Patent Application: US-65775507-A

Abstract:
this invention describes a novel family of polysaccharide prodrugs with enhanced colonic delivery advantage . the prodrugs are synthesized by chemically linking a parent compound with a specially selected polysaccharide containing galactose residues . its characteristics are that it is synthesized by chemically linking polysaccharides with the parent compound through different bridge links for targeted colonic delivery ; that the polysaccharides contain galactose residues . because of this , the polysaccharide component can protect the parent compound from absorption in the upper gastrointestinal tract and deliver a high concentration of the bound compound to the colonic area . upon reaching the colon , the active component of the parent drug will be released locally from the polysaccharide via enzymatic hydrolysis , allowing it to act locally for colonic disease such as inflammation or infection and / or taking advantage of the favorable microenvironment in the colon for steady and stable colonic drug absorption .

Description:
the purpose of this invention is to develop a method and process for preparing a family of prodrugs for enhanced colonic delivery and its application as indicated in fig1 where r is the linker and z is a parent compound . the following terms are used as defined herein . the use of these terms does not preclude the use of other terms not defined herein that are essentially synonymous with the defined terms . the term prodrug refers to a compound whose efficacy is greatly enhanced after one or more conversion step ( s ) that occurs in vivo after administering the compound to a subject or patient . the term galactose - containing polysaccharide refers to a polysaccharide having at least one galactose residue . a galactose - containing polysaccharide may be naturally occurring or may be prepared by modifying a different polysaccharide . further , a galactose - containing polysaccharide may comprise unmodified galactose residues or modified galactose - derived residues . the term galactose - containing fragment refers to a portion of the galactose - containing polysaccharide that may arise from being acted on by various enzymes . enzymes that will generate galactose - containing fragments are largely expected in the colon . these enzymes are largely bacterial in nature . the term therapeutic parent compound refers to a compound having therapeutic and / or diagnostic properties in a form prior to its linkage to a galactose - containing polysaccharide . this term is synonymous with parent compound and parent therapeutic compound . the term derivatize or derivatizing refers to modifying a compound , e . g ., galactose - containing polysaccharide or a therapeutic parent compound , by adding one or more reactive groups to the compound by reacting the compound with a functional group - adding reagent . as used herein , the term also refers to the attachment of cross - linkers to the compounds . the cross - linkers may be bifunctional , thus reacting with both compounds . a cross - linker possesses spacer arms that vary in size in different cross - linking compounds . this may be useful if one elects to have a known fixed distance between the galactose - containing polysaccharide and therapeutic parent compound . the term linkage or linking bond , refers to the covalent bond connecting , or linking , the galactose - containing polysaccharide and the therapeutic parent compound . this bond may be formed by attaching one or more functional groups to either of , or both of , the therapeutic parent compound and galactose - containing polysaccharide . the galactose - containing polysaccharide and / or the therapeutic parent compound may be derivatized by the addition of various functional groups . the term conjugate as used herein refers to the prodrug of the structural formula galactose - containing polysaccharide - r - z . the purpose of this invention is to develop a method and process for preparing a family of prodrugs for enhanced colonic delivery and its application as indicated in fig1 where r is the linker and z is a parent compound . first , the prodrug directs the parent compound to the colon , and then the parent - compound - galactose is bound to an appropriate protein on the surface of the colonic cells . this design can increase the parent compound selectivity , enhance its therapeutic effects , and reduce systemic toxicities , as well as with improved systemic absorption ( if so desired ). the technical proposal of this invention is a novel family of prodrugs with enhanced colonic delivery advantage and its application with the characteristic of the chemical formula as indicated in fig1 . the structure shown is the prodrug for the colonic delivery resulted from the linkage of polysaccharides with a parent compound through various bridge linkages . several unique features are : the polysaccharides are the polysaccharides with galactose residues . the polysaccharides with galactose include natural gums . the natural gums are pectin , guar gum , and carob bean gum , and the vegetative polysaccharides include aloe polysaccharide , medlar polysaccharide , and rhubarb polysaccharide . the compound resulted is polysaccharide - r - z , in which r is a linkage that is cleavable by bacterial enzymes present in the colon . the cleavable linkage can be replaced with any one of the following functional groups : r =—( ch 2 ) n —, — co —, — co ( ch 2 ) n —, — co ( ch 2 ) n co —, and n = 1 , 2 , 3 , 4 ; and z is a parent compound , includes but not limited to , anticancer drugs , steroids , non - steroidal anti - inflammatory drugs ( nsaids ), antibiotics , antidiabetic drugs , etc . the pectin , guar gum , and carob bean gum are hydrolyzed first with alkali ( ph = 9 - 10 ) then with acid ( ph = 3 - 5 ), and precipitated in alcohol and dialyzed to produce natural gums of target molecular weights of 10 5 - 10 7 da , containing galactose for linkage with the parent compound . the extraction method for the aloe polysaccharide , medlar polysaccharide , and rhubarb polysaccharide is as follows : first , pulverize the aloe ( or medlar , rhubarb ) plant materials and boiled with ethanol for three eight - hour - periods . the components dissolved in ethanol were extracted . the residue was boiled with water for another three eight - hour - periods in order to extract polysaccharides . all the water extractions were then collected . the polysaccharide - enriched fractions were obtained by precipitation with 5 volumes of ethanol for 3 times . after removing proteins , dialysis , separate and purify with gel filtration chromatography , polysaccharide components are obtained with molecular weights of 10 5 - 10 7 da . during the extraction process , the following analytical instrumentation and techniques are implemented : a ). high - performance liquid chromatography ( hplc ) for purity analysis ; b ). ultraviolet ( uv ) and infrared spectroscopic identification for qualitative examination ; c ). measurement of sugar and glycuronic acid contents respectively by vitriol - phenol and vitriol - carbazole methods ; and d ). measurement of monosaccharide compositions of the polysaccharides of different molecular weights and their component ratio with chromatographic techniques and gas chromatography . the synthetic method for the prodrug is to bind the parent compound at the free hydroxyl group of the galactose residues contained in the polysaccharide via the formation of an ester or ether linkage through modifying the free hydroxyl group to a reactive carboxyl group ( e . g . an acyl chloride ). then , the — nh 2 or similar functional groups ( including — nhnh 2 , — onh 2 , — nhc ═( o ) nhnh 2 , — oh , — co 2 h , and — sh ) of the parent compound reacts the modified carboxyl group of the galactose residues contained in the polysaccharides via the formation of an ester or ether linkage . the following is but one illustrative embodiment of a method for linking the galactose - containing polysaccharide with a parent therapeutic compound , z . a hydroxyl group in the 2 position of galactose is activated by chloroacetic acid . then the activated carbonyl linker group reacts with the amine group of z . this synthesis is exemplified below , using 5 - fu as the embodiment of z . alternatively , an amine group in an appropriate embodiment of z reacts with chloroacetic acid to create a carboxylic acid linker group . subsequent dcc ( dicyclohexylcarbodiimide ) coupling will link z &# 39 ; s newly added carboxylic acid linker group to the hydroxyl group in the 2 position of galactose in a galactose - containing polysaccharide . this method of linking is shown below . the formation of the ester linkage is made through acyl chloride method or n , n ′- dicyclohexylcarbodiimide ( dcc ) method . the formation of the ester linkage is carried out through condensation . the formation of the acyl - amine linkage is derived from aminolysis of acyl chloride . an embodiment of the drug delivery system involved in this invention is as follows : pectin : a polysaccharide composed of straight chains of galacturonic acid . the symbols “**” and “*” indicate the position of β ( 1 - 4 ) glycosidic linkages and n is from 1 to about 12 , 500 . guar gum : a non - ionic polysaccharide mainly polymerized with galactose and mannose , belonging to natural galactomannan with mannose as its main chain and β ( 1 - 4 ) glycoside link as the linkage between d - mannopyranose units . meanwhile , galactopyranose is connected to the mannose main chain through α ( 1 - 6 ) link . the molar ratio between mannose and galactose is 2 : 1 . carob bean gum : a colorless and flavorless polysaccharide refined from plant endosperm , mainly containing mannose and galactose with an average molecular weight of 300 kda . it is currently known that the natural occurring gums containing galactose residues , such as pectin , guar gum , and carob bean gum have the functions of regulating the bacterial colonies in the intestinal tract as well cholesterol lowering . in addition , aloe polysaccharides , medlar polysaccharides , and rhubarb polysaccharides are rich in galactose with known immunoregulation functions , which have not , as of yet , been fully explored for pharmaceutical development . the preparation methods of the prodrug involved in this invention and its characteristic of release in the colon will be described below . the new uses of the current colonic drug delivery and potential therapeutic use will also be discussed . however , this invention is not limited to the examples described below . preparation method of the novel family of prodrugs for the targeted delivery in the colon the natural gums containing galactose residues are hydrolyzed first with alkali ( ph = 9 - 10 ) then with acid ( ph = 3 - 5 ), and precipitated with alcohol and dialyzed to obtain natural gums of targeted molecular weights ( 10 5 - 10 7 da ) containing galactose residues . the extraction method for galactose such as aloe polysaccharide , medlar polysaccharide , and rhubarb polysaccharides is to pulverize aloe , medlar , and rhubarb plant materials , and boiled with ethanol for three eight - hour - periods . the components dissolved in ethanol were extracted . the