USE OF GROWTH HORMONE FOR NITROGEN RETENTION UNDER HYPOCALORIC CONDITIONS

Abstract: A method and composition for effecting protein accretion in an animal. The composition comprises a hypocaloric dietary component comprising a metabolizable source of nitrogen, carbohydrate, and, optionally, at least one of a triglyceride source, minerals, and vitamins, in conjunction with growth hormone. The method comprises administering an effective amount of the hypocaloric dietary component and growth hormone, orally or parenterally, to effect a positive nitrogen balance under hypocaloric conditions.

Description: USE OF GROWTH HORMONE FOR NITROGEN RETENTION UNDER HYPOCALORIC CONDITIONS Cross Reference to Related Application This application is a continuation-in-part of United States Application Serial No. 817,263, filed January 9, 1986. Technical Field This invention relates to a method and composition for accreting protein in an animal under hypocaloric conditions. The method comprises administering a hypocaloric dietary component to the animal being treated in conjunction with growth hormone. The composition comprises growth hormone and a hypocaloric dietary component. The practice of this invention effects protein buildup in the treated animal where the animal, by design or necessity, is being maintained on a diet comprising from 10 to 95 percent of the resting metabolic requirement (RMR) . Background Art Pituitary growth hormone (GH) is an anabolic protein which promotes growth of tissue and is involved in the regulation of other phases of protein metabolism as well as fat, carbohydrate, and mineral metabolism. Growth hormones from various species differ in their antigenicities, in the range of animals in which they can produce biological responses, in their isoelectric points, N- terminal and C- ter inal amino acid residues and a ino acid composition. Their molecular weights range from 21,500 for human growt hormone (HGH) , to 47,400 for bovine growth hormone. An growth hormone appears to demonstrate a degree of species specificity. It is known, however, the humans respond t growth hormone of human or monkey origin. Growth hormone has been isolated from bovine anterio pituitary, Li, et al.. J. Biol. Chem. 159. 353 (1945) ; Wilhelmi et al.. J. Biol. Chem. 176 735 (1948); Li, U.S. Patent No. 3,118,815 (1964). Human growth hormone has bee isolated from human pituitary, Lewis et al.. U.S. Patent No. 2,974,088 (1961); Reisfeld, et al.. Endocrinology 71, 559 (1962). Until very recently, isolation of HGH from huma cadaver pituitary was the only source for the protein. Accordingly, lack of available material was a primar deterrent for the continuing investigation and definition o therapeutic roles for HGH. Recently, however, a practical method for synthesizin biologically active growth hormone via recombinant DN technology has been developed, The Medical Letter, vol. 27, 101-102 (1985) . Further, recombinant HGH has now bee approved by the Food and Drug Administration for use i humans. See, for example, Genetic Engineering News, Vol. 5, No. 10, pp 1,8 (1985) . Growth hormone plays a prominent role in protei metabolism and the regulation of growth. This i accomplished by accelerating the rate of transfer of amin acids from the extracellular to the intracellular compartmen and incorporating the transferred amino acids into cel proteins. Evidence that growth hormone stimulates th synthesis of messenger RNA, ribosomal RNA, and transfer RN in liver has led to the hypothesis that growth hormon promotes protein synthesis via gene activation. Se PHYSIOLOGY, Third Edition, Edited by Selkurt, Little, Brow and Company, page 730 (1971) . The overall effect of growt hor one on protein metabolism is evident in the well- docu ented increase in linear growth resulting from the administration of growth hormone to GH-deficient dwarves. I has been shown that this increase in body cell mass (a reflected by total body potassium) is at the expense o adipose tissue. Collipp, T.J. et al.. Metabolism 2214, 589 595 (1973). Similar changes in burn patients treated with growt hormone have been demonstrated as well. Soroff, H.S. et al. , Ann. Surg. 166. 739-752 (1967) . Metabolic studies on norma subjects have consistently shown overall retention o nitrogen and potassium as well as other cellular constituent upon administration of growth hormone. Beck, J.C. et al. Metabolism 8 , 699-737 (1960); Bergenstall, D.M. , et . al. , ___ Clin. Endo. and Metab.. 20-11. 1427-1436 (1960) ; and MR Panel, Lancet, 1., 7-12 (1959) . The action of growth hormon in stressed states has been studied extensively in burne patients. Prudden et al. administered bovine growth hormon to four burned patients and demonstrated an anabolic effec dependent on food intake; growth hormone only improve nitrogen balance at high levels of nitrogen intake. Below certain level of nitrogen intake it appeared to have deleterious effect, resulting in a worse balance of nitroge and essential elements. Surg. Gvn. Obs.. 102, 695-70 (1956) . Burned rats receiving adequate nutrition and growt hormone did not sustain a cataboliσ response, but when th burned rats were starved, they lost weight at a greater rat than controls. Gump, F.E. et al. , Am. J. Med. Sci. , 239, 27 32 (1960) . Based on this observation, Gump et al. propose the "critical point" hypothesis which stated that "adequate calories and nitrogen are necessary for growth hormone t exert its anabolic effect. This hypothesis was reinforced b the work of Felig, P. et al.. J. Clin. Invest.. 