Patent Application: US-11397008-A

Abstract:
polyamine cationic lipids have been synthesized that have the ability to be transported into cells having an active polyamine transport system . accordingly , these lipids may be conjugated with various agents and , thereby , act as vectors for transporting the agent into the cell aided by the cell &# 39 ; s own polyamine transport system . a method of delivering an agent into a cell includes associating the agent with a polyamine cationic lipid selected from compounds 25 , 26 , 27 , 28 , their pharmaceutically acceptable salts and combinations thereof and contacting the cell therewith .

Description:
the present invention will now be described more fully hereinafter with reference to the accompanying drawings , in which preferred embodiments of the invention are shown . unless otherwise defined , all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains . although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention , suitable methods and materials are described below . any publications , patent applications , patents , or other references mentioned herein are incorporated by reference in their entirety . in case of conflict , the present specification , including any definitions , will control . in addition , the materials , methods and examples given are illustrative in nature only and not intended to be limiting . accordingly , this invention may , however , be embodied in many different forms and should not be construed as limited to the illustrated embodiments set forth herein . rather , these illustrated embodiments are provided so that this disclosure will be thorough and complete , and will fully convey the scope of the invention to those skilled in the art . other features and advantages of the invention will be apparent from the following detailed description , and from the claims . as used herein , the term “ pharmaceutically acceptable salt or prodrug ” is intended to describe any pharmaceutically acceptable form ( such as a salt of these amine systems with an organic carboxylic acid like acetic acid or toluene - sulfonic acid or methane sulfonic acid or an inorganic acid such as hcl , hbr , phosphoric acid , or a related group or prodrug ) of a compound of the invention , which , upon administration to a subject , provides the mature or base compound ( e . g ., the lipophilic polyamine compound ). pharmaceutically acceptable salts include those derived from pharmaceutically acceptable inorganic or organic bases and acids . suitable salts include those derived from alkali metals such as potassium and sodium , alkaline earth metals such as calcium and magnesium , among numerous other acids well known in the pharmaceutical art . pharmaceutically acceptable prodrugs refer to a compound that is metabolized , for example hydrolyzed or oxidized , in the host to form the compound of the present invention . typical examples of prodrugs include compounds that have biologically labile protecting groups on a functional moiety of the active compound . prodrugs include compounds that can be oxidized , reduced , aminated , deaminated , hydroxylated , dehydroxylated , hydrolyzed , dehydrolyzed , alkylated , dealkylated , acylated , deacylated , phosphorylated , dephosphorylated to produce the active compound . a series of derivatives were synthesized with a variety of linear polyamine headgroups . these cationic headgroups contained from two to four positive charges . the convergent synthetic route involved : a ) the synthesis of the hydrophobic lipid moiety , b ) the synthesis of the boc - protected amine head groups , c ) coupling of the two separate components followed by d ) the deprotection of the polyamine moiety to furnish the final compounds as hcl salts . the 3 , 4 - disubstituted benzene containing lipids were chosen due to the significant success of these lipids in earlier transfection studies by safinya and others . 10 , 23 , 24 this structural element is also present in dospa 4 and dotma 5 ( fig2 ). the unsaturated unit within the c18 chain was shown to prevent side chain recrystallisation and conferred a high degree of flexibility upon the cationic liposomes . these side chains were attached to the central benzene core via ether linkages . an aldehyde or acid chloride in position 1 allowed for attachment of the polyamine component . the polyamine scaffolds and controls were identified in earlier investigations of pat - mediated drug delivery . 25 - 29 the first step was the synthesis of 3 , 4 - di ( oleyloxy ) benzaldehyde 8a by the o - alkylation of 3 , 4 - dihydroxy benzaldehyde 7a with oleyl bromide ( scheme 1 as shown in fig3 ). similarly , the bis - o - alkylation of ethyl 3 , 4 - dihydroxybenzoate 7b with oleyl bromide was used to form ethyl 3 , 4 - di ( oleyloxy ) benzoate 8b using the method of safinya et al ., 23 , 24 subsequent cleavage of 8b with koh was carried out to give the acid 8c ( scheme 1 ) followed by treatment with oxalyl chloride to give the desired acid chloride 8d . as shown in scheme 2 ( fig4 ), the next step in the synthesis involved the generation of the polyamine moieties : n 1 -( tert - butoxycarbonyl ) putrescine 9 , n 1 , n 4 - di -( tert - butoxycarbonyl ) homospermidine 12 34 and n 1 , n 4 , n 8 - tri -( tert - butoxycarbonyl ) homospermine 14 ( scheme 2 ). the mono boc protection of diaminobutane gave the amine 9 in a good yield . sequential addition of bromobutyronitrile , boc protection of the newly formed secondary amine , followed by reduction of the nitrile with raney ni gave masked triamine 12 . repetition of these three steps on 12 gave tri - boc - protected tetraamine 14 . the coupling of the aldehyde 8a to a range of polyamines was based on previous procedures for coupling of amines to benzaldehyde derivatives . 25 - 28 as shown in scheme 3 ( shown in fig5 ), the reductive amination of 8a was achieved in two steps via in situ generation of the imine ( with a series of amines ) followed by reduction using nabh 4 to give the respective 2 ° amines ( 15 - 19 ). as shown in scheme 4 ( shown in fig6 ), the respective polyamines 12 and 14 were coupled to acid chloride 8d to provide the boc - protected benzamide systems , 20 and 21 . the final step to produce the desired lipid - polyamine conjugates involved acidification of the amines with a solution of anhydrous hcl in ethyl acetate . 23 , 24 as shown in scheme 5 ( shown in fig7 ), treatment of the penultimate compounds 15 - 21 with anhydrous hcl / etoac provided the target hcl salts , 22 - 28 . by design , the lipid portion of the conjugate ( boxed structure in scheme 5 ) was held constant throughout the series . this feature allowed for later comparisons and an understanding of how the polyamine component influenced dna delivery . before conducting the transfection studies , the series of conjugates ( 22 - 28 ) were first evaluated for cytotoxicity in chinese hamster ovary ( cho ) cells . this was an important step in determining what dose of lipid - polyamine conjugate could be tolerated by the cell line . ideally , one would use a dose of the conjugate , which is not cytotoxic to the cell line to be transfected . this is an important caveat in evaluating dna delivery systems . as expected , there were significant differences in the aqueous solubility of these new conjugates . dmso was added in portions to provide aqueous solutions of 22 - 28 . since dmso itself is toxic to cho cells above 40 μm , stock solutions of each conjugate were made in such a manner so that the total dmso concentration remained below 40 μm . this constraint limited the amount of conjugate that could be dosed . poorly - soluble materials required higher dmso levels , which in turn limited the amount of material that could be dosed to cells in our toxicity screen . taking these factors into account , cytotoxicity screens were performed to investigate the relative toxicity of each system . most materials were relatively non - toxic with ic 50 values ≧ 20 μm . armed with this insight , cells were treated with ≦ 6 . 