l hp cht c chit xut t cy thng .

Do c kh nng c ch s phn r mng li vi th ca thoi nhim sc, kch thch qu trnh ghp cc dimer ca vi ng thnh mng li vi th v n nh mng li vi th bng cch ngn chn qu trnh tho xon. T c ch s ti t chc nng lng bnh thng ca mng li vi th, mt hin tng chc nng sng ch yu ca t bo trong tin trnh gian k v gin phn. V th Taxol co th lam gian oan s phn chia va s phat trin cua t bao, ngn cn s pht trin ca t bo ung th. Kt hp vi c tinh khng gy c taxol tr thanh cht hang u trong iu tri ung th t nhng thp nin 90 v c ch nh cho bnh nhn ung th v di cn hoc bnh nhn ung th bung trng giai on mun. Nhu cu v taxol rt ln m ham lng Taxol trong thin nhin co gii han. Thc t cho thy phai dung 6 cy thng 100 tui mi co u lng Taxol iu tri cho mt bnh nhn. Hin nay c nm con ng cng nghip ha sn xut taxol: chit tch t v cc loi thng , tng hp ton phn (1), bn tng hp (2-6), nui cy m, t bo thc vt (7-13), cng ngh chuyn ha v ln men nm ni sinh (14-32). bi ny s tp trung vo vic phn lp nm ni sinh thc to taxol, nhng ng gp ca cng ngh gen v nhng gii hn ang gp phi trong vn sn xut taxol t nm. 1. Phn lp v pht hin nm ni sinh thc vt sn xut taxol Trong sut qu trnh ng tin ha, mi quan h thn thin c hnh thnh gia tng sinh vt ni sinh v cy ch. Mt vi sinh vt ni sinh c kh nng sn xut nhng hot cht nh hoc tng t vi nhng hp cht c sinh ra t cy ch. Tht vy, nm 1993 Taxomyces andreanae l nm ni sinh thc vt (fungal endophyte) c phn lp t mch libe bn trong v cy thng Pacific. Nm ny thuc lp nm bt ton v thuc nhm hyphomyceteous, khi c nui cy trn mi trng lng bn tng hp s sn xut taxol v cc hp cht lin quan. Taxol c pht hin bng sc k, khi ph v phn ng vi khng th n dng c hiu cho taxol (14). Sau , dng nm TF5, thuc Tubercularia sp., c phn lp t Taxus mairei c nui cy trn mi trng lng dextrose khoai ty to ra c taxol. Chng c hot tnh khng phn bo mnh khi c th trn dng t bo ung th P388 v KB. ng thi, taxol t dng nm ny cng thc y s n nh ca cc vi ng (microtube) v kch thch s polymer ha invitro tng t nh taxol t thng (16). Fusarium solani c phn lp t nhng mu thn Taxus celebica sau khi nui cy trn mi trng lng c th sn xut 1,6ug/l. Paclitaxel c sn xut t nm ny gy apoptosiss trn t bo Jurkat ti 60% trong khi paclitaxel chun ch c 51%. Ngoi paclitaxel, nm ny cn sn xut ra cc taxane, baccatin III v 10 deacetyl baccatin III, nhng tin cht dng trong bn tng hp paclitaxel cng nghip (18). Metarhizium anisopliae, H-27, mt trong 13 loi nm c sng lc t 115 dng nm c phn lp t v cy thng bc (Taxus chinensis) c th sn xut ra 846,1 g/ l taxol (29).

Aspergillus candidus MD3 (hnh 1) c phn lp t v Taxus x media c th to ra 112mg/g taxol (tnh theo khi lng kh ca si nm) (31). Trong khi , dng Aspergillus niger var. taxi HD86-9 (hnh 2) phn lp t Taxus cuspidata c th sn xut 273,46 ug/l (32).

Hnh 1: hnh thi ca Aspergillus candidus MD3. (a) khun lc sau 14 ngy 250C trn mi trng CYA. (b) mt di ca khun lc; (c,d) nh di knh hin vi in t qut ca ht bo t ; (e-h) nh di knh hin vi ca bo t v cung bo t. phng i: e 100X; f,g 400X; h 1000X.

