CA1206415A - Pyridine soluble extract of a microorganism - Google Patents
Pyridine soluble extract of a microorganismInfo
- Publication number
- CA1206415A CA1206415A CA000430665A CA430665A CA1206415A CA 1206415 A CA1206415 A CA 1206415A CA 000430665 A CA000430665 A CA 000430665A CA 430665 A CA430665 A CA 430665A CA 1206415 A CA1206415 A CA 1206415A
- Authority
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- Canada
- Prior art keywords
- composition
- amount
- weight
- extract
- pyridine
- Prior art date
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-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicinal Chemistry (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Molecular Biology (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Epidemiology (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
ABSTRACT OF THE DISCLOSURE
A pharmaceutical composition is disclosed comprising a purified pyridine-soluble extract obtained from a microorganism which contains between about 7 and 20% by weight of protein, be-tween about 10 and 16% by weight of sugar, and between about 35 and 55% by weight of fatty acids which when combined with trehalose dimycolate and an acetone precipitated by-product of endotoxic glycol lipids extracted with chloroform-methanol in a pharmaceuti-cally acceptable medium is useful as an anti-tumor agent in the treatment of animals and humans.
A pharmaceutical composition is disclosed comprising a purified pyridine-soluble extract obtained from a microorganism which contains between about 7 and 20% by weight of protein, be-tween about 10 and 16% by weight of sugar, and between about 35 and 55% by weight of fatty acids which when combined with trehalose dimycolate and an acetone precipitated by-product of endotoxic glycol lipids extracted with chloroform-methanol in a pharmaceuti-cally acceptable medium is useful as an anti-tumor agent in the treatment of animals and humans.
Description
~2~6~S
U-Wp-2661-(3) RIBI
~ACKGROUND OF THE INVENTION
. .
The present invention is directed to a pyridine-soluble extract of a microorgani~m which, when combined with trehalose dimycola~e (TDM) and an acetone precipitated by-product of endotoxic glycolipid~ extracted with chloroform-methanol~ACP), provldes a pharmaceutical composition posse~sing anti-tumor propertie Bacteria such as Corynebacterium parvum have been the subject of experiment~l work to isolate and characteri2e the component responsible for inducing inhibition of tumor growth lsee, for example, Anti Tumor Activity and Lymphoreticular Stimulation Properties of Fractions Isolated from C. parvum;
Cantrell~ et al, Cancer Research 39, pgs. 3554-3563 (September~
1979)]. Apart from anti-tumor activity, C _E~yg~ has shown to be a potent ~imulator of the lymphoreticular sy~tem`resulting in undesirable increases in ~pleen a~d liver weights and blastogenesi Applicant has discovered that a pyridine-soluble extract o~ a microorganism possesses potent anti-tumor proper-tie~ without the undesirahle toxic effects as~ociated with the prior art pro~uct~
~ rehalo~e dimy~olates ~TDM), may be obtained ~rom organ-i~ms suah a~, ~or example, M~aviuml M phlei, M.tuberculosis ~Strain H 37 ~V and.~yoma B), M.~ovis BCG, ~ ~m~gm~3~, M.kansa~ii, Nocardia rubra, M.bovinis and Corynebacterium i~htheriae , Bacteria such as M.avium are grown, harvested and then s heat killed. The cell mass is then extracted with several solvents and then an active, solvent soluble, fraction is e~tracted. This extract is further puriEied by a series of solvent extractions to provide crude TDM ~See BioloqicallV
Active Components from Mycobacterial Cell Walls. I. Isolation and Composition of Cell Wall Skeleton and Component P3;
Azuma, et al, Journal of the National Cancer Institute, Volume 52, pgs. 95-101, 1974). ~s disclosed in Azuma et al, crude TDM may then be further purified by centrifugal microparticulate silica gel chromatography to give purified TDM.
An acetone precipitated by-product of endotoxic glycolipids extracted with chloroform-methanol (ACP) does not possess tumor regressive properties when used alone or in combination with trehalose dimycolate. For a more complete discussion of ACP, its properties and methods of produc~ion, reference is made to Peptides as Requirement for Immuno-th _~eY _ ~ne C~inea-Pig Line-10 Tumor with Endotoxins;
_ Ribi, e~ al, Cancer Immunol. Immunother., Volume 7, pg~.
