGloSensor-42F plasmid binds to cGMP, and this binding induces a conformational shift of GloSensor luciferase, activating an otherwise inactive enzyme. The active GloSensor luciferase can convert luciferin to oxyluciferin, a process that produces bioluminescence. Luciferase activity can be quantified by detecting luminescence during cell lysis and luciferase substrate delivery. Therefore, adding an hNPR2 activating compound, followed by the addition of a detection reagent to reporter cells, induces luminescence generation in a dose-dependent manner. The maximum half-volume effective concentration (EC50) can be determined by testing several different concentrations of the hNPR2 activating compound. This measurement, also known as in vitro potency, is reported for all compounds tested. In all in vitro activity experiments, CNP22 was used as a positive in-plate control for hNPR2 activation. To reduce inter-assay variability, the potency of each compound relative to the potency of the CNP22 control on each plate is also reported.

[0289] Procedure: 35 HEK293-hNPR2 / GloSensor clone cells were continuously cultured in selective medium (DMEM containing 10% FCS, 1% P / S, 200 μg / mL G418, and 100 μg / mL hygromycin). To perform the assay, cells were detached using TrypLE Express (Gibco #12604013), passed through a 70 μm cell strainer, counted, and seeded overnight in 30 μL / well assay medium (DMEM without phenol red and containing 1% P / S) at a density of 10,000 cells / well in white 384-well plates (Perkin Elmer #6007680). The following day, the test compounds were serially diluted in dilution medium (DMEM without phenol red, containing 1% OVA (Sigma #A5505 Lot 04M7001V), 0.01% Tween20 (Roche #33766700), and 1% P / S), and then 10 μL / well of the diluted test compound was added to the cells. For each compound, 10 different concentrations ranging from 0.25 pM to 10 μM were tested. After incubation at 37°C and 5% CO2 for 30 minutes, 40 μL / well of Bright-Glo detection reagent (Promega #E2650) was added. Immediately afterward or 15 minutes later, luminescence was detected using an EnVision Multimode Plate Reader (PerkinElmer). To determine the in vitro potency or EC50 of each test compound, 4-parameter logistic regression was performed on the raw data of each test compound using the Python package SciPy optimize. The average EC50 values ​​are listed in Table 2 as "hNPR2 0% HSA.[EC50(nM)]". Relative potency is calculated using the following formula: EC50(CNP22 in-plate control) / EC50(test compound)*100 and is shown in Table 2 as "hNPR2 0% HSA.[EC50 % relative CNP]". At least two copies were measured for each test compound. The reported values ​​are the average of the copies and are listed in Table 2. [Table 7-1] [Table 7-2] [Table 7-3] [Table 7-4]

[0290] These data demonstrate that the tested CNP compounds exhibit a wide range of NPR2 in vitro activation levels, and that negatively charged CNP compounds commonly show reduced in vitro NPR2 potency compared to positively charged CNP compounds.

[0291] [Example 3] In vitro neprilysin stability of CNP compounds Natural CNPs have been reported to be metabolized by neprilysin (neutral endopeptidase, NEP), a protease widely present in the body. Therefore, an in vitro neprilysin stability assay was established to test the neprilysin-mediated degradation of selected CNP compounds.

[0292] Procedure: In short, the assay buffer consisted of PBS buffer (pH 7.4, ThermoFisher Scientific) with tween 20 (Sigma-Aldrich) at a final concentration of 0.005% v / v. Recombinant human neprilysin (rhNEP; R&D Systems) and CNP compounds were dissolved in the assay buffer. Natural CNP22 (0065, Chem.1) was included as a positive control on each day of incubation. First, the CNP compound (final concentration was 1000 nM) was pre-incubated in the assay buffer at 37°C for 10 minutes. The assay was initiated by adding either rhNEP (final concentration was 2 μg of protein per 1 mL) or the assay buffer (adsorption experiment), and incubation was performed at 37°C. The reaction was terminated at selected time points (0.5 min, 5 min, 15 min, 30 min, 60 min, 90 min, and 120 min) by adding 1 volume of the incubation mixture to 3 volumes of ethanol (containing 1% v / v formic acid). The mixture was centrifuged at 13,000 rpm for 20 minutes at 4°C. After centrifugation, 1 volume of the supernatant was mixed with 1 volume of Milli-Q water.

[0293] Analysis by LC-MS: The mixture was analyzed by LC-MS using an Acquity UPLC Peptide CSH C18 analytical column from Waters (130 Å, 1.7 μm, 1 × 50 mm) operated at 60°C. Gradient elution was performed using a Nexera UHPLC system (Shimadzu) with mobile phase A (consisting of Milli-Q water with 0.1% formic acid) and mobile phase B (consisting of acetonitrile with 0.1% formic acid). The flow rate was 0.3 ml / min. A zenoTOF 7600 mass spectrometer (Sciex) was used as the detector and operated in positive electrospray ionization mode. Data were recorded in full scan mode (m / z 300~1700).

[0294] Calibration curves were prepared in assay buffer and treated as test samples. These samples were used to calculate the concentrations of relevant CNP compounds in in vitro samples, including quality control samples. For at least 75% of the standards and QC samples, the deviation between nominal and calculated concentrations was less than 20%.

[0295] All incubations were performed in double cycles. Data were reported as the percentage remaining at 90 minutes (based on calculated concentration) compared to the average concentration of the sample at 0.5 minutes. Examples of seven compounds and CNP22 (0065, Chem.1) are shown in Table 3. [Table 8]

[0296] These data demonstrate that the CNP compound of the present invention is substantially stable in the presence of recombinant human neprilysin compared to CNP22, which is rapidly degraded.

[0297] [Example 4] Peptide Net Charge Calculation The charge of ionizable groups in a peptide at a given pH can be calculated using the Henderson-Hasselbalch equation, using empirically determined acid dissociation constants (pKa) for each amino acid residue or modifying group known in the art, for example, by the methods described by B. Skoog and A. Wichman (Trends in Analytical Chemistry, 1986, vol. 5, pp. 82-83) and L. Kozlowski (Biology Direct, 2016, 11:55). This is shown for negatively charged amino acids or modifying groups in Equation 1 and for positively charged amino acids and modifying groups in Equation 2, where pKn is the pKa of the negatively charged group, pKp is the pKa of the positively charged group, and pH in the equation is the pH of interest. The net charge of the peptide at a given pH is the sum of the negative and positive charges of all ionizable groups in the peptide.

number

[0298] For natural amino acid residues, tabular pKa constants are used for the free ends and ionizable sites in the side chains. The pKa values ​​of modified amino acids and modifying groups were estimated using the ACD / Labs ver.12 package and are also shown in Table 4.

