paclitaxel ( ptx ) is 
 one of the first - line chemotherapeutics used 
 to treat patients with breast , ovarian , nonsmall cell lung cancer , 
 and advanced forms of kaposi s sarcoma . the mechanism involves 
 interfering with the normal breakdown of microtubules during cell 
 division . successful application of ptx 
 in the clinic has been limited by its poor water solubility and the 
 systemic toxicity . taxol is a cremophor el / ethanol formulation of 
 ptx that has been used in the clinic . however , cremophor el can cause 
 hyperactivity reactions , neuropathy , and other serious side effects . thus , there is a need to develop an alternative 
 delivery system for ptx . various macromolecular delivery systems such 
 as liposomes , dendrimers , and nanoparticles are under investigation , 
 among which polymeric micelles have gained considerable attention 
 owing to ease in preparation and their very small sizes ( 10100 
 nm ) . recent studies have substantiated that sub-100 nm was critical for 
 a delivery system to achieve effective tumor targeting . our group has previously developed peg - fts as a dual - functional 
 carrier for the delivery of poorly water - soluble anticancer drugs . this system was constructed by coupling two 
 molecules of s - trans , trans - farnesylthiosalicylic acid ( fts ) to poly(ethylene glycol ) ( peg , mw = 5000 ) through an ester linkage ( peg5k - fts2 ) . different from most reported delivery 
 systems that use  inert  excipients , our system employ 
 water - insoluble drug fts as the hydrophobic region of polymeric micelles . it can inhibit both 
 oncogenically activated ras and growth factor receptor - mediated ras 
 activation , resulting in the inhibition of ras - dependent tumor growth . preliminary study showed that the antitumor activity of fts was 
 well retained following coupling to peg5k . furthermore , 
 peg5k - fts2 readily formed small - sized micelles 
 ( 2030 nm ) that are effective in loading and delivering ptx . in vivo study demonstrated that the antitumor activity of 
 the ptx - loaded peg5k - fts2 micelles was significantly 
 higher than that of taxol . recent 
 studies from us and others have shown that the vitamin e - based 
 micellar system could be significantly improved via modulating the 
 peg motifs and the molar ratio of peg / vitamin e. in another 
 study with peg - embelin system , we showed that a conjugate with two 
 embelin molecules linked to peg was significantly more effective than 
 the conjugate with one embelin molecule coupled to peg . this has prompted us to carry out a similar study with peg - fts system . four peg - fts conjugates that vary in the molecular weight of peg ( peg2k vs peg5k ) and the molar ratio of peg / fts ( 1/2 
 vs 1/4 ) have been developed . we demonstrated that peg5k - fts4 formed the most stable mixed micelles with ptx among 
 the four peg - fts conjugates . furthermore , ptx formulated in peg5k - fts4 micelles was the most effective formulation 
 in inhibiting the tumor growth in vivo . phosphate buffered saline ( dpbs ) was purchased from lonza ( md , u. 