~3-58 (l979). ~CP can be prepared from an Enterobacteriaciae includinq, but not limited to, the following genera:
Salmonella, Shiqella, Escherichia, Brucella, Bordetella, -Citrobacter, Pseudomonas, Pa~turella, ~eisseria, Proteus, _ ~lebsiella, and Serratia.
~_ ___ qlhe ~ollowing ~pecles are typically employ~d:
ml~ne~o~a, ~ um/ ~. pertussi~, B abor~u~, S.
en~erit_di~ oli, S. typhi, S. marce_cens~ S~ typhosa, Shi~e~la flexnl, and It is, therefore, an objec~ of the present inven-tlon to provide a pharmaceutical composition containing a pyridine-soluble extract of a microorqanism in combination with trehalose ~,1,,, ~Z~641S
dimycolate and an acetone-precipitated by-product of ~ACP) endo-toxic gl~colipids extracted with chloroform-methanol.
It is another object of the invention to provide a method of treating tumors in warm blooded animals and humans using the composition containing the pyridine-soluble extract of a ~icro-organism,. TDM and ACP.
The present invention relates to pharmaceutical composi-tion~ comprising a pyridine-soluble extract of a microorganism containing between about 7 and 20% by weight of protein about 10 to 16% by weight of sugar, and about 35 to 55% by weight of fatty acids, in combination with TDM and ACP. The pycidine soluble extract preferably contains about 12% by weight of each of protein a~d sugar and about 45% by weight of fatty acids.
Any microorganism may be used to obtain the pyridi~e-soluble extract including, for example~ M. bovis BCG, M. phlei, 9~ , M. kansasii,.NocardiA rubra., Cory~_bacterium diphtheriae and Corynebacterium parvum. :Corynebacterium ~arvum is especially pxeferred.~ ~
. Whole cells of the bacteria preferably i~ the form of.a paste, are mixed vith pyridine. The resulting mixture is separated to obtain a supernatant frac~ion whi~h contains the pyridine-soluble extract and a pyridine residue~ Optionally, the pyridine:residue may be subjected to repea~ed separation procedures as described above using pyridine to remove furthex quantikies of the desired extract The pyridine is then removed from the ex~ract and the dried extract is dialyzed against a suitable liguid such as dis-tilled water. The absence of whole cells or cell fragment s contaminants is confirm~d by electron microscopy. The resulting purified extract may then be lyophilized by known methods to obta~n a stable product.
The pyridine-soluble extract produced in accordance with thi~ invention ~ay be combined with TDM and ACP to produce a composition having poten~ an~i-tumor activity without stimu-lating the induction of spleen and liver enlargements, The cancers which may be treate~ by this composition include animal tumor~ such as bovine squamous cell carcinoma, bovine fibro-sarcoma, equine sarcoid, equine melanoma, equine squamous cell carcinoma, canine mammary tumors, canine adenoma and canine melanoma and human tumors such as breast tumoes, lung tumors, colon tumors, malignant melanoma, squamous cell carcinomas and ovarian tumors.
The composition is preferably administered by injection in a pharmaceutically acceptable medium such as an oil-droplet emulsion directly into the tumor under conditions more partic-ularly described below. The aforesaid composition may be stab~ ed as,~or example, by a lyophilization procedure and ~h~n reconsti~ut~d without los5 of potency.
The amount of the p~ridine-soluble extract in a single lnjection for the treatment oE animals is between~about 375 and 2500 microqrams/milliliter. The amount of each of TDM and ACP
i9 b~tween about 375 and 1250 micrograms/milliliter.
~ he number of milliliters o~ the biologic injected into ~h~ tumor i~ d~termined by the size o~ the tumor in accordance with the ~ollowlng table:
-4- ~
~.
~2~6~
Animal Dosage Accordinq to Tumor Size Diameter of Tumor (cm) Amount of Biologic Injected (ml) .. ... _ _ .