Hnh 2: hnh thi HD86-9. (a) khun lc trn mi trng PDA 280C sau 3 ngy. (b,c) bo t nh di knh hin vi in t qut (2000X v 5000X); (d,e) cung bo t di knh hin vi (1000X). 2. Phn lp nm t cc ngun khc Trong khi hu ht cc nghin cu u phn lp nm ni sinh t thng sn xut taxol th Kumaran v cng s (2008) li phn lp nm Phyllosticta citricarpa (McAlp.) Van der Aa, l loi nm gy bnh m l, t l cy Citrus medica L. (Rutaceae) b bnh Chennai, n . Loi nm ny c th sn xut lng taxol kh cao, ln ti 265ug/l trn mi trng M1D, cao hn 5.3103 ln so vi lng taxol t nm T. andreanae (14) v 137ug/l trn mi trng PDB (potato dextrose broth) (22). V sau ba dng nm Phomopsis khc cng c phn lp t l cy Taxus cuspidata (thng Nht), Ginkgo biloba (cy bch qu) v Larix leptolepis (thng rng l Nht). Nm ny c th to 418 ug/l taxol (27). Nm Phyllosticta tabernaemontanae, nm m l phn lp t l cy Wrightia tinctoria b bnh, khi nui cy trn mi trng M1D c th to ra 461 ug/l taxol v 150ug/l trn mi trng PDB (28). iu ny cho thy nm sn xut ra taxol khng ch c tm thy trn thng m c nhng thc vt khc v c bit bao gm c nhng loi nm gy bnh. Noh v cng s (1999) phn lp c dng nm Pestalotia heterocornis (hnh 3) t t khu rng trng thng (15). Sau khi nui cy trong mi trng bn tng hp h thu c 31ug/l paclitaxel sau 5-7 ngy ln men 230C.

Hnh 3: hnh thi bo t P. heterocornis.

Nm Chaetomella raphigera (dng TAC-15) hnh 4 c phn lp t cy dc liu Terminalia arjuna c th to ra 79.6 g taxol/L (25) hnh. Cng trn l cy ny Gangadevi v cng s phn lp c dng Pestalotiopsis terminaliae c th to ra 211g/l taxol. Khi th nghim trn cc dng t bo ung th ngi BT220, H116, Int 407, HL 251 v HLK 210 cho thy hot tnh khng phn bo ca taxol t dng nm ny rt mnh (26).

Hnh 4: (a) Hnh thi nm C. raphigera ; (b) bo t (40X) Nm Pestalotiopsis pauciseta (dng CHP-11) c phn lp t l cy Cardiospermum helicacabum c th sn xut 113,3ug/l taxol. Khi kim tra trn cc trn cc dng t bo ung th viz., BT 220, H116, Int 407, HL 251 and HLK 210 cho thy taxol to ra t nm ny c hot tnh khng ung th rt mnh thng qua con ng apoptosis (20). 3. S dng cng ngh sinh hc trong phn lp, sng lc v nng cao hiu qu to taxol Cc gen m ha cho 10-deacetylbaccatin III-10-O-acetyl transferase v C-13 phenylpropanoid side chain-CoA acyltransferase c dng lm marker phn t sng lc cc nm ni sinh sn xut taxol. Sau khi sng lc 90 dng nm c phn lp t Taxus x media v Taxus yunnanensis bng k thut PCR, Zhang v cng s (2008) tm ra ba dng dng tnh vi hai gen trn. Khi nui cy cc dng ny trong 300ml mi trng lng dextrose khoai ty 25C sau 10 ngy h thu c t 100 ti 160ug taxol/g (tnh theo trng lng kh ca si nm thu c sau nui cy) (23). Kt qu cho thy vic dng cc marker phn t sng lc s nhanh hn rt nhiu v d dng chn lc c nhng dng nm c kh nng sn xut taxol cao. S dng k thut genome shuffling, Zhao v cng s (2008) lai to c dng nm Nodulisporium sylviform sn xut ti 516.37 g/L taxol, cao hn 31.52%44.72% so vi dng b m (24). 4. Mt s kh khn v cc kha cnh lin quan trong vic sn xut taxol t nm Hin nay cc dng nm c phn lp ch sn xut Taxol vi lng nh. l nguyn nhn v sao cha th sn xut taxol cng nghip t cc dng nm ny. Mt trong nhng l do m taxol thu c khng cao l khi taxol tit ra mt lng gii hn trong mi trng nui cy c ch s sinh trng v tit taxol t nm. Nu c th hiu r hn v con ng c ch ngc ny c th tm ra nhng gii php nh c ch cc enzyme dn truyn tn hiu gy c ch ny.