[0299] As an example, to calculate the net charge of compound 9482 at pH 7.4 using the pKa values ​​shown in Table 4, the sum of the negative charges supported by the positions of Cys37 (C-terminal acid), Asp27, Glu2, gGlu(1), gGlu(2), gGlu(3), gGlu(4), gGlu(5), and the C18d fatty acid is -8.995 Fd. The sum of the positive charges supported by the positions of Arg28, Lys15, Lys14, Lys10, Lys8, Arg7, and His3 is 6.162 Fd. Therefore, the net charge is the sum of the above, i.e., -2.833.

[0300] Table 5 shows the calculated charge values ​​for the CNP compound of the present invention at selected pH levels (pH 7.4 and pH 6.5) using the above principle. [Table 9] [Table 10-1] [Table 10-2] [Table 10-3] [Table 10-4]

[0301] [Example 5] Solubility of CNP compounds To evaluate the suitability of CNP compounds for soluble liquid formulations, the solubility of CNP compounds in pharmaceutically relevant buffers was measured. Since pH is expected to affect solubility, solubility was investigated at two different pH levels (pH 4.0 and pH 6.5).

[0302] Preparation of pH 4.0 samples: pH 4.0 samples of the CNP compound were prepared by mixing the lyophilized peptide with 5 mM sodium acetate and 250 mM glycerol buffer at pH 4.0 to a nominal concentration of at least 4000–5000 nmol / mL at room temperature. The pH of the sample was measured and adjusted to pH 4.0. If the sample was not completely dissolved, it was equilibrated at room temperature until the next day.

[0303] Preparation of pH 6.5 Sample: A pH 6.5 sample of the CNP compound was prepared by dissolving the lyophilized peptide in water. While stirring, a portion of the peptide solution was transferred to a buffer consisting of 41 mM sodium phosphate and 118 mg / mL glycerol, pH 7.5–8.0, followed by the addition of diluted sodium hydroxide. Another portion of the peptide solution was then added, and further diluted sodium hydroxide was added. This procedure was repeated until all of the peptide solution was added, reaching a nominal concentration of at least 4000–5000 nmol / mL, with a buffer composition of 8 mM sodium phosphate and 23 mg / mL glycerol. The pH of the sample was measured and adjusted to pH 6.5. If the sample was not completely dissolved, it was allowed to equilibrate at room temperature until the next day.

[0304] Procedure: Samples were prepared at pH 4.0 as described above, and the compound concentration in the supernatant was determined using CAD (Method F, CAD02). The compound concentration at pH 4.0 represents the solubility at pH 4.0, and the measured values ​​are shown in Table 6. [Table 11]

[0305] Samples were prepared at pH 6.5 as described above, and the peptide concentration in the supernatant was determined by CAD (Method F, CAD02). The peptide concentration at pH 6.5 represents the solubility at pH 6.5, and the measured values ​​are shown in Table 7. [Table 12]

[0306] Roughly speaking, only samples containing more than 4000 μM were subjected to further testing as described below.

[0307] These results demonstrate that the CNP compound of the present invention can be formulated in a pharmaceutically relevant buffer at a concentration suitable for subcutaneous administration.

[0308] Example 6 - Thioflavin T (ThT) assay The physical stability of CNP compounds and their suitability for liquid formulations were evaluated using the ThT assay. Low physical stability of peptides can lead to amyloid fibril formation, which is observed in the sample as an ordered, thread-like polymer structure that can eventually lead to gel formation. This has traditionally been examined by visual inspection of the sample. However, such measurements are highly subjective and vary from observer to observer. Therefore, the application of small molecule indicator probes is far more advantageous. Thioflavin T (ThT) is such a probe and exhibits a distinct fluorescent signature when bound to fibrils [Naiki et al. (1989) Anal. Biochem. 177, 244-249, LeVine (1999) Methods. Enzymol. 09, 274-284].

[0309] Procedure: Thioflavin T was added to the supernatant samples from Example 5 (pH 4.0 and 6.5) using aqueous ThT stock solution to a final concentration of approximately 1 μM in the sample. 150 μL sample aliquots were placed in a 96-well microtiter plate (Packard OptiPlate®-96, white polystyrene). Typically, 2-3 copies of each sample were applied to the plate. The plate was sealed and placed in a fluorescence plate reader (Fluoroskan Ascent FL) and incubated at 37°C with circular shaking (960 rpm, amplitude 1 mm). Fluorescence measurements were performed every 20 minutes using excitation through a 444 nm filter and emission through a 485 nm filter. Between each measurement, the plate was shaken and heated as described above, and the assay was terminated after 45 hours. Fluorescence measurements from each microtiter plate well were plotted against time, and the lag time (time until an increase in ThT fluorescence was observed) was estimated.

[0310] To assess the loss of dissolved peptides after 45 hours of incubation, peptide recovery was measured as the ratio of the total peak area before and after incubation using the RP-UPLC method described below.

[0311] Measurement of lag time and peptide recovery % using RP-UPLC: RP-UPLC was performed using an Acquity UPLC BEH C18 1.7 μm (2.1 × 30 mm) Waters column (eluent A: 0.1% v / v% TFA in water, eluent B: 4:1 acetonitrile / 0.1% v / v TFA in water) with gradient elution (0 min: 95% A, 2 min: 20% A, 2.3 min: 20% A, 2.4 min: 95% A, 9 min: 95% A) at a flow rate of 0.9 mL / min and a column temperature of 30°C. Total peptide area was evaluated using UV detection at 215 nm.

[0312] Samples at pH 4.0 were prepared as described in Example 5 and subjected to the ThT assay described above. The lag time and peptide recovery values ​​are shown in Table 8. [Table 13]

[0313] Samples at pH 6.5 were prepared as described in Example 5 and subjected to the ThT assay described above. The lag time and peptide recovery values ​​are shown in Table 9. [Table 14]

[0314] In conclusion, this experiment demonstrates that most of the CNP compounds of the present invention can be formulated in pharmaceutically relevant buffers with a low tendency to form fibrils.

[0315] Example 7 - Measurement of High Molecular Weight Proteins (HMWPs) To evaluate the tendency of CNP compounds to form covalent dimers or oligomers when formulated, the high molecular weight protein content of CNP compound formulations was determined at pH 4.0 and 6.5.