0-l up to 0.5 0.5 to 2~5 ~-3 2.5 to 5 3-5 5 to lO
5-8 lO to 15 greater ~han 8 15 to 20 The maximum dose per injection is about 40 milligrams for ~he pyridine-soluble extract~ about 6 milligrams for TDM and about 20 milligrams for ACP. The course of ~reatment comprises up to six injections administered a~ abou~ two week i~tervals.
Ths present composition in a suitable injection medium such as an oil-drople~ emulsion is administered directly into human tumors. The amount o the pyridine-soluble extract in a ~ingle iniection is between about ~00 and 5000 micrograms, pre~erably between about 800 and l200 micrograms. The amount of TPM is between about 50 and lO00 micrograms and the amount of ACP is between about 150 and lO00 micrograms. The preferred singl~ dosage level for each of TDM and ACP is between about 475 and 525 microgram~. All of the above-mentioned dosage levels b~ed ~n a typi~l 70 kilogram adul~ p~tier~ he in~ec-~i~n~ ara admin~5tered ahou~ once every week for up to a ~o~al ~P 15 inj~ction~, A~ descr1bed ahove, ~he composition for treatment o~ warm ~lQoded animals and humans may be used in ~he form oE an oil droplet emul~ion. ~he amount of oil used is in the range of between about 0.5 and 3.0 percent by volume based on the total il ~Z~6~S
volume of the composition. It is preferred -to use between about 0.75 and 1.5 percent by volume of the oil. ~xamples of such oils include light mineral oil, squalane, squalene, 7-n-hexyloctadecane, Conoco, (a trademark) superoil and Drakeol 6 VR (a trademark) mineral oil (produced by the Penreco Company, Butler, Pennsylvania).
The homogeni2ed oil containing mixture is then combined with a detergent which may optionally be dissolved in a saline solutivn prior to mixing. The amount of detergent is typically between ahout 0.02 and 0.25 percent by volume and preferably betwe~n about 0.10 and 0.20 percent by volume based on the total volume of the composition. ~ny common detergent material may be used including Tween-80 (a trademark) and Arlacel, ~a trademark) ~produced by the Atlas Chemical Company).
The mixture resulting from the addition of detergent is th~ homogenized to form a suspension which has a high percentage o~ oil droplets coated with the active components as determined by obs~rvation under a microscope.
The following examples are for il~ustrative purposes only and are not intencled to ]imit or in any way redefine the invcntiorl as claimed in the claims appended hereto.
EX~MPI,~ 1 ~ Praparation of Pyridine-Soluble Extract rom nebacterlllm Parvum Cor~nebact~rium_~axvum tP.acnes,Strain 4182) was grown nd ha~vas~d at 37C ln MI~I-khiocJlycolAte broth Eor betw~en ancl 7~h~urs ko ob~Aln a wholc cell paste. r~he past~ was -th~n wa~he~l with .500 ml~ o~ distilled water. 90 ~rams (wet wol~ht~) o~ t:h~ w~hecl paste was mixed with 200 ml. o~ neat p~r:Ldisle and c~ntriEued at 1700 x g ~or one hour at ~ C. A
pyrldin~-soluble extract was removed as a supernatant ~raction.
--The remaining residue was extracted with additional pyridine under identical conditions as described above. Following filtration, using Whatman No~ 1 paper, the pyridine extrac~s w~re pooled and the solvent was removed by evapora~ion at 50 C.
in a ~uchi Rotavapor (Brinkmann Insteuments, Westbury, New York)O The dried pyridine extract was extensively dialyzed against distilled water and then lyophilized. The resulting puriied pyridine extract contained about 12% by weight of protein, about 12% by weight of sugar and about 45% of fatty acids. The extract was examined under an electron microscope and found to be free of contaminating whole cells and cell wall ragmentsO The yield of the pyridine-soluble extract wa5 9%
(8.1 ~.).
EX~MPLE 2 - Preparakion of Pyridine-Soluble Extract from M.bovis Strain ~CG
M! bovis Strain BC~ was yrown and harvested in Sautons medium at 37~ for 3 to 4 weeks to obtain a washed whole cell paste produced in the same manner disclosed in Example ls 50 grams ~we~ weight~ of the washed paste was then treated in the same manner as Example 1 to produce a yield of the pyridine-soluble extract oi 7% (3~59). The extract contained 15% ~y weight of protein, 10~ by weight of sugar and 52~ by weight of ~atty acid~.