Bng nhiu n lc ca mnh vi mc ch nng cao hm lng taxol t nm, qua gn hai thp k k t khi dng nm sn xut taxol u tin c phn lp (14), hm lng taxol c nng cao ng k t 70ng/l ti 1000ug/l (bng 1). Nhiu dng nm c phn lp vi hiu qu ngy cng cao v c bit khng ch cc dng ni sinh trn thng m cn t t hay c nhng loi tho dc khc. ng lu l hin nay cc dng c phn lp t cc loi tho dc li c kh nng to taxol cao hn rt nhiu so vi cc dng c phn lp t thng . Bng 1: Bng lit k cc dng nm ni sinh thc vt sn xut taxol (33).

Nm ni sinh c th to ra lng taxol cao tng t k ch, tuy nhin y l loi cng sinh nn vi nhng mi trng phn lp dng tng hp hay bn tng hp nh hin gi cng kh c th nui cy v sng lc ht nhng dng nm c th cho sn lng cao hn. C ngha l nhiu dng vi sinh cha bit c tim nng to taxol cao th khng c nui cy v sng lc. Mc d lng taxol c sn xut t nm cn thp so vi chit xut t thng nhng vi t l tng trng cao, thi gian sinh trng ngn hn rt nhiu so vi cy thng th s nghin cu sn xut taxol t nm l vic lm c ngha. Cng vi thnh tu ca cng ngh ln men hin i th hng nghin cu ny thc s mang nhiu ha hn. Nghin cu tng hp paclitaxel vn ang c cc nhm nghin cu trn th gii tin hnh nhm tm ra nhng con ng bn tng hp mi v ngn hn cng nh to ra nhng hot cht mi da trn paclitaxel m c hot tnh sinh hc cao hn hoc /v d tng hp hn. Dc ng hc : - Hp thu: Sau khi c truyn tnh mch, nng ca paclitaxel trong huyt tng gim dn theo mt ng cong biu din gm hai pha. Dc ng ca paclitaxel c nghin cu trong iu kin truyn tnh mch thi gian l 3 gi v 24 gi, v liu 135 v 175mg/m2. Thi gian bn hy trung bnh ca pha cui t 3 n 52,7 gi v gi tr trung bnh ca thanh thi ton phn t 11,6 n 24l/gi/m2 S thanh thi ton phn dng nh gim khi nng ca paclitaxel trong huyt tng cao. - Phn b: Th tch phn phi trung bnh tnh trng cn bng dao ng t 198 n 688l/m2 iu ny cho thy c s khuch tn ngoi mch v/hoc gn vo m ng k ca paclitaxel. Khi tng 30% liu, t 135 n 175mg/m2, cc gi tr Cmax v din tch di ng cong tng tng ng vi 75% v 81% khi truyn trong 3 gi, nh vy vic tng liu khng c lin quan tuyn tnh vi vic tng cc thng s dc ng. S khc bit gia ngi ny v ngi khc khi dng paclitaxel ng ton thn rt t. Khng ghi nhn c hin tng tch t paclitaxel khi dng nhiu t iu tr lin tip. Cc nghin cu in vitro c thc hin trn proteine huyt thanh ngi cho thy t l gn

vi proteine trung bnh t 89 n 98%. Khi dng ng thi vi cimetidine, ranitidine, dexamethasone hay diphenhydramine, t l gn vi proteine khng b thay i. - Chuyn ho v thi tr: Th thc o thi ca paclitaxel ngi hin cha c bit r. T l o thi qua thn di dng khng i t 1,3 n 12,6% liu dng, cho thy c s thanh thi ng k ngoi thn. Cc cht chuyn ha chnh c tm thy di dng hydroxyl ha. S chuyn ha qua gan v o thi qua mt dng nh l c ch o thi chnh ca paclitaxel. Cha c nghin cu v nh hng trn s chuyn ha thuc khi dng truyn tnh mch trong 3 gi cho bnh nhn b suy gan hay suy thn. Cc thng s dc ng thu c bnh nhn chy thn c truyn Paclitaxel 135mg/m2 trong 3 gi th tng t nh bnh nhn bnh thng.

