US3577540A

US3577540A - Method of promoting osteogenesis

Info

Publication number: US3577540A
Application number: US775162A
Authority: US
Inventors: Dirck V Myers
Original assignee: ER Squibb and Sons LLC
Current assignee: ER Squibb and Sons LLC
Priority date: 1968-11-12
Filing date: 1968-11-12
Publication date: 1971-05-04
Anticipated expiration: 1988-05-04

Abstract

A METHOD OF PROMOTING OSTEOGENESIS IN ANIMAL HOSTS WHICH COMPRISES APPLYING TO THE AREA OF A BONE DEFECT A QUARTERNARY AMMONIUM COMPOUND OF THE FORMULA

(R-)2-N(+)(-(CH2)N-)-((CH2)N)-N(+)(-R)2 (X(-))2

WHEREIN R REPRESENTS LOWER ALKYL, X REPRESENTS CHLORINE OR BROMINE, AND N IS AN INTEGER FROM SIX TO TWELVE; OR OF THE FORMULA;

R1-N(+)(-R2)(-R3)-R4 X(-)

WHEREIN R4 REPRESENTS AN ALKYL GROUP CONTAINING TWELVE TO TWENTY CARBON ATOMS; R1, R2 AND R3 EACH REPRESENT LOWER ALKYL, AND R1, R2 AND R3 TAKEN TOGETHER WITH THE NITROGEN ATOM REPRESENT PYRIDIUM; AND X REPRESENTS CHLORINE OR BROMINE.

Description

United States Patent 3,577,540 METHOD OF PROMOTING OSTEOGENESIS Dirck V. Myers, Atlanta, Ga., assignor to E. R. Squibb & Sons, Inc., New York, NY. No Drawing. Filed Nov. 12, 1968, Ser. No. 775,162 Int. Cl. A61k 27/00 US. Cl. 424-244 8 Claims ABSTRACT OF THE DISCLOSURE A method of promoting osteogenesis in animal hosts which comprises applying to the area of a bone defect a quaternary ammonium compound of the formula wherein R represents lower alkyl, X represents chlorine or bromine, and n is an integer from six to twelve; or of the formula 31 R 1 IR X- wherein R represents an alkyl group containing twelve to twenty carbon atoms; R R and R each represent lower alkyl, and R R and R taken together with the nitrogen atom represent pyridinium; and X represents chlorine or bromine.

the formula R (CH2)n R I wherein R represents lower alkyl, X represents chlorine or bromine, and n is an integer from six to twelve; or

of the formula R Ir wherein R represents an alkyl group containing twelve to twenty carbon atoms; R R and R each represent lower alkyl, and R R and R taken together with the nitrogen atom represent pyridinium; and X represents chlorine or bromine.

The term lower alkyl as employed herein includes both straight and branched chain radicals of less than eight carbon atoms. Preferred compounds for use in accordance with this invention comprise compounds of Formula I in which all four R groups represent methyl, and compounds in accordance With Formula 11 wherein R represents a straight chain alkyl group.

Compounds of Formula I may be prepared by a twostep process. In the first step, a polymethylene dichloride or dibromide containing six to twelve carbon atoms is reacted with a di(lower alkyl)amine in the presence of a solvent such as absolute alcohol to produce a corresponding polymethylene-bis-di(lower alkyl) amine. This is then reacted in the second step of the process with an equimolar amount of the polymethylene dihalide to produce the quaternary ammonium compounds of this invention.

For use, the quaternary ammonium compounds of this invention are preferably suspended in a sterile physiological saline, e.g., about 2.0 to about 60 mg./ml. This is administered by injection into the area immediately adjacent or surrounding the site in the animal host (e.g., rats, dogs, cats, cattle, horses), at which it is desired to promote or stimulate bone growth, for example, at the site of bone implants, fractures, or in various metabolic bone defects (osteoporosis), or application to the periodontal tissue. In general, about 1 to 30 mg., preferably about 2 to 5 mg. of octeogenic agent per kg. of body weight is injected. This may be repeated one or more times as required. Alternatively, in the case of bone implants, a solution (for instance, 1 mg./ml.) can be made in sterile water and implant material can be saturated therewith as by immersion before being used in the patient.

The following examples illustrate the invention, all temperatures being in degrees centigrade.

EXAMPLE 1 (a) Bis-1,10-dimethylaminodecane Into a pint-sized pressure bottle were placed 30.0 g. of 1,10-decarnethylene dibromide, 18.0 g. dimethylamine, and 150 ml. absolute alcohol. The bottle was sealed and allowed to stand for five days. It was heated intermittently to 50. The contents were poured into a 2.1. Erlenmeyer flask and diluted with 750 ml. anhydrous ether. A crystalline material was obtained, dimethylamine hydrobromide. It was removed by filtration and the filtrate was concentrated. The residue was distilled to give 17.5 g. of product, B.P. 143-146 (8 mm.)

(b) 1,1,12,12-tetramethyl-1,12-diazacyclodocosane dibromide Into a pressure bottle were placed 16.9 g. of the product of Example 1(a), 24.3 g. decamethylene dibromide and 150 ml. acetone. The mixed materials were allowed to stand twelve days. The product was filtered and recrystallized from 200 ml. absolute alcohol and 1200 ml. acetone to give 27.9 g. of product, M.P. 193-194". The product is quite hygroscopic.

