JP2006298938A - Use of glucagon-like peptide-1 (glp-1) or analog thereof to prevent catabolic change after surgery - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method of improving recovery after surgery by preventing the catabolic reaction and insulin resistance caused by surgical trauma. <P>SOLUTION: The method of attenuating post-surgical catabolic change and insulin resistance involves administering a compound selected from GLP-1, a GLP-1 analog, a GLP-1 derivative, and a pharmaceutically acceptable salt thereof to a patient requiring the attenuation. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to methods for improving post-surgical recovery by preventing catabolic reactions and insulin resistance resulting from surgical injury (trauma).

  In Western countries, approximately 20-25,000 surgical procedures per million are performed during the year. Surgery similar to injury causes significant changes in metabolism [Non-patent document 1; Non-patent document 2; Non-patent document 3; The promotion of synthesis of glucose, the primary fuel for tissue repair, is an important substance metabolism after surgery and occurs at the expense of body protein and stored energy [Non-Patent Document 5; Non-Patent Document 6].

  These modulations have traditionally been attributed to gluco-regulated stress hormones and other catabolic factors such as cytokines that are released in response to injury. The more prominent the modulation toward catabolism is, the greater the patient morbidity and the slower the recovery [7].

  Post-operative catabolic states can be treated with anabolic hormones, particularly growth hormone and IGF-1 [Non-patent document 9; Non-patent document 10; Non-patent document 11]. Several studies have shown that insulin treatment is clearly beneficial in catabolic injury patients [Non-patent document 12; Non-patent document 13; Non-patent document 14; Non-patent document 15; Reference 17].

  However, another study has shown that the postoperative benefits of insulin are often impaired by insulin resistance. With insulin resistance, normal concentrations of insulin usually elicit less than the following responses. Insulin resistance may be due to a decrease in the binding between insulin and cell surface receptors or a change in intracellular metabolism. The first type, characterized as a decrease in insulin sensitivity, can usually be overcome by increasing insulin concentration. The second type, characterized as a decrease in insulin responsiveness, cannot be overcome by large amounts of insulin.

  Since insulin resistance after injury can be overcome by the amount of insulin depending on the degree of insulin resistance, this seems to be a decrease in insulin sensitivity [Non-patent document 18; Non-patent document 19]. The decrease in insulin sensitivity after selective abdominal surgery lasts for at least 5 days, but does not exceed 3 weeks, and this decrease is most severe on the first postoperative day and returns to normal 3 weeks may be spent [Non Patent Literature 20].

  The cause of transient insulin resistance observed after injury is not fully understood. Both cortisol and glucagon can contribute to the catabolic response to injury [Non-patent document 21; Non-patent document 22; Non-patent document 23]. However, postoperative insulin resistance studies failed to show a correlation between the modulation of these protein catabolic hormones and changes in insulin sensitivity after surgery [20]; Non-patent document 25].

  Increased lipid availability after injury may induce insulin resistance via the glucose-fatty acid cycle [26]. Even when insulin was co-injected, increased availability of free fatty acids (FFA) induced insulin resistance and altered substrate oxidation from glucose to fat [27]; [28] Reference 29; Non-patent document 30; Non-patent document 31].

  Selective surgical procedures are typically performed after an overnight fast to reduce the risk of anesthesia. This established practice of fasting patients overnight (10-16 hours) prior to surgery increases catabolic development and exacerbates insulin resistance. Studies in rats loaded with stresses such as hemorrhage and endotoxemia show that fasting within 24 hours significantly exacerbates the catabolic response to injury [Non-Patent Document 32; Non-Patent Document 33; Ljungqvist , O., et al., Am. J. Physiol., 22: E692-98 (1990) (Non-patent Document 34) Even if the fasting time before the injury is short, the carbohydrate storage amount is markedly reduced in rats. , The hormonal environment changes significantly, the stress response increases, and most importantly the mortality increases [32].

  When glucose is administered orally [35] before surgery or by infusion, insulin resistance after surgery is reduced compared to fasted patients. Patients who received glucose infusion (5 mg / kg / min) overnight prior to selective abdominal surgery lost an average of 32% postoperative insulin sensitivity, and patients who entered surgery after a regular overnight fast were insulin Sensitivity is lost by an average of 55% [Non-patent Document 36].

  In addition to the adverse effects of fasting on recovery from surgery, patient fixation and low calorie nutrition during and after surgery also increases insulin resistance after surgery. In healthy individuals, 24-hour immobilization and low calorie nutrition have been shown to increase peripheral insulin resistance in healthy volunteers by 20-30%. Thus, the reported postoperative insulin resistance [37] after preoperative glucose infusion may be due in part to the additional effects of postoperative bed rest and low calorie nutrition.

  Given the prevalence of surgery, it is important to minimize negative side effects such as catabolic responses and insulin resistance, improve healing and reduce mortality. Postoperative insulin resistance leads to failure of treatment of catabolic conditions with insulin. Pre-operative fasting, a hard medical practice, exacerbates postoperative catabolism and insulin resistance. Thus, there is a need for treatments that overcome both catabolic conditions and insulin resistance.

  One treatment described herein to overcome both catabolic conditions and insulin resistance is to co-administer glucose and insulin before, during and after surgery. However, insulin infusion can cause hypoglycemia, where blood glucose levels (blood glucose) are defined as 0.3 mM or less. Hypoglycemia increases the risk of ventricular arrhythmias and is a dangerous consequence of insulin infusion. In order to prevent hypoglycemia, an algorithm for insulin injection for diabetes has been developed [Non-patent Document 38]. However, even under this algorithm, 21% of patients caused hypoglycemia. In another glucose control test after myocardial infarction, 18% of patients caused hypoglycemia when insulin and glucose were infused [39].

  Further monitoring of blood glucose levels is necessary for insulin infusion so that the onset of hypoglycemia can be detected and treated as quickly as possible. In patients who received insulin infusion in the above test [Non-Patent Document 40], blood glucose was measured at least every 2 hours and the infusion rate was adjusted accordingly. Thus, the safety and efficacy of insulin-glucose infusion therapy in patients with myocardial infarction depends on the ease and speed of access to blood glucose data. This strong need to monitor blood glucose places a heavy burden on health care workers and increases the inconvenience and cost of the procedure. As a result, pre-surgical clinics often do not take steps to monitor and optimize pre-surgical blood glucose levels that would be obtained by intravenous administration of insulin. Given the risks and burdens inherent in insulin infusion, new ways of controlling the catabolic response to injury before / after surgery are sought.