The pyridine soluble extract in combination with TDM and A~P in a pharmaceutically acceptable medium is significantly effective in the treatment of tumors to obtain total regression of the tumor in most cases.
U-Wp-2661-(3) RIBI
~ACKGROUND OF THE INVENTION
. .
The present invention is directed to a pyridine-soluble extract of a microorgani~m which, when combined with trehalose dimycola~e (TDM) and an acetone precipitated by-product of endotoxic glycolipid~ extracted with chloroform-methanol~ACP), provldes a pharmaceutical composition posse~sing anti-tumor propertie Bacteria such as Corynebacterium parvum have been the subject of experiment~l work to isolate and characteri2e the component responsible for inducing inhibition of tumor growth lsee, for example, Anti Tumor Activity and Lymphoreticular Stimulation Properties of Fractions Isolated from C. parvum;
Cantrell~ et al, Cancer Research 39, pgs. 3554-3563 (September~
1979)]. Apart from anti-tumor activity, C _E~yg~ has shown to be a potent ~imulator of the lymphoreticular sy~tem`resulting in undesirable increases in ~pleen a~d liver weights and blastogenesi Applicant has discovered that a pyridine-soluble extract o~ a microorganism possesses potent anti-tumor proper-tie~ without the undesirahle toxic effects as~ociated with the prior art pro~uct~
~ rehalo~e dimy~olates ~TDM), may be obtained ~rom organ-i~ms suah a~, ~or example, M~aviuml M phlei, M.tuberculosis ~Strain H 37 ~V and.~yoma B), M.~ovis BCG, ~ ~m~gm~3~, M.kansa~ii, Nocardia rubra, M.bovinis and Corynebacterium i~htheriae , Bacteria such as M.avium are grown, harvested and then s heat killed. The cell mass is then extracted with several solvents and then an active, solvent soluble, fraction is e~tracted. This extract is further puriEied by a series of solvent extractions to provide crude TDM ~See BioloqicallV
Active Components from Mycobacterial Cell Walls. I. Isolation and Composition of Cell Wall Skeleton and Component P3;
Azuma, et al, Journal of the National Cancer Institute, Volume 52, pgs. 95-101, 1974). ~s disclosed in Azuma et al, crude TDM may then be further purified by centrifugal microparticulate silica gel chromatography to give purified TDM.
An acetone precipitated by-product of endotoxic glycolipids extracted with chloroform-methanol (ACP) does not possess tumor regressive properties when used alone or in combination with trehalose dimycolate. For a more complete discussion of ACP, its properties and methods of produc~ion, reference is made to Peptides as Requirement for Immuno-th _~eY _ ~ne C~inea-Pig Line-10 Tumor with Endotoxins;
_ Ribi, e~ al, Cancer Immunol. Immunother., Volume 7, pg~.
~3-58 (l979). ~CP can be prepared from an Enterobacteriaciae includinq, but not limited to, the following genera:
Salmonella, Shiqella, Escherichia, Brucella, Bordetella, -Citrobacter, Pseudomonas, Pa~turella, ~eisseria, Proteus, _ ~lebsiella, and Serratia.
~_ ___ qlhe ~ollowing ~pecles are typically employ~d:
ml~ne~o~a, ~ um/ ~. pertussi~, B abor~u~, S.
en~erit_di~ oli, S. typhi, S. marce_cens~ S~ typhosa, Shi~e~la flexnl, and It is, therefore, an objec~ of the present inven-tlon to provide a pharmaceutical composition containing a pyridine-soluble extract of a microorqanism in combination with trehalose ~,1,,, ~Z~641S
dimycolate and an acetone-precipitated by-product of ~ACP) endo-toxic gl~colipids extracted with chloroform-methanol.
It is another object of the invention to provide a method of treating tumors in warm blooded animals and humans using the composition containing the pyridine-soluble extract of a ~icro-organism,. TDM and ACP.