4 μm of the polyamine conjugate so as to avoid significant toxic effects from the delivery agent itself . indeed , at this dose ≧ 90 % of cells survived transfection for all compounds except 28 . indeed , compound 28 was very toxic ( after 24 h incubation ) and even at the lowest dose used for transfection ( 0 . 5 μg / ml ; 0 . 5 μm 28 ) killed 95 % of the cells . as such , 28 was too toxic for efficient transfection at the doses surveyed and its data is strongly biased by the few remaining cells , which survived . armed with knowledge of the cytotoxicity range of the series 22 - 28 , cho - k1 cells were evaluated for dna uptake using a fluorescently labeled dna alexa fluor - 488 - dna ). as shown in fig8 , cells were dosed with the alexa fluor - 488 - dna in the presence of increasing concentrations ( i . e ., 0 . 5 , 2 . 5 , 5 μg / ml ) of the respective conjugate , 22 - 28 . each conjugate was as good ( 24 , 25 ) or better ( 26 , 27 ) than the lipofectamine control ( lfa , except 22 and 23 ) and facilitated uptake of the fluorescent dna probe in a concentration - dependent manner ( μg / ml ). while the molecular weights in this series do range from 784 ( 22 ), 840 ( 23 ), 844 ( 24 ), 891 ( 25 ), 999 ( 26 ), 869 ( 27 ), and 976 g / mol ( 28 ), they are relatively close ( within 10 - 22 %) and allow for general comparisons , especially in light of the large differences observed in activity . for example , tetraamine 26 , which has the highest molecular weight of the series ( 999 g / mol ), was at a slightly lower concentration ( 5 μm ) than 22 ( 6 . 4 μm ) at the 5 μg / ml dose . nevertheless , dna uptake experiments with tetraamine 26 had over a 5 - fold increase in dna uptake ( as measured by fluorescence of the imported dna probe ) than those conducted with diamine 22 . clearly , compound 26 was more efficient in facilitating dna delivery to cells . interestingly , conjugate 27 , which represents a butanediamine motif similar to 22 except with the diamine placed further away from the lipid tail , was a more efficient dna delivery agent than 22 . in contrast , compound 23 , which separated the ammonium centers via an octanediamine had similar or lower activity as 22 ( depending on the dose ). insertion of the polyether motif present in 24 maintained this eight - atom spacer , yet nearly doubled the dna delivery ( dna probe fluorescence ) observed . the pat - selective homospermidine motif present in 25 resulted in a three - fold increase in dna delivery . however , this outcome may simply be due to the presence of the additional charge provided by the triamine motif present in 25 . indeed , significant increases in dna delivery were observed across the homologous series 22 , 25 , and 26 which at 5 μg / ml gave 94 , 281 and 504 fluorescence absorbance units ( a . u . ), respectively . fig8 . shows dna uptake activity by novel cationic lipids 22 - 28 ( compared with lipofectamine , lfa ). cho k1 cells were grown to confluence in 24 - well plates , rinsed with medium and incubated for 4 h at 37 ° c . with 1 μg / ml alexa fluor - 488 - dna complexed with 0 . 5 μg / ml ( white bars ), 2 . 5 μg / ml ( grey bars ), or 5 μg / ml ( black bars ) of the respective cationic lipid , as indicated above each entry . cells were then extensively washed with pbs , treated with trypsin , and cell pellets were again extensively washed with pbs 1 % bsa to completely remove non - specific fluorescence from the cell exterior . finally , cells were analyzed for alexa fluor - 488 - dna uptake by flow cytometry on a facscalibur ( bd biosciences ) operated by cell - quest software . (*) during this relatively short incubation ( 4 h ), cells remained viable with 28 . note : in converting from μg / ml to μm , the concentrations used were typically approximately 0 . 6 , 3 . 0 , and 6 μm , respectively and are tabulated specifically in the supporting information . in fig9 we see the results obtained from the experiment described by fig8 , which were analyzed using a gated channel to determine the percentage of cells that had internalized significant amounts of dna as compared with cells incubated with dna only . a gate was set just above the threshold signal for control cells ( dna only with no lipid conjugate ). note : alexa fluor - 488 - dna complexed with 0 . 5 μg / ml ( white bars ), 2 . 5 μg / ml ( grey bars ), or 5 μg / ml ( black bars ) of the respective cationic lipid was used , as indicated in fig8 . data are presented as the mean ± sd . while the average amount of dna taken up / cell shows wide variations between the different compounds ( fig8 ), all compounds were able to deliver significant amounts of dna to virtually the entire cell population ( fig9 ). however , in terms of gene therapy , simple dna delivery to the cell is insufficient . there are other cellular barriers , which must be traversed . in order for the ‘ therapy ’ to be effective , the dna must escape from the endosome and enter the cell &# 39 ; s nucleus , be transcribed to a regulatory , non coding rna ( rnai ) or to mrna that is translated into its coded protein . therefore , we investigated the conjugate - assisted expression of an egfp dna plasmid encoding for the green fluorescent protein ( gfp ). cho cells , which were successfully transfected , were easily identified by their green fluorescence . control experiments conducted with only the egfp dna plasmid ( and no lipid carrier ) gave virtually no fluorescence . a gate or instrumental threshold was set based upon this low background fluorescence . fluorescence detected above this background was considered a positive response . cho k1 cells were grown to approx . 50 % sub - confluence in 24 - well plates , rinsed with medium and incubated for 4 h at 37 ° c . with 1 μg / ml egfp dna plasmid complexed with 0 . 5 μg / ml ( white bars ), 2 . 5 μg / ml ( grey bars ), or 5 μg / ml ( black bars ) of the respective cationic lipid , as indicated above . medium was then changed , and cells were incubated for another 24 h to allow for gfp plasmid expression . cells were detached by trypsin treatment and analyzed for gfp expression by facs . lipofectamine is denoted as lfa . the value obtained with dna plasmid only was subtracted from each entry above ( almost negligible ). the results obtained from the experiment described in fig1 were analyzed using a gated channel to determine the percentage of cells that expressed significant levels of gfp as compared with cells incubated with dna plasmid only . a gate was set just above the threshold measured with no dna plasmid and no conjugate added ( almost negligible ). all cells which exhibited a fluorescence signal above that level were considered positive . data are presented as the mean ± sd . these are shown in fig1 . the interpretation of the data in fig1 and 11 can be quite subtle . for example , fig1 relates the amount of gfp expression per cell and is a summary measurement of how well the dna ‘ message ’ was delivered , read and protein ( gfp ) produced . while the magnitudes are different the relative trends are consistent with those observed in fig8 . in this regard , dna delivery correlated with gfp expression . fig1 revealed the % gfp positive cells observed in the total cell population after the transfection experiment . this information is a direct measure of transfection efficiency . using the 5 μg / ml dose for comparisons , 22 ( 38 %), 25 ( 49 %), and 27 ( 40 %) were all comparable to the lfa control ( 41 %) in terms of transfection efficiency . both 23 ( 23 %) and 24 ( 25 %) gave lower values . the lone standout was tetraamine 26 ( 67 %), which had over 50 % higher transfection efficiency than the lfa control ( 41 %). cells treated with conjugate 23 at the high dose ( 5 μg / ml , black bars in fig9 ) gave 84 % positive cells of the total cell population remaining after the dna uptake experiment ; whereas all the others ( lfa , 22 , and 24 - 28 ) gave typically & gt ; 94 % positive cells . using these data as benchmarks of dna import via each conjugate , the significantly lower % gfp positive cells ( 23 - 67 %) observed in fig1 suggests that intracellular processing and nuclear delivery of dna also depend on the structure of the cationic lipid . a closer analysis of the data revealed just how important these latter two parameters ( i . e ., intracellular processing and nuclear delivery ) are for successful gene delivery . for example , although 26 was capable of delivering 3 - fold more dna to the cell than the control lfa ( black bars in fig8 : 26 : 504 ; lfa : 158 a . u . ), the relative gfp expression / cell was only two fold higher in fig1 ( 25 vs . 