  Endocrine hormone glucagon-like peptide 1 (hereinafter abbreviated as GLP-1) is processed from proglucagon in the gastrointestinal tract and increases nutrient-induced insulin release [Non-patent Document 41]. Various decapitated forms of GLP-1 are known to stimulate insulin secretion (insulinotropic action) and cAMP production [see, eg, Non-Patent Document 42]. Correlation between various in vitro research experiments and insulin secretory response in mammals, particularly humans, to externally administered GLP-1, GLP- (7-36) amide and GLP-1 (7-37) acid is established [See, for example, Non-Patent Document 43; Non-Patent Document 44; Non-Patent Document 45; and Non-Patent Document 46]. GLP-1 (7-36) amide exhibits a clear anti-diabetic effect in insulin-dependent diabetic patients by stimulating insulin sensitivity and increasing glucose-induced insulin release at physiological concentrations [non-patented Reference 44]. When GLP-1 (7-36) amide is administered to non-insulin dependent diabetic patients, insulin release is stimulated, glucagon secretion is reduced, gastric emptying is inhibited, and glucose utilization is increased [Non-Patent Document]. 47; Non-patent document 48; Non-patent document 49].

  The use of GLP-1 type molecules for long-term therapy has been hampered by the very short serum half-life of such peptides. For example, GLP-1 (7-37) has a serum half-life of 3-5 minutes. GLP-1 (7-36) amide has a half-life of about 50 minutes when administered subcutaneously. Thus, in order to obtain long-term effects, these GLP molecules must be administered by continuous infusion [Non-patent Document 50].

Shaw, J.H.F., et al., Ann. Surg., 209 (1): 63-72 (1989) Little, R.A., et al., Prog. Clin. Biol. Res. 263A: 463-475 (1987) Frayn, K.N., Clin. Endorinol. 24: 577-599 (1986) Brandi, L., et al., Clin. Sci. 85: 525-35 (1993) Gump, F.E., et al., J. Trauma, 14: 378-88 (1974) Black, P.B., et al., Ann. Surg. 196: 420-35 (1982) Thorell, A., et al., Eur. J. Surg., 159: 593-99 (1993) Chernow, B., et al., Arch. Intern. Med., 147: 1273-8 (1987) Hammarkvist, F., et al., Ann. Surg., 216 (2): 184-190 (1991) Ziegler, T., et al., Annu. Rev. Med., 45: 459-80 (1994) Ziegler, T.R., et al., J. Parent. Ent. Nutr. 14 (6): 547-81 (1990) Hinton.P., Et al., Lancet, 17 (April): 767-69 (1971) Shizgal, H., et al., Am. J. Clin. Nutr., 50: 1355-63 (1989) Woolfson, A.M.J., et al., N.Clin.Nutr., 50: 1355-63 (1989) Woolfson, A.M.J., et al., N.Engl.J.Med. 300: 14-17 (1979) Brooks, D., et al., J. Surg. Res. 40: 395-405 (1986) Sakurai, Y., et al., Ann. Surg. 222: 283-97 (1995) Brandi, L.S., et al., Clin. Science 79: 443-450 (1990) Henderson, A.A., et al., Clin. Sci. 80: 25-32 (1990) Thorell, A. et al. (1993) Alberti, J.G.M.M., et al., J.Parent.Ent.Nutr.4 (2): 141-46 (1980) Gelfand, R.A., et al., J. Clin. Invest. 74 (December): 2238-2248 (1984) Marliss, E.B., et al., J. Clin. Invest. 49: 2256-70 (1970) Thorell A., Karolinska Hospital and Institute, 104 01 Stockholm, Sweden (1993) Thorell, A, et al., Br. J. Surg. 81: 59-63 (1994) Randle, P.J., et al., Diab. Metab. Rev. 4 (7): 623-38 (1988) Ferrannini, E., et al., J. Clin. Invest. 72: 1737-47 (1983) Bevilacqua, S. et al., Metabolism 36: 502-6 (1987) Bevilacqua, S. et al., Diabetes 39: 383-89 (1990) Bonadonna, R.C., et al., Am. J. Physiol. 259: E736-50 (1990) Bonadonna, R.C., et al., Am. J. Physiol. 257: E49-56 (1989) Alibegovic, A., et al., Circ. Shokc, 39: 1-6 (1993) Eshali, A.H., et al., Eur.J. Surg., 157: 85-89 (1991) Ljungqvist, O., et al., Am. J. Physiol., 22: E692-98 (1990) Nygren, J., et al., Ann. Surg. 222: 728-34 (1995) Ljngqvist, O., et al., J. Am. Coll. Surg. 178: 329-36 (1994) Ljungqvist, O., (1994) Hendra, T.J., et al., Diabetes Res. Clin. Pract., 16: 213-220 (1992) Malmberg, K.A., et al., Diabetes Care, 17: 1007-1014 (1994) Malmberg, 1994 Krcymann B., et al., Lancet 2: 1300-1303 (1987) Mojsov, S., Int. J. Peptide Protein Reserch, 40: 333-343 (1992) Nauck, M.A., et al., Diabetologia, 36: 741-744 (1993) Gutniak, M., et al., New England J. of Medicine, 326 (20): 1316-1322 (1992) Nauck, M.A., et al., J. Clin. Invest., 91: 301-307 (1993) Thorens, B., et al., Diabetes, 42: 1219-1225 (1993) Nauck, 1993 Gutniak, 1992 Nauck, 1993 Gutniak M. et al., Diabetes Care 17: 1039-1044 (1994)

In the present invention, patients are usually hospitalized before surgery, where the solution is administered parenterally continuously before, during, and after surgery, resulting in a short half-life of GLP-1 and consequent The need for such continuous administration is not inconvenient.
Accordingly, the present invention is a method for attenuating post-surgical catabolic modulation and insulin resistance, in which GLP-1, GLP-1 homologues, GLP-1 derivatives and their pharmaceuticals are used in patients in need thereof. The present invention provides for the first time a method characterized by administering a compound selected from the group consisting of pharmaceutically acceptable salts.

（配列番号１） “GLP-1” means GLP-1 (7-37). It is customary in the art to assign the number 7 to the amino terminus of GLP-1 (7-37) and the number 37 to the carboxy terminus. The amino acid sequence of GLP-1 (7-37) is well known to those skilled in the art, but for convenience, the sequence is shown below:

(SEQ ID NO: 1)

A “GLP-1 homologue” is defined as a molecule having one or more amino acid substitutions, deletions, inversions or additions compared to GLP-1. GLP-1 homologs known to those skilled in the art include, for example, GLP-1 (7-34) and GLP-1 (7-35), GLP-1 (7-36), Gln ⁹ -GLP-1 ( 7-37), D-Gln ⁹ -GLP-1 (7-37), Thr ¹⁶ -Lys ¹⁸ -GLP-1 (7-37), and Lys ¹⁸ -GLP-1 (7-37). Preferred GLP-1 homologues are GLP-1 (7-34) and GLP-1 (7-35), which are incorporated herein by reference, US Pat. No. 5,118,666. Also preferred is GLP-1 (7-36), which is a biologically processed form of GLP-1 having insulin secretion properties. Other GLP-1 analogs are described in US Pat. No. 5,545,618, which is hereby incorporated by reference.