The present invention relates to pharmaceutical composi-tion~ comprising a pyridine-soluble extract of a microorganism containing between about 7 and 20% by weight of protein about 10 to 16% by weight of sugar, and about 35 to 55% by weight of fatty acids, in combination with TDM and ACP. The pycidine soluble extract preferably contains about 12% by weight of each of protein a~d sugar and about 45% by weight of fatty acids.
Any microorganism may be used to obtain the pyridi~e-soluble extract including, for example~ M. bovis BCG, M. phlei, 9~ , M. kansasii,.NocardiA rubra., Cory~_bacterium diphtheriae and Corynebacterium parvum. :Corynebacterium ~arvum is especially pxeferred.~ ~
. Whole cells of the bacteria preferably i~ the form of.a paste, are mixed vith pyridine. The resulting mixture is separated to obtain a supernatant frac~ion whi~h contains the pyridine-soluble extract and a pyridine residue~ Optionally, the pyridine:residue may be subjected to repea~ed separation procedures as described above using pyridine to remove furthex quantikies of the desired extract The pyridine is then removed from the ex~ract and the dried extract is dialyzed against a suitable liguid such as dis-tilled water. The absence of whole cells or cell fragment s contaminants is confirm~d by electron microscopy. The resulting purified extract may then be lyophilized by known methods to obta~n a stable product.
The pyridine-soluble extract produced in accordance with thi~ invention ~ay be combined with TDM and ACP to produce a composition having poten~ an~i-tumor activity without stimu-lating the induction of spleen and liver enlargements, The cancers which may be treate~ by this composition include animal tumor~ such as bovine squamous cell carcinoma, bovine fibro-sarcoma, equine sarcoid, equine melanoma, equine squamous cell carcinoma, canine mammary tumors, canine adenoma and canine melanoma and human tumors such as breast tumoes, lung tumors, colon tumors, malignant melanoma, squamous cell carcinomas and ovarian tumors.
The composition is preferably administered by injection in a pharmaceutically acceptable medium such as an oil-droplet emulsion directly into the tumor under conditions more partic-ularly described below. The aforesaid composition may be stab~ ed as,~or example, by a lyophilization procedure and ~h~n reconsti~ut~d without los5 of potency.
The amount of the p~ridine-soluble extract in a single lnjection for the treatment oE animals is between~about 375 and 2500 microqrams/milliliter. The amount of each of TDM and ACP
i9 b~tween about 375 and 1250 micrograms/milliliter.
~ he number of milliliters o~ the biologic injected into ~h~ tumor i~ d~termined by the size o~ the tumor in accordance with the ~ollowlng table:
-4- ~
~.
~2~6~
Animal Dosage Accordinq to Tumor Size Diameter of Tumor (cm) Amount of Biologic Injected (ml) .. ... _ _ .
0-l up to 0.5 0.5 to 2~5 ~-3 2.5 to 5 3-5 5 to lO
5-8 lO to 15 greater ~han 8 15 to 20 The maximum dose per injection is about 40 milligrams for ~he pyridine-soluble extract~ about 6 milligrams for TDM and about 20 milligrams for ACP. The course of ~reatment comprises up to six injections administered a~ abou~ two week i~tervals.
Ths present composition in a suitable injection medium such as an oil-drople~ emulsion is administered directly into human tumors. The amount o the pyridine-soluble extract in a ~ingle iniection is between about ~00 and 5000 micrograms, pre~erably between about 800 and l200 micrograms. The amount of TPM is between about 50 and lO00 micrograms and the amount of ACP is between about 150 and lO00 micrograms. The preferred singl~ dosage level for each of TDM and ACP is between about 475 and 525 microgram~. All of the above-mentioned dosage levels b~ed ~n a typi~l 70 kilogram adul~ p~tier~ he in~ec-~i~n~ ara admin~5tered ahou~ once every week for up to a ~o~al ~P 15 inj~ction~, A~ descr1bed ahove, ~he composition for treatment o~ warm ~lQoded animals and humans may be used in ~he form oE an oil droplet emul~ion. ~he amount of oil used is in the range of between about 0.5 and 3.0 percent by volume based on the total il ~Z~6~S
volume of the composition. It is preferred -to use between about 0.75 and 1.5 percent by volume of the oil. ~xamples of such oils include light mineral oil, squalane, squalene, 7-n-hexyloctadecane, Conoco, (a trademark) superoil and Drakeol 6 VR (a trademark) mineral oil (produced by the Penreco Company, Butler, Pennsylvania).