13 a . u .) and the % of gfp positive cells was only 1 . 5 fold higher in fig1 ( 67 % vs . 41 %). it cannot be excluded that maximum gfp expression occurs at varying time points post - transfection for the different compounds . nevertheless , the data reflect an interesting correlation between intracellular processing of internalized dna and the cationic lipid structure . in summary , while a significant number of cells imported the fluorescent dna probe in the presence of the synthetic conjugates ( 22 - 28 ), the amount of the dna probe entering each cell varied depending upon the conjugate used . the % gfp positive cells of the total cell population roughly correlated with the gfp expression / cell . cells with low % gfp positive cells ( fig1 : 23 , 23 %; 24 , 25 %) had low gfp expression / cell ( fig1 : 23 , 5 ; 24 , 7 ). cells with high % gfp positive cells ( fig1 : 26 , 67 %) had high gfp expression / cell ( fig1 : 26 , 25 ). indeed , the additional ‘ intracellular barriers ’ associated with successful gfp expression ( as shown in fig1 in terms of % gfp positive cells ) seemed to moderate the large differences seen in the earlier dna delivery study ( fig8 ). we theorized that 26 may be using the polyamine transporter , pat , for cellular entry . indeed , as shown in fig1 , tetraamine 26 was shown to be a potent inhibitor of spermine uptake ( ic 50 of approximately 10 μm ) using a 14 c - radiolabeled spermine competition assay . this observation implied that 26 was able to compete for the polyamine recognition sites on the cell surface ( e . g ., pat ). therefore , it is possible that certain lipophilic polyamines , which present the correct polyamine ‘ message ’, may be able to recognize and target cells expressing high levels of polyamine transporters on their cell surface ( e . g ., cancer cells , rapidly dividing tumors , etc .). however , as shown in this report , delivery into the cell is just one step in successful gene transfection . nevertheless , the findings disclosed herein are an important first step in developing ‘ smart ’ transfection agents . proof - of - principle for this novel therapeutic approach was provided in the following journal article which utilized an anti - cancer toxin that consisted of a chemical conjugate between the human urokinase plasminogen activator and saporin : lipopolyamine treatment increases the efficacy of intoxication with saporin and an anticancer saporin conjugate . geden s . e . ; gardner r . a . ; fabbrini m . s . ; ohashi m . ; phanstiel , iv , o . ; teter k . febs j . 2007 , 274 , 4825 - 4836 ( published online aug . 22 , 2007 ). this journal article is incorporated herein by reference in its entirety . we have also documented lipopolyamine - induced toxin sensitization with two other saporin - based toxins : ( i ) fgf - saporin , a chemical conjugate between fibroblast growth factor and saporin ; and ( ii ) the okt10 immunotoxin which consists of a chemical conjugate between saporin and an antibody that recognizes the cd38 cancer antigen . du145 human prostate cancer cells exposed to lipopolyamine 26 and fgf - saporin were 85 - fold more sensitive to intoxication than cells incubated with fgf - saporin alone . the commercial transfection agent lipofectamine generated significant cellular sensitization to fgf - saporin as well . however , lipofectamine also sensitized a non - target cell population ( hela cells , which do not express the fgf receptor ) to fgf - saporin . in contrast , hela cells exposed to lipopolyamine 26 and fgf - saporin were no more sensitive to intoxication than cells incubated with fgf - saporin alone . our reagent thus exhibited a level of specificity that was not seen with lipofectamine . we have also shown that ramos human leukemia cells exposed to lipopolyamine 26 and okt10 are 100 - fold more sensitive to intoxication than cells incubated with okt10 alone . ramos cells exposed to lipofectamine and okt10 were no more sensitive to intoxication than cells exposed to okt10 alone . collectively , these results demonstrate the utility of our lipopolyamine compounds over the commercially available lipofectamine reagent . this investigation demonstrated that the number and position of the positive charges along the polyamine scaffold plays a key role in dna delivery and in determining the intracellular outcome of the dna import event , i . e . the transfection efficiency . although the lipofectamine ( lfa : 75 % 4 and 25 % 6 mixture ) has five positive charges ( presumably a dna delivery enhancing feature ) in one of its components ( e . g ., 4 ), it also has a neutral component dope 6 present , which makes structural comparisons to this control difficult beyond the dose to dose comparisons made above . future studies will look at the effect of neutral lipids ( like 6 ) in terms of transfection efficiency of polyamine - lipid conjugates . indeed , dope 6 has been shown to play a role in endosome disruption and to facilitate transfection . 35 in addition , this paper presented the first study , which probed the transfection phenomena with a pat - selective homospermidine motif 28 attached to a lipid cargo , e . g ., 25 . as expected compound 25 showed a 3 - fold enhancement in dna delivery over its butanediamine analogue 22 . unfortunately , the dna delivery enhancement observed with 25 seemed to be muted by the latter steps of the transfection process ( e . g ., intracellular trafficking , nuclear delivery , etc ). as a result , 25 had only slightly higher transfection efficiency than 22 ( e . g ., 25 and 22 gave 49 % and 38 % gfp positive cells in fig1 , respectively ). this phenomenon was also observed with the two diamines , 27 and 22 . compound 27 showed greater than 4 - fold enhancement in dna delivery over its butanediamine analogue 22 . evidently , moving the diamine ‘ message ’ further away from the lipid component enhanced the delivery characteristics of the conjugate ( fig8 ). again , this potential delivery enhancement by 27 was tempered by the latter steps of the transfection process . alas , nearly identical transfection efficiency was observed for 27 and 22 ( 40 % and 38 % gfp positive cells in fig1 , respectively ). clearly , cellular dna delivery alone is insufficient for successful transfection . understanding how different polyamine structures 21 ′ 36 and neutral lipids like 6 35 enhance the intracellular trafficking and nuclear delivery of plasmid dna is critical for the future design of efficient polyamine transfection agents . silica gel ( 32 - 63 μm ) and chemical reagents were purchased from commercial sources and used without further purification . all solvents were distilled prior to use . 