A “GLP-1 derivative” has the amino acid sequence of GLP-1 or a homologue of GLP-1, but one of the amino acid side chain groups, α-carbon atom, terminal amino group or terminal carboxylic acid group or It is defined as a molecule that additionally has chemical modifications. Chemical modifications include, but are not limited to, adding chemical moieties, creating new bonds, and deleting chemical moieties. Modifications of amino acid side groups include acylation of lysine ε-amino groups, N-alkylation of arginine, histidine or lysine, alkylation of glutamic acid or aspartic acid carboxylic acid groups, and deamidation of glutamine or asparagine. However, it is not limited to these. Terminal amino modifications include, but are not limited to, des-amino, N-lower alkyl, N-di-lower alkyl, and N-acyl modifications. Modifications of the terminal carboxy group include, but are not limited to, amide, lower alkyl amide, dialkyl amide and lower alkyl ester modifications. Lower alkyl is _C 1 -C ₄ alkyl. Furthermore, one or more side chain groups or terminal groups may be protected by protecting groups known to protein chemists with ordinary knowledge. The α-carbon of the amino acid may be mono- or di-methylated.

（配列番号２）
［式中、Ｒ_１はＬ−ヒスチジン、Ｄ−ヒスチジン、デスアミノ−ヒスチジン、２−アミノ−ヒスチジン、β−ヒドロキシ−ヒスチジン、ホモヒスチジン、α−フルオロメチル−ヒスチジン、およびα−メチル−ヒスチジンの中から選ばれ、
ＸはＡｌａ、Ｇｌｙ、Ｖａｌ、Ｔｈｒ、Ｉｌｅおよびα−メチル−Ａｌａの中から選ばれ、
ＹはＧｌｕ、Ｇｌｎ、Ａｌａ、Ｔｈｒ、ＳｅｒおよびＧｌｙの中から選ばれ、
ＺはＧｌｕ、Ｇｌｎ、Ａｌａ、Ｔｈｒ、ＳｅｒおよびＧｌｙの中から選ばれ、そして
Ｒ_２はＮＨ_２およびＧｌｙ−ＯＨの中から選ばれる。ただし、該化合物の等電点は約６.０から約９.０であり、また、Ｒ_１がＨｉｓ、ＸがＡｌａ、ＹがＧｌｕおよびＺがＧｌｕの場合、Ｒ_２はＮＨ_２でなければならない。］
で示される分子およびその製薬的に許容される塩から構成される。 A preferred group of compounds of GLP-1 homologues or derivatives used in the present invention is of the formula:

(SEQ ID NO: 2)
Wherein R ₁ is selected from among L-histidine, D-histidine, desamino-histidine, 2-amino-histidine, β-hydroxy-histidine, homohistidine, α-fluoromethyl-histidine, and α-methyl-histidine. Chosen,
X is selected from Ala, Gly, Val, Thr, Ile and α-methyl-Ala;
Y is selected from Glu, Gln, Ala, Thr, Ser and Gly,
Z is selected from among Glu, Gln, Ala, Thr, Ser and Gly, and R ₂ is selected from among NH ₂ and Gly-OH. However, the isoelectric point of the compound is from about 6.0 to about 9.0, and when R ₁ is His, X is Ala, Y is Glu and Z is Glu, R ₂ is not NH ₂ Don't be. ]
And a pharmaceutically acceptable salt thereof.

など［例えばＷＯ９１／１１４５７号を参照］がある。 Many GLP-1 homologues and derivatives with isoelectric points in this range have already been described, including, for example,

[See, for example, WO91 / 11457].

Another preferred group of active compounds for use in the present invention is described in WO 91/11457, and has GLP-1 (7-34), GLP- having at least one modification selected from the following modification groups: 1 (7-35), GLP-1 (7-36) or GLP-1 (7-37) or their amides and their pharmaceutically acceptable salts:
(a) substitution with glycine, serine, cysteine, threonine, asparagine, glutamine, tyrosine, alanine, valine, isoleucine, leucine, methionine, phenylalanine, arginine or D-lysine in place of lysine at positions 26 and / or 34; or Substitution of glycine, serine, cysteine, threonine, asparagine, glutamine, tyrosine, alanine, valine, isoleucine, leucine, methionine, phenylalanine, lysine or D-arginine in place of arginine at position 36;
(b) substitution of an oxidation resistant amino acid in place of tryptophan at position 31;
(c) 16th position valine to tyrosine, 18th position serine to lysine, 21st position glutamic acid to aspartic acid, 22nd position glycine to serine, 23rd position glutamine to arginine, 24th position alanine to arginine and 26th position lysine At least one substitution selected from among glutamine;
(d) glycine, serine or cysteine in place of alanine at position 8; aspartic acid, glycine, serine, cysteine, threonine in place of glutamic acid at position 9; asparagine, glutamine, tyrosine, alanine, valine, isoleucine, leucine, methionine or phenylalanine; At least one substitution selected from serine, cysteine, threonine, asparagine, glutamine, tyrosine, alanine, valine, isoleucine, leucine, methionine or phenylalanine in place of glycine at position 10; and glutamic acid in place of aspartate at position 15; and
(e) Substitution of glycine, serine, cysteine, threonine, asparagine, glutamine, tyrosine, alanine, valine, isoleucine, leucine, methionine or phenylalanine or D- or N-acylated or alkylated histidine instead of histidine at position 7 ;
Here, in the substitution of (a), (b), (d) and (e), the substituted amino acid may be D-type, and the amino acid substituted at the 7-position is N-acylated or N- It may be an alkylated type.

Since dipeptidyl-peptidase IV enzyme (DPPIV) can be involved in the observed rapid in vivo inactivation of GLP-1 [eg Mentlein, R., et al., Eur. J. Biochem., 214: 829- 835 (1993)], it is preferred to administer GLP-1 homologues and derivatives protected from this DPPIV activity, Gly ⁸ -GLP-1 (7-36) NH ₂ , Val ⁸ -GLP-1 (7- 37) Administering OH, α-methyl-Ala ⁸ -GLP-1 (7-36) NH ₂ , and Gly ⁸ -Gln ²¹ -GLP-1 (7-37) OH or a pharmaceutically acceptable salt thereof More preferably.

（配列番号３）
［式中、ＸはＬｙｓおよびＬｙｓ−Ｇｌｙから選ばれる］
を有するペプチドおよび該ペプチドの誘導体から構成される群から選ばれ、該ペプチドは該ペプチドの製薬的に許容される酸付加塩、該ペプチドの製薬的に許容されるカルボキシレート塩、該ペプチドの製薬的に許容される低級アルキルエステル、およびアミド体、低級アルキルアミド体および低級ジアルキルアミド体から選ばれる該ペプチドの製薬的に許容されるアミド体の中から選ばれる。 The molecules claimed in US Pat. No. 5,188,666 (which is expressly incorporated herein by reference) are preferably used in the present invention. Such molecules have the following amino acid sequences:

(SEQ ID NO: 3)
[Wherein X is selected from Lys and Lys-Gly]
Wherein the peptide is a pharmaceutically acceptable acid addition salt of the peptide, a pharmaceutically acceptable carboxylate salt of the peptide, a pharmaceutical of the peptide Selected from pharmaceutically acceptable lower alkyl esters and pharmaceutically acceptable amides of the peptide selected from amides, lower alkylamides and lower dialkylamides.