50, 411-42 (1971) who administered growth hormone to fasting, obes volunteers and reported reduction in urea excretion which wa associated with market ketoacidosis. This acidosis resulted in increased renal ammonia generation and excretion so that net nitrogen loss was unchanged from previous control periods. Soroff, H.S. et al.. Surg. Gvn. Obs.. 111. 259-273 (I960), working with burned patients, was unable to show a beneficial effect from administration of bovine growth hormone. The same authors did show a positive effect in similar study using human growth hormone during the anabolic phase of burn recovery. Ann. Surg.. 166. 739-752 (1967) . Liljedhal, S. et al. , Acta. Char. Scand. , 122, 1-14 (1961) and Wilmore, D.W. et al.. Surg. Gyn. Obs., 138, 875-884 (1973) demonstrated significant improvement in nitrogen and potassium balance in the post-burn period, the latter specifically with high calorie and protein intake. Moreover, a mood-elevating and appetite stimulating effect was reported which, in the Liljedhal et al. study where ad libitum intake was allowed, led to greater substance intake. Roe, C.F. et al., Sur . Forum. 13, 369-371 (1962), demonstrated an alteration in substrate utilization in postoperative orthopedic patients given growth hormone, with a fall in respiratory quotient (RQ) and a shift to lipid substrates. Johnston, R. D. A. et al.. Lancet. 584-586 (March 16, 963) showed no improvement in nitrogen balance after herniorrhaphy in patients treated with growth hormone and in matched controls. However, nitrogen intakes were low, caloric provisions were not measured, and only the immediate postoperative period was studied. Administration of growth hormone to postoperative patients receiving only 5% dextrose has been described by Ward, H.C. et al. , in an abstract presented at the Annual European Society of Enteral and Parenteral Nutrition, Abstract Book No. 0.24 (Sept. 1984). The individuals studied excreted less nitrogen and had lower respiratory quotients than the placebo-matched controls. Ward et al. investigated the effect of human GH treatments (0.1 mg/kg/day) given in conjunction with hypocaloric (400 kcals as 2 liters of 5% dextrose/day) peripheral intravenous infusion. No amino acid supplement was present in the feedings. Dietary control of the nitrogen balance has been studied as well. Howard, U.. Patent No. 4,009,265, disclosed formulations for the treatment of obesity, describing the use of a low calorie (160-600 Kcal.) diet containing 15-75 grams of amino acids per day to prevent nitrogen loss. Howard, U.S. Patent No. 4,298,601, described a diet for maintaining nitrogen balance and controlling ketosis and water retention. The daily intake comprised at least 15 grams of amino acids, in the proportion required by humans, and from 15-75 grams of carbohydrates, with a total caloric value of 160-600 kcals. Dietz et al.. U.S. Patent No. 4,283,392, disclosed infusions for low-calorie parenteral nutrition containing 10-200 grams of essential and non-essential amino acids and 50-10,000 ug of kinin per liter of infusion solution. This infusion mixture was designed to replace approximately 1,000 calories per day. According to Dietz et al. , the presence of kinin provided a definite improvement in the amino acid utilization in the sense of a anabolic effect relating to the build-up of body-produced proteins. Description of the Invention There are many medical situations which require parenteral feedings. In most such situations, anabolis characterized by protein build-up in the body is essential to achieve maximum recovery in the patient. Unfortunately, the dietary requirements associated with an anabolic state (high protein, high calorie intake) demand the infusion of such large quantities of nutrient that the parenteral infusion process necessitates the use of a central venous catheter to accommodate the volume of infusate. Intravenous administration of calories, nitrogen, an other nutrients in sufficient quantities to achieve tissu synthesis and anabolis is called total parenteral nutrition. The normal calorie requirement for an adult is approximatel 2,500 per day. If these were to be provided totally b Dextrose Injection 5%, approximately 16 liters would b required. Each liter contains 50 grams of dextrose, equivalent to 170 calories. However, it is only possible t administer 3 or 4 liters per day without causing flui overload. To reduce this fluid volume, the concentration o dextrose would have to be increased. By increasing th dextrose to 25 percent, it would be possible to administe five times the calories in 1/5 the volume. Dextros Injection 25% is hypertonic, however; it cannot b administered in large amounts into a peripheral vein withou sclerosing the vein. Total parenteral nutrition is indicate in patients who are unable to ingest food due to carcinoma o extensive burns; patients who refuse to eat, as in the cas of depressed geriatrics or young patients suffering fro anorexia nervosa; and surgical patients who should not be fe orally. Remington's Pharmaceutical Sciences. 