1 h and 13 c nmr spectra were recorded at 300 and 75 mhz , respectively , unless otherwise noted . tlc solvent systems are based on volume % and nh 4 oh refers to concentrated aqueous nh 4 oh . high - resolution mass spectrometry was performed at the university of florida mass spectrometry facility . ulysis alexafluor - 488 ( molecular probes ) was used for labeling of dna as recommended by the manufacturer . all fine chemicals were from sigma . lipofectamine 2000 was purchased from invitrogen . wild - type chinese hamster ovary cells ( cho - k1 ) were obtained from the atcc ( manassas , va ., usa ). cho cells were routinely cultured in f12k nutrient mixture supplemented with 10 % ( v / v ) fetal calf serum , 2 mm l - glutamine , 100 units / ml penicillin , and 100 μg / ml streptomycin ( growth medium ) in a humidified 5 % co 2 , 37 ° c . incubator . dna uptake studies . alexafluor - 488 labeled dna with or without cationic lipid was mixed in serum - free f12k and added to extensively rinsed cells grown in 24 - well plates . cells were then incubated for 4 h at 37 ° c . after removal of the incubation medium and rinsing with pbs , cells were detached with trypsin followed by extensive washing with ice - cold pbs , 1 % ( w / v ) bovine serum albumin ( bsa ) to remove non - specific fluorescence . finally , cells were suspended in pbs , 1 % bsa and analyzed for dna uptake by flow cytometry on a facscalibur ( bd biosciences ) operated by cell - quest software . cells remained viable with all compounds tested , including 28 . gfp transfection experiments — egfp encoding dna plasmid with or without cationic lipid was mixed in f12k and incubated with pre - rinsed , sub - confluent cells in 24 - well plates for 4 h , followed by another incubation period of 24 h in growth medium . cells were then washed with pbs , detached by trypsin treatment , dissolved in pbs , 1 % bsa , and analyzed by facs for gfp expression . note : severe toxicity was noted for compound 28 after the 24 h incubation period . cho cells were grown in rpmi medium supplemented with 10 % fetal calf serum , glutamine ( 2 mm ), penicillin ( 100 u / ml ), streptomycin ( 50 μg / ml ). cells were grown at 37 ° c . under a humidified 5 % co 2 atmosphere . aminoguanidine ( 2 mm ) was added to the culture medium to prevent oxidation of the drugs by the enzyme ( bovine serum amine oxidase ) present in calf serum . trypan blue staining was used to determine cell viability before the initiation of a cytotoxicity experiment . l1210 cells in early to mid log - phase were used . cell growth was assayed in sterile 96 - well microtiter plates ( becton - dickinson , oxnard , calif ., usa ). cho cells were plated at 2e 3 cells / ml . drug solutions ( 10 μl per well ) of appropriate concentration were added after an overnight incubation for the cho cells . after exposure to the drug for 48 hr , cell growth was determined by measuring formazan formation from 3 -( 4 , 5 - dimethylthiazol - 2 - yl ) 2 , 5 - diphenyltetrazolium using a titertek multiskan mcc / 340 microplate reader for absorbance ( 540 nm ) measurements . 37 these experiments allowed for a quick assessment of conjugate cytotoxicity and facilitated determination of the proper dosing for the latter transfection and delivery experiments . to a solution of oleyl bromide ( 1 . 8 g , 5 . 4 mmol ) in cyclohexanone ( 20 ml ) was added protocatachualdehyde ( 0 . 341 g , 2 . 5 mmol ), potassium carbonate ( 1 . 02 g , 7 . 4 mmol ) and potassium iodide ( 0 . 05 g , 0 . 3 mmol ). the suspension was stirred at 100 ° c . for 18 hours under nitrogen . due to the light sensitivity of the oleyl bromide the flask was covered in aluminum foil . tlc ( 10 % meoh / chcl 3 ) showed the reaction was complete . the hot reaction mixture was filtered to remove some of the particulates and the solvent was removed in vacuo . the residue was dissolved in chcl 3 ( 100 ml ) and washed with water ( 2 × 75 ml ). the chcl 3 layer was dried over anhydrous sodium sulphate , filtered and the solvent removed in vacuo to give a dark yellow oil . the oily residue was purified by flash column chromatography ( 5 % etac / hexane ) to yield the product 8a as a white solid ( 1 . 08 g , 69 %), r f = 0 . 3 ( 5 % etac / hexane ). 1 h nmr ( cdcl 3 ) d 9 . 80 ( s , 1h , cho ), 7 . 37 ( m , 2h , phenyl ), 6 . 93 ( d , 1h , phenyl ), 5 . 33 ( t , 4h , olefinic ), 4 . 06 ( t , 2h , och 2 ), 4 . 04 ( t , 2h , och 2 ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 85 ( m , 4h , c h 2 ch 2 o ), 1 . 50 - 1 . 18 ( m , 44h , 22 × ch 2 ), 0 . 88 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 191 . 00 , 155 . 34 , 149 . 95 , 130 . 14 , 130 . 12 , 129 . 97 , 126 . 79 , 111 . 89 , 111 . 04 , 69 . 36 , 32 . 29 , 30 . 16 , 29 . 92 , 29 . 90 , 29 . 88 , 29 . 72 , 29 . 64 , 29 . 62 , 29 . 44 , 29 . 36 , 27 . 60 , 26 . 37 , 26 . 34 , 23 . 09 , 14 . 53 ; hrms ( fab ): theory for c 43 h 75 o 3 ( m + 1 ), 639 . 5716 ; found ( m + 1 ), 639 . 5755 . anal . ( c 43 h 74 o 3 ): c , h . to a solution of oleyl bromide ( 1 . 8 g , 5 . 4 mmol ) in cyclohexanone ( 20 ml ) was added ethyl 3 , 4 - dihydroxybenzoate 7b ( 0 . 45 g , 2 . 5 mmol ), potassium carbonate ( 1 . 02 g , 7 . 4 mmol ) and potassium iodide ( 0 . 05 g , 0 . 3 mmol ). the suspension was stirred at 100 ° c . for 18 hours under nitrogen . due to the light sensitivity of the oleyl bromide the flask was covered in aluminum foil . the hot reaction mixture was filtered to remove some of the particulates and the solvent was removed in vacuo . the residue containing ester 8b was dissolved in a solution of ethanol ( 20 ml ) containing potassium hydroxide ( 0 . 8 g , 20 mmol ) and refluxed for 4 h . the hot reaction mixture was added to water ( 30 ml ) and acidifying with 1m hcl to ph 1 resulted in the precipitation of a white solid . the solid 8c was filtered off and washed several times with water . the crude acid was re - crystallized from ethanol ( 10 ml ) to give the product 8c as a white solid ( 1 . 14 g , 70 %); 1 h nmr ( 500 mhz , cdcl 3 ) d 7 . 73 ( dd , 1h , phenyl ), 7 . 58 ( d , 1h , phenyl ), 6 . 89 ( d , 1h , phenyl ), 5 . 35 ( t , 4h , olefinic ), 4 . 05 ( 2 × t , 4h , 2 × och 2 ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 85 ( m , 4h , c h 2 ch 2 o ), 1 . 48 ( quin , 4h , 2 × ch 2 ), 1 . 41 - 1 . 19 ( m , 40h , 20 × ch 2 ), 0 . 88 ( t , 6h , ch 3 ). 1 , 4 - diaminobutane ( 4 . 4 g , 0 . 05 mol ) was dissolved in a solution of triethylamine and methanol ( 10 % tea in meoh , 110 ml ). a solution of di - tert - butyl dicarbonate ( 3 . 63 g , 0 . 017 mol ) in methanol ( 10 ml ) was added dropwise to this mixture with vigorous stirring . the mixture was stirred at rt overnight . the tert - butoxy - carbonylation was complete as evidenced by tlc ( 4 % nh 4 oh / meoh ). the excess 1 , 4 - diaminobutane , methanol and tea were removed in vacuo to yield an oily residue that was dissolved in dichloromethane ( 100 ml ) and washed with a solution of sodium carbonate ( 10 % aq , 2 × 100 ml ). the organic layer was dried over anhydrous sodium sulfate , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) to give the product 9 as a clear oil ( 2 . 23 g , 71 %), r f = 0 . 38 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ). 1 h nmr ( cdcl 3 ) d 4 . 72 ( br s , 1h , nhco ), 3 . 12 ( q , 2h , ch 2 ), 2 . 70 ( t , 2h , ch 2 ), 1 . 57 - 1 . 30 ( m , 13h , 2 × ch 2 , 3 × ch 3 ). to a solution of the boc protected diamine 9 ( 2 . 10 g , 0 . 01 mol ) in anhydrous acetonitrile ( 50 ml ) was added potassium carbonate ( 5 . 14 g ) and the suspension was stirred at rt for 10 minutes . a solution of 4 - bromobutyronitrile ( 1 . 65 g , 0 . 01 mol ) in acetonitrile ( 25 ml ) was added and the resulting mixture stirred at 50 ° c . for 24 hours . tlc ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) showed that the reaction was 95 % complete . the mixture was filtered to remove most of the inorganic salts and the acetonitrile was removed in vacuo to give a solid / oily residue that was purified by flash column chromatography ( 1 : 5 : 94 nh 4 oh : meoh : chcl 3 ) to yield the product 10 as a clear oil ( 1 . 74 g , 61 %), r f = 0 . 5 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ); 1 h nmr ( cdcl 3 ) d 4 . 79 ( br s , 1h , nhco ), 3 . 12 ( q , 2h , ch 2 ), 2 . 74 ( t , 2h , ch 2 ), 2 . 60 ( t , 2h , ch 2 ), 2 . 46 ( t , 2h , ch 2 ), 1 . 81 ( quin , 2h , ch 2 ) 1 . 57 - 1 . 37 ( m , 13h , 2 × ch 2 , 3 × ch 3 ). the amino - nitrile 10 ( 1 . 74 g , 6 . 8 mmol ) was dissolved in a solution of triethylamine and methanol ( 10 % tea in meoh , 40 ml ). a solution of di - tert - butyl dicarbonate ( 3 . 63 g , 0 . 