（配列番号４）
［式中、Ｒ^１は４−イミダゾプロピオニル、４−イミダゾアセチル、または４−イミダゾ−α，α−ジメチルアセチルであり、
Ｒ^２はＣ_６−Ｃ_１０非分枝アシルの中から選ばれるか、または不存在であり、
Ｒ^３はＧｌｙ−ＯＨまたはＮＨ_２であり、
ＸａａはＬｙｓまたはＡｒｇである］
で示される米国特許第５，５１２，５４９号（これは引用によって本明細書に明らかに包含される）にて特許請求されている化合物およびその製薬的に許容される塩から構成され、これらを本発明に使用することができる。 Another preferred group of compounds used in the present invention has the formula:

(SEQ ID NO: 4)
[Wherein R ¹ is 4-imidazopropionyl, 4-imidazoacetyl, or 4-imidazo-α, α-dimethylacetyl;
R ² is selected from C ₆ -C ₁₀ unbranched acyl or is absent,
R ³ is Gly-OH or NH ₂ ;
Xaa is Lys or Arg]
Consisting of the compounds claimed in US Pat. No. 5,512,549 (which is expressly incorporated herein by reference) and pharmaceutically acceptable salts thereof, Can be used in the present invention.

A more preferred compound represented by SEQ ID NO: 4 used in the present invention is a compound wherein Xaa is Arg and R ² is C ₆ -C ₁₀ unbranched acyl.
An even more preferred compound represented by SEQ ID NO: 4 used in the present invention is a compound wherein Xaa is Arg, R ² is C ₆ -C ₁₀ unbranched acyl, and R ³ is Gly-OH. .

A more preferred compound represented by SEQ ID NO: 4 used in the present invention is that Xaa is Arg, R ² is C ₆ -C ₁₀ unbranched acyl, R ³ is Gly-OH, and R ¹ is It is a compound that is 4-imidazopropionyl.
The most preferred compound represented by SEQ ID NO: 4 used in the present invention is that Xaa is Arg, R ² is C ₈ unbranched acyl, R ³ is Gly-OH, and R ¹ is 4-imidazo A compound that is propionyl.

（配列番号１）
を有するペプチドおよび該ペプチドの誘導体から構成される群から選ばれ、該ペプチドは該ペプチドの製薬的に許容される酸付加塩、該ペプチドの製薬的に許容されるカルボキシレート塩、該ペプチドの製薬的に許容される低級アルキルエステル、ならびにアミド体、低級アルキルアミド体および低級ジアルキルアミド体から選ばれる該ペプチドの製薬的に許容されるアミド体の中から選ばれる。 More preferably, the molecules claimed in US Pat. No. 5,120,712 (which is expressly incorporated herein by reference) are used in the present invention. Such molecules have the following amino acid sequences:

(SEQ ID NO: 1)
Wherein the peptide is a pharmaceutically acceptable acid addition salt of the peptide, a pharmaceutically acceptable carboxylate salt of the peptide, a pharmaceutical of the peptide Selected from pharmaceutically acceptable lower alkyl esters and pharmaceutically acceptable amides of the peptide selected from amides, lower alkylamides and lower dialkylamides.

（配列番号５）
である。 In the present invention, it is most preferable to use GLP-1 (7-36) amide or a pharmaceutically acceptable salt thereof. The amino acid sequence of GLP-1 (7-36) amide is:

(SEQ ID NO: 5)
It is.

  Methods for preparing the active compounds used in the present invention, ie, GLP-1, GLP-1 homologues or GLP-1 derivatives used in the present invention are well known and are described in US Pat. No. 5,118,666. 5,120,712 and 5,523,549, which are hereby incorporated by reference.

  The amino acid part of the active compound or precursor thereof used in the present invention is produced either by 1) solid phase synthetic chemistry, 2) purification of GLP molecules from natural sources or 3) recombinant DNA technology.

  Solid phase chemical synthesis of polypeptides is well known to those skilled in the art and is described by Dugas, H. and Penney, C., Bioorganic Chemistry, Springer-Verlag, New York (1981), pp54-92, Merrifield, JM, Chem. Soc. , 85: 2149 (1962), and common textbooks in the industry such as Stewart and Young, Solid Phase Peptide Synthesis, Freeman, San Francisco (1969) pp. 24-66.

For example, the amino acid moiety is a 430A peptide synthesizer (PE-Applied Biosystems, Inc., 850 Lincoln Center Drive, Foster City, CA 94404) supplied by PE-Applied Biosystems and a synthetic cycle. It can be synthesized by the phase method. BOC-amino acids and other reagents are commercially available from PE-Applied Biosystems and other chemical suppliers. For the preparation of C-terminal carboxamides, continuous Boc chemistry using a double couple protocol is applied to the starting p-methylbenzhydrylamine resin. For the production of C-terminal acids, the corresponding PAM resin is used. Asn, Gln and Arg are coupled using a previously prepared hydroxybenzotriazole ester. The following can be used as side chain protecting groups:
Arg, tosyl Asp, cyclohexyl Glu, cyclohexyl Ser, benzyl Thr, benzyl Tyr, 4-bromocarbobenzoxy.

  Boc deprotection can be performed using trifluoroacetic acid in methylene chloride. When the synthesis is complete, the peptide is deprotected with anhydrous hydrogen fluoride (HF) containing 10% meta-cresol and cleaved from the resin. The cleavage of the side chain protecting group and the peptide from the resin are performed at -5 ° C to 5 ° C, preferably on ice for 60 minutes. After removal of HF, the peptide / resin is washed with ether and the peptide is extracted with glacial acetic acid and lyophilized.

The active compounds used in the present invention can be prepared using techniques well known to those skilled in the art of recombinant DNA technology. Indeed, the recombinant DNA technique is preferred because of its high yield. The basic process of this recombinant production consists of the following steps:
a) isolating the natural DNA sequence encoding the GLP-1 molecule, or constructing a synthetic or semi-synthetic DNA coding sequence of the GLP-1 molecule,
b) transferring the resulting coding sequence into an expression vector in a manner suitable for expressing the protein alone or as a fusion protein;
c) transforming an appropriate eukaryotic or prokaryotic host cell with this expression vector,
d) culturing the transformed host cells under conditions that allow expression of the GLP-1 molecule; and e) recovering and purifying the recombinantly produced GLP-1 molecule.

As described above, the coding sequence may be synthesized as a whole or may be the product of a modification of a larger native glucagon-encoding DNA. The DNA sequence encoding preproglucagon is described in Lund et al., Proc. Natl. Acad. Sci. USA 79 : 345-349 (1982), which is modified by the present invention by modification to obtain the desired product. Can be used as starting materials in the semi-synthetic production of these compounds.

  Synthetic genes in in vitro or in vivo transcription and translation leading to production of GLP-1 molecules can be constructed by techniques well known to those skilled in the art. Due to the natural degeneracy of the genetic code, those skilled in the art will appreciate that a fairly large but limited number of DNA sequences can be constructed that all encode the GLP-1 molecule.