 s. a. ) . poly(ethylene glycol ) methyl ether ( meo  peg  oh , mw = 2000 , 5000 kda ) , dimethyl sulfoxide ( dmso ) , 
 succinate anhydride , diethanolamine , 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl 
 tetrazolium bromide ( mtt ) , trypsin - edta solution , triton x-100 , and 
 dulbecco s modified eagle s medium ( dmem ) were all purchased 
 from sigma - aldrich ( mo , u. s. a. ) . fetal bovine serum ( fbs ) and penicillin n - hydroxysuccinimide ( nhs ) and dicyclohexylcarbodiimide ( dcc ) were 
 purchased from alfa aesar ( ma , u. s. a. ) . 4-(dimethylamino ) pyridine 
 ( dmap ) was purchased from calbiochem - novabiochem corporation ( ca , 
 u. s. a. ) . all cell lines were cultured in dmem 
 containing 5% fbs and 1% penicillin - streptomycin at 37 c in 
 a humidified 5% co2 atmosphere . peg5k - fts4 was prepared 
 via solution phase condensation reactions ( figure 1 ) . we started to synthesize two hydroxyl group terminated 
 peg monomethyl ether ( meo  peg5k peg5k(oh)2 by a facile chemical reaction with succinic anhydride and 
 dmap . to obtain four hydroxyl groups the polymer was precipitated and washed by ice - cold diethyl 
 ether and ethanol twice , respectively , and concentrated under vacuum . oh4 , fts , dcc , and dmap 
 were then dissolved in chloroform and allowed to react overnight at 
 room temperature . the solution was filtered and precipitated in ice - cold 
 diethyl ether and ethanol twice respectively , and concentrated under 
 vacuum . the powder was then dissolved in water and filtered through 
 a filter with a pore size of 0.22 m . ptx 
 ( 10 mm in chloroform ) and four peg - fts conjugates ( 10 mm in chloroform ) 
 were mixed at various carrier / drug ratios . the organic solvent was 
 removed by nitrogen flow to form a thin film of drug / carrier mixture . the film was dried under vacuum for 1 h to remove the remaining solvent . dpbs was added to hydrate the thin film and the drug - loaded micelles 
 were formed . unincorporated ptx ( precipitate ) was removed by filtering 
 through a syringe filter ( pore size : 0.22 m ) . the particle size and 
 zeta potential of micelles were measured by 
 a zetasizer ( dls ) ( zetasizer nano zs instrument , malvern , worcestershire , 
 u. k. ) . the morphology and size distribution of drug - free or drug - loaded 
 peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles were observed using transmission electron microscopy 
 ( tem ) . the copper grid was immersed 
 in a drop of sample solution and stained with 1% uranyl acetate . imaging 
 was performed at room temperature on jeol jem-1011 . the critical 
 micelle concentrations ( cmc ) of four peg - fts micelles were determined 
 by using pyrene as a fluorescence probe . four peg - fts conjugates , peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 , were prepared in chloroform at 1.2 mg / ml , 
 and various amounts were added to nine separate vials . then 10 l 
 of 1.8  10 m of pyrene in chloroform was 
 added to each vial and the solution was mixed well . the organic solvent 
 was removed by oil pump , and then 3 ml of milli - q water was added 
 to each vial . the final pyrene concentration was 6  10 m with the four peg - fts conjugate concentrations ranging from 0.0001 
 to 0.5 mg / ml . the vials were kept on a shaker for 24 h at 37 c 
 to reach equilibrium before fluorescence measurement . the fluorescence 
 intensities of samples were measured at the excitation wavelength 
 of 334 nm and emission wavelength of 390 nm by synergy h1 hybrid multi - mode 
 microplate reader ( winooski , vt , u. s. a. ) . the cmc is determined 
 from the threshold concentration , where the sharp increase in pyrene 
 fluorescence intensity is observed . the ptx loading efficiency 
 was quantified by high performance liquid 
 chromatography ( hplc ) ( alliance 2695 - 2998 system ) as described previously . drug loading capacity ( dlc ) and drug loading 
 efficiency ( dle ) were calculated according to the following equation : the in vitro ptx release kinetics for the four 
 peg - fts micelles 