017 mol ) in methanol ( 20 ml ) was added dropwise to this mixture with vigorous stirring . the mixture was stirred at it overnight . tlc ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) showed the tert - butoxycarbonylation was complete . the methanol and tea were removed in vacuo to yield an oily residue that was dissolved in dichloromethane ( 100 ml ) and washed with a solution of sodium hydroxide ( 2 . 5 m , 3 × 30 ml ) and water ( 2 × 30 ml ). the organic layer was dried over anhydrous na 2 so 4 , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 40 % etac / hexane ) to give the product 11 as a clear oil ( 2 . 10 g , 87 %), r f = 0 . 45 ( 40 % etac / hexane ); 1 h nmr ( cdcl 3 ) d 4 . 60 ( br s , 1h , nhco ), 3 . 28 ( t , 2h , ch 2 ), 3 . 14 ( m , 4h , 2 × ch 2 ), 2 . 35 ( t , 2h , ch 2 ), 1 . 88 ( quin , 2h , ch 2 ), 1 . 58 - 1 . 28 ( m , 22h , 2 × ch 2 , 6 × ch 3 ). the nitrile 11 ( 2 . 00 g , 5 . 6 mmol ) was dissolved in ethanol ( 100 ml ). nh 4 oh ( 10 ml ) and raney nickel ( 8 g ) were added and ammonia gas was bubbled through the solution for 20 minutes at 0 ° c . the suspension was hydrogenated at 50 parr for 24 hours . air was bubbled through the solution and the raney nickel was removed by filtering through a sintered glass funnel keeping the raney nickel residue moist at all times . the ethanol and nh 4 oh were removed in vacuo and the oily residue dissolved in ch 2 cl 2 and washed with 10 % aq , na 2 co 3 ( 3 × 50 ml ). the organic layer was dried over anhydrous na 2 so 4 , filtered and the solvent removed in vacuo to give the product 12 as a clear oil without further purification ( 1 . 92 g , 95 %), r f = 0 . 4 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ); 1 h nmr ( cdcl 3 ) d 4 . 65 ( br s , 1h , nhco ), 3 . 14 ( m , 6h , 3 × ch 2 ), 2 . 69 ( t , 2h , ch 2 ), 2 . 35 ( t , 2h , ch 2 ), 1 . 58 - 1 . 26 ( m , 26h , 4 × ch 2 , 6 × ch 3 ). to a solution the amine 12 ( 1 . 32 g , 3 . 68 mmol ) in anhydrous acetonitrile ( 20 ml ) was added potassium carbonate ( 1 . 7 g ) and the suspension was stirred at rt for 10 minutes . a solution of 4 - bromobutyronitrile ( 0 . 54 g , 3 . 68 mmol ) in acetonitrile ( 10 ml ) was added and the resulting mixture stirred at 50 ° c . under nitrogen for 24 hours . tlc ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) showed the reaction was 95 % complete . the mixture was filtered to remove most of the inorganic salts and the acetonitrile was removed in vacuo to give a solid / oily residue . the oil was dissolved in anhydrous thf ( 70 ml ). a solution of di - tert - butyl dicarbonate ( 1 . 21 g , 5 . 5 mmol ) in thf ( 20 ml ) was added dropwise to this mixture with vigorous stirring . the mixture was stirred at rt overnight . tlc ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) showed the tert - butoxycarbonylation was complete . the thf was removed in vacuo to yield an oily residue that was dissolved in dichloromethane ( 100 ml ) and washed with a solution of sodium hydroxide ( 2 . 5 m , 3 × 30 ml ). the organic layer was dried over anhydrous sodium sulfate , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( chcl 3 / meoh 99 : 1 ) to give the product 13 as a clear oil ( 1 . 46 g , 75 %), r f = 0 . 5 ( chcl 3 / meoh 99 : 2 ); 1 h nmr ( cdcl 3 ) d 4 . 63 ( bs , 1h , nhco ), 3 . 28 ( t , 2h , ch 2 ), 3 . 14 ( m , 8h , 4 × ch 2 ), 2 . 64 ( t , 2h , ch 2 ), 1 . 88 ( quin , 2h , ch 2 ), 1 . 58 - 1 . 35 ( m , 35h , 4 × ch 2 , 9 × ch 3 ). the nitrile 13 ( 1 . 33 g , 2 . 5 mmol ) was dissolved in ethanol ( 100 ml ). nh 4 oh ( 10 ml ) and raney nickel ( 6 g ) were added and ammonia gas was bubbled through the solution for 20 minutes at 0 ° c . the suspension was hydrogenated at 50 psi for 24 hours . air was bubbled through the solution and the raney nickel was removed by filtering through a sintered glass funnel keeping the raney nickel residue moist at all times . the ethanol and nh 4 oh were removed in vacuo and the oily residue dissolved in ch 2 cl 2 and washed with sodium carbonate ( 10 %, aq , 3 × 50 ml ). the organic layer was dried over anhydrous sodium sulphate , filtered and the solvent removed in vacuo . the oil was purified by flash column chromatography ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) to give the product 14 as a clear oil ( 0 . 85 g , 64 %), r f = 0 . 5 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ). 1 h nmr ( cdcl 3 ) d 4 . 65 ( s , 1h , nhco ), 3 . 15 ( m , 10h , 5 × ch 2 ), 2 . 70 ( t , 2h , c h 2 nh 2 ), 1 . 58 - 1 . 39 ( m , 39h , 6 × ch 2 , 9 × ch 3 ); hrms ( fab ): theory for c 27 h 55 n 4 o 6 ( m + 1 ), 531 . 4122 ; found ( m + 1 ), 531 . 4111 ; anal . calcd ( c 27 h 54 n 4 o 6 0 . 1h 2 o ): c , h , n . to a vigorously stirred solution of the amine 9 ( 0 . 071 g , 0 . 38 mmol , 1 . 2 equiv ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ) was added a solution of the aldehyde 8a ( 0 . 20 g , 0 . 33 mmol ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ), dropwise over 20 minutes . the resulting mixture was stirred at it under an atmosphere of nitrogen overnight . 1 h nmr showed the reaction to be complete when there was no aldehyde peak present . the solvent was removed in vacuo and the crude imine dissolved in ch 2 cl 2 / meoh ( 1 : 1 , 10 ml ). the solution was cooled to 0 ° c . and nabh 4 ( 60 mg , 1 . 58 mmol ) was added in 15 mg portions over 30 minutes . tlc ( 10 % etac / hexane ) showed the reaction to be complete after stirring overnight . the solvent was removed in vacuo and the crude oil dissolved in ch 2 cl 2 ( 50 ml ) and washed with sodium carbonate ( 10 % aq , 3 × 40 ml ). the ch 2 cl 2 layer was dried over na 2 co 3 , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 4 % meoh / chcl 3 ) to give the product 15 as a clear oil ( 0 . 18 g , 78 %), r f = 0 . 2 ( 3 % meoh / chcl 3 ). 1 h nmr ( 500 mhz , cdcl 3 ) d 6 . 85 ( s , 1h , aryl ), 6 . 80 ( m , 2h , aryl ), 5 . 34 ( t , 4h , olefinic ), 4 . 78 ( br s , 1h , nhco ), 3 . 97 ( q , 4h , och 2 ), 3 . 69 ( s , 2h , benzylic ), 3 . 13 ( m , 2h , ch 2 ) 2 . 63 ( t , 2h , ch 2 nh ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 80 ( m , 4h , c h 2 ch 2 o ), 1 . 55 ( m , 4h ), 1 . 49 - 1 . 12 ( m , 57h , 24 × ch 2 , 3 × ch 3 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 156 . 17 , 149 . 34 , 148 . 29 , 133 . 15 , 130 . 11 , 130 . 00 , 120 . 67 , 114 . 18 , 114 . 08 , 79 . 23 , 69 . 69 , 69 . 52 , 54 . 12 , 49 . 22 , 40 . 80 , 32 . 28 , 30 . 16 , 29 . 92 , 29 . 82 , 29 . 71 , 29 . 67 , 28 . 81 , 28 . 27 , 27 . 68 , 27 . 60 , 26 . 44 , 23 . 09 , 14 . 54 ; anal . ( c 52 h 94 n 2 o 4 ): c , h , n . to a vigorously stirred solution of diaminooctane ( 0 . 23 g , 1 . 56 mmol , 5 equiv ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ) was added a solution of the aldehyde 8a ( 0 . 20 g , 0 . 33 mmol ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ), dropwise over 1 hour . the resulting mixture was stirred at rt under an atmosphere of nitrogen overnight . nmr showed the reaction to be complete when there was no aldehyde peak present in the nmr spectrum . the solvent was removed in vacuo and the crude imine dissolved in ch 2 cl 2 / meoh ( 1 : 1 , 10 ml ). the solution was cooled to 0 ° c . and sodium borohydride ( 60 mg , 1 . 58 mmol ) was added in 15 mg portions over 30 minutes . tlc ( 10 % etac / hexane ) showed the reaction to be complete after stirring overnight . the solvent was removed in vacuo and the crude solid dissolved in ch 2 cl 2 ( 50 ml ) and washed with sodium carbonate ( 10 % aq , 3 × 40 ml ). the ch 2 cl 2 layer was dried over sodium sulfate , filtered , the solvent removed in vacuo and the solid residue purified by flash column chromatography ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) to give the product 16 as a white solid ( 0 . 15 g , 63 %), r f = 0 . 5 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ). 1 h nmr ( cdcl 3 ) d 6 . 85 ( s , 1h , aryl ), 6 . 80 ( m , 2h , aryl ), 5 . 34 ( t , 4h , olefinic ), 3 . 97 ( q , 4h , och 2 ), 3 . 69 ( s , 2h , benzylic ), 2 . 67 ( t , 2h , c h 2 nh 2 ), 2 . 60 ( t , 2h , c h 2 nh 2 ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 80 ( m , 4h , c h 2 ch 2 o ), 1 . 54 - 1 . 