  Methods for constructing synthetic genes are well known to those skilled in the art. See Brown et al. (1979) Methods in Enzymology, Academic Press, N.Y., 68: 109-151. The genetic code is used to design a DNA sequence from the desired amino acid sequence, which can be easily ascertained by a biologist in the art. If designed, the sequence itself can be synthesized by a conventional DNA synthesizer, such as a 380A or 280B model synthesizer (PE-Applied Biosystems, Inc., 850 Lincoln Center Drive, Foster City, CA 94404).

  In order to express the amino acid portion of the compounds used in the present invention, the engineered synthetic DNA sequence is inserted into a number of suitable recombinant DNA expression vectors by using an appropriate restriction endonuclease. For a review, see Maniatis et al. (1989) Molecular Cloning; A Laboratory Manual, Cold Springs Harbor Laboratory Press, N.Y., Vol. 1-3. The restriction endonuclease cleavage site is isolated from an amplification and expression vector well known to those skilled in the art and engineered to be at either end of the GLP-1 molecule encoding DNA so that it can be incorporated into it. The particular endonuclease used will vary depending on the restriction endonuclease cleavage pattern of the parent expression vector used. The restriction sites are selected so that the coding sequence is correctly oriented relative to the control sequence, so that the protein of interest is correctly placed and expressed in the open reading frame. The coding sequence must be located in the correct reading frame relative to the promoter and ribosome binding site of an expression vector that functions in the host cell in which the protein is to be expressed.

  In order for a synthetic gene to transcribe efficiently, it must be operably associated with a promoter-operator region. Thus, the promoter-operator region of the synthetic gene is placed sequentially in the same orientation relative to the ATG start codon of the synthetic gene.

A variety of expression vectors useful for transforming prokaryotic and eukaryotic cells are well known to those of skill in the art. The Promega Biological Research Products Catalog (1992) (Promega Corp., 2800 Woods Hollow Road, Madison, WI, 53711-5399); and The Stratagene Cloning Systems Catalog (1992) (Stratagene Corp., 11011 North Torrey Pines Road, La Jolla , CA, 92037). Further, US Pat. No. 4,710,473 discloses a circular DNA plasmid transformation vector useful for expressing foreign genes at a high rate in E. coli. This plasmid is useful as a transformation vector for recombinant DNA techniques, and
(a) imparting autonomous replication ability to a plasmid in a host cell;
(b) controlling autonomous plasmid replication in relation to the temperature at which the host cell culture is maintained;
(c) stabilizing the integrity of the plasmid in the host cell aggregate;
(d) synthesizing a protein product that serves as an indicator of plasmid integrity in the host cell population;
(e) providing a series of restriction endonuclease recognition sites unique to the plasmid; and
(f) Terminate mRNA transcription.
This circular DNA plasmid is useful as a vector for a recombinant DNA technique for expressing a foreign gene at a high level.

  Once an expression vector for the amino acid portion of the compound used in the present invention is constructed, the next step is to construct a recombinant host cell useful for expressing the polypeptide by placing the vector in a suitable cell. It is. Techniques for transforming cells with recombinant DNA vectors are well known to those skilled in the art and can be found in general texts such as Maniatis et al. (Supra). The host cell to be created is constructed from either eukaryotic or prokaryotic cells.

  Prokaryotic host cells generally produce proteins in culture relatively easily and in a relatively high rate. Proteins expressed in high level bacterial expression systems are characterized by aggregation in the form of particles or inclusion bodies, including high levels of overexpressed proteins. Such protein aggregates usually must be recovered, solubilized, denatured and refolded by techniques well known to those skilled in the art. Kreuger, et al. (1990) in Protein Folding, Gierash and King, eds, pg136-142, Amerian Association for the Advancement of Science Publication No. 89-18S, Washington, DC; and U.S. Pat.No. 4,923,967. See

  To prepare the desired GLP-1 homologue or GLP-1 derivative, the precursor GLP-1 or GLP-1 homologue amino acid modifications are modified by the following well-known methods: Chemicals of GLP-1 precursor Modification, enzymatic modification or a combination of chemical and enzymatic modification. Traditional liquid phase and semi-synthetic techniques are also useful in preparing the GLP-1 molecules used in the present invention. Methods for preparing the GLP-1 molecules of the present invention are well known to those skilled in the art of peptide chemistry.

Acyl groups can be added to the ε-amino group of Lys ³⁴ by any of a variety of methods known to those skilled in the art. Bioconjugate Chem. “Chemical Modifications of Proteins: History and Applications”, pages 1–2-12 (1990) and Hashimoto et al., Pharmaceutical Res. 6 (2): 171-176 (1989).
For example, N-hydroxy-succinimide ester of octanoic acid can be added to lysyl-ε amine using 50% acetonitrile in borate buffer. The peptide can be acylated either before or after adding the imidazole group. Furthermore, if the peptide is prepared recombinantly, it may be acylated prior to enzymatic cleavage. Alternatively, lysines of GLP-1 derivatives can be acylated as taught in WO 96-29342, which is incorporated herein by reference.

  Many protected, unprotected and partially protected natural and non-natural functional homologues in GLP-1 (7-36) amide and GLP-1 (7-37) molecules and The presence and preparation of the derivatives are described in the literature of the art [eg, US Pat. Nos. 5,120,712 and 5,118,666, which are hereby incorporated by reference), and See Orskov, C., et al., J. Biol. Chem., 264 (22): 12826-12829 (1989) and WO 91/11457 (Buckley, DI, et al., Issued August 8, 1991)] .

  If desired, the amino and carboxy terminal amino acid residues of the GLP-1 derivative may be protected, or either one may be protected. Reactions to form and remove such protecting groups are described in standard literature such as “Protective Groups in Organic Chemistry”, Plenum Press, London and New York (1973); Green, TH, “Protective Groups in Organic Synthesis”, Whiley, New York (1981); and “The Peptides”, Vol. I, Schroder and Lubke, Academic Press London and New York (1965). Representative amino protecting groups include, for example, formyl, acetyl, isopropyl, butoxycarbonyl, fluorenylmethoxycarbonyl, carbobenzyloxy, and the like. Representative carboxy protecting groups include, for example, benzyl ester, methyl ester, ethyl ester, t-butyl ester, p-nitrophenyl ester and the like.

GLP-1 derivatives of lower alkyl esters at the carboxy terminus used in the present invention are prepared by reacting the desired (C ₁ -C ₄ ) alkanol with the desired polypeptide in the presence of a catalytic acid such as hydrochloric acid. The Suitable conditions for such alkyl ester formation are a reaction temperature of about 50 ° C. and a reaction time of about 1 hour to about 3 hours. Similarly, alkyl ester derivatives of Asp and / or Glu residues can be formed.

  Preparation of carboxamide derivatives of the compounds used in the present invention is carried out as described, for example, in Stewart, J.M., et al., Solid Phase Peptide Synthesis, Pierce Chemical Company Press, 1984.