[0316] Procedure: The relative amount of covalently bound HMWP was evaluated using SEC-UPLC. SEC-UPLC was performed using a Waters UPLC Protein BEH SEC column, 125 Å, 4.6 × 150 mm, 1.7 μm, with homogeneous concentration elution (0.3 M NaCl, 10 mM NaH2PO4, and 5 mM H3PO4, 50% (v / v) isopropanol, pH 2.4), at a flow rate of 0.3 mL / min, column temperature of 50 °C, and UV detection at 215 nm. The total area of ​​peaks eluting before the monomeric major peak is referred to as HMWP and is expressed on a percentage scale relative to the total peptide peak area.

[0317] Results: HMWP formation after 2 weeks of dormant incubation at 37°C: Samples were prepared at pH 4.0 as described in Example 5, and the amount of HMWP was determined as described above. The samples were incubated at 37°C for 2 weeks, and the amount of HMWP was determined again. The increase in HMWP over 2 weeks of incubation at 37°C is reported in Table 10 (if the incubation period deviated from 2 weeks, the increase in HMWP was normalized to 2 weeks assuming a constant formation rate). [Table 15]

[0318] Samples at pH 6.5 were prepared as described in Example 5, and the amount of HMWP was determined as described above. The samples were incubated at 37°C for 2 weeks, and the amount of HMWP was determined again. The increase in HMWP over 2 weeks of incubation at 37°C is reported in Table 11 (if the incubation period deviated from 2 weeks, the increase in HMWP was normalized to 2 weeks assuming a constant formation rate). [Table 16]

[0319] In conclusion, this experiment demonstrates that the CNP compound of the present invention can be formulated in a pharmaceutically relevant buffer with a low tendency to form HMWPs.

[0320] Example 8 - Accelerated chemical stability of CNP compounds To determine the chemical stability of CNP compounds in the formulation, samples of CNP compounds in buffer were exposed to chemical degradation accelerated by heating. The stability of a given peptide was evaluated by measuring the recovery of intact peptides by quantitative analysis using RP-UPLC. Samples were also analyzed by LC-MS to evaluate the chemical degradation pattern.

[0321] Sample preparation: Samples for a two-week study were prepared according to the principle described in Example 5.

[0322] Samples for the 6-week study were prepared by dissolving lyophilized peptides in a buffer solution (8 mM phosphate, 250 mM glycerol, pH 7.4) to a nominal peptide concentration of 5000 nmol / ml, followed by pH adjustment to 6.5 with 0.1 N HCl.

[0323] Incubation: For the 2-week study, time zero (TZ) samples were removed and stored at -18°C, while the remaining volume was incubated at 37°C. After 2 weeks of incubation, the samples were removed and stored at -18°C. The frozen samples were thawed, diluted to approximately 200 nmol / mL, and the degree of chemilysis was determined using LC-MS as described.

[0324] For a 6-week study, time-zero (TZ) samples were taken and stored at -18°C, while the remaining volume was incubated at 37°C. Samples were taken weekly and stored at -18°C until the final sample collection point at 6 weeks. Frozen samples were thawed, diluted to approximately 200 nmol / mL, and the degree of chemilysis was determined using RP-UPLC and LC-MS as described.

[0325] Analysis of peptide recovery by RP-UPLC: The RP-UPLC method was performed using a Waters ACQUITY H-CLASS UPLC system equipped with a PDA detector. Separation was performed using an ACQUITY UPLC BEH C8 column (130 Å, 1.7 μm, 2.1 mm × 150 mm) operating at a column temperature of 50 °C with eluent A (19.59 mM NH4H2PO4, 41.71 mM (NH4)2HPO4, pH 6.5, and MeCN, 9:1 v:v) and eluent B (80% MeCN in water). UV detection was performed at 214 nm. Analysis was performed by injecting 3 μL of sample into the column and eluting it over 20 minutes using a one-step linear gradient of eluent A and B with 27–30% B, followed by washing with 95% B over 2 minutes, and then a re-equilibrium step of 7 minutes at a constant flow rate of 0.3 mL / min. This allowed for an analysis time of 30 minutes per sample. The peak area corresponding to intact CNP compounds is shown as a percentage of the total peptide peak area.

[0326] Analysis of chemical decomposition patterns by LC-MS: Samples were analyzed using a Thermo Scientific VANQUISH UPLC with a Waters Acquity C18 reversed-phase column, 300 Å, 1.7 μM particles, and a 1 mm × 150 mm array. Compounds were separated over 48 minutes by a linear gradient of 5%–55% solvent B using solvent A: water, 0.1% formic acid containing 0.02% TFA, and solvent B: acetonitrile, 0.1% formic acid containing 0.02% TFA. Mass spectra were acquired using a Q Exactive Plus hybrid Quadrupole-Orbitrap mass spectrometer, BioPharma option (Thermo Scientific), interfaced with an H-ESI II ion source. Data acquisition was performed with a scan range of m / z 500–2000 and a resolution of 30000.

[0327] The data list was consolidated into a total table of compounds. The table header was defined as follows:

[0328] "Recovery": Recovery of the compound peak (i.e., the major peak intensity at time zero with the compound monoisotopic mass) normalized to the compound peak at a given time point. "Isomer": As the total intensity of peaks with isobaric mass relative to the major compound, isolated in the RT dimension. "18amu loss": i.e., the total intensity of peaks with a mass 18amu lower than the CNP compound peak, isolated in the RT dimension. Fragments: As isolated in the RT dimension, the total intensity of peaks with a mass 50amu lower than the compound peak and peaks with a mass 18amu higher than the compound mass, assumed to occur over time and result from hydrolyzed peptide backbone cleavage. Finally, "Other": Includes the total intensity of peaks with a 32amu addition or dimer of the major peak, occurring over time.

[0329] Results: The results of LCMS analysis of a 2-week accelerated degradation study of several CNP compounds of the present invention are summarized in Table 12. The results of RP-UPLC and LCMS analysis of a 6-week accelerated degradation study of CNP compound 9482 (Chem. 137) are summarized in Tables 13 and 14, respectively. [Table 17] [Table 18] [Table 19]

[0330] These data demonstrate that the CNP compounds according to the present invention exhibit substantial chemical stability in formulations.

[0331] Example 9 - Local resistance to CNP compounds in rats The purpose of this study was to evaluate the local tissue response to subcutaneous injection of CNP compounds in rats.