16th ed.. Mac Publishing Company, Easton, PA., pg. 1496 (1980). The use of a central venous catheter requires stric protocol and trained personnel to minimize metabolic septi complications and optimized patient benefits. Further, i many cases, even under the best of circumstances, a centra vein parenteral nutrition results in increased body fat an water retention but decreased body protein. Additionally, body protein is essential for norma function and tissue repair. Accordingly, treatment o obesity by dietary restrictions resulting in decreases i body protein, especially where dietary restrictions ar imposed for long periods of time, are undesirable. Under th present invention, the subjects would eat small quantities o energy but utilize the body fat, in conjunction with th hypocaloric dietary component and growth hormone, to support maintenance or an increase in body protein, primarily skeletal muscle. The present invention was developed as the result of a recognition of the need for parenteral solutions which would provide protein build-up while still being suitable for peripheral vein or central venous catheter introduction. Further, there has long been a need for a method for treating obese patients which would permit concomitant weight reduction and protein accretion. The invention comprises a method and composition for maintaining a positive nitrogen balance (protein accretion or build-up) under hypocaloric conditions. It comprises co- administration of growth hormone and a dietary component comprising a etabolizable source of nitrogen and carbohydrate, the total caloric content of which comprises 10 to 95% of the caloric intake determined to be required to maintain an existing nitrogen level, said dietary component and said growth hormone each administered in a amount effective to produce a positive nitrogen balance. Best Mode of Carrying Out the Invention By the term "animal" is included all members of the animal kingdom wherein growth hormone plays a role in protein metabolism; particularly, the term encompasses mammals and birds. The invention is especially suited for the treatment of humans, particularly humans requiring parenteral feeding and humans suffering from obesity. By the term "coadministering," regardless of tense, is intended the administration of the dietary component and the growth hormone in time-sequence relationship such that the period of biological activity of the growth hormone and the dietary component overlap. Thus in the case of parenteral feeding, the growth hormone may be administered along with the dietary component and drip-infused through a peripheral vein. Alternatively, the growth hormone may be injected periodically, for example, once daily by subcutaneous injections, or drip-infused as a separate component during the parenteral feeding. Where the dietary component is administered orally, the growth hormone may also be administered orally or may be administered parenterally, i.e., by subcutaneous injection. By the term "hypocaloric dietary component" is intended a dietary component comprising, as a minimum requirement, (1) 4-16 grams of a metabolizable source of nitrogen and (2) carbohydrate, the caloric content of said dietary component being between 10 and 95 percent of the RMR caloric intake. Optionally, up to 50% of the carbohydrate calorie content of the dietary component may be replaced by (3) a triglyceride source, such as fat emulsion. Additionally, the hypocaloric dietary component may contain vitamins, minerals, and/or trace elements as a particular case might indicate. By the term "a metabolizable source of nitrogen" is intended any dietary source of nitrogen from which protein anabolism in the presence of growth hormone will result. Typically, such sources of nitrogen include dietary proteins, essential amino acids, non-essential amino acids, peptides and the alpha-keto analogues of amino acids. Preferably the metabolizable source of nitrogen is one in which all of the amino acids, both the essential and non- essential ones, are present in the relative proportions required by the animal in question. The amino acid profile essential to man is well known to those skilled in the art, and, for example, is set out in Rose, W.C. et al. , J. Biol. Chem. , 217. 987 (1955) , incorporated by reference herein. convenient and inexpensive method of producing the require amino acid mixture is disclosed in U.S. Patent 4,009,265 to Howard, incorporated by reference herein. A protein hydrolysate produced by any conventional procedure, such as by subjecting a protein to hydrolysis using an acid or a proteolase as the hydrolytic agent is analyzed to determin its amino acid profile. It is then adjusted to the optimu profile, either by addition of required quantities of th pure amino acids to make up for any deficiencies and/or b passing the hydrolysate through suitable combinations o anionic and cationic exchange resins to eliminate an excesses. Further, where indicated, the amino acid profil may be altered to accommodate the increase in body protein. Typical dietary sources of natural protein include eg protein, milk, soy beans, peanuts, fish, and plasma protein The hypocaloric dietary component is produced in a manne such that its daily administration provides between 4 and 2 grams of nitrogen. A preferred range for the metabolizabl source of nitrogen is from 7-21 grams of nitrogen per day with 8-16 grams of nitrogen most