Taxol(paclitaxel) v ung th
u nhng nm 1960, vin Ung th Quc gia M tch ra c mt hp cht c hot tnh t cy Thng gi l taxol. - 1967, t v thn Thng Taxus brevifolia (Taxaceae) -> Taxol tinh khit = Paclitaxel -> ch phm Taxol (Bristol-Myers Squibb) Taxol cn c / Taxus cuspidata, T. wallichiana h Taxaceae - Nm 1969 : t 1250 kg v thn chit xut ra 28 kg cao ton phn v 10 g Taxol tinh khit Taxol c tm thy trong v v r cy thng nhng hm lng trong v cy nhiu hn. Hm lng khong 1000mg/kg v cy kh (0,01%). Mt cy thng khong 100 tui cho gn 3kg v cy.Vic thu thp v lm cy nhanh lo ho v cht, do vy cn nghin cu phng php tng hp taxol. - Cc nh ho hc tm thy quy trnh bn tng hp taxol t 10-DAB, mt tin cht cho s tng hp sinh hc, c th chuyn ho thnh taxol vi hiu sut rt cao. 10-DAB c tm thy trong nn thng v l thng.Vo thp nin 1980 P.Poitier chit t l cy thng 10-DAB vi hm lng kh cao(0,1%). - 1971 :Wall v Wani cng b cu trc Taxol. - 1989 : Robert A Holton thuc trng i hc Florida pht trin thnh cng phng php bn tng hp Taxol. - 1994: Tng hp ton phn Taxol - Spencer v Faulds(1994) xem xt tnh dc ng hc ca taxol v tim nng trong vic cha bnh ung th.Taxol l mt trong nhng dc phm cho kt qu y ha hn trong ho tr liu cc bnh ung th v, ung th bung trng; ngn chn s mt kim sot trong qu trnh phn chia ca t bo ung th. Dc l v c ch tc dng Paclitaxel, hot cht c trong v cy thng Taxux brevifolia, l mt thuc chng ung

th. Paclitaxel lm tng qu trnh trng hp cc dime tubulin to thnh cc vi qun v lm n nh cc vi qun do c ch qu trnh gii trng hp. S n nh ny c ch s t chc li bnh thng ca mng vi qun rt quan trng gian k ca qu trnh phn bo gim nhim, v c vi hot ng ca ty lp th. Paclitaxel cng gy to thnh cc cu trc bt thng trong cc vi qun trong qu trnh phn bo. Tuy cha c nghin cu k nhng do c ch tc dng ca n, paclitaxel phi c coi l cht gy ung th v c i vi gen. Cc th nghim in vivo v in vitro cho thy paclitaxel c tc dng gy t bin gen trn t bo ng vt c v. Nng thuc trong huyt tng t l thun vi liu c truyn vo tnh mch v gim theo th c 2 pha. T l gn vi protein l 89% (in vitro) v khng b thay i khi dng cng vi cimetidin, ranitidin, dexamethason, hoc diphenhydramin. giai on n nh, th tch phn b l 5 6 lt/kg th trng (68 162 ml/m2), cho thy thuc khuch tn nhiu ra ngoi mch v/hoc gn nhiu vi cc thnh phn ca m. Ngi ta cn cha bit r hon ton s phn b v chuyn ha thuc trong c th. Na i trong huyt thanh l 6 13 gi. Sau khi truyn tnh mch, c khong 2 13% lng thuc c thi qua nc tiu di dng ban u, nh vy l ngoi thn cn c nhng ng o thi khc. Trn ng vt th nghim, paclitaxel c chuyn ha ti gan. thanh thi dao ng t 0,3 n 0,8 lt/gi/kg (hay 6,0 15,6 lt/gi/ m2). CH NH Ung th biu m bung trng : - Tr liu u tay ung th bung trng tin xa hoc bu tn lu ( 1 cm) sau phu thut, dng phi hp vi cisplatine. - Trong tr liu th nh, ung th bung trng di cn, sau khi tht bi vi phng php iu tr c in bng cc dn xut ca platine. Ung th biu m v : - Trong iu tr h tr ung th giai on sm, c hch dng tnh, dng lin tip sau phc c cha doxorubicine. - Trong iu tr ung th v di cn tht bi sau mt t ha tr hoc ti pht sau khi iu tr h tr. Thng cc iu tr trc c cha anthracycline, tr khi no c chng ch nh. Ung th phi khng phi t bo nh : - Taxol dng chung vi cisplatine, c c ch nh u tay iu tr bnh nhn ung th phi khng phi t bo nh, giai on khng th phu thut v/hoc x tr c. Ngoi ra, Taxol cn dng iu tr Ung th Kaposi c lin quan n bnh AIDS. CHNG CH NH - Chng ch nh Taxol cho nhng bnh nhn c phn ng qu mn cm nng vi

paclitaxel hoc vi bt c thnh phn no ca thuc, c bit l vi huile de ricin polyoxyethylene (Crmophor EL). - Khng c iu tr bng Taxol nhng bnh nhn c s lng bch cu trung tnh di 1.500/mm3. - Ph n c thai hay cho con b : xem thm phn Lc c thai v Lc nui con b.