[0332] Procedure: The CNP compound was formulated according to the general preparation method - Method G. Each Sprague Dawley rat (male, 10-11 weeks old upon arrival, Janvier, France) was subcutaneously administered 250 or 350 μL of the same compound (formulation concentration of 1000 nmol / mL) 120 hours prior to tissue sampling, with a total of two injections per animal (Table 15). The injection site (flank or neck) was alternated between animals within each group and at each time point. [Table 20]

[0333] The animals were given free access to tap water and diet (Altromin 1324). The group size was n=3 for the vehicle and positive control, and n=4 for the test compound. The vehicle and compound were formulated with either 5 mM sodium acetate, 250 mM glycerol, pH 4.0, or 8 mM sodium phosphate, 250 mM glycerol, pH 7.4, as shown in Table 16.

[0334] The animals were anesthetized with isoflurane. A square area of ​​fur (2cm x 2cm) was removed from either the neck or flank region of the rat via an electric shaver. For administration, a new injection needle (25G cannula, 1ml syringe) was placed in the middle of the square boundary closest to the animal's head, and the needle was inserted into the skin from the boundary toward the center of the square. As a result, the needle tip was located in the center of the square, the position was marked with an oil-based pen, and the preparation was injected into the center of the square. On the day of tissue sampling, the rats were deeply anesthetized with isoflurane and euthanized by bleeding. The injection sites were examined and scored for lesions. The injection sites were evaluated, any changes (location, color, shape, and size) were noted, and samples were taken for histological preparation.

[0335] The injection site was removed and fixed in a sealed plastic container in 4% phosphate-buffered neutral formaldehyde.

[0336] Next, each injection site was cut into three sections at 0.8 cm, 1.0 cm, and 1.2 cm from the square boundary where the injection needle was placed. These sections represented the injection site and adjacent tissue, respectively. The tissue was processed, embedded in paraffin, cut into thin sections of 2–4 μm, stained with hematoxylin and eosin (H&E), and microscopically evaluated using a light microscope. Histopathological changes were graded on a 5-level scale (minor (Min), mild (Mil), moderate (Mod), prominent (Mar), and severe (Sev)). The study is summarized in Table 16. [Table 21]

[0337] Results: The results of the local resistance study are summarized in Table 17. [Table 22]

[0338] Conclusion: Net positive charge control 0776 showed moderate to significant focal subcutaneous necrosis 5 days after subcutaneous injection in rats. In comparison, compounds 1351, 9384, 9407, and 1420 showed no local subcutaneous necrosis or only mild local subcutaneous necrosis 5 days after subcutaneous injection in rats, and were all evaluated as acceptable for human subcutaneous administration.

[0339] Example 10 - Local CNP resistance test in LYD pigs The purpose of this study was to evaluate the local tissue response to subcutaneous injection of CNP compounds in pigs.

[0340] Procedure: In domestic pigs, injection sites were subjected to histopathological examination after subcutaneous administration. The injection site was placed mid-dorsally, between the shoulders and rump on both sides of the dorsal midline. One or two days prior to injection, a 2 x 2 cm area was shaved and marked with oil-based ink under light anesthesia.

[0341] Each injection consisted of 100 or 200 μl of a formulation at a concentration of approximately 4400 μM. The CNP compound formulations were prepared according to the general preparation method—principles of Method G—using one of the following: A) 8 mM sodium phosphate, 250 mM glycerol, pH 6.5; B) 5 mM sodium acetate, 250 mM glycerol, pH 4.0; or C) 5 mM sodium acetate, 240 mM propylene glycol, pH 4.0, with physiological saline included as a negative control.

[0342] On day 1, the pigs were lightly anesthetized. An insulin pen (NovoPen 4) and insulin needle (NovoTwist 32G / 5mm) were used, with the needle held perpendicular to the skin, to precisely deposit the CNP preparation into the subcutaneous adipose tissue. On day 5 (4 days after injection) or day 6 (5 days after injection), the pigs were euthanized, the injection site was removed, and the tissue was fixed in 4% phosphate-buffered neutral formaldehyde in a sealed plastic container. The tissue blocks were pre-fixed in 10% buffered formalin for 2–4 hours, cut into 2 mm thick plates with a multipurpose knife, examined for macroscopic changes, and finally fixed overnight in a cassette. Two tissue pieces with the greatest tissue reaction or located in the central part of the injection site were selected for evaluation by light microscopy. Three–four 2–4 μm sections were cut at three levels at a distance of 100 μm, stained with hematoxylin and eosin (H&E), and evaluated by light microscopy. Histopathological changes were graded on a 5-level scale ((min (Min), mild (Mil), moderate (Mod), prominent (Mar), and severe (Sev))). The study is summarized in Table 18. [Table 23]

[0343] Results: The results are summarized in Table 19. *Necrosis was used as a marker for local subcutaneous reactions; **NAD: No abnormalities detected. [Table 24]

[0344] In conclusion, the net positive charge control compound 0776 showed moderate to significant local subcutaneous necrosis 5 days after subcutaneous injection in pigs, while the net positive charge CNP peptide 0106, which was not bound to a fatty acid albumin binder, showed no observable injection site reaction.

[0345] In comparison, compounds 9480 and 9482 did not show local subcutaneous necrosis 5 days after subcutaneous injection in pigs. Compounds 9384 and 9407 showed no local subcutaneous necrosis or minimal local subcutaneous necrosis 5 days after subcutaneous injection in pigs, compounds 9435 and 1235 showed minimal to mild local subcutaneous necrosis 5 days after subcutaneous injection in pigs, and compound 9483 showed no local subcutaneous necrosis or moderate local subcutaneous necrosis 5 days after subcutaneous injection in pigs. Compounds 9384, 9407, 9435, 9480, and 9482 were all evaluated as acceptable for human subcutaneous administration.

[0346] Example 11 - PK / PD of CNP compound after intravenous and subcutaneous administration to rats To evaluate the intravenous and subcutaneous PK / PD profiles of the CNP compound of the present invention, formulations of the CNP compound were administered to rats, blood samples were collected, and exposure and cGMP response were measured.

[0347] cGMP biomarker assay: The in vivo biological activity of CNP compounds was evaluated by measuring cGMP in plasma samples using plasma protein precipitation followed by quantification by LC-MS / MS.

[0348] To evaluate the cyclic guanosine monophosphate (cGMP) biomarker response in biological samples, CNP compounds were administered to test animals, followed by a plasma protein precipitation procedure and then liquid chromatography using tandem mass spectrometry (LC-MS / MS). As a slowdown measure, the concentration of cGMP in the test plasma samples was measured using a stable isotope-labeled (SIL) surrogate analyte as a reference. 13 C 10 15The determination was made using a standard curve prepared with N5cGMP. Substitute analytes were spiked in a pooled true blank substrate to cover the analytical range of 5 nM to 2000 nM. 8-methoxymethyl-3-isobutyl 1-methylxanthine (MMPX) was used as an internal standard. Plasma protein precipitation was performed using an organic solvent to precipitate plasma proteins, leaving a supernatant containing cGMP molecules, which was then analyzed by LC-MS / MS instrumentation.