preferred. By the term "carbohydrate" is intended any digestibl and/or metabolizable source of carbohydrate. Typica digestible carbohydrates include monosaccharides, for exampl glucose, glucoselactone, fructose, or lactose, disaccharide such as sucrose or maltose, or polysaccharide, for exampl edible starch or dextrin. Where the carbohydrate is to b provided through parenteral feeding, typical sources o carbohydrates include dextrose solutions, fructose solutions glycerol solutions, and sorbitol solutions. One preferre carbohydrate source is dextrose solutions. Optionally, the hypocaloric dietary component may als contain a triglyceride component, typically in the form of fat emulsion. Where present, the triglyceride componen replaces a portion, up to 50% in terms of calories, of th carbohydrate component. Typical sources for the triglycerid component include soybean or safflower emulsion and mediu chain triglycerides (MCT) , the latter commercially availabl as 8-carbon or 10-carbon triglycerides ( emington 1 s, supra at page 973) . Additionally, and optionally, the hypocaloric dietary component may contain minerals and vitamins as required. The minerals required by humans are sodium, potassium, calcium, magnesium, manganese, iron, copper, zinc, chloride, phosphorous, sulfur, iodine, and other trace elements. The central vitamins are vitamin A, vitamin B12, vitamin C, vitamin D, vitamin E, vitamin K, riboflavin, peridoxin, niacin, inositol, calcium pantothenate, biotin, folic acid, choline, and thiamin hydrochloride. The so-called minimum daily requirements for each of these minerals and vitamins are well known and may be found in any standard work on nutrition, such as, for example, Davidson, S. et al.. "Human Nutrition and Dietetics," 4th Edition, published by E. and S. Livingstone, Edinburgh, pages 244 and 255 (1969) . The hypocaloric dietary component is formulated to contain less than the calories equivalent to the RMR requirements for the individual to whom or to which it is to be administered. Typically, the hypocaloric dietary component will contain 10- 95 percent, preferably 20-90 percent, most preferably 30-80 percent of the RMR requirement based upon total calories. By the term RMR is intended that calorie equivalent considered to be the maintenance level of caloric consumption for the individual being treated. This figure may be determined from standard height and weight charts, adjusted for age, or may be determined experimentally. A typical experimental determination of energy expenditure is by indirect calorimetry using measurements of respiratory gas exchange. From these measurements energy requirements, i.e., RMR can be determined. Various methods for determining respiratory gas exchange are known, Assessment of Energy Requirements In Health and Disease , published by Ross Laboratories, Columbus, Ohio (1980) . A typical experimental determination is provided by the Beckman Metabolic Measurement Cart, described therein. By the term "growth hormone" is intended either natural or recombinant pituitary growth hormone, regardless of the source. The term is limited only in that the material must demonstrate pituitary growth hormone biological activity in the recipient. Therefore, it also applies to physiologically active equivalents, fragments, or portions of the complete growth hormone molecule. Included within the term is naturally occurring growth hormone which has been isolated from cadavers using techniques well known in the art. Typical techniques for isolation of human growth hormone are disclosed by Lewis et al. , supra, and Reisfeld et al.. supra. Isolation of growth hormone from bovine anterior pituitary is disclosed by Li et al... supra, and Wilhelmi et al.. supra. Also included is recombinant growth hormone; preparation of recombinant growth hormone is disclosed by Goeddell, D.V. et al.. Nature (London), Vol. 281, pp. 544-548 (1979). In one embodiment, the recombinant growth hormone includes an additional ethionine at the N-terminus which is not found on the natural molecule. In a different embodiment, the recombinant growth hormone may be the "mature" form, i.e., having the same N-terminus as the natural growth hormone. In one embodiment of the invention, the hypocaloric dietary component and growth hormone are combined to form a unitary composition. This composition comprises the subcomponents of the hypocaloric dietary component i.e., as a minimum, carbohydrate in a hypocaloric equivalent, and from 4-16 grams of metabolizable nitrogen, optionally further including at least one of triglycerides as a replacement for up to 50% of the carbohydrate calories, vitamins, and minerals, and an amount of growth hormone effective to provide a positive nitrogen balance under hypocaloric conditions. The specific amount of growth hormone required by each individual will be dependent upon the age, health, and weight of the recipient, kind of concurrent treatment, if any, frequency of treatment, and the nature of the effect desired. Generally, daily dosages of growth hormone will be from about 0.05 to 0.2 mg per kg of body weight. Normally, from 0.07 to 0.15 mg per kg per day, in one or more applications per day is effective to obtain the desired result. In an alternative approach, the growth hormone, particularly where formulated in a timed-release form, may be