18 ( m , 56h , 28 × ch 2 ), 0 . 88 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 149 . 31 , 148 . 20 , 133 . 55 , 130 . 11 , 130 . 00 , 120 . 59 , 114 . 14 , 114 . 07 , 69 . 70 , 69 . 50 , 54 . 24 , 49 . 83 , 42 . 60 , 34 . 20 , 32 . 28 , 30 . 46 , 30 . 16 , 29 . 92 , 29 . 81 , 29 . 71 , 29 . 67 , 27 . 73 , 27 . 60 , 27 . 22 , 26 . 44 , 23 . 08 , 14 . 53 ; hrms ( fab ): theory for c 51 h 95 n 2 o 2 ( m + 1 ), 767 . 7394 ; found ( m + 1 ), 767 . 7377 . anal . ( c 51 h 94 n 2 o 2 . 0 . 3h 2 o ): c , h , n . to a vigorously stirred solution of diaminodioxaoctane ( 0 . 23 g , 1 . 56 mmol , 5 equiv ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ) was added a solution of the aldehyde 8a ( 0 . 20 g , 0 . 33 mmol ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ), dropwise over 1 hour . the resulting mixture was stirred at rt under an atmosphere of nitrogen overnight . nmr showed the reaction to be complete when there was no aldehyde peak present in the nmr spectrum . the solvent was removed in vacuo and the crude imine dissolved in ch 2 cl 2 / meoh ( 1 : 1 , 10 ml ). the solution was cooled to 0 ° c . and nabh 4 ( 60 mg , 1 . 58 mmol ) was added in 15 mg portions over 30 minutes . tlc ( 10 % etac / hexane ) showed the reaction to be complete after stirring overnight . the solvent was removed in vacuo and the crude oil dissolved in ch 2 cl 2 ( 50 ml ) and washed with sodium carbonate ( 10 % aq , 3 × 40 ml ). the ch 2 cl 2 layer was dried over anhydrous na 2 so 4 , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ) to give the product 17 as a clear oil ( 0 . 168 g , 70 %), r f = 0 . 45 ( 1 : 10 : 89 nh 4 oh : meoh : chcl 3 ). 1 h nmr ( cdcl 3 ) d 6 . 87 ( s , 1h , aryl ), 6 . 81 ( m , 2h , aryl ), 5 . 34 ( t , 4h , olefinic ), 3 . 97 ( q , 4h , och 2 ), 3 . 72 ( s , 2h , benzylic ), 3 . 61 ( m , 6h , och 2 ), 3 . 61 ( t , 2h , och 2 ), 2 . 86 ( t , 2h , c h 2 nh 2 ), 2 . 81 ( t , 2h , c h 2 nh 2 ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 80 ( m , 4h , c h 2 ch 2 o ), 1 . 54 - 1 . 17 ( m , 44h , 22 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 149 . 31 , 148 . 25 , 133 . 11 , 130 . 09 , 129 . 99 , 120 . 73 , 114 . 26 , 114 . 07 , 73 . 57 , 70 . 85 , 70 . 57 , 70 . 52 , 69 . 69 , 69 . 51 , 54 . 02 , 48 . 93 , 42 . 01 , 32 . 28 , 30 . 15 , 29 . 91 , 29 . 80 , 29 . 70 , 29 . 67 , 27 . 59 , 26 . 45 , 26 . 42 , 23 . 07 , 14 . 53 ; hrms ( fab ): theory for c 49 h 91 n 2 o 4 ( m + 1 ), 771 . 6979 ; found ( m + 1 ), 771 . 6995 ; anal . ( c 49 h 90 n 2 o 4 ): c , h , n . to a vigorously stirred solution of the amine 12 ( 0 . 135 g , 0 . 38 mmol , 1 . 2 equiv ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ) was added a solution of the aldehyde 8a ( 0 . 20 g , 0 . 33 mmol ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ), dropwise over 20 minutes . the resulting mixture was stirred at rt under an atmosphere of nitrogen overnight . nmr showed the reaction to be complete when there was no aldehyde peak present in the nmr spectrum . the solvent was removed in vacuo and the crude imine dissolved in ch 2 cl 2 / meoh ( 1 : 1 , 10 ml ). the solution was cooled to 0 ° c . and sodium borohydride ( 60 mg , 1 . 58 mmol ) was added in 15 mg portions over 30 minutes . tlc ( 10 % etac / hexane ) showed the reaction to be complete after stirring overnight . the solvent was removed in vacuo and the crude oil dissolved in ch 2 cl 2 ( 50 ml ) and washed with sodium carbonate ( 10 % aq , 3 × 40 ml ). the ch 2 cl 2 layer was dried over sodium sulphate , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 3 % meoh / chcl 3 ) to give the product 18 as a clear oil ( 0 . 24 g , 78 %), r f = 0 . 3 ( 3 % meoh / chcl 3 ). 1 h nmr ( cdcl 3 ) d 6 . 85 ( s , 1h , aryl ), 6 . 80 ( m , 2h , aryl ), 5 . 34 ( t , 4h , olefinic ), 4 . 60 ( br s , 1h , nhco ), 3 . 97 ( q , 4h , och 2 ), 3 . 69 ( s , 2h , benzylic ), 3 . 13 ( m , 6h , ch 2 ) 2 . 63 ( t , 2h , c h 2 nh ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 80 ( m , 4h , c h 2 ch 2 o ), 1 . 67 - 1 . 15 ( m , 70h , 26 × ch 2 , 6 × ch 3 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 156 . 17 , 155 . 71 , 149 . 33 , 148 . 25 , 133 . 31 , 130 . 11 , 130 . 00 , 120 . 62 , 114 . 15 , 114 . 08 , 79 . 45 , 69 . 70 , 69 . 51 , 54 . 13 , 49 . 42 , 47 . 34 , 46 . 89 , 40 . 58 , 32 . 28 , 30 . 14 , 29 . 91 , 29 . 80 , 29 . 70 , 29 . 67 , 28 . 85 , 28 . 79 , 27 . 79 , 27 . 70 , 27 . 59 , 26 . 42 , 23 . 07 , 14 . 53 ; hrms ( fab ): theory for c 61 h 112 n 3 o 6 ( m + 1 ), 982 . 8551 ; found ( m + 1 ), 982 . 8510 ; anal . ( c 61 h 111 n 3 o 6 ): c , h , n . to a vigorously stirred solution of the amine 14 ( 0 . 20 g , 0 . 38 mmol , 1 . 2 equiv ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ) was added a solution of the aldehyde 8a ( 0 . 20 g , 0 . 33 mmol ) in ch 2 cl 2 / meoh ( 3 : 1 , 5 ml ), dropwise over 20 minutes . the resulting mixture was stirred at rt under an atmosphere of nitrogen overnight . nmr showed the reaction to be complete when there was no aldehyde peak present in the spectrum . the solvent was removed in vacuo and the crude imine dissolved in ch 2 cl 2 / meoh ( 1 : 1 , 10 ml ). the solution was cooled to 0 ° c . and sodium borohydride ( 60 mg , 1 . 58 mmol ) was added in 15 mg portions over 30 minutes . tlc ( 10 % etac / hexane ) showed the reaction to be complete after stirring overnight . the solvent was removed in vacuo and the crude oil dissolved in ch 2 cl 2 ( 50 ml ) and washed with sodium carbonate ( 10 % aq , 3 × 40 ml ). the ch 2 cl 2 layer was dried over sodium sulphate , filtered , the solvent removed in vacuo and the oily residue purified by flash column chromatography ( 3 % meoh / chcl 3 ) to give the product 19 as a clear oil ( 0 . 319 g , 88 %), r f = 0 . 25 ( 3 % meoh / chcl 3 ). 1 h nmr ( cdcl 3 ) d 6 . 85 ( s , 1h , aryl ), 6 . 80 ( m , 2h , aryl ), 5 . 34 ( t , 4h , olefinic ), 4 . 60 ( br s , 1h , nhco ), 3 . 97 ( q , 4h , och 2 ), 3 . 70 ( s , 2h , benzylic ), 3 . 14 ( m , 10h , 5 × ch 2 ) 2 . 64 ( t , 2h , c h 2 nh ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 80 ( m , 4h , c h 2 ch 2 o ), 1 . 59 - 1 . 14 ( m , 83h , 28 × ch 2 , 9 × ch 3 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 156 . 12 , 155 . 69 , 149 . 33 , 148 . 25 , 133 . 28 , 130 . 10 , 129 . 99 , 120 . 61 , 114 . 14 , 114 . 07 , 79 . 40 , 69 . 69 , 69 . 50 , 54 . 15 , 49 . 47 , 47 . 24 , 46 . 98 , 40 . 57 , 32 . 27 , 30 . 14 , 29 . 90 , 29 . 80 , 29 . 69 , 29 . 66 , 28 . 85 , 28 . 79 , 27 . 77 , 27 . 58 , 26 . 44 , 23 . 07 , 14 . 53 ; hrms ( fab ): theory for c 70 h 129 n 4 o 8 ( m + 1 ), 1153 . 9810 ; found ( m + 1 ), 1153 . 9840 ; anal . calcd ( c 70 h 128 n 4 o 8 ): c , h , n . a solution of 3 , 4 - bis - octadec - 9 - enyloxy - benzoic acid 8c ( 0 . 2 g , 0 . 31 mmol ) in 2 : 1 dichloromethane / benzene ( 15 ml ) was stirred at 0 ° c . for 10 min . anhydrous dmf ( 2 drops ) and oxalyl chloride ( 0 . 3 ml ) were added in sequence and the mixture was stirred for 1 h at 0 ° c . the solution was concentrated in vacuo to give the crude acid chloride , 8d . crude 8d was dissolved in ch 2 cl 2 ( 10 ml ) and added dropwise to a solution of the amine 12 ( 0 . 132 g , 0 . 37 mmol , 1 . 2 equiv .) dissolved in ch 2 cl 2 ( 10 ml ) and 1m naoh ( 10 ml ) that had been cooled to 0 ° c . for 15 minutes . the reaction was stirred overnight under n 2 at room temperature . the water layer was separated off . the ch 2 cl 2 layer was washed with na 2 co 3 ( 10 % aq . 3 × 20 ml ), dried over na 2 so 4 , filtered , removed in vacuo and the oily residue purified by flash column chromatography ( 30 % etac / hexane ) to give the product 20 as a clear oil ( 0 . 261 g , 86 %), r f = 0 . 25 ( 30 % etac / hexane ); 1 h nmr ( 500 mhz , cdcl 3 ) d 7 . 50 - 7 . 20 ( m , 2h , phenyl ), 6 . 90 ( br s , 0 . 5h , nhco ), 6 . 81 ( d , 1h , phenyl ), 6 . 41 ( br s , 0 . 5h , nhco ), 5 . 35 ( t , 4h , olefinic ), 4 . 68 ( m , 1h , nhco ), 3 . 99 ( 2t , 4h , 2 × och 2 ), 3 . 44 ( m , 2h , ch 2 ), 3 . 11 ( m , 6h , 3 × ch 2 ), 2 . 00 ( m , 8h , ch 2 c ═ c ), 1 . 79 ( m , 4h , c h 2 ch 2 o ), 1 . 