The homogeni2ed oil containing mixture is then combined with a detergent which may optionally be dissolved in a saline solutivn prior to mixing. The amount of detergent is typically between ahout 0.02 and 0.25 percent by volume and preferably betwe~n about 0.10 and 0.20 percent by volume based on the total volume of the composition. ~ny common detergent material may be used including Tween-80 (a trademark) and Arlacel, ~a trademark) ~produced by the Atlas Chemical Company).
The mixture resulting from the addition of detergent is th~ homogenized to form a suspension which has a high percentage o~ oil droplets coated with the active components as determined by obs~rvation under a microscope.
The following examples are for il~ustrative purposes only and are not intencled to ]imit or in any way redefine the invcntiorl as claimed in the claims appended hereto.
EX~MPI,~ 1 ~ Praparation of Pyridine-Soluble Extract rom nebacterlllm Parvum Cor~nebact~rium_~axvum tP.acnes,Strain 4182) was grown nd ha~vas~d at 37C ln MI~I-khiocJlycolAte broth Eor betw~en ancl 7~h~urs ko ob~Aln a wholc cell paste. r~he past~ was -th~n wa~he~l with .500 ml~ o~ distilled water. 90 ~rams (wet wol~ht~) o~ t:h~ w~hecl paste was mixed with 200 ml. o~ neat p~r:Ldisle and c~ntriEued at 1700 x g ~or one hour at ~ C. A
pyrldin~-soluble extract was removed as a supernatant ~raction.
--The remaining residue was extracted with additional pyridine under identical conditions as described above. Following filtration, using Whatman No~ 1 paper, the pyridine extrac~s w~re pooled and the solvent was removed by evapora~ion at 50 C.
in a ~uchi Rotavapor (Brinkmann Insteuments, Westbury, New York)O The dried pyridine extract was extensively dialyzed against distilled water and then lyophilized. The resulting puriied pyridine extract contained about 12% by weight of protein, about 12% by weight of sugar and about 45% of fatty acids. The extract was examined under an electron microscope and found to be free of contaminating whole cells and cell wall ragmentsO The yield of the pyridine-soluble extract wa5 9%
(8.1 ~.).
EX~MPLE 2 - Preparakion of Pyridine-Soluble Extract from M.bovis Strain ~CG
M! bovis Strain BC~ was yrown and harvested in Sautons medium at 37~ for 3 to 4 weeks to obtain a washed whole cell paste produced in the same manner disclosed in Example ls 50 grams ~we~ weight~ of the washed paste was then treated in the same manner as Example 1 to produce a yield of the pyridine-soluble extract oi 7% (3~59). The extract contained 15% ~y weight of protein, 10~ by weight of sugar and 52~ by weight of ~atty acid~.
The pyridine soluble extract in combination with TDM and A~P in a pharmaceutically acceptable medium is significantly effective in the treatment of tumors to obtain total regression of the tumor in most cases.
Claims (5)
1. A pharmaceutical composition comprising a therapeutically effective amount of a purified pyridine-soluble extract obtained from a microorganism, said extract containing between about 7 and 20% by weight of protein, between about 10 and 16% by weight of sugar, and between about 35 and 55% by weight of fatty acids, trehalose di-mycolate, and an acetone precipitated by-product of endotoxic glycolipids extracted with chloroform-methanol and a pharma-ceutically acceptable carrier.
2. The composition of claim 1 wherein the micro-organism is M. bovis BCG, M.phlei, M. smegmatis, M.Kansasii, Nocardia rubra, Corynebacterium diphtheriae or Corynebacter-ium parvum and preferably Corynebacterium parvum.
3. The composition of claim 1 wherein the extract contains about 12% by weight of each-of protein and sugar and about 45% by weight of fatty acids, the amount of the extract is up to about 40 milligrams, the amount of trehalose dimycolate is up to about 6 milligrams, and the amount of said acetone precipitated by-product is up to about 20 milligrams, and the composition is in lyophilized form or in the form of an oil droplet emulsion.