 was determined by a dialysis method according to our published protocol . briefly , ptx loaded peg - fts micelles at a concentration 
 of 0.5 mg ptx / ml were placed into a dialysis bag ( mw cutoff 14000 ) . the dialysis bag was incubated in 200 
 ml pbs containing 0.5% ( w / v ) tween 80 with gentle shaking at 37 c . the concentrations of ptx remaining in the dialysis bag at designated 
 time points were measured by hplc . hemolysis assay 
 was performed using fresh blood collected through cardiac puncture 
 from rats . red blood cells ( rbcs ) were 
 collected by centrifugation and washed with pbs three times . then 
 rbcs were diluted in pbs with a final concentration of 2% w / v . a total 
 of 1 ml of diluted rbc suspension was mixed with different concentrations 
 ( 0.2 and 1.0 mg / ml ) of four peg - fts micelles and pei , respectively , 
 and then incubated at 37 c in an incubator shaker for 4 h. the 
 mixtures were centrifuged , and supernatants were transferred into 
 a 96-well plate . the release of hemoglobin was determined at 540 nm 
 absorbance using a microplate reader . rbcs incubated with pbs and 
 triton x-100 ( 2% ) were used as the negative and positive controls , 
 respectively . the percentage of hemolysis of rbcs was calculated as 
 ( odsample  odnegative  control)/(odpositive  control  odnegative  control )  100% . the cytotoxicity 
 of ptx formulated in peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles was assessed with several cancer cell lines 
 and compared to free ptx in dmso , respectively . briefly , 4t1.2 ( 1000 
 cells / after 24 h of 
 incubation in dmem with 5% fbs and 1% streptomycin  penicillin , 
 the old medium was removed and the cells were incubated for 72 h in 
 the presence of indicated concentrations of ptx ( free or formulated 
 in four peg - fts micelles ) . a total of 100 l of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium 
 bromide ( mtt ) in dpbs ( 0.5 mg / ml ) was added to each well and cells 
 were further incubated for 2 h. mtt formazan was solubilized by dmso . 
 the absorbance in each well was measured by a microplate reader with 
 wavelength at 550 nm and reference wavelength at 630 nm . cell viability was calculated as [ ( odtreat  odblank)/(odcontrol  
 odblank )  100% ] . the cytotoxicity of peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles alone 
 was similarly tested in 4t1.2 , mcf-7 , and hct-116 cell line as described 
 above . ras protein expression level in hct-116 
 cells was evaluated by western blotting following our published method . briefly , hct-116 cells with 6070% confluency 
 in a 6-well plate were treated with four peg - fts conjugates for 20 
 h at a fts concentration of 40 m . female 
 balb / c mice , 46 weeks in age , 
 were purchased from charles river ( davis , ca , u. s. a. ) . all animals 
 were housed under pathogen - free conditions according to aaalac guidelines . all animal - related experiments were performed in full compliance with 
 institutional guidelines and approved by the animal use and care administrative 
 advisory committee at the university of pittsburgh . a syngeneic 
 murine breast cancer model ( 4t1.2 ) was used to examine the therapeutic 
 effect of different formulations of ptx . a total of 2  10 4t1.2 cells in 200 l of pbs were inoculated s.c . at 
 the right flank of female balb / c mice . treatments were started when 
 tumors in the mice reached a tumor volume of 50 mm and this day was designated as day 1 . on day 1 , these mice were 
 randomly divided into six groups ( n = 5 ) and administered 
 i.v . with pbs ( control ) , taxol ( 10 mg ptx / kg ) , ptx - loaded peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles 
 ( 10 mg of ptx / kg ) , respectively on days 1 , 3 , 5 , 8 , 10 , and 12 . tumor 
 sizes were measured with digital caliper three times a week and calculated 
 by the formula : ( l  w)/2 , where l is the longest and w is the shortest of the tumor diameters ( mm ) . to compare between 
 groups , relative tumor volume ( rtv ) was calculated at each measurement 
 time point ( where rtv equals to the tumor volume at a given time point 