  In the present invention, pharmaceutically acceptable salt forms of GLP-1, GLP-1 homologues or GLP-1 derivatives can be used. Acids commonly used to form acid addition salts are inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and p-toluenesulfonic acid, methanesulfonic acid, Organic acids such as acid, p-bromophenylsulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid and acetic acid. Examples of such salts are sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogen phosphate, dihydrogen phosphate, metaphosphate, pyrophosphate, chloride, Bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caproate, heptanoate, propiolate, oxalate, malonic acid Salt, succinate, suberate, sebacate, fumarate, maleate, butyne-1,4-diate, hexyne-1,6-diate, benzoate, chlorobenzoate , Methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, sulfonate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate , Lactate, γ Hydroxybutyrate, glycolate salts include, tartrate, methanesulfonate, propanesulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate and other salts. Preferred acid addition salts are salts formed from mineral acids such as hydrochloric acid and odorous acid, especially hydrochloric acid.

  Base addition salts include those derived from ammonium or inorganic bases such as hydroxylated, carbonic acid, alkali bicarbonate or alkaline earth metals. Useful bases for preparing the salts of the present invention include sodium hydroxide, potassium hydroxide, ammonium hydroxide, potassium carbonate and the like. The salt form is particularly preferred.

The GLP-1, GLP-1 homologue or GLP-1 derivative used in the present invention can be formulated with one or more excipients prior to use in the present invention. For example, the active compounds used in the present invention can be complexed with divalent metal cations by well-known methods. Such metal cations include, for example, Zn ⁺⁺ , Mn ⁺⁺ , Fe ⁺⁺ , Co ⁺⁺ , Cd ⁺⁺ , Ni ^++, and the like.

  The active compounds used in the present invention can be combined with pharmaceutically acceptable buffers, if necessary, pH adjusted to provide acceptable stability, and suitable for parenteral administration The pH can be set. One or more pharmaceutically acceptable salts can be added to adjust ionic strength or tonicity. One or more excipients can be added to further adjust the isotonicity of the formulation. Glycerin is an example of an isotonicity adjusting excipient.

  Administration is by any route that the physician recognizes as effective. Parenteral administration is preferred. Parenteral administration is commonly understood in the medical literature as injecting the dosage form into the body via a sterile syringe or some other mechanical device such as an infusion pump. Parenteral administration includes intravenous, intramuscular, subcutaneous, intraperitoneal, intrathecal, intrathecal, intraventricular, intraarterial, subarachnoid, and epidural. Intravenous, intramuscular and subcutaneous routes of administration are more preferred for the compounds used in the present invention. Intravenous and subcutaneous routes are even more preferred for the compounds of the present invention. For parenteral administration, the active compound used in the invention is combined with distilled water at a suitable pH.

  Moreover, the action time can be controlled by another pharmaceutical method. Controlled release preparation can be carried out by using polymers capable of complexing or absorbing the active compounds used in the present invention. The extended period selects the appropriate macromolecule, such as polyesters, polyamino acids, polyvinylpyrrolidone, ethylene vinyl acetate, methylcellulose, carboxymethylcellulose or protamine sulfate, and the macromolecule concentration and incorporation method; This is achieved by prolonging the release. Another possible method for prolonging the activity is to use the active compounds used in the present invention for polymeric materials such as polyesters, polyamino acids, hydrogels, poly (lactic acid) or ethylene vinyl acetate copolymers. Incorporate into the particles. Alternatively, instead of incorporating the compound into these polymer particles, the compound used in the present invention can be encapsulated in a microcapsule, which is prepared, for example, by coacervation or interfacial polymerization, For example, hydroxymethylcellulose or gelatin-microcapsules, respectively, or the compounds can be encapsulated in colloidal drug delivery systems such as liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules, or microemulsions. . Such techniques are described in Remington's Pharmaceutical Sciences (1980).

  In the procedure of the present invention, the patient needs the compound used in the present invention for 1-16 hours prior to the surgical procedure, during the surgical procedure, and for a period not exceeding about 5 days after the surgical procedure.

  As noted above, the time prior to surgery to begin administering the compound used in the present invention is from about 16 hours to about 1 hour prior to the start of the surgery. The pre-surgical time at which the compounds used in the present invention to reduce catabolism and insulin resistance should be administered is a factor known to the physician, such as most importantly pre-surgical preparation Depends on whether the patient is fasting or is receiving glucose infusions or drinks or other forms of nutrition during the period, and the gender, weight and age of the patient, the degree of lack of glycemic control, It depends on, but is not limited to, the cause of the deficiency, the expected severity of the injury from the surgical procedure, the route of administration and bioavailability, the body's endurance, the formulation, and the active intensity of the administered compound. A preferred time interval for initiating administration of the compounds used in the present invention is from about 1 hour to about 10 hours prior to the start of the surgical procedure. The most preferred interval for initiating dosing is 2 to 8 hours prior to the start of the surgical procedure.

  As already explained, insulin resistance after individual forms of surgical procedures, selective abdominal surgery is most severe on the first day after surgery, which lasts at least 5 days and 3 weeks to normalize It takes a while [Thorell, A., et al. (1993)]. Thus, post-operative patients require that the compounds used in the present invention be administered for a period of time following a surgical injury depending on the factors determined by the physician. Among these factors are patients being fasted after surgery, glucose infusions or drinks or other forms of nutrition being provided, and also the patient's gender, weight and age, lack of glycemic control Depends on the extent of the disease, the cause of the lack of glycemic control, the actual severity of the injury caused by the surgical procedure, the route of administration and bioavailability, the body's endurance, the formulation, and the intensity of activity of the administered compound. It is not limited. The preferred administration time for the compounds used in the present invention is within 5 hours after the surgical procedure.

  The term “post-surgical catabolic modulation” is well known to surgeons [Shaw, JHF et al., Ann. Surg. (1989); Little, RA, et al. (1987); Freyn, KN (1986); Brandi, L. et al. (1993)], defined herein as a metabolic state resulting from surgical injury that can be characterized by one or more of the following phenomena: Loss of body nitrogen Negative nitrogen equilibrium with [Wernerman, J. et al., J. Parent. Enter. Nutr. 10: 578-82 (1986); Tashiro, T., et al., J. Paretn. Enter. Nutr. 9: 452-5 (1985)], peripheral utilization of lipids in preference to glucose with reduced respiratory quotient [Frayn, KN, et al., Arch. Emerg. Med. 4: 91-9 (1987); Stjernstrom, H. et al., Clin. Physiol. .1: 59-72 (1981)], and endogenous glucose production from body protein and energy storage despite hyperglycemia [Gump, FE et al. (1974); Black, RB et al. (1982); Frayn, KN, et al. (1987); Freyn, KN Br. Med. Bull. 41 (3): 232-9 (1985)].

  The term “insulin resistance” is also well known to clinicians and is defined herein as a physiological condition in which a normal concentration of insulin can elicit a response that is less than a normal response. Insulin resistance may be due to decreased insulin binding to cell surface receptors or changes in intracellular metabolism. The first type, characterized as a decrease in insulin sensitivity, can usually be overcome by increasing insulin concentration. The second type, characterized as a decrease in insulin responsiveness, cannot be overcome by large amounts of insulin. Since insulin resistance after injury can be overcome by the amount of insulin depending on the degree of insulin resistance, this is thought to be a decrease in insulin sensitivity [Brandi, LS, et al., Clin. Science 79: 443-450 ( 1990); Henderson, AA et al., Clin. Sci. 80: 25-32 (1990)]. The decrease in insulin sensitivity after selective abdominal surgery lasts for at least 5 days but does not exceed 3 weeks, this decrease being most severe on the first postoperative day and 3 to return to normal May spend weeks [Thorell, A. et al. (1993)]. The cause of transient insulin resistance observed after injury is not fully understood.