[0349] To prepare standard samples, frozen true blank plasma substrates were thawed and then centrifuged at 4°C and 4000 RPM for 5 minutes. A 2000 nM standard was prepared from the blank plasma using stock SIL-cGMP and then transferred to a 1 mL Eppendorf 96-well plate (Patent No. 8,636,965). This was placed in a liquid handler (TECAN Fluent 78) and standard and QC (5, 10, 20, 50, 100, 200, 500, 1000, 2000 nM) were prepared by serial dilution with blank plasma in a 96-well microplate (Chimney Well 651201). Blank plasma samples were prepared on the same plate. The study samples were thawed by ventilation at ambient temperature for 10 minutes, shaken on a benchtop shaker at ambient temperature for 5 minutes, and then centrifuged at 4°C and 4000 RPM for 5 minutes. Next, the samples were placed in a liquid handler and the plasma protein precipitation procedure was performed. A certain amount of standard sample, QC sample, zero sample, and study sample were precipitated using four volumes of acetonitrile (OPTIMA® LC / MS grade) containing 30 nM MMPX (Sigma #M2547). The samples were then shaken on a benchtop shaker at ambient temperature for 4 minutes, followed by centrifugation at 4°C and 4000 RPM for 30 minutes. The supernatant was then transferred to a new 96-well microplate. The plate was then evaporated using an Eppendorf concentrator plus® at 30°C for 60 minutes. The residue was reconstituted with a certain amount of 5% acetonitrile and 0.1% formic acid (RATHBURN LC / MS Grade) and shaken on a benchtop shaker at ambient temperature for 2 minutes, followed by centrifugation at 4°C for 10 minutes. Next, the prepared samples were analyzed using an LC-MS / MS instrument (Sciex 6500+ Triple quadrupole-bound Sciex ExionLC®). Gradient elution was applied to reverse-phase LC using 0.1% formic acid as mobile phase A and 95% acetonitrile and 0.1% formic acid as mobile phase B. A Waters Acquity UPLC® CSH® Fluoro-Phenyl 1.7 μm, 2.1 × 100 mm column was used as the stationary phase and heated to 60°C.SIL-cGMP, endogenous cGMP, and MMPX were detected by selective reaction monitoring using cation-mode MS / MS.

[0350] Acceptance Criteria for Analysis Execution: The SIL-cGMP standard curve was established with a weighting of 1 / x2 and evaluated in terms of linearity, precision, and accuracy. At least 75% of the calibration standards had to meet the following criteria when working backward: Calibration standards had to be within ±15% of the nominal concentration of each calibration standard, with the exception of LLOQ, which had to be within ±20% of the nominal concentration of the LLOQ standard. Values ​​outside these limits had to be discarded unless the established model was changed. If discarding one calibration standard caused another calibration standard to fall outside the boundary, that calibration standard was also discarded.

[0351] At least two-thirds of the QC samples had to be within 15% of their respective nominal values. One-third of the QC samples could be outside 15% of their respective nominal values, but they had to be no more than 50% of the QC samples at the same nominal concentration.

[0352] Quantitative plasma analysis of CNP compounds: Plasma concentrations of CNP compounds were assayed by plasma protein precipitation and analyzed by turbo-flow liquid chromatography-mass spectrometry (TF-LC-MS). Standards were prepared by spiking plasma from either a blank rat or a miniature pig with the relevant CNP compound in the range of 0.5–2000 nM. For TF-LC-MS by protein precipitation, standards, plasma blanks, or study samples were prepared by adding either 3 or 4 volumes of ethanol or methanol containing a 20 nM internal standard to 1 volume of the sample. For some compounds, formic acid was added to the precipitation reagent (final concentration was 1% v / v). After adding the precipitation reagent, the mixture was centrifuged at 6200 rpm, 4°C for 30 minutes. After centrifugation, 1 volume of supernatant was mixed with 2 volumes of water (containing 1% formic acid). The mixture was analyzed by TF-LC-MS using a Cyclone turboflow column (0.5 × 50 mm, ThermoFisher Scientific) and either an XBridge Protein BEH C4 3.5 μm 300 Å or an XBridge Protein BEH C18 3.5 μm 130 Å analytical column (both 50 × 2.1 mm, obtained from Waters). Gradient elution was performed using mobile phase A (consisting of milli-Q water containing 1% formic acid and 5% methanol / acetonitrile (50 / 50)) and mobile phase B (consisting of methanol / acetonitrile (50 / 50) containing 1% formic acid and 5% milli-Q water). The mass spectrometer was operated in cationization mode. A TSQ Altis mass spectrometer (ThermoFisher Scientific) was used as the detector in selected reaction monitoring mode (specific transitions were optimized for each CNP compound). A linear calibration curve (weighted 1 / x2) was used to calculate concentrations in plasma samples. Quality control samples were included. The deviation between nominal and calculated concentrations in the standard material and quality control samples was less than 15%.

[0353] Animals and Administration: CNP compounds, formulated in either A) 8 mM sodium phosphate, 250 mM glycerol, 0.007% polysorbate 20, pH 7.4 or B) 5 mM sodium acetate, 250 mM glycerol, 0.007% polysorbate 20, pH 4.0, were administered to groups of 4-6 rats per compound and route of administration. Subcutaneous administration was 300 nmol / kg at a concentration of 1000 nmol / L, resulting in a dosage volume of 0.3 mL / kg, while intravenous administration was 100 nmol / kg at a concentration of 100 nmol / L, resulting in a dosage volume of 1 mL / kg.

[0354] Sampling: 250 μL sublingual blood samples were collected from unanesthetized rats in EDTA-coated vials (Microvette 600K3E, see #15.1673.100 Sarstedt) at baseline (pre-administration - 1 hour) and at 5 minutes (intravenous administration group only), 2 hours, 6 hours, 24 hours, 48 ​​hours, and 72 hours (exposure analysis only) after administration. Blood samples were rapidly stored on ice and centrifuged within 10 minutes of collection (4°C, 8000 RPM, 5 minutes). 50 μL plasma samples were transferred to individual microtubes for either exposure or cGMP analysis, held on dry ice, and stored in a freezer (-20°C). Rats were free to consume food (altromin) and tap water.