 divided by the tumor volume prior to first treatment ) . mice were sacrificed 
 before the tumor reached 2000 mm . to monitor the 
 potential toxicity , the body weights of all mice from different groups 
 were measured every three days . in addition , serum levels of transaminases 
 ( ast , alt ) in the mice with different treatments were investigated 
 at the completion of the study . in all statistical analysis , the 
 significance level was set at a probability of p < 
 0.05 . all results were reported as the mean  standard deviation 
 ( sd ) . unless otherwise indicated . statistical analysis was performed 
 by student s t test for two groups , and one - way 
 anova for multiple groups , followed by newman  keuls test if p < 0.05 . peg5k - fts4 conjugate , containing 
 four molecules of fts coupled 
 to one molecule of peg5k via a labile ester linkage , was 
 developed by solution phase condensation reactions . h nmr spectra of peg5k - fts4 conjugate are shown 
 in supporting information figure s1d . the 
 intense peak at 3.66 ppm was attributable to the methylene protons 
 of peg . carbon chain signals and benzene ring signals of fts were 
 located at 1.52.2 ppm and 78 ppm , respectively . maldi - tof 
 suggested that four fts were successfully attached to peg5k ( supporting information figure s2d ) . we also synthesized peg2k - fts2 , peg2k - fts4 , and peg5k - fts2 conjugates , 
 which were confirmed by h nmr spectra and maldi the four peg - fts 
 conjugates readily formed micelles in aqueous solution with the particle 
 sizes of 2030 nm ( table 1 ) . dynamic 
 light scattering ( dls ) measurements showed that peg5k - fts4 micelles had hydrodynamic sizes around 27 nm at the concentration 
 of 20 mg / ml ( figure 2a ) . the size observed by tem shows good agreement with that 
 determined by dls ( supporting information figures s3 and s4 ) . the spherical shape and size distribution 
 were well retained when ptx was loaded into micelles at a drug concentration 
 of 1 mg / ml and a carrier / drug ratio of 2.5/1 ( m / m ) ( figure 2c and figure 2d ) . similar 
 results were shown for the other three micelles ( supporting information figures s3 and s4 ) . particle size distribution 
 and morphology of drug - free and ptx - loaded 
 peg5k - fts4 micelles . the size and size distribution 
 of drug - free peg5k - fts4 micelles ( a ) and ptx - loaded 
 peg5k - fts4 micelles ( c ) were evaluated by dls . the morphology of drug - free peg5k - fts4 micelles 
 ( b ) and ptx - loaded peg5k - fts4 micelles ( d ) was 
 examined by tem . the cmc of peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles 
 were measured using pyrene as a fluorescence probe ( table 1 ) . when the concentration of the peg - fts reached 
 the cmc , the fluorescence intensity of pyrene would change dramatically 
 due to the transfer of pyrene from polar microenvironment to nonpolar 
 surroundings caused by the formation of micelles . the cmcs of peg5k - fts2 and peg5k - fts4 conjugates 
 were determined to be 0.34 and 0.29 m , respectively , which 
 were lower than those of peg2k - fts2 ( 1.22 m ) 
 and peg2k - fts4 ( 1.43 m ) ( supporting information figure s5 ) . the lower 
 cmcs of peg5k - fts micelles may be attributed to the longer 
 peg hydrophilic chain and more effective stabilizing effect for the 
 micelles . the ptx loading of peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles with different 
 carrier to drug molar ratios was determined by hplc ( table 2 ) . the sizes of these ptx loaded peg - fts micelles 
 were also evaluated under corresponding conditions . peg5k - fts4 micelles could effectively solubilize ptx in aqueous 
 solution at a molar ratio as low as 1:1 ( m / m ) with particle size around 
 50 nm ( table 2 ) . however , these ptx - loaded 
 peg5k - fts4 micelles were only stable for 2.5 
 h. with an increase in carrier / drug ratio to 2.5/1 , they formed mixed 
 micelles with ptx that were stable for about 1 day . for the other 
 three micelles , a minimal carrier / drug ratio of 2.5/1 was required 
 to solubilize ptx . overall , the conjugates with four molecules of 