The dose of GLP-1, GLP-1 homologue or GLP-1 derivative effective to normalize a patient's blood glucose level depends on a number of factors such as the patient's gender, weight and age, lack of ability to regulate blood glucose Depending on the extent, the cause of the lack of glycemic control, whether to administer glucose or other carbohydrate sources at the same time, the route of administration and bioavailability, the body's endurance, the formulation, the intensity of activity of the administered compound, It is not limited to these. In the case of continuous administration, a suitable administration rate is 0.25 to 6 pmol / kg body weight / min, preferably about 0.5 to about 1.2 pmol / kg / min. When administered intermittently, the dose in a single dose should take into account the interval between doses, the bioavailability of GLP-1, GLP-1 homologues or GLP-1 derivatives, and the levels necessary to provide normoglycemia It is. The dosage and rate of administration of GLP-1, GLP-1 homologues or GLP-1 derivatives to obtain the desired clinical effect will be determined by the ordinary physician.
In order to make the present invention more straightforward, specific examples are set forth below, which are illustrative and not limiting of the present invention.

Example 1
Thirteen patients scheduled for selective orthopedic surgery (hipartroplasty) were subjected to this study. None of the patients has a history or signs of metabolic disease, liver damage or diabetes. Fasting blood glucose, CRP and liver tests (bilirubin, alkaline phosphatase, AST and ALT) were normal across all 13 patients.

Seven patients (insulin group, age 56 ± 5 years; BMI, 25 ± 1 kg / m ² ) were tested from 08:00 after an overnight fast. Samples are taken for blood glucose and hormone measurements and after an initial basal period of 30 minutes of indirect calorie measurement, insulin (Actrapid®, Novo, Copenhagen) is constant at 0.8 mU / kg / min. During the intravenous infusion, glucose (200 mg / ml) was variably injected intravenously to maintain blood glucose at a constant level (4.5 mM). After 1 hour of steady state conditions, all patients received a standardized surgical procedure (Hypertroplasty). Surgery started 290 ± 23 minutes after the start of insulin infusion. The high insulin, normoglycemic clamp was then maintained during the surgical procedure and continued for another 3-4 hours after the surgical procedure. The data is presented according to the following items:
Basic 30 minutes before the start of insulin infusion,
Preoperative clamp Steady state high insulin, normoglycemic clamp 60 minutes before surgery Initial op 10 to 40 minutes after start of surgery Late op Last 30 minutes of surgical procedure Postoperative clamp From 143 ± 30 minutes after start of surgical procedure A 60-minute steady state high insulin, normoglycemic clamp.

Same for 7 days prior to surgery for second group of patients (control group, n = 6, age 59 ± 3 years; BMI, 26 ± 1 kg / m ² ) compatible with the above insulin groups for age and BMI A preoperative protocol (basic and preoperative clamp) was applied. On the day of surgery, the control group did not receive a basal or preoperative clamp. However, at the same time as the surgical procedure, insulin infusion (0.8 mU / kg / min) was given to each patient in the control group, and a high insulin / normoglycemic (4.5 mM) clamp (postoperatively) was started.

  Indirect calorie measurement (Deltatrac®, Dansjoo, Sweden) [Frayn, KNJ Appl. Physiol. 55 (2): 628-34 (1983); Takala, J., et al., Crit. Care Med. 17 (10) : 1041-47 (1989)] was performed for 30 minutes during the basal period, twice during surgery (early op and late op) and the last 30 minutes of pre- and post-operative clamps. Urine was sampled periodically to analyze urinary urea excretion. After calibrating changes in urine storage [Tappy, L. et al., Diabetes 37: 1212-16 (1988)], non-protein energy expenditure (EE), respiratory quotient (RQ) and substrate oxidation rate were calculated.

  Blood samples were repeatedly taken from the veins of the warmed hands during the basal, preoperative, early op, late op and postoperative periods. Blood glucose was measured immediately after collection by the glucose oxidase method (Yellow Springs Instruments, Yellow Springs, Ohio) [Hugget, A.S., et al., Lancet 2: 368-70 (1957)]. Using radioimmunoassay (RIA), insulin [Grill, V. et al., Metabolism 39: 251-58 (1990)]; C-peptide (Novo Research, Bagsvaerd, Denmark); cortisol [Harris, V., et al., In Jaffe, BM & Behrman, HR, eds. Methods of Hormone radioimmunoassay, Academic Press, New York and London (1979) pp.643-56]; and glucagon (Euro-Diagnostica AB, Malmo, Sweden) [Faloona, GR, et al. , Glucagon radioimmunoassay technique. Vol.1: Academic Press, New York (1974)].

  All values are individual values or mean ± SEM (standard error of the mean). Mathematical significance is accepted at p <0.05 using the Wilcoxon signed rank test and Mann-Whitney U test for paired and unpaired data, respectively. Serum insulin levels at the time of post-operative clamp tended to be lower in the control group than in the insulin group (p = 0.06), so the GIR during the clamp period was also the 60 minute stationary phase GIR and the average serum insulin level Was calibrated for normal insulin levels.

  Serum insulin levels were similar between the two groups at basal and preoperative clamp periods. In the insulin group, insulin levels remained approximately 60 μU / ml during the surgical procedure and post-operative clamp phase. In the control group, insulin levels were unchanged compared to basal levels during the surgical procedure. Insulin levels at the time of postoperative clamp in the control group were not significantly different from those at the preoperative clamp period, and were not different from those at the postoperative clamp stage in the insulin group.

  C-peptide levels (Table I) were similar between the two groups at basal and pre- and post-operative clamp periods. C-peptide levels at the time of surgery in the insulin group were lower compared to the control group.

  Serum glucagon levels decreased after surgery in both groups (p <0.05) (Table I). However, the relative change after surgery (% before surgery) was greater in the insulin group (vs. control p <0.01).

  Serum cortisol levels (Table I) decreased after surgery in the insulin group and showed a tendency to increase in the control group (p = 0.1). Cortisol postoperative levels were lower in the insulin group than in the control group (p <0.05).

^＊Ｐ＜０.０５ Ｗilcoxonの符号つき順位検定による術前との比較
^†Ｐ＜０.０５ Ｍann-Ｗhitney Ｕ-検定による対照との比較 Table I
Hormonal levels of patients receiving hypertroplasty after overnight fast (control, n = 6) or after 4 hours of physiologically high insulin (insulin, n = 7)

^* P <0.05 Comparison with preoperative Wilcoxon signed rank test
^† P <0.05 Mann-Whitney U-test compared to control

  Although glucagon levels decreased in both groups after surgery, the greatest decrease (%) was found in the insulin group (vs. control <0.01). In the insulin group, cortisol levels decreased after surgery (versus preoperative p <0.05), while the levels in the control group tended to increase (p = 0.1). Thus, after surgery, cortisol levels are significantly lower in the insulin group than in the control group (p <0.05).