administered less frequently, i.e., every other day or every third day. The composition may be employed in dosage form such as tablets, capsules, powder packets, or liquid solutions, suspensions or elixirs, for oral .administration, or sterile liquid for formulations such as solutions or suspensions for parenteral use. In such compositions, the hypocaloric dietary component and growth hormone will be present in such amount as to provide the recipient with the prescribed per day dosage of each component. Nitrogen balance may be calculated by determining, per 24 hour period, the total nitrogen ingested, the total nitrogen excreted, and subtracting the latter from the former. Where this number is positive, i.e. "a positive nitrogen balance," protein anabolism is occurring. Where this number is negative, protein catabolis is occurring. Alternative methods for determining nitrogen balance are within the skill of the art. Where the composition is to be administered orally, the growth hormone component must be enterically coated in order to prevent gastric digestion or decomposition of the growth hormone. As is known in the art, enteric coatings do not permit release of a significant quantity of drug until the dosage form passes into the small intestine. Enteric coating compositions are well known to the art, enteric coatings do not permit release of a significant quantity of drug in the stomach but rapidly and completely release the drug when the dosage form passes into the intestine. Enteric coating compositions are well known to the art and generally may be subdivided into three groups, namely, mixtures of fats an fatty acids, shellac and shellac derivatives, and th cellulose acetate phthalates. This last group of compounds, the cellulose acetate phthalates, are preferred, but any o the enteric coatings known and in common use throughout th pharmaceutical industry are suitable for the practice of th invention. In one aspect of the invention, the hypocaloric dietar component and growth hormone are introduced by periphera vein infusion. Accordingly, the composition for peripheral vein infusion comprises the hypocaloric dietary component an growth hormone as a unitary composition. In this form, the composition comprises the hypocaloric dietary component an the growth hormone dissolved and/or suspended in a appropriate pharmaceutical carrier. Any carrier in commo use and known to the prior art is suitable for this purpose. Typical carriers include Sterile Water for Injection, Sodiu Chloride Injection, Ringers Injection, Dextrose and Sodiu Chloride Injection, Lactated Rigners Injection, and the like. In a different embodiment, the hypocaloric dietar component may be administered by peripheral vein infusion, with the growth hormone administered separately. In thi embodiment, the appropriate dosage of growth hormone may b administered parenterally, i.e., subcutaneously intravenously, intramuscularly, or intraperitoneally Typically, growth hormone is administered parenterally as 10 mg subcutaneous injection daily. The materials for use in the method of the invention ar ideally suited for the preparation of a kit. Such a kit ma comprise a carrier means being compartmentalized to receiv in close confinement one or more container means such a vials, test tubes, and the like, each of set container mean comprising one of the separate elements to be used in th method. For example, one or more of said container means ma comprise the hypocaloric dietary component, compounded fo oral or parenteral administration. A further container means may comprise a daily dose of growth hormone, with this daily dose of growth hormone formulated for oral consumption (i.e., enterically coated) or for parenteral administration. Having generally described the invention, the same will be further illustrated by means of specific examples which are presented herewith for purposes of illustration only, and are not intended to be limiting thereof, unless otherwise specified. EXAMPLES EXAMPLE I Two normal male volunteers were studied for two seven- day periods separated by two-to-three week intervals. They weighed 76 kg and 78 kg, respectively. During each admission, they received parenteral nutrition by peripheral vein infusion containing 165 grams of dextrose, 75 grams of amino acid (12.1 grams N) and 30 grams of triglyceride (given as 10% Intralipid) daily. This mixture provided 1,133 kσal/day. In one 6-day period the subjects received growth hormone (10 mg daily by subcutaneous injection) , and during the other period they received saline injections. Every morning respiratory gases were measured, all urine and stool were collected and analyzed for nitrogen, and blood was drawn daily. At the end of the study period, an oral glucose tolerance test was performed, combined with serial forearm flux studies and respiratory gas exchange measurements. Data collected as a result of this experiment appears in Table I below. TABLE I Control Growth Hormone Mean N-bal, g/day -1.8 +2.2 6-day cumulative N-bal, g -10.7 +13.2 6:00 a.m. metabolic rate, k :ccaall//ddaayy 1355 1260 Plasma glucose, mg/dl 94 103 Blood urea nitrogen, mg/dl 16 10 Insulin, U/ml 17 34 Growth hormone, U/ml 1.1 18 The difference in protein accretion between the control and growth hormone period was approximately 150 grams in 6 days. This occurred while the subjects were negative caloric balance and utilizing body fat as fuel. Glucose tolerance tests demonstrated slight glucose resistance associated with marked hyperinsulinemia during the growth hormone period. A marked protein anabolic effect of growth hormone was observed while the subjects received hypocaloric intravenous feeding. EXAMPLE 2 In this example, the subjects were studied for at least two separate periods, each of seven day's duration. The studies were paired with caloric level the same for the two study periods. The subjects received growth hormone or placebo during each of the studies. There were at least two week's interval between studies. The treatments were 1. Caloric feeding, with and without growth hormone. 