 fts worked better than the conjugates with two molecules of fts and 
 peg5k conjugates were more effective than the peg2k counterparts in forming stable drug loaded micelles . the four conjugates 
 were ranked as peg5k - fts4 > peg2k - fts4 > peg5k - fts2 > peg2k- fts2 with respect to their efficiency in forming 
 stable 
 mixed micelles with ptx . ptx concentration in micelles was 
 kept at 1 mg / ml . measured 
 by dynamic light scattering 
 particle sizer . data mean that 
 there was no noticeable 
 size change during the follow - up period . the profile 
 of ptx release from the four peg - fts micelles was examined by a dialysis 
 method . as shown in figure 3 , peg5k - fts4/ptx mixed micelles showed relatively slower kinetics 
 of ptx release compared to other ptx - loaded peg - fts micelles . cumulative 
 ptx release profile from four ptx - loaded peg - fts micelles . pbs containing 0.5% ( w / v ) tween 80 was used as the release medium . ptx concentration was fixed at 0.5 mg ptx / ml . values reported are 
 the means  sd for triplicate samples . as a delivery system for 
 intravenous application , the potential detrimental interaction of figure 4 shows the hemolytic activities of 
 drug - free peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles . polyethylenimine ( pei ) , a cationic polymer known 
 to have significant hemolytic effect , was included as a positive control . in contrast , all of the four drug - free peg - fts micelles displayed 
 only negligible levels of hemolytic activities , suggesting that peg - fts 
 micelles are mild surfactants suitable for in vivo delivery of hydrophobic anticancer drugs . in vitro hemolysis assay of four peg - fts micelles 
 compared with pei . rat rbcs were treated with the four different peg - fts 
 micelles or pet at 0.2 and 1 mg / ml , respectively . the lysis of rbcs 
 was determined by measuring the release of hemoglobin spectrophotometrically 
 (  = 540 nm ) . rbcs incubated with pbs and triton x-100 ( 2% ) 
 were used as the negative and positive controls . the fts2 conjugates ( peg2k - fts2 and peg5k - fts2 ) were more effective 
 than fts4 conjugates ( peg2k- fts4 and peg5k - fts4 ) in inhibiting the tumor cell 
 proliferation . similar results were found when the four conjugates 
 were examined in hct-116 ( figure 5b ) and 4t1.2 
 cells ( figure 5c ) . cytotoxicity of four 
 drug - free peg - fts micelles in mcf-7 human 
 breast carcinoma cell line ( a ) , hct-116 human colon carcinoma cell 
 line ( b ) , and 4t1.2 mouse breast cancer cell line ( c ) . cells were 
 treated with different micelles for 72 h and cytotoxicity was determined 
 by mtt assay . figure 6 shows 
 the protein expression level of total ras 20 h following treatment 
 of hct-116 cells with the four different conjugates at a fts concentration 
 of 40 m . however , the conjugates with two fts molecules 
 were more active than the counterparts with four fts molecules in 
 reducing the protein expression levels of ras in hct-116 cells . effects of 
 peg - fts micelles on total ras protein expression in 
 hct-116 cells . hct-116 cells were treated with four different peg - fts 
 micelles for 20 h ( at a fts concentration of 40 m ) . figure 7 shows the cytotoxicity of free 
 ptx ( in dmso ) and ptx formulated in peg2k - fts2 , peg2k - fts4 , peg5k - fts2 and peg5k - fts4 micelles in 4t1.2 cell line . delivery 
 of ptx via the four different peg - fts micelles led to varied levels 
 of improvement . nonetheless , ptx - loaded peg5k - fts4 micelles were more potent than ptx formulated in the other micellar 
 formulations ( peg2k - fts2 , peg2k - fts4 , and peg5k - fts2 ) in inhibiting the 
 tumor cell growth . the ic50 of free ptx and the four micellar 
 formulations of ptx are summarized in supporting 
 information table s1 . cytotoxicity of ptx - loaded peg - fts micelles 
 in 4t1.2 mouse breast 
 cancer cell line . cells were treated with free ptx or different micellar 
 formulations of ptx for 72 h and cytotoxicity was determined by mtt 
 assay . ptx - loaded peg5k - fts4 micelles showed 