residue was boiled with water for another three eight - hour - periods in order to extract polysaccharides . all the water extractions were then collected . the polysaccharide - enriched fractions were obtained by precipitation with 5 volumes of ethanol for 3 times . after removing proteins , dialysis , separate and purify with gel filtration chromatography , polysaccharide components are obtained with molecular weights of 10 5 - 10 7 da . during the extraction process , hplc for purity analysis , uv and infrared spectroscopic identification for qualitative examination , measurement of sugar and glycuronic acid contents respectively by vitriol - phenol and vitriol - carbazole methods , and measurement of monosaccharide compositions of the polysaccharides of different molecular weights and their component ratio with chromatographic techniques and gas chromatography are performed . link the above - mentioned polysaccharides ( including those prepared from natural gums ) with the parent compound . the linkage method first changes the free hydroxyl group to reactive carboxyl group of the aforementioned polysaccharides and then connecting them with the parent compound under different conditions as per implementation example 1 ; the linkage method can also be modifying the parent compound first , and then connect it with the aforementioned polysaccharides under different conditions as per implementation examples 2 and 3 . this method includes connecting the parent compound with the hydroxyl group of polysaccharides through derivation to form an ester or ether linkage , or chemically linking polysaccharides with the — nh part of the parent compound to form an acyl - amine linkage through derivation . the formation of the ester linkage is carried out through acyl chloride method or n , n ′- dicyclohexylcarbodiimide ( dcc ) method . the forming of the ester linkage is carried out through condensation , and the formation of the acyl - amine linkage is derived from aminolysis of acyl chloride . in view of the foregoing disclosure several embodiments of the prodrug and its methods of preparation are disclosed . the following embodiments are presented for illustrative purposes only and are not meant to limit the scope of the claimed subject matter . persons of ordinary skill in the art may be able to describe further embodiments based on the guidance set forth in the foregoing disclosure , the examples below and knowledge in the art . a desirable embodiment is a prodrug that can deliver a therapeutic parent compound with target specificity toward the colon . an additional embodiment is a prodrug capable of delivering a therapeutic parent compound to cells and / or tumors expressing the galactose binding lectin , galectin - 3 . embodiments directed to the methodologies for preparing and using the prodrug are also disclosed herein and are encompassed by invention . an additional embodiment is encompassed and illustrated by the isolation of the appropriate galactose - containing polysaccharides and their linkage to a parent therapeutic compound . further desirable embodiments are encompassed and illustrated by the prodrug having a structural formula , polysaccharide - r - z , wherein r is a covalent linkage and z is a therapeutic parent compound . further desirable embodiments are encompassed and illustrated by the prodrug having a structural formula , polysaccharide - r - z , wherein r is a covalent linkage and z is an anti - cancer drug , a non - steroidal anti - inflammatory agent , a steroid , an antibiotic , an anti - diabetes agent , etc . it is also desirable to provide embodiments of a prodrug for the targeted treatment of galectin - 3 expressing cancers wherein the galactose - containing polysaccharide is prepared from natural gums or plant material . these embodiments of the prodrug may have a galactose - containing polysaccharide prepared from pectin , guar gum , and carob bean gum , and the plant materials aloe , medlar and rhubarb . however , virtually any plant material having galactose - containing polysaccharides would make a suitable starting material for isolating said galactose - containing polysaccharide . additional embodiments of the prodrug for targeting galectin - 3 expressing cancers may have the parent compound 5 - fu linked to a galactose - containing polysaccharide . the linkage , e . g ., between 5 - fu and a galactose - containing polysaccharide that is mediated by a bifunctional cross - linker having spacer - arms of varying lengths . alternatively , linkages may be formed between a derivatized or underivatized galactose - containing polysaccharide and a derivatized or underivatized 5 - fu . in other embodiments of the methods for preparing the prodrug the 5 - fu may be derivatized . it is understood that derivatizing as referred to herein describes the addition of reactive groups to a galactose - containing polysaccharide or a 5 - fu molecule without introducing the spacer arms that characterize commercially available cross - linkers . additional embodiments of the prodrug polysaccharides - r - z are encompassed and illustrated by a covalent linkage comprising any of the following functional groups : —( ch 2 ) n —, — co —, — co ( ch 2 ) n —, — co ( ch 2 ) n co —, and n = 1 , 2 , 3 , or 4 . an embodiment of the prodrug may e . g ., result from forming a covalent linkage between the 5 - fu and free hydroxyl groups of the galactose residues in the polysaccharides . this linkage may be achieved via the formation of ester or ether linkages through derivatization . an illustrative example would be to form the bond between the — nh of 5 - fu at the free