Nghiên cứu đánh dấu kháng thể đơn dòng kháng CD20 với đồng vị phóng xạ i 131 dùng trong điều trị bệnh ung thư máu

BO KHOA HOC VA
CONG
NGHE
VIEN NANG
Ll/ONG
NGUYEN
TlT
VIET NAM
VlfN NGHIEN ctru
HAT NHAN
BAO CAO TONG KET
THlTC
HIEN DE TAI
cAP
BO NAM 2007 - 2008
TEN DE
TAI
NGHIEN
ClTU
DANH
DAU
KHANG THE BON DONG
KHANG CD20
V6l
DONG VI PHONG XA
1-131
DUNG
TRONG DIEU TRI BENH UNG
THU*
MAU
Ma s6: 05/07-09/NLNT

Chu
nhifm
de tai: ThS. Nguyen
Thj
Thu
THANG 12/2009
DANH SACH
NHlTNG NGircn
THAM GIA
THlTC
HIEN DE TAI
Nguyen Thi Thu ThS. NCV
Duong Van Dong ThS. NCVC
V6
Thi Cam Hoa CN, NCV
Chu Van Khoa
Mai Phudc Tho
ThS,
NCV
Biii
Van
Cucmg
CN, NCV
CN, NCV
Pham Ngoc Dien CN, NCV
Mai Trong Khoa PGS. TS.
Le Quang Huan PGS. TS.
10
Le Ngoc Ha
TS.

BS
Trung Tam Nghien cuu va dieu che
Dong vi Phong xa,
Vien Nghien cuu Hat nhan
Trung Tam Nghien cuu va dieu che
Dong vi Phong xa,
Vien Nghien ciru Hat nhan
Trung Tam Nghien cuu va dieu che