 an improvement in cell growth inhibition compared with other formulations . * p < 0.05 ( ptx / peg5k - fts4 vs ptx ) , p < 0.05 ( ptx / peg5k - fts4 vs ptx / peg5k - fts2 or ptx / peg2k - fts4 ) , p < 
 0.05 ( ptx / peg5k - fts4 vs ptx / peg2k - fts2 , ptx / peg5k - fts2 , or ptx / peg2k - fts4 ) . the in vivo therapeutic effectiveness of ptx formulated in peg2k - fts2 , peg2k - fts4 , peg5k - fts2 , and peg5k - fts4 micelles 
 was evaluated , respectively , in a syngeneic murine breast cancer model 
 ( 4t1.2 ) , and compared to taxol . as shown in figure 8a , ptx - loaded peg2k - fts2 micelles exhibited 
 a similar tumor growth inhibitory effect compared to taxol treatment 
 group . in contrast , ptx formulated in peg5k - fts2 and peg5k - fts4 micelles demonstrated a significantly 
 enhanced antitumor activity compared to taxol ( p < 
 0.01 ) . furthermore , ptx formulated in peg5k - fts4 showed a trend of improvement in antitumor activity compared with 
 ptx formulated in peg5k - fts2 ( p = 0.09 ) . no significant changes in body weight were noticed in all 
 treatment groups compared to pbs control group ( figure 8b ) . in addition , serum levels of transaminases in the mice 
 with different treatments were comparable to those in pbs control 
 group ( table 3 ) . babl / c 
 mice were inoculated s.c . with 4t1.2 cells 
 ( 2  10 cells / mouse ) . six days later , mice received 
 various treatments twice a week and tumor growth was monitored and 
 plotted as relative tumor volume ( a ) . significant improvement in antitumor 
 activity was found for the ptx - loaded peg5k - fts4 group compared with the taxol group ( * * p < 0.01 ; n = 5 ) and the group of ptx - loaded peg2k - fts2 ( * p < 0.05 ; n = 5 ) . changes in body weights of mice in different treatment groups were 
 also monitored ( b ) . we 
 have systematically compared the physicochemical property and 
 the in vitro and in vivo ptx delivery 
 efficiency of four peg - fts conjugates that vary in the length of peg 
 motif ( peg2k vs peg5k ) and the molar ratio of 
 peg / fts ( 1/2 vs 1/4 ) . all of the four peg - fts micelles possessed very 
 small sizes of 2030 nm . doxil ( doxorubicin hcl liposome ) and 
 abraxane ( albumin - bound paclitaxel ) are two fda - approved formulations . the particle sizes of doxil and abraxane are 150 and 130 nm , 
 respectively . their relatively large sizes may limit the diffusion 
 in the tumor , and thus limit their therapeutic effectiveness . it has been reported that nanoparticles need 
 to be smaller than 100 nm in order to circumvent macrophage clearance 
 in the lungs and that particles of further reduced size ( 64 nm ) are needed 
 for effective penetration through neovasculatures to reach tumor cells . the small sizes 
 of our peg - fts micelles shall ensure efficient passive targeting to 
 the solid tumors . in addition to size , drug loading capacity 
 and formulation stability 
 are two other important features for an effective micellar system . our data showed that the peg - fts conjugates with four fts molecules 
 were significantly more effective than the conjugates with two molecules 
 of fts in forming stable drug - loaded micelles . previous studies have 
 shown that increasing the ratio of hydrophobic / hydrophilic blocks 
 is associated with increased drug loading capacity and enhanced formulation 
 stability . the improved performance of 
 peg - fts conjugates with an increased number of core forming units 
 may be attributed to a similar mechanism . fts , as a hydrophobic core 
 of micelles , has a lipid chain and benzene ring structure . the lipid 
 chain of fts contributes to the loading of hydrophobic drug through 
 hydrophobic interaction . at the same time , the benzene ring of fts 
 is capable of forming  interaction with drug 
 carrying aromatic ring structure . in the peg - fts micellar system , 
  stacking and hydrophobic interaction are likely 
 to work cooperatively to promote both drug / carrier and carrier / carrier 
 interactions . such interactions are also expected to be further enhanced 