2. Hypocaloric feeding, with and without growth hormone. 3. Fat-free hypocaloric feeding, with and without growth hormone. The order of growth hormone administration was determined by sequential randomization. The subjects were admitted to the Clinical research Center in the afternoon of the first day (day 0) and further familiarized with the study procedures. A normal evening meal was allowed, but no further oral intake permitted thereafter, except water, which was freely available. Adequate calorie intake consisted of calories equivalent to RMR (obtained from standard height and weight tables) plus 25%, and six gm nitrogen/m 2 . Half of the non-protein calories were administered as carbohydrate and half as fat emulsion. Hypocaloric intake consisted of 50% of the calorie level as calculated above, equally divided between fat emulsion and carbohydrate, with the same nitrogen intake. Fat-free hypocaloric intake excluded the fat emulsion from the hypocaloric intake; thus only 25% of the energy needs were provided. Where the study involved adequate calorie-parenteral nutrition, a central line was placed under fluoroscopic visualization via the anticubital route using strict aseptic conditions. During the hypocaloric studies, a peripheral venous line was utilized. Normal saline was passed through these lines until the first bag of nutrient solution was hung at 6:00 a.m. the following morning (day 1). The parenteral nutrition was started at 6:00 a.m. on day 1 and continued through six 24 hour periods. Peripheral lines were changed every 48 hours and administration set for the central lines at the same frequency. Blood pressure was monitored daily and pulse and temperature 4-hourly. The catheter entry sites were regularly inspected and dressings changed every two days. Parenteral solutions were provided, the same volume for both intake levels (approximately 3 liters) . The subjects were allowed minimal activity through the day, including reading, watching television, and smoking, if this was their usual habit. During growth hormone treatment periods, at 8:30 each morning, lO g of growth hormone, manufactured by a recombinant DNA methodology, was injected subcutaneously. During control periods, a similar volume of saline was injected. The subjects were blinded from the substance injected. 1. Blood Chemistry Each morning, immediately before the injection of growth hormone or saline, blood was drawn for serum determinations. The following determinations were made: Every day: Potassium, sodium, free fatty acids, ketones, blood urea nitrogen, glucose. Days 1, 4, 7: Total protein and fractions, transaminases, insulin, growth hormone, amino acid profile, glycerol, and phosphate. Day 4: Magnesium, complete blood count and differential. Serum growth hormone levels were assayed four hours after the growth hormone injection on days 4 and 7. 2. Substrate Balance The metabolic day began and ended at 6:00 a.m. Urine and stool were collected and each pooled for each 24-hour period. The following measurements were conducted: Urine measurements: potassium, total nitrogen, urea, creatinine, ammonia, ketone bodies, and C-peptide. Stool measurements: potassium, total nitrogen. 3. Metabolic Rate Each subject's day started at 7:30 a.m. when he was awakened, weighed, and oxygen consumption and carbon dioxide production measured with the Beckman Metabolic Measurement Cart. Respiratory quotient (RQ) ad RMR were calculated. The subject's exhaled respiratory gases were analyzed by breathing through a mouthpiece and tubing with a noseclip in place. 4. Nitrogen Flux Nitrogen turnover was measured using N 15 -labeled glycine. A dose of N 15 glycine of 0.5 mg/kg/24 hours was administered during the last three 24-hour periods of each study, administered every three hours in eight equal doses. Three-hourly urine samples were collected during these periods, with aliquots from the last day analyzed by mass spectroscopy for N 15 enrichment of urinary urea nitrogen, and turnover calculated. 