[0355] Analysis: PK parameters were calculated for each animal by non-compartmental analysis (NCA) using the software Phoenix WinNonlin (ver. 8.1 or later, Certara). Clearance (Cl) was calculated as follows: dose / AUCinf (linear up, log down). Half-life (t1 / 2) was calculated as ln(2) / λ, where λ was estimated by linear regression of time vs. log concentration over a time range of typically 2–48 hours (e.g., 6–48 hours). Bioavailability was calculated as dose-normalized AUC: (AUC(sc) / dose(sc)) / (AUC(iv) / dose(iv)). The values ​​in the table are the mean from individual animals (n=4–6 individual animals). The subsequent plot of cGMP plasma concentration vs. time shows (Y-axis) mean cGMP concentration after IV dosing (n=4–6 individual animals) and (X-axis) nominal plasma sampling time.

[0356] Results: PK data including clearance (CL), half-life T1 / 2, and bioavailability (F%) are summarized in Table 20. Plasma cGMP concentrations after intravenous administration of CNP compounds are shown in Figures 1-27, and the doses are shown in Table 20. Plasma cGMP is reported as an average. [Table 25]

[0357] In conclusion, this experiment demonstrates that the CNP compounds of the present invention exhibit extended half-life and reduced clearance in an in vivo rat model. Furthermore, this experiment demonstrates that the CNP compounds are biologically active based on their ability to elicit a cGMP response in administered animals (Figures 1-27).

[0358] Example 12 - PK / PD of CNP compounds after intravenous and subcutaneous administration to miniature pigs and LYD pigs The purpose of this study was to investigate the pharmacokinetic and pharmacodynamic properties after intravenous (iv) and subcutaneous (sc) administration to Göttingen miniature pigs and domestic LYD pigs, and to estimate bioavailability after subcutaneous administration. Quantitative plasma analysis of CNP and cGMP levels was performed as described in Example 11.

[0359] Animals and Administration: Pharmacokinetic and pharmacodynamic studies in Göttingen miniature pigs or domestic LYD pigs were conducted at CRO Minerva Imaging or Novo Nordisk A / S, respectively. CNP compounds were administered to normal female Göttingen miniature pigs, 1) 6–12 months old with an average weight of 22 kg, or 2) 4–5 months old with an average weight of 80 kg. Permanent central venous catheters were implanted at least one week prior to administration for blood sampling and intravenous (iv) administration. For subcutaneous (sc) administration, the pigs were ultrasound-scanned and the optimal injection site, located laterally above the neck, was marked with a permanent tattoo. Using insulin pens (NovoPen Echo or NovoPen 4) and insulin needles (NovoFine 32G / 4mm or NovoTwist 32G / 5mm), the needle was held perpendicular to the skin to precisely deposit the CNP preparation into the subcutaneous adipose tissue.

[0360] The CNP compound was formulated according to the general preparation method—principles of Method G—in one of the following formulations: A) 8 mM sodium phosphate, 250 mM glycerol, pH 6.5; B) 5 mM sodium acetate, 250 mM glycerol, pH 4.0; C) 5 mM sodium acetate, 240 mM propylene glycol, pH 4.0; D) 8 mM sodium phosphate, 250 mM glycerol, pH 7.5; or E) 20 mM sodium phosphate, 223 mM propylene glycol, pH 6.0.

[0361] For intravenous administration, the CNP compound was administered using formulations A to E at a dose of 30 to 60 nmol / kg to miniature pigs and 15 to 43 nmol / kg to domestic pigs. For subcutaneous administration, the CNP compound was administered using formulations A to D at a dose of 55 to 62 nmol / kg to miniature pigs and 20 to 30 nmol / kg to domestic pigs.

[0362] Sampling: For cGMP and CNP compound analysis by LC-MS, approximately 1 mL of blood sample was collected in EDTA-coated tubes (8 mM) before administration and up to 14 days after administration. In a few cases, cGMP was analyzed only up to 48 hours after intravenous administration in domestic LYD pigs. Blood samples were held on wet ice for up to 30 minutes before centrifugation at 4°C and a minimum of 1500G for 10 minutes, and plasma samples were stored at -20°C until analysis.

[0363] Plasma concentration-time profiles were analyzed by non-compartmental PK analysis using Phoenix WinNonlin 8.1, Pharsight Inc., Mountain View, CA, USA. Area under the plasma concentration-time curve (AUC) was calculated using the "linear up log down" method, and uniform weighting was used to estimate the terminal rate constant (λz). Subcutaneous bioavailability (F) was calculated by dividing the dose-normalized AUC (AUC / dose) after subcutaneous administration by the AUC / dose after intravenous administration.

[0364] Results: PK data with clearance (CL), half-life T1 / 2, and bioavailability (F%) in miniature pigs are summarized in Table 21. PK data with clearance (CL), half-life T1 / 2, and bioavailability (F%) in LYD pigs are summarized in Table 22. Plasma cGMP concentrations after intravenous or subcutaneous administration of CNP compounds in miniature pigs and domestic pigs are shown in Figures 28-48, with doses shown in Tables 21 and 22. Plasma cGMP is reported as the average of n=2-4 over 14 days for miniature pigs and as the average of n=2-4 over 2 days for domestic LYD pigs. [Table 26] [Table 27]

[0365] Conclusion: This experiment demonstrates that the CNP compounds of the present invention exhibit extended half-life and reduced clearance in both miniature pigs and domestic LYD pigs. Furthermore, this experiment demonstrates that the CNP compounds are biologically active based on their ability to elicit a cGMP response in administered animals (Figures 28-48).

[0366] Furthermore, this experiment demonstrates subcutaneous bioavailability (F) of over 40% of the compounds of the present invention (Tables 21 and 22). On the other hand, the non-active, positively charged examples (compound IDs 0776 and 0312) show low subcutaneous bioavailability (7-13%). The non-active example with a nearly neutral charge (compound ID 1235) shows moderate subcutaneous bioavailability of 25%.

[0367] Example 13 - Telemetry measurement of heart rate and blood pressure The purpose of this study was to investigate the effects of acute administration of CNP compound 9482 (Chem.137) on mean arterial pressure (MAP) and heart rate (HR) in awake rats.

[0368] Procedure: The transducer was implanted in Sprague Dawley rats (Charles River), and the rats were allowed to recover for 2-3 weeks. Before surgery, the device was placed in the receiver plate and registered with the software by powering it on. Before the start of surgery, saline solution was added to the transducer catheter, and then the transmitter was powered on. The transmitter was placed in the abdomen, and the sensor was placed in the abdominal aorta. Isoflurane was used as an anesthetic, and during and after surgery, the rats were given Temgesic (0.05 mg / kg subcutaneously), Norodyl (5 mg / kg subcutaneously), and Baytril (10 mg / kg subcutaneously) for pain relief and to prevent bacterial infection.