4. The composition of claim 3 wherein the oil is a light mineral oil, squalane, squalene, 7-n-hexyloctadecane, Conoco superoil or Drakeol 6VR mineral oil, the oil being present in an amount between about 0.5 and 3.0% by volume based on the total volume of the composition and there may be present in the composition a detergent in an amount between about 0.02 and 0.25% by volume based on the total volume of the composition.
5. The composition according to claim 1, wherein the amount of the extract is between about 200 and 5000 micrograms, the amount of trehalose dimycolate is between about 50 and 1000 micrograms and the amount of the acetone precipitated by-product is between about 150 and 1000 micrgrams.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US39382382A | 1982-06-30 | 1982-06-30 | |
| US393,823 | 1982-06-30 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1206415A true CA1206415A (en) | 1986-06-24 |
Family
ID=23556394
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000430665A Expired CA1206415A (en) | 1982-06-30 | 1983-06-17 | Pyridine soluble extract of a microorganism |
Country Status (6)
| Country | Link |
|---|---|
| JP (1) | JPS5917991A (en) |
| CA (1) | CA1206415A (en) |
| DE (1) | DE3323092A1 (en) |
| FR (1) | FR2529462B1 (en) |
| GB (1) | GB2122897B (en) |
| IT (1) | IT1163624B (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS59137408A (en) * | 1983-01-27 | 1984-08-07 | Taisho Pharmaceut Co Ltd | Ointment |
| US4663306A (en) * | 1983-09-23 | 1987-05-05 | Ribi Immunochem Research, Inc. | Pyridine-soluble extract-refined detoxified endotoxin composition and use |
| BE1007823A3 (en) * | 1993-12-10 | 1995-10-31 | Anda Biolog Sa | Use of a composition containing at least one antigen and / or one or more fragments of this antigen for obtaining a drug for treating and / or preventing cancer. |
| ZA984650B (en) * | 1997-06-19 | 1998-12-08 | Orion Corp | Intratumoral administration of triphenylethylenes for the treatment of cancer |
| CN100341574C (en) * | 2005-07-08 | 2007-10-10 | 中国药科大学 | Novel use of cell wall skeleton of red nocar-ray-fungus for treating liver diseases |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| FR2189021A1 (en) * | 1972-06-20 | 1974-01-25 | Anvar | Non-arthrogenic immunological adjuvants - obtained by extraction of lipid-free mycobacterial residues |
| US3976544A (en) * | 1973-06-19 | 1976-08-24 | The Agence Nationale De Valorisation De Le Recherche | Water-soluble immunological adjuvants, in particular for vaccines, obtained from mycobacteria and related microorganisms and process for their extraction |
| JPS5428813A (en) * | 1977-08-09 | 1979-03-03 | Yuuichi Yamamura | Solid preparation containing cell membrane extract substance used as suspension when using same |
-
1983
- 1983-06-17 CA CA000430665A patent/CA1206415A/en not_active Expired
- 1983-06-27 DE DE19833323092 patent/DE3323092A1/en active Granted
- 1983-06-28 FR FR8310673A patent/FR2529462B1/en not_active Expired
- 1983-06-29 IT IT21854/83A patent/IT1163624B/en active
- 1983-06-29 JP JP58116250A patent/JPS5917991A/en active Granted
- 1983-06-30 GB GB08317744A patent/GB2122897B/en not_active Expired
Also Published As
| Publication number | Publication date |
|---|---|
| IT1163624B (en) | 1987-04-08 |
| JPS6254774B2 (en) | 1987-11-17 |
| GB2122897A (en) | 1984-01-25 |
| GB8317744D0 (en) | 1983-08-03 |
| FR2529462A1 (en) | 1984-01-06 |
| FR2529462B1 (en) | 1987-02-13 |
| IT8321854A0 (en) | 1983-06-29 |
| GB2122897B (en) | 1986-03-26 |
| DE3323092A1 (en) | 1984-01-05 |
| DE3323092C2 (en) | 1987-02-05 |
| JPS5917991A (en) | 1984-01-30 |
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