  The glucose infusion rate (GIR) was not significantly different between the insulin group and the control group during the preoperative clamp period. In the control group, the average GIR required to maintain normoglycemia during the postoperative clamp phase compared to the preoperative clamp was reduced (−39 ± 5%, p <0.05). In contrast, the insulin group maintained GIR during the surgical procedure and even showed a tendency to increase GIR on average during the postoperative clamp phase (+ 16 ± 20%, p = 0.2). Most importantly, surprisingly, the mean GIR in the postoperative clamp phase in the insulin group was significantly higher than in the control group (p <0.05) (see FIG. 1). All fluctuations in GIR during pre-operative and post-operative clamps were statistically significant (p <0.05), even if the GIR was calibrated for mean serum insulin levels at stationary phase.

  Glucose and lipid oxidation rates before surgery were similar in both groups. At the time of surgery, the glucose oxidation rate was significantly higher in the insulin group and the lipid oxidation rate was significantly lower (vs. control p <0.05). At the time of postoperative clamping in the insulin group, there was no change in the substrate oxidation rate compared with the preoperative clamping. Resting energy expenditure (EE) before and after surgery was not different between the two groups, and both groups remained the same after surgery compared to preoperative clamps.

  Fasting glucose levels were similar in the insulin and control groups. In the stationary phase at the time of insulin infusion, normoglycemia was maintained, and the average between the coefficients of glucose fluctuation was 4.6% in the control group and 6.2% in the insulin group.

  There is no doubt that the above findings show that patients undergoing selective surgery in the fasted state show insulin resistance after surgery and increase lipid oxidation. Furthermore, these findings indicate that if a patient is placed under surgical stress with elevated insulin levels that are maintained throughout the operation, post-surgery catabolism is completely blocked, and the hormonal response to stress is completely eliminated. It also shows for the first time that it is attenuated.

Example 2
GLP-1 (7-36) amide was administered to 5 patients with non-insulin dependent diabetes mellitus (NIDDM) by subcutaneous injection at a dosing rate of 1.2 pmol / kg / hr over 10 hours at night. As a control, insulin was continuously infused into the same five patients on a different day than the day of GLP-1 (7-36) amide infusion. The rate of insulin infusion was adjusted every 2 hours for optimal control to prevent hypoglycemia. As shown in the data in Table II and FIG. 2, subcutaneous infusion of GLP-1 (7-36) amide almost normalized blood glucose without inducing hypoglycemia in any patient. Metabolic control by GLP-1 (7-36) amide is better than that achieved by insulin, and mean blood glucose levels at 23:00, 0:00 and 1:00 are GLP-1 (7-36) The amide treatment was statistically significantly lower than the control.

Table II
Mean blood glucose level in 5 NIDDM patients who were continuously infused with GLP-1 (7-36) amide for 10 hours at night. In controls tested with the same patient on different days, insulin was administered by continuous infusion.

Example 3
GLP-1 (7-36) amide was injected during the day into 5 NIDDM patients at breakfast, lunch and dinner for 3 hours. The injection time was 7: 30-10: 30 (at breakfast), 10: 30-1: 30 (at lunch), and 4: 30-7: 30 (at dinner) as shown in FIG. It was. In a control experiment with the same 5 NIDDM patients performed on different days, insulin was injected subcutaneously immediately before the meal as shown in FIG. During the infusion of GLP-1, the postprandial glucose excursion observed by insulin infusion was eliminated, and normal blood glucose levels were maintained. Immediately after the end of each infusion of GLP-1 (7-36) amide, blood glucose levels increased significantly. No inappropriate side effects of GLP-1 (7-36) amide were observed. These data show that GLP-1 (7-36) amide infusion controls postprandial glucose levels more effectively than insulin injection and is effective as long as this control continues with GLP-1 (7-36) amide infusion. It shows that there is.

Table III
Mean blood glucose level of 5 NIDDM patients infused with GLP-1 (7-36) amide for 3 hours starting from the start of each meal. In a control experiment performed on the same day using the same patient, insulin was administered by subcutaneous injection immediately before each meal. Meals started at 7:30, 10:30 and 4:30.

FIG. 1 shows post-operative versus pre-operative (pre-operative) clamp period in six control patients (O) and seven patients (■) who underwent high insulinic normoglycemic infusion before and sometimes selective surgery (Postoperative) It is a graph of the change percentage of the glucose infusion rate (GIR) of a clamp period. FIG. 2 is a graph showing the effect (− ■ −) of continuous infusion of GLP-1 (7-36) amide on mean blood glucose concentration (mM) at night in 5 non-insulin dependent diabetes mellitus (NIDDM) patients. . The graph also shows the effect of continuous infusion of insulin (--O--) on another nocturnal mean blood glucose concentration in the same 5 NIDDM patients. FIG. 3 shows the effect of GLP-1 (7-36) amide on the mean blood glucose concentration (mM) of 5 NIDDM patients when injected for 3 hours from the start of each of 3 meals during the day (− ■ It is a graph which shows-). The graph also shows the effect of subcutaneous injection of insulin immediately before each meal (--O--) on another day's mean blood glucose concentration in the same 5 NIDDM patients.

Claims (13)

  A method for attenuating post-surgical catabolic modulation and insulin resistance to a patient in need thereof for GLP-1, GLP-1 homologues, GLP-1 derivatives and pharmaceutically acceptable salts thereof A method comprising administering a compound selected from the group consisting of:   The method of claim 1, wherein the compound is administered intravenously.   The method of claim 1, wherein the compound is administered subcutaneously.   4. A method according to claim 2 or 3, wherein the administration is continuous.   5. The method of claim 4, wherein the compound administration rate is 0.25 to 6 pmol / kg / min.   6. The method of claim 5, wherein the administration rate of the compound is 0.5 to 2.4 pmol / kg / min.   6. The method of claim 5, wherein the rate is from about 0.5 to about 1.2 pmol / kg / min.   The method of claim 2, wherein intravenous administration is intermittent.   3. The method of claim 2, wherein the compound is administered intravenously and further by other parenteral routes.   10. The method of claim 9, wherein the other parenteral route is the subcutaneous route.   2. The method of claim 1, wherein the compound to be administered is GLP (7-36) amide or a pharmaceutically acceptable salt thereof.   A method for attenuating hormonal responses to catabolic modulation and stress after surgery, wherein GLP-1, GLP-1 homologues and GLP-1 derivatives exhibit their insulinotropic activity to patients in need thereof A method comprising administering a compound that exhibits insulinotropic activity by interacting with the same receptor or group of receptors that interact with each other.   A method for attenuating hormonal responses to catabolic modulation and stress after surgical treatment, wherein GLP-1, GLP-1 homologues and GLP-1 derivatives interact to increase insulin sensitivity and Administering a compound that increases insulin sensitivity by interacting with the same.

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