[0369] Study Design: Rats will be housed in cages of two (one with equipment, one as a social partner). All animals will have free access to water and food (althromin) throughout the study. Rats will be housed in a light cycle of 6am to 6pm and 6pm to 6am. Eight rats will be divided into two groups according to a crossover study design.

[0370] Telemetry: Each rat was equipped with a transducer (DSI HD-S10) capable of measuring blood pressure, heart rate, and activity using Ponemah software with a frequency of 100 Hz. To minimize data variability during measurement, the room in which the rats were housed was kept undisturbed. A 4-hour baseline recording was performed before the start of the study to capture the mean arterial pressure (MAP) and heart rate (HR) patterns of untreated, undisturbed rats. The recording period after drug administration was 24 hours.

[0371] Drug administration scheme: Rats were administered in a crossover study design using 4 rats per group. Each rat was given a 1-week washout period before the next administration event. Drug administration was performed intravenously (IV) into the tail vein. 9482 doses were tested in separate experiments at intravenous doses of 30, 100, and 300 nmol / kg. The vehicle consisted of 8 mM phosphate, 250 mM glycerol, and pH 7.4.

[0372] Data analysis and statistics were performed using Ponemah software and Graphpad Prism. For statistical analysis, paired t-tests were applied with a 95% confidence level and a p-value of <0.05 for statistical significance.

[0373] Results: Within the first 30 minutes after administration, no differences in MAP and HR were observed with CNP compound 9482 at 30, 100, or 300 nmol / kg compared to the vehicle (Tables 23 and 25). The mean over the first 12 hours did not result in any difference in MAP between the vehicle and the three tested doses of 9482 (Table 24). Increases of 8.8±10.1% and 6.4±6.6% in HR (P<0.05) were detected at 100 and 300 nmol / kg during the same period (Table 26). No change in HR was observed at the 30 nmol / kg dose (Table 26). [Table 28] [Table 29] [Table 30] [Table 31]

[0374] Conclusion: Acute IV infusion of CNP compound 9482 was not associated with any major hemodynamic changes over 12 hours, as indicated by no changes in MAP and only a slight increase in HR at the two highest doses.

[0375] Example 14 - Mouse metatarsal incision and culture: The objective of this study was to evaluate bone growth after administration of CNP compounds in an ex vivo mouse model.

[0376] Procedure: 1-day-old mouse pups (NMRI, Javier Labs France) were euthanized before excision. All animal care and use guidelines applicable internationally and internally from Novo Nordisk were followed. Three metatarsal bones were excised from each hind limb. The excised bones were placed in 24-well cell culture plates and cultured in 400 μl of α-minimum essential medium supplemented with 0.2% BSA and 1% Pen / Strep at 37°C and 5% CO2. Bones from all animals were randomized into a control group and a group treated with six CNP compounds (100 nM, n=8). The medium was changed every 2-3 days. All compounds were formulated according to the general preparation method - Method G. Compounds 9435 and 9483 were formulated in A) 5 mM sodium acetate, 250 mM glycerol, pH 4.0, while compounds 9384, 9407, 9480, and 9482 were formulated in B) 8 mM sodium phosphate, 250 mM glycerol, pH 7.4.

[0377] Vehicle alone did not affect bone growth compared to culture medium alone.

[0378] Digital photographs were taken on days 0, 4, 7, 11, and 14 using a digital camera attached to a Nikon microscope. The length of each metatarsal bone was measured using ImageJ software, and the increase in bone length was expressed as the percentage change (mean (SD)) from the length measured on the day of resection.

[0379] Results: All CNP compounds significantly increased bone length compared to the control group 11 days after treatment (p<0.05, two-way ANOVA). The overall increase in bone length was 28–48% higher after treatment with CNP compounds compared to the control group 14 days after culture. The results of the study are summarized in Table 27. [Table 32]

[0380] Table 27 shows that metatarsals were isolated from mice and cultured for up to 14 days. Length was measured every 3-4 days throughout the experiment and expressed as the percentage change from day 0 (mean (SD)). Statistically significant differences were shown compared to the control group (two-way ANOVA).

[0381] Conclusion: This experiment demonstrates that the CNP compound of the present invention can significantly increase bone length in an ex vivo mouse model.

[0382] Example 15 - Mouse Growth Study The objective of this study was to evaluate growth in mice treated with two doses of CNP compound.

[0383] Procedure: Mice (C57BL / 6J, male, 4 weeks old, n=10) were administered subcutaneously with compound 9482 or vehicle daily for 35 days. The formulations were prepared according to the general preparation method - Method G (30 and 70 nmol / kg, 10 mL / kg; formulation buffer 8 mM sodium phosphate, 250 mM glycerol, 0.007% polysorbate 20, pH 7.4). Body weight, as well as tail length and body length, were measured weekly throughout the study. Growth increases were expressed as the percentage change from day 0 (mean (SD)).

[0384] Results: Both doses of CNP compound 9482 (30 nmol / kg and 70 nmol / kg) significantly increased tail length and body length at 7 and 18 days, respectively, compared to the control group (p<0.05, two-way ANOVA). At 35 days, compared to the control group, the change in tail length (from day 0) increased by 45% and 83% after treatment with 30 nmol / kg and 70 nmol / kg of CNP compound. Body length was 30% and 67% higher, respectively, after 35 days of treatment with 30 nmol / kg and 70 nmol / kg of CNP compound compared to the control group. The results of the study are summarized in Tables 28 and 29. [Table 33]

[0385] Table 28 shows the increase in tail length as the percentage change from day 0 (mean (SD)). A statistically significant difference is shown compared to the control group (two-way ANOVA). [Table 34]

[0386] Table 29 shows the increase in body length as the percentage change (mean (SD)) from day 0. A statistically significant difference is shown compared to the control group (two-way ANOVA).

[0387] Conclusion: This experiment demonstrates that the CNP compound of the present invention, when administered subcutaneously in a pharmaceutically relevant formulation, significantly increases the tail length and body length of mice. Inclusion by referencing the sequence list This application is submitted together with an electronic sequence listing. The entire contents of the sequence listing are incorporated herein by reference.