5. Forearm Substrate Flux On the morning of the final day (day 7) the parenteral nutrition was discontinued at 6:00 a.m. The allotted bloods were drawn and a final dose of growth hormone administered. At 11:00 a.m. a retrograde anticubital venous catheter was placed and a further catheter positioned in a vein on the back of the contralateral hand. This hand was warmed on a pad and arterialized-venous blood drawn from it. The basal forearm blood flow was measured using venous occlusion plethysmography (initiating two cuts on the arm; one at the wrist and at the upper arm) . Basal levels of venous and arterialized-venous glucose, glycerol, free fatty acids, amino acids, and carbon dioxide were then drawn, and using the blood flow, fluxes calculated. Sixty minutes later, the metabolic chart was used to measure basal levels of oxygen consumption and carbon dioxide production. A further thirty minutes later, a 100-gram oral does of glucose was provided and the metabolic response to this load assessed by measurement of forearm fluxes (as described above, from blood flow and venous-arterialized- venous differences), and respiratory gas exchanged at 30, 60, 120, and 180 minutes. The results appear in Table II below. TABLE H THE EFFECTS OF HUMAN GROWTl . J_W _EN BAl LANCE DURING INI AVΪNCUS FEEDINGS fMEAN OF DAYS 2-6 OF EACH STUDY PERIOD. BODY CALORIES NITROGEN NΓIKG-_N CALORIES NITROGEN Nl'I WEIGHT in in BALANCE in in BAL SUBJECT (kσ. (kcal/d. (g/d. . fq/d) (kcal/d. fg/d. f (Adequate Calories Provided) 01 76.3 2262 11.6 +0.5 2262 11.6 +2 02 78.2 2300 12.2 -0.3 2300 12.2 + (Hypocaloric feeding, 50% of caloric req rements provided) 01 76.3 1131 11.6 -2.5 1131 11.6 +3 02 78.2 1150 12.6 -1.8 1150 12.2 +2 03 71.0 1020 10.9 -3.5 1020 10.7 - 04 63.4 985 11.0 -2.8 985 10.8 +2 (Fat-free hypocaloric feeding, 30%-40% of caloric requirements provided) 02 78.2 810 12.2 -4.4 810 12.2 +2 04 63.4 697 11.1 -2.8 697 10.6 + *Given as methioπyl-HGH, 10 mg subcutaneously/day x 7 days, in all studies excep receiving 50% of caloric intake. These individuals received human GH at the same dos As may be seen from Table II, administration of growth hormone results in a positive nitrogen balance, indicating protein accretion, under hypocaloric conditions, even where only 30-40% of the adequate calories are supplied. EXAMPLE 3 In this example, seven patients with various medical conditions (see Table III) were studied during two separate periods, each of seven days duration, except for one patient (No. 3) who was tested twice using this protocol. The patients received either growth hormone (10 mg, injected subcutaneously) or placebo in addition to 1.3 to 1.7 g protein/kg and a caloric intake equal to 60-100% of RMR. The order of growth hormone-placebo administration was determined by sequential randomization. Nutrition was administered by peripheral vein infusion, except in one case, where the patient received total oral nutrition (see Table IV) . All urine and stool were collected and pooled for each 24 hour period. These samples were analyzed for nitrogen, potassium and phosphorous. The balance of these elements are reported in Table V. Blood was drawn on days 1, 4, and 7 of each week and analyzed for blood urea nitrogen (BUN) , glucose and insulin levels. These data are reported in Table VI. As can be seen in Table V, six of the seven patients receiving growth hormone experienced a positive nitrogen balance that was larger than that observed during the control period. Five of seven patients experienced a positive potassium balance larger than observed during the control period. In addition, all six of the patients tested for phosphorous experienced a positive phosphorous balance that was larger than during the control period. Taken all together, these data indicate protein accretion for patients receiving growth hormone, even when receiving a hypocaloric diet. All seven patients showed a lower BUN when administered growth hormone compared to the control period. Blood glucose tended to be higher in patients receiving growth hormone (4 of 7) compared to the control period. In addition, insulin levels were significantly higher for all patients, compared to the control period. TABLE Ill PATIENT DATA PATIENT AGE SEX ENTRY IDEAL DIAGNOSIS WEIGHT WEIGHT (kg) (kg) 1 46 F 81.5 68 Gastric fistula 2 35 M 63.6 69 S. Bowel fistula Chrom's disease 3a 78 F 67.9 61 S. Bowel fistula postoperative 3b 78 68.7 61 S. Bowel fistula postoperative 19 M 66.2 70 75% Total body surface burns 5 78 M 64.2 75 Cardiac cachexia 6 23 F 62.0 65 Short bowl syndrome 7 53 M 61.0 67 S. Bowel fistula intestinal obstruc¬ tion TABLE IV NUTRIENT INTAKE GROWTH PATIENT CALORIES NITROGEN ROUTE LEVEL OF HORMONE (Kcal/d) (g/α.) FEEDING THERAPY 1 1100 16 IV hypocaloric 1st 2 1100 17 IV hypocaloric 2nd 3a 1100 14 IV hypocaloric 2nd 3b 1900 14 IV eucaloric 1st 4 3000 26 IV eucaloric 2nd 5 2400 11.5 oral eucaloric 1st 6 1100 15 IV hypocaloric 1st 7 1100 12 IV hypocaloric 2nd TABLE V NITROGEN, POTASSIUM, PHOSPHOROUS BALANCE DATA NHROGEN (g d) POTASSIUM (mEq/d) JH-SFKROUS (mg/d Patient Control GH Control Control GΉ. Control Control (_H Con 1 - +5.4 +5.1 - +19 +23 - +150 +9 2 +1.4 +4.4 - +49 +44 - -78 +105 - 3a +2.5 +4.3 - +22 +56 - -88 +214 - 3b - +6.3 +4.9 - +48- +32 - +351 +3 4 -0.9 +5.9 - +7 +36 - -38 +291 - 5 - +4.7 +7.3 - -5 -25 - - - 6 - +3.8 +0.5 - +45 +25 - +382 +13 7 -1.1 +2.5 - -3.7 +23 - -146 +127 _ TABLE VI BDXD CHEMICAL VALUES BUN (mg dl) GiprπsE (mg/dl) INSULIN (ucJJ/ml) Patient Osntrol __£ Control Control G.H Control Control G.H Cont 1 - 21 26 - 104 91 110 2 2 12 10 - 124 106 - 41 63 - 3a 19 16 - 105 118 - 7 16 - 3b - 16 19 - 141 107 - 36 4 20 13 - 99 125 - 5 22 - 5 - 16 29 - 113 118 - 55 1 6 - 11 13 - 120 116 - 35 2 7 16 11 >. Ill 114 _ 16 20 Having now fully described this invention, it will be apparent to one of ordinary skill in the art that the same may be carried out with minor modifications which do not affect the content or spirit thereof.