53 - 1 . 17 ( m , 70h , 26 × ch 2 , 6 × ch 3 ), 0 . 88 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 167 . 32 , 156 . 18 , 151 . 84 , 148 . 98 , 130 . 11 , 130 . 09 , 127 . 32 , 113 . 02 , 112 . 39 , 79 . 62 , 79 . 32 , 69 . 52 , 69 . 36 , 47 . 09 , 40 . 49 , 39 . 92 , 32 . 26 , 30 . 13 , 29 . 89 , 29 . 76 , 29 . 68 , 29 . 65 , 29 . 62 , 29 . 59 , 29 . 50 , 28 . 84 , 28 . 77 , 27 . 79 , 27 . 58 , 26 . 38 , 26 . 36 , 23 . 06 , 14 . 51 ; hrms ( fab ): theory for c 61 h 109 n 3 o 7 ( m + na ), 1018 . 8158 ; found ( m + na ), 1018 . 8123 ; anal . ( c 61 h 109 n 3 o 7 ): c , h , n . a solution of 3 , 4 - bis - octadec - 9 - enyloxy - benzoic acid 8c ( 0 . 2 g , 0 . 31 mmol ) in 2 : 1 dichloromethane / benzene ( 15 ml ) was stirred at 0 ° c . for 10 min . anhydrous dmf ( 2 drops ) and oxalyl chloride ( 0 . 3 ml ) were added in sequence and the mixture was stirred for 1 h at 0 ° c . the solution was concentrated in vacuo to give the crude acid chloride 8d . the acid chloride 8d was dissolved in ch 2 cl 2 ( 10 ml ) and added dropwise to a solution of the amine 14 ( 0 . 195 g , 0 . 37 mmol , 1 . 2 equiv .) dissolved in ch 2 cl 2 ( 10 ml ) and 1m naoh ( 10 ml ) that had been cooled to 0 ° c . for 15 minutes . the reaction was stirred overnight under n 2 at room temperature . the water layer was separated off . the ch 2 cl 2 layer was washed with na 2 co 3 ( 10 % aq . 3 × 20 ml ), dried over na 2 so 4 , filtered , removed in vacuo and the oily residue purified by flash column chromatography ( 30 % etac / hexane ) to give the product 21 as a clear oil ( 0 . 295 g , 83 %), r f = 0 . 20 ( 30 % etac / hexane ); 1 h nmr ( 500 mhz , cdcl 3 ) d 7 . 42 - 7 . 20 ( m , 2h , phenyl ), 6 . 82 ( br s , 0 . 5h , nhco ), 6 . 80 ( d , 1h , phenyl ), 6 . 39 ( br s , 0 . 5h , nhco ), 5 . 35 ( t , 4h , olefinic ), 4 . 65 ( m , 1h , nhco ), 3 . 99 ( 2 × t , 4h , 2 × och 2 ), 3 . 42 ( m , 2h , ch 2 ), 3 . 13 ( m , 10h , 5 × ch 2 ), 2 . 00 ( m , 8h , ch 2 c ═ c ), 1 . 79 ( m , 4h , c h 2 ch 2 o ), 1 . 53 - 1 . 17 ( m , 83h , 28 × ch 2 , 9 × ch 3 ), 0 . 88 ( t , 6h , ch 3 ); 13 c nmr ( cdcl 3 ) d 167 . 35 , 156 . 16 , 155 . 73 , 151 . 88 , 148 . 99 , 130 . 13 , 130 . 11 , 127 . 37 , 113 . 05 , 112 . 41 , 79 . 54 , 69 . 54 , 69 . 38 , 47 . 03 , 40 . 58 , 39 . 97 , 32 . 27 , 30 . 14 , 29 . 90 , 29 . 77 , 29 . 69 , 29 . 66 , 29 . 63 , 29 . 60 , 29 . 51 , 28 . 85 , 28 . 79 , 27 . 78 , 27 . 59 , 26 . 40 , 26 . 37 , 23 . 07 , 14 . 53 ; hrms ( fab ): theory for c 70 h 126 n 4 o 9 ( m + na ), 1189 . 9407 ; found ( m + na ), 1189 . 9365 ; anal . ( c 70 h 126 n 4 o 8 ): c , h , n . a concentrated solution of the amine 15 ( 0 . 158 g , 0 . 19 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 4 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 22 as an off white powder ( 0 . 15 g , 98 %); 1 h nmr ( 500 mhz , cdcl 3 ) d 9 . 50 ( br s , 2h , r 2 n + h 2 ), 8 . 22 ( br s , 3h , rn + h 3 ), 7 . 24 ( s , 1h , aryl ), 7 . 05 ( d , 1h , aryl ), 6 . 82 ( d , 1h , aryl ), 5 . 34 ( m , 4h , olefinic ), 4 . 02 ( q , 4h , och 2 ), 3 . 93 ( t , 2h , benzylic ), 3 . 07 ( m , 2h , ch 2 ) 2 . 81 ( m , 2h , ch 2 ), 2 . 01 ( m , 8h , ch 2 c ═ c ), 1 . 95 ( m , 2h , ch 2 ), 1 . 85 ( m , 2h , ch 2 ), 1 . 78 ( m , 4h , c h 2 ch 2 o ), 1 . 45 ( m , 4h , 2 × ch 2 ), 1 . 38 - 1 . 20 ( m , 40h , 20 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( 300 mhz , cdcl 3 ) d 150 . 00 , 149 . 37 , 130 . 12 , 130 . 09 , 129 . 95 , 122 . 58 , 115 . 81 , 113 . 49 , 69 . 63 , 69 . 38 , 51 . 44 , 51 . 06 , 45 . 80 , 39 . 63 , 32 . 29 , 30 . 24 , 30 . 21 , 30 . 16 , 30 . 00 , 29 . 93 , 29 . 71 , 27 . 61 , 26 . 63 , 26 . 49 , 24 . 83 , 23 . 75 , 23 . 08 , 14 . 54 ; hrms ( fab ): theory for c 47 h 87 n 2 o 2 ( m + 1 ), 711 . 6768 ; found ( m + 1 ), 711 . 6827 ; anal . ( c 47 h 88 cl 2 n 2 o 4 . 1 . 2h 2 o ): c , h , n . a concentrated solution of the amine 16 ( 0 . 097 g , 0 . 13 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 4 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 23 as an off white powder ( 0 . 104 g , 98 %); 1 h nmr ( cdcl 3 ) d 7 . 24 ( s , 1h , aryl ), 7 . 05 ( d , 1h , aryl ), 6 . 85 ( d , 1h , aryl ), 5 . 35 ( m , 4h , olefinic ), 4 . 07 ( q , 4h , och 2 ), 3 . 95 ( t , 2h , benzylic ), 3 . 07 ( m , 2h , ch 2 ) 2 . 79 ( m , 2h , ch 2 ), 2 . 05 ( m , 8h , ch 2 c ═ c ), 1 . 82 ( m , 8h , c h 2 ch 2 o , 2 × ch 2 ), 1 . 58 - 1 . 21 ( m , 52h , 26 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( 300 mhz , cdcl 3 ) d 149 . 36 , 148 . 78 , 130 . 85 , 130 . 33 , 129 . 74 , 122 . 23 , 115 . 56 , 113 . 45 , 69 . 56 , 69 . 32 , 51 . 56 , 51 . 15 , 45 . 96 , 39 . 86 , 32 . 53 , 30 . 44 , 30 . 44 , 30 . 23 , 30 . 12 , 30 . 00 , 29 . 71 , 27 . 61 , 27 . 51 , 27 . 10 , 26 . 83 , 26 . 69 , 24 . 82 , 23 . 77 , 23 . 08 , 14 . 54 ; hrms ( fab ): theory for c 51 h 95 n 2 o 2 ( m + 1 ), 767 . 7394 ; found ( m + 1 ), 767 . 7419 ; anal . ( c 51 h 96 cl 2 n 2 o 2 . 0 . 3h 2 o ): c , h , n . a concentrated solution of the amine 17 ( 0 . 14 g , 0 . 18 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 4 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 24 as an off white powder ( 0 . 15 g , 98 %); 1 h nmr ( cdcl 3 ) d 7 . 27 ( s , 1h , aryl ), 7 . 00 ( d , 1h , aryl ), 6 . 82 ( d , 1h , aryl ), 5 . 33 ( m , 4h , olefinic ), 4 . 14 ( m , 2h , benzylic ), 4 . 01 ( t , 2h , och 2 ), 3 . 95 ( t , 2h , och 2 ), 3 . 87 ( t , 2h , ch 2 ), 3 . 78 ( t , 2h , ch 2 ), 3 . 70 ( t , 4h , ch 2 ), 3 . 25 ( t , 2h , ch 2 ) 2 . 93 ( m , 2h , ch 2 ), 2 . 02 ( m , 8h , ch 2 c ═ c ), 1 . 89 ( m , 4h , c h 2 ch 2 o ), 1 . 53 - 1 . 18 ( m , 44h , 22 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( 300 mhz , cdcl 3 ) d 149 . 91 , 149 . 62 , 130 . 12 , 130 . 00 , 123 . 28 , 122 . 61 , 115 . 46 , 113 . 63 , 70 . 22 , 70 . 09 , 69 . 64 , 69 . 40 , 66 . 50 , 65 . 98 , 51 . 06 , 45 . 08 , 40 . 11 , 32 . 28 , 30 . 15 , 29 . 92 , 29 . 70 , 27 . 60 , 26 . 54 , 26 . 43 , 23 . 07 , 14 . 54 ; hrms ( fab ): theory for c 49 h 91 n 2 o 4 ( m + 1 ), 771 . 6973 ; found ( m + 1 ), 771 . 6967 ; anal . ( c 49 h 92 cl 2 n 2 o 4 . 0 . 4h 2 o ): c , h , n . a concentrated solution of the amine 18 ( 0 . 17 g , 0 . 17 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 4 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 25 as a white powder ( 0 . 151 g , 98 %); 1 h nmr ( cdcl 3 / ch 3 od 9 : 1 ) d 7 . 13 ( s , 1h , aryl ), 7 . 02 ( d , 1h , aryl ), 6 . 85 ( d , 1h , aryl ), 5 . 33 ( m , 4h , olefinic ), 4 . 05 ( m , 2h , benzylic ), 3 . 97 ( t , 2h , och 2 ), 3 . 04 ( m , 6h , 3 × ch 2 ) 2 . 93 ( m , 2h , ch 2 ), 2 . 02 ( m , 8h , ch 2 c ═ c ), 1 . 90 ( m , 8h , 4 × ch 2 ), 1 . 81 ( m , 4h , c h 2 ch 2 o ), 1 . 47 ( m , 4h , 2 × ch 2 ), 1 . 40 - 1 . 17 ( m , 40h , 20 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( 300 mhz , d 2 o ) δ 148 . 94 , 148 . 33 , 129 . 26 , 129 . 19 , 123 . 64 , 122 . 52 , 113 . 10 , 68 . 50 , 68 . 39 , 49 . 96 , 45 . 98 , 45 . 43 , 39 . 86 , 31 . 28 , 29 . 18 , 29 . 11 , 28 . 93 , 28 . 86 , 28 . 70 , 28 . 62 , 26 . 64 , 26 . 59 , 25 . 68 , 25 . 62 , 24 . 00 , 22 . 54 , 22 . 46 , 22 . 11 , 13 . 89 ; hrms ( fab ): theory for c 51 h 96 n 3 o 2 ( m + 1 ), 782 . 7497 ; found ( m + 1 ), 782 . 7495 ; anal . ( c 51 h 95 cl 3 n 3 o 2 . 0 . 6h 2 o ): c , h , n . a concentrated solution of the amine 19 ( 0 . 253 g , 0 . 22 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 5 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 26 as a white powder ( 0 . 215 g , 98 %); 1 h nmr ( cd 3 od ) δ 7 . 13 ( s , 1h , aryl ), 7 . 04 ( d , 1h , aryl ), 6 . 98 ( d , 1h , aryl ), 5 . 33 ( t , 4h , olefinic ), 4 . 12 ( m , 2h , benzylic ), 4 . 03 ( t , 2h , och 2 ), 3 . 99 ( t , 2h , och 2 ), 3 . 08 ( m , 10h , 5 × ch 2 ), 2 . 98 ( t , 2h , ch 2 ), 2 . 02 ( m , 8h , ch 2 c ═ c ), 1 . 89 ( m , 16h , 8 × ch 2 ), 1 . 47 ( m , 4h , 2 × ch 2 ), 1 . 42 - 1 . 19 ( m , 40h , 20 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); 13 c nmr ( 300 mhz , d 2 o ) δ 149 . 74 , 148 . 90 , 129 . 62 , 129 . 46 , 123 . 92 , 123 . 71 , 69 . 36 , 68 . 88 , 51 . 12 , 46 . 68 , 38 . 99 , 32 . 14 , 29 . 