EXAMPLE 2 1,1,8,8-tetramethyl-1,8-diazacyclotetradecane dibromide Following the procedure of Example 1(b) but substituting 17 g. of bis-1,6-dimethylaminohexane for the bis-1,10- dimethylaminodecane and 24.5 g. of hexamethylene dibromide for the decamethylene dibromide, there is obtained 27 g. of product, M.P. 2525-2535".

EXAMPLE 3 Sr assay of osteogenic agents The osteogenic agents of this invention are assayed by suspending a 60 mg. sample in 1 ml. of physiological saline and the pH is adjusted within the range of 6 to 7.

The ostegoenic agent thus suspended is injected (0.1 ml., 6 mg. dose) along the periosteal surface of the radiusulna complex of male rats weighing 180200 g. The contrallateral radius-ulna complex serves as the control. The rats are maintained for one week at which time 2 micro curies of radioactive Sr preferably as Sr nitrate, is injected intrathoracically. On the following day the animals are sacrificed and the Sr content of the treated and control radius-ulna complex is measured. The assay depends upon the enhanced ability of newly formed bone (matured osteoid) to bind Sr in addition to that bound by the pre-existing bone. The extent of bone growth induced within the week by osteogenic material is determined by calculating the difierence in uptake of Sr as calculated by the following expression:

A value of 100 corresponds to no excess incorporation of Sr in the treated complex as compared with the control. Values above 100 give a measure of the degree of osteogenic potency of the materials (per mg. of dose) injected.

New bone production is always accompanied by a soft tissue reaction. This reaction varies with the material used to induce osteogenesis and may be separated histologically into an acceptable and an unacceptable category.

An acceptable tissue reaction is described as follows: Eight days following the injection of the osteogenic factor preparation there exists shrinkage and lysis of some muscle bundles. There is an influx of large numbers of fibroblasts associated with considerable new blood vessel formation. These fibroblasts (precursor cells) are non-differentiated; however, some do differentiate into osteoblasts in the area of new bone formation. A few giant cells are usually present in these areas and no indication of abscess formation exists. The new bone formation is quite extensive and occurs radially around the radius and ulna. The new bone is not compact but appears to be cancellous in nature with the formation of many discrete bone marrow cavities. This is the type of histological response which accompanies normal bone formation (e.g., repair of fractures).

Abnormal undesirable tissue reactions are described as follows: Eight days following the injection of certain sub stances there is shrinkage, lysis and necrosis of many muscle bundles. Abscess formation exists accompanied by massive tissue destruction. Fibroplasia does occur; however, it is not extensive. New bone formation occurs radially around the radius and the ulna. Resorption of the original shaft of the radius and ulna exists, often accompanied by rechanneling of the marrow cavity.

The Sr ratio and histological response observed in rat legs injected with compounds of this invention are tabulated below. In all cases, the histological response is acceptable, the new bone formation being cancellous in nature with the formation of many discrete bone marrow cavities. No resorption of the original bone shaft has taken place.

wherein R represents lower alkyl, X is selected from the group consisting of chlorine and bromine, and n is an integer from six to twelve, and compounds of the formula a- R wherein R represents an alkyl group containing twelve to twenty carbon atoms; R R and R are each lower alkyl; R R and R taken together with a nitrogen atom is pyridinium; and X is selected from the group consisting of chlorine and bromine.

2. A method in accordance with claim 1 wherein the compound administered has the formula R\ )CHQH R H N N R/ (C2)n R wherein R, X and n are as set forth in claim 1.

3. A method in accordance with claim 1 wherein the compound administered has the formula R1 R2 Is -N R3 wherein R R R and R and X are as set forth in claim 1.

4. A method in accordance with claim 1 wherein the quaternary ammonium compound is administered in an amount of from about 1 to about 30 mg. per kg. of body weight.

5. A process in accordance with claim 2 wherein the compound administered is 1,1,12,12 tetramethy1-l,12- diazacyclodocosane dibromide.

6. A method in accordance with claim 2 wherein the compound administered is 1,1,8,8-tetramethyl-l,8-diazacyclotetradecane dibromide.

7. A method in accordance with claim 3 wherein the compound administered is hexadecyltrimethylammonium bromide.

8. A method in accordance with claim 3 wherein the compound administered is l-hexadecylpyridinium chloride.

Dose New bone Undifieren- Giant Abcess Compound (mg/kg.) (Sr ratio) Fibroplasia tiated cells cells Necrosis formation 0E3 )CEzho/CHQ N N -2 Br- 2.0 267 Extensive." Yes Yes None None. CH3 (OHz)1o CH3 CH3 (OHM CH3 -2 Br- 2.3 Do. CH3 (CHz)o CH3 CH:4(CHz) 5-N -Cl- 3.7 D0.

CHs( 2)1s s)a-B 2- D0.

Same as above. 2. 5 D0. D0 1.7 D0. D0 0.8 Do. Do 0.4 Do.

References Cited UNITED STATES PATENTS 2,899,357 8/1959 Cavallito et al. 424-329 2,977,280 3/1961 Forsyth et al 424329 3,034,965 5/1962 Drake et a1. 424329 3,227,614 1/1966 Scheuer 424-329 STANLEY J. FRIEDMAN, Primary Examiner D. M. STEPHENS, Assistant Examiner U.S. Cl. X.R. 424-329