Claims: CLAIMS 1. A method for effecting protein accretion in an animal comprising administering to said animal: (a) a hypocaloric dietary component comprising (i) a metabolizable source of nitrogen and (ii) carbohydrate; and (b) growth hormone; said hypocaloric dietary component (a) and growth hormone (b) each administered in an amount effective to produce in said animal a positive nitrogen balance. 2. The method of claim 1 wherein said hypocaloric dietary component further comprises at least one of a triglyceride, vitamins and minerals. 3. The method of claim 1 wherein the metabolizabl source of nitrogen is selected from the group consisting o dietary proteins, essential and non-essential amino acids, peptides, and the alpha-keto analogues of amino acids. 4. The method of claim 1 wherein the hypocalori dietary component provides 10-95% of the RMR calorie intake. 5. The method of claim 4 wherein the hypocalori dietary component provides 30-90% of the RMR calorie intake. 6. The method of 1 wherein the growth hormone i selected from the group consisting of natural and recombinan growth hormone, and the biologically active fraction thereof. 7. The method of claim 6 wherein said growth hormon is human growth hormone. 8. The method of claim 6 wherein said growth hormone is administered in an amount of 0.05 to 0.2 mg per kg of body weight per day. 9. The method of claim 1 wherein said hypocaloric dietary component and said growth hormone are administered as a unitary composition. 10. The method of claim 1 wherein said growth hormone is administered separately. 11. The method of claim 1 wherein said hypocaloric dietary component and said growth hormone are administered parenterally or orally. 12. The method of claim 11 wherein said hypocaloric dietary component and said growth hormone are administered parenterally. 13. The method of claim 11 wherein said hypocaloric dietary component and said growth hormone are administered orally. 14. The method of claim 12 wherein said hypocaloric dietary component and said growth hormone are administered by means of peripheral venous infusion. 15. The method of claim 1 wherein said animal is a human. 16. A composition comprising (a) a hypocaloric dietary component comprising (i) a metabolizable source of nitrogen, and (ii) carbohydrate; and (b) growth hormone; said hypocaloric dietary component (a) and growth hormone (b) each contained in an amount effective to produce a positive nitrogen balance in an animal consuming same. 17. The composition of claim 16 wherein said hypocaloric dietary component further comprises at least one of a triglyceride, vitamins, and minerals. 18. The composition of claim 16 wherein said metabolizable source of nitrogen is selected from the group consisting of dietary proteins, essential and non-essential amino acids, peptides, and the alpha-keto analogues of amino acids. 19. The composition of claim 16 wherein said hypocaloric dietary component provides 10-95% of the RMR calorie intake for an animal consuming same. 20. The composition of claim 19 wherein said hypocaloric dietary component provides 30-60% of the RM calorie intake. 21. The composition of claim 16 wherein said growt hormone is selected from the group consisting of natural growth hormone and recombinant growth hormone, and the biologically active fraction thereof. 22. The composition of claim 21 wherein said growt hormone is human growth hormone. 23. The composition of claim 21 wherein said growt hormone is contained in an amount sufficient to provide fro 0.05 to 0.2 mg per kg of body weight per day for an animal receiving same. 24. The composition of claim 16 as a parenterally injectable solution. 25. The composition of claim 16, in liquid or solid form, for oral consumption. 26. The composition of claim 16 wherein said growth hormone comprises an enteric coating. 27. A kit comprising a carrier means, said carrier means being compartmentalized to receive, in close confinement, a first container means comprising a hypocaloric dietary component and a second container means comprising a growth hormone. 28. A method of parenteral feeding of an animal comprising administering through a peripheral vein a protein accreting composition comprising (a) A hypocaloric dietary component comprising (i) a metabolizable source of nitrogen, and (ii) carbohydrate; and (b) growth hormone; said hypocaloric dietary component (a) and growth hormone (b) each administered in an amount effective to produce in said animal a positive nitrogen balance. 29. The method of Claim 28 wherein said hypocaloric dietary component further comprises at least one of a triglyceride, vitamins, and minerals. 30. The method of Claim 28 wherein the metabolizable source of nitrogen is selected from the group consisting of dietary proteins, essential and non-essential amino acids, peptides, and the alpha-keto analogues of amino acids. -31- 31. The method of Claim 28 wherein the hypocaloric dietary component provides 25-95% of the RMR calorie intake. 32. The method of Claim 31 wherein the hypocaloric dietary component provides 30-60% of the RMR calorie intake. 33. The method of Claim 28 wherein the growth hormone is selected from the group consisting of natural and recombinant growth hormone, and the biologically active fraction thereof. 34. The method of Claim 33 wherein said growth hormone is human growth hormone. 35. The method of Claim 33 wherein said growth hormone is administered in an amount of 0.05 to 0.2 mg per kg of body weight per day. 36. The method of Claim 28 wherein said animal is a human.

Inventors: WILMORE DOUGLAS W, WILMORE, DOUGLAS, W.
Applicants: BRIGHAM & WOMENS HOSPITAL, BRIGHAM AND WOMEN'S HOSPITAL
Application year: 1987
Main classification: A61K37
IPC codes: A61K31, A61K38, A61P3, A61P15