97 , 29 . 81 , 29 . 60 , 27 . 36 , 26 . 53 , 24 . 69 , 24 . 22 , 23 . 14 , 23 . 06 , 22 . 84 , 14 . 05 ; hrms ( fab ): theory for c 55 h 105 n 4 o 2 ( m + 1 ), 853 . 8238 ; found ( m + 1 ), 853 . 8264 ; anal . ( c 55 h 109 cl 4 n 4 o 2 . 0 . 6h 2 o ): c , h , n . a concentrated solution the amide 20 ( 0 . 22 g , 0 . 22 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 5 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 27 as a white powder ( 0 . 19 g , 98 %); 1 h nmr ( cdcl 3 ) d 8 . 95 ( br s , 2h , r 2 n + h 2 ), 8 . 20 ( br s , 3h , rn + h 3 ), 7 . 45 ( m , 2h , aryl ), 6 . 78 ( m , 1h , aryl ), 5 . 33 ( m , 4h , olefinic ), 3 . 95 ( m , 4h , och 2 ), 3 . 38 ( m , 2h , ch 2 ), 3 . 11 ( m , 2h , ch 2 ), 3 . 01 ( m , 4h , 2 × ch 2 ), 2 . 08 - 1 . 89 ( m , 14h , 7 × ch 2 ), 1 . 71 ( m , 6h , 3 × ch 2 ), 1 . 47 - 1 . 18 ( m , 44h , 20 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); hrms ( fab ): theory for c 51 h 94 n 3 o 3 ( m + 1 ), 796 . 7290 ; found ( m + 1 ), 796 . 7249 ; anal . ( c 51 h 95 cl 2 n 3 o 3 . 1 . 6h 2 o ): c , h , n . a concentrated solution of the amide 21 ( 0 . 22 g , 0 . 19 mmol ) in ethyl acetate was added cooled to 0 ° c . a total of 5 ml of a freshly prepared saturated solution of hcl in ethyl acetate was added dropwise and the solution stirred for 1 h at room temperature during which time a white precipitate formed . the ethyl acetate was removed in vacuo and the residue co - evaporated with ethyl acetate and chloroform to give the product 28 as a white powder ( 0 . 18 g , 98 %); 1 h nmr ( cdcl 3 / ch 3 od 9 : 1 ) d 7 . 42 ( m , 2h , aryl ), 6 . 78 ( m , 1h , aryl ), 5 . 33 ( m , 4h , olefinic ), 4 . 03 ( m , 4h , och 2 ), 3 . 41 ( m , 2h , ch 2 ), 3 . 01 ( m , 10h , 5 × ch 2 ), 2 . 08 - 1 . 66 ( m , 24h , 12 × ch 2 ), 1 . 71 ( m , 6h , 3 × ch 2 ), 1 . 56 - 1 . 13 ( m , 44h , 20 × ch 2 ), 0 . 89 ( t , 6h , ch 3 ); hrms ( fab ): theory for c 55 h 105 n 4 o 3 ( m + 1 ), 867 . 8025 ; found ( m + 1 ), 867 . 8022 ; anal . calcd for ( c 55 h 105 cl 3 n 4 o 3 . 1 . 5h 2 o ): c , h , n . advantages of the present invention include the surprising discovery that compound (# 26 ) is much more efficient in delivering dna into a target cell than the commercial lipofectamine ™ transfection agent . 39 this efficiency , coupled with the straightforward synthesis we developed for this material makes this product commercially viable to pursue . uses of compound # 26 and other compounds herein disclosed include use as smart anticancer agents which recognize and selectively deliver “ cargoes ” to specific cell types . the cargoes could be either ‘ corrective ’ dna which repairs or replaces the cancerous genetic lesion and allows for the cell to heal itself . alternatively , one could deliver toxic compounds such as the plant toxin saporin , which targets cell protein synthesis ability . blocking protein synthesis is a toxic event for the cell . therefore , one could envision delivering toxic compounds selectively to cancer cells to kill them , even in the presence of normal healthy cells . coadministration of the agent saporin with a lipopolyamine was shown to promote entry of the agent into cells by the cell &# 39 ; s own polyamine transporter system . these results were published in febs j ., 274 ( 2007 ) 4825 - 4836 , a paper which is noted above and which has been incorporated herein by reference in its entirety . a deficiency in previously known compounds is their lower efficiency of transfection and their inability to target specific cell types via the polyamine transport system . r 1 and r 2 may be the same or may be different and comprise a long hydrocarbon chain which is between six and eighteen carbons long and either contains or does not contain units of unsaturation . for example , the chain could be derived from the c 18 oleic acid and attached via an ester linkage or via an ether linkage and an oleyl group . the chains can range from lauryl , stearic , myristic and oleic acids . r 3 is a polycation comprising a polyamine side chain . these aliphatic polyamine side chains can range from mono amines to octaamines with carbon spacer groups ranging from two to six in between the nitrogen centers . these can also be branched systems as well . for example : — nh —( ch 2 ) a nh ( ch 2 ) b nh ( ch 2 ) c nh ( ch 2 ) d nh ( ch 2 ) e nh ( ch 2 ) f nh 2 — nh —( ch 2 ) a nh ( ch 2 ) b nh ( ch 2 ) c nh ( ch 2 ) d nh ( ch 2 ) e nh ( ch 2 ) f nh ( ch 2 ) g nh 2 note : other polycationic moieties may be used in place of the polyamine . more biofriendly cations like polyhistine , polyarginine , polylysine or polyornithine peptides could be used . these could be in the all l , all d or mixed l , d forms . in addition , the hydrochloride salts as well as other pharmaceutically acceptable salt forms of compound 1 could be used . r 1 and r 2 are the same or different and are comprised of a long hydrocarbon chain which is between six and eighteen carbons long and either contains or does not contain units of unsaturation . for example , the chain could be derived from the c 18 oleic acid and attached via an ester linkage or via an ether linkage and an oleyl group . the chains can range from lauryl , stearic , myristic and oleic acids . r 3 is a polycations comprised of a polyamine side chain . these aliphatic polyamine side chains can range from mono amines to octaamines with carbon spacer groups ranging from two to six in between the nitrogen centers . these can also be branched systems as well . — nh —( ch 2 ) a nh ( ch 2 ) b nh ( ch 2 ) c nh ( ch 2 ) d nh ( ch 2 ) e nh ( ch 2 ) f nh 2 — nh —( ch 2 ) a nh ( ch 2 ) b nh ( ch 2 ) c nh ( ch 2 ) d nh ( ch 2 ) e nh ( ch 2 ) f nh ( ch 2 ) g nh 2 note : other polycationic moieties may be used in place of the polyamine . more biofriendly cations like poly - histine , polyarginine , polylysine or polyornithine peptides could be used . these could be in the all l , all d or mixed l , d forms . in addition , the hydrochloride salts as well as other pharmaceutically acceptable salt forms of 2 could be used as well . iii . pharmaceutical compositions of either 1 and / or 2 along with a neutral lipid ( e . g ., dope ) and a therapeutic agent such as a dna plasmid , sirna ( gene silencing agent ), or toxic compound . the dna plasmid could be a corrective gene to repair or replace a damaged gene within the cell . the sirna could be a rna molecule designed to silence a bad gene to provide a positive therapeutic outcome . the toxic compound could be a toxic drug that kills the cell like methotrexate , doxorubicin , cis platin , taxol , or other toxins like saporin or ricin which target specific critical cell functions . summary . the invention will allow for the selective delivery of therapeutic agents to cancer cells via the polyamine transport system . the technology is predicated upon the fact that certain cancer cells readily import polyamines from their environment . this property provides an opportunity to target these cells via their selective uptake of these cationic materials . the patented materials can also delivery other therapeutic agents to cells such as dna plasmids , rna for gene silencing therapies , and proteins which repair or destroy cells . the technology also allows for enhanced transfection of cells by utilizing this special uptake pathway to deliver new genes , nucleic acids and proteins into cells . accordingly , in the drawings and specification there have been disclosed typical preferred embodiments of the invention and although specific terms may have been employed , the terms are used in a descriptive sense only and not for purposes of limitation . the invention has been described in considerable detail with specific reference to these illustrated embodiments . it will be apparent , however , that various modifications and changes can be made within the spirit and scope of the invention as described in the foregoing specification and as defined in the appended claims . 1 . lander , e . s . ; 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