hydroxyl groups of the galactose residues in the polysaccharides via the formation of an acylamide linkage . embodiments of the methods for preparing the prodrug may use as starting material for galactose - containing polysaccharide isolation , pectin , guar gum , or carob bean gum . either material is first hydrolyzed with alkali ( ph = 9 - 10 ) then with acid ( ph = 3 - 5 ), and followed by precipitation with alcohol and dialysis . these methods yield galactose - containing polysaccharides of molecular weights from approximately 10 5 da to approximately 10 7 da . additional embodiments of the methods for preparing the prodrug may comprise isolating the galactose - containing polysaccharides from aloe , medlar or rhubarb as follows : pulverizing the aloe / medlar / rhubarb plant material , and boiling with ethanol for three eight - hour - periods . the components dissolved in ethanol are extracted . the ethanol insoluble residue is boiled with water for another three eight - hour - periods in order to extract polysaccharides . all the water extractions are finally collected . the polysaccharide - enriched fractions are obtained by precipitation with 5 volumes of ethanol for 3 times . after removing proteins , dialysis , separate and purify with gel filtration chromatography , polysaccharide components are obtained with molecular weights of 10 5 to 10 7 da . during the extraction process , high - performance liquid chromatography ( hplc ) for purity analysis , ultraviolet ( uv ) and infrared spectroscopic identification for qualitative examination , measurement of sugar and glycuronic acid contents respectively by vitriol - phenol and vitriol - carbazole methods , and measurement of monosaccharide compositions of the polysaccharides of different ( weight - average ) molecular weights and their component ratio with chromatographic techniques and gas chromatography are performed . additional embodiments of the invention encompass linking a galactose - containing polysaccharide and 5 - fu by the formation of the ester linkage made through acyl chloride method or n , n ′- dicyclohexylcarbodiimide ( dcc ) method . the embodiments of the prodrug illustrated above are effective for the treatment of galectin - 3 expressing cancers , including but not limited to the following : breast , lung , prostate , bladder , thyroid , other head and neck , lymphoma , colon , pancreas and other gastrointestinal cancers . the examples described below provide illustrative embodiments of methods of preparing the inventive prodrug . it should be readily appreciated that these examples taken together with knowledge in the art would allow persons in the art to practice related embodiments that are clearly encompassed by the subject matter disclosed and claimed herein . a prodrug with target specificity against colorectal cancer and its preparation methods . the major characteristic of this novel compound is that it is a prodrug synthesized by chemically linking a uniquely prepared polysaccharide with 5 - fluorouracil ( 5 - fu ), irinotecan , capecitabine , and camptothecin through various bridge links . the r group in the following examples is galactose residues with linker groups . to treat inflammatory bowel disease , ulcerative colitis , and crohn &# 39 ; s disease , steroids , such as dexamethasone , hydrocortisone , prednisolone , and fluorocortisone , can be modified to become new prodrugs to achieve colonic specificity . non - steroid anti - inflammatory drugs ( nsaids ), such as aspirin and ibuprofen . there are actually many nsaids ( more than 10 ), aspirin and ibuprofen are included as illustrative examples only . the r group in the following examples is galactose residues with linker groups . to achieve targeted colonic delivery and reduce side effects of antibiotics , drugs like penicillins , cephalosporins , quinolones , and metronidazole , can be linked to polysaccharides . the r group in the following examples is galactose residues with linker groups . to achieve colonic delivery and minimize side effects of other drugs that can benefit from colon delivery . one example is the anti - diabetic drugs in the class of biguanides , sulfonylurea , thiazolidinedione , etc can be linked to polysaccharides . the r group in the following examples is galactose residues with linker groups . to achieve systemic absorption via colon , any protein - based drug , vaccine or drug with poor upper gi absorption can be linked to polysaccharides . vaccine prod - drugs via colonic delivery have the added advantage in that the colonic environment is rich in lymphoid tissues with abundant immune cells . such approach can have the advantage of enhanced immune responses for the vaccine products . this invention is not limited to the implementation examples as described in these specifications . the implementation examples are for illustration only . the actual pharmaceutical forms of this invention can be any suitable formulation in any vehicle to be used for patients . without intent to limit the scope of the invention , exemplary methods and their related results according to the embodiments of the present invention are given above . note that titles or subtitles are used in the examples for convenience of a reader , which in no way should limit the scope of the invention . moreover , certain theories are proposed and disclosed herein ; however , in no way they , whether they are right or wrong , should limit the scope of the invention . 1 . abu shamat m . the role of gastrointestinal microflora in the metabolism of drugs . int j pharm 1993 ; 97 : 1 - 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