 with an increase in the number of fts molecules in peg - fts conjugates . a prominent 
 advantage of peg modification is to impart the in vivo longevity to drug carriers . torchilin s group has reported that peg / core ratio affected the 
 performance of the micelles . furthermore , the length of peg also acted 
 on the cmc , which in turn influenced the performance of the micelles , 
 particularly in vivo . our data showed that peg5k - conjugates 
 had a lower cmc and were more effective than peg2k - conjugates 
 in forming stable micelles with ptx . one unique feature for 
 peg - fts micellar delivery system is its 
 intrinsic antitumor activity . the four different peg - fts conjugates 
 showed varied levels of antitumor activity by themselves in three 
 cancer cell lines . the conjugates with two fts molecules showed higher 
 levels of cytotoxicity compared to the counterparts with four fts 
 molecules . the higher levels of cytotoxicity for the conjugates with 
 two fts molecules are unlikely due to a more active surface activity 
 of the double chain conjugates as all of the four conjugates showed 
 minimal hemolytic activity ( figure 4 ) . it is 
 likely that active fts is more readily cleaved from the conjugates 
 with two fts than the ones with four fts molecules due to less steric 
 hindrance to intracellular esterases . we also compared the in vitro cytotoxicity of the four ptx - loaded peg - fts micelles 
 ( figure 7 ) . ptx - loaded peg5k - fts4 micelles showed better cytotoxicity than free ptx and ptx 
 formulated in other three micelles in 4t1.2 cells . this is likely 
 due to a more efficient intracellular delivery of ptx via peg5k - fts4 micelles because peg5k - fts4 formed the most stable micelles with ptx among the four micellar 
 systems tested . in vivo therapy study clearly 
 showed a significantly 
 higher level of antitumor activity for ptx formulated in peg5k - fts4 micelles compared to either taxol or ptx formulated 
 in peg2k - fts2 ( figure 8a ) . this is likely due to the significantly improved stability for 
 peg5k - fts4 micelles , which contributes to more 
 effective delivery of ptx to tumor tissue in vivo . in summary , we have shown that peg5k - fts4 formed the most stable mixed micelles with ptx among 
 four peg - fts 
 micelles . furthermore , ptx formulated in peg5k - fts4 micelles was more active in cytotoxicity than free ptx and 
 ptx formulated in other three peg - fts micelles . in vivo , ptx - loaded peg5k - fts4 led to an improved 
 tumor growth inhibitory effect in comparison to ptx formulated in 
 peg2k - fts2 , peg2k - fts4 , and peg5k - fts2 as well as taxol in a syngeneic 
 mouse model of breast cancer ( 4t1.2 ) . more studies on the structure 
 and activity relationship are needed to further improve the peg - fts - based 
 delivery system .
we 
 have recently designed and developed a dual - functional drug 
 carrier that is based on poly(ethylene glycol ) ( peg)-derivatized farnesylthiosalicylate 
 ( fts , a nontoxic ras antagonist ) . 
 peg5k - fts2 readily form micelles ( 2030 nm ) and hydrophobic drugs such 
 as paclitaxel ( ptx ) could be effectively loaded into these micelles . 
 ptx formulated in peg5k - fts2 micelles showed 
 an antitumor activity that was more efficacious than taxol in a syngeneic 
 mouse model of breast cancer ( 4t1.2 ) . in order to further improve 
 our peg - fts micellar system , 
 four peg - fts conjugates were developed 
 that vary in the molecular weight of peg ( peg2k vs peg5k ) and the molar ratio of peg / fts ( 1/2 vs 1/4 ) in the conjugates . 
 these conjugates were characterized including cmc , drug loading capacity , 
 stability , and their efficacy in delivery of anticancer drug ptx to 
 tumor cells in vitro and in vivo . 
 our data showed that the conjugates with four fts molecules were 
 more effective than the conjugates with two molecules of fts and that 
 fts conjugates with peg5k were more effective than the 
 counterparts with peg2k in forming stable mixed micelles . 
 ptx formulated in peg5k - fts4 micelles was the 
 most effective formulation in inhibiting the tumor growth in vivo .