Claims

1. A CNP compound comprising a CNP peptide and a modifying group, wherein the net charge of the compound at physiological pH is 0 or negative, and the CNP peptide is of formula I: AA ０１ -AA ０２ -AA ０３ -AA ０４ -AA ０５ -AA ０６ -AA ０７ -AA ０８ -AA ０９ -AA １０ -AA １１ -AA １２ -AA １３ -AA １４ -AA １５ -AA １６ -AA １７ -AA １８ -AA １９ -AA ２０ -AA ２１ -AA ２２ -AA ２３ -AA ２４ -AA ２５ -AA ２６ -AA ２７ -AA ２８ -AA ２９ -AA ３０ -AA ３１ -AA ３２ -AA ３３ -AA ３４ -AA ３５ -AA ３６ -AA ３７ (In the formula, AA ０１ However, it is either Gln or absent, AA ０２ However, it is either Glu or absent, AA ０３ However, His is either absent, AA ０４ However, it is either Pro or absent. AA ０５ However, it is either Asn, Gln, or Glu, or absent. AA ０６ However, it is either Ala or absent, AA ０７ However, it is either Arg, His, or Ala, or absent. AA ０８ However, it is either Lys, Ser, or His, or absent. AA ０９ However, it is either Tyr or Glu, or absent. AA １０ However, it is either Lys, Glu, Gln, or His, or is absent. AA １１ However, it is Gly, AA １２ However, it is Ala, AA １３ However, it is Glun or Asn or Glu, AA １４ However, it is Lys or His or Glu, AA １５ However, it is Lys or Ser or Glu or Thr or His, AA １６ However, it is Gly, AA １７ However, it is Leu or Gly or Ser or Val, AA １８ However, it is either Ser or His. AA １９ However, it is Gln or Ser or Lys or His, AA ２０ However, it is Gly, AA ２１ However, it is Cys, AA ２２ However, it is Phe, AA ２３ However, it is Gly, AA ２４ However, it is Leu, AA ２５ However, it is Pro or Lys, AA ２６ However, it is Leu, AA ２７ However, it is either Asp or Glu. AA ２８ However, it is Arg, AA ２９ However, it is Ile, AA ３０ However, it is Gly, AA ３１ However, it is Ser, AA ３２ However, it is Leu or Nle or Met, AA ３３ However, it is Ser, AA ３４ However, it is Gly, AA ３５ However, it is Leu, AA ３６ However, it is Gly, AA ３７ However, it contains an amino acid sequence that follows Cys. The modifying group comprises Chem. A, Chem. B, and Chem. C. Chem. A 【Chemistry 1】 (In the formula, p is an integer in the range of 14 to 20, * is selected from the group consisting of (where * represents the amide bond connecting Chem. A and Chem. B), Chem. B, 【Chemistry 2】 (In the formula, q is an integer in the range of 1 to 8, * represents the amide bond connecting Chem. A- and Chem. B-. ** is selected from the group consisting of (where ** represents an amide bond connecting Chem. B- and Chem. C-), Chem. C 【Transformation 3】 (In the formula, r is an integer in the range of 0 to 4, s is an integer in the range of 0 to 3, t is an integer in the range of 0 to 1. u is an integer in the range of 0 to 3. ** represents the amide bond connecting Chem. B- and Chem. C-. A CNP compound selected from the group consisting of *** (where *** represents an amide bond connecting Chem. C- and the N-terminal alpha-amine on the CNP peptide).

2. The AA of the CNP peptide １３ However, it is Gln and AA １９ However, it is Gln or Ser, and AA ２５ However, it is Pro and AA ３２ The CNP compound according to claim 1, wherein the compound is Leu.

3. The AA of the CNP peptide ５ However, it is Gln and AA １３ However, it is Gln and AA １９ However, it is Gln or Ser, and AA ２５ However, it is Pro and AA ３２ The CNP compound according to claim 1, wherein the compound is Leu.

4. The CNP compound according to claim 1, wherein the CNP peptide comprises one of the following amino acid sequences. Table 1-1 Table 1-2 Table 1-3 Table 1-4 Table 1-5

5. The CNP compound according to claim 1, wherein the CNP peptide has one of the following amino acid sequences. Table 2-1 Table 2-2 Table 2-3 Table 2-4 Table 2-5 Table 2-6

6. The aforementioned CNP peptide has the following amino acid sequence: GAQKKGSSQGCFGLPLDRIGLSGLGC (Sequence No. 136), ARKYKGAQKKGLSQGCFGLPLDRIGLSGLGC (Sequence No. 77), YKGAQKKGGGSQGCFGLPLDRIGLSGLGC (Sequence No. 88), YKGAQKKGLSQGCFGLPLDRIGLSGLGC (Sequence No. 103), QEHPQARKYKGAQKKGLSSGCFGLPLDRIGLSGLGC (Sequence ID 67), and The CNP compound according to claim 1, comprising any one of QEHPQARKYKGAQKKGLSSGCFGLPLERIGSGLGC (Sequence ID 104).

7. The aforementioned CNP peptide has the following amino acid sequence: The CNP compound according to claim 1, comprising QEHPQARKYKGAQKKGLSSGCFGLPLDRIGLSGLGC (Sequence ID 67).

8. The CNP compound according to any one of claims 1 to 7, wherein the Chem. B of the modifying group has q values ​​from 1 to 5.

9. The modifying groups are: Chem. E, Chem. F, Chem. G, Chem. H, Chem. I, and Chem. J 【Chemistry 4-1】 【Chemistry 4-2】 A CNP compound according to any one of claims 1 to 8, selected from (wherein the formula, the dotted line defines the amide bond of the CNP peptide to the N-terminal alpha-amine).

10. Formula II: 【Transformation 5】 A CNP compound that conforms to [the specified formula].

11. Formulas II, III, IV, V, VI, and VII: 【Chemistry 6-1】 【Chemistry 6-2】 A CNP compound described in any one of the following.

12. A CNP compound comprising a CNP peptide and a modifying group, wherein the compound is one of the compounds disclosed in Table 1. Table 3-1 Table 3-2 Table 3-3 Table 3-4 Table 3-5 Table 3-6

13. A pharmaceutical composition comprising a compound according to any one of claims 1 to 12 and one or more pharmaceutically acceptable excipients.

14. A compound according to any one of claims 1 to 12 or a composition according to claim 13, for use in the treatment or prevention of cardiovascular metabolic diseases, including heart failure, and growth disorders, including chondrodysplasia.

15. A method for treating or preventing heart failure, comprising administering to a patient in need of such treatment an effective amount of a compound according to any one of claims 1 to 12, in combination with one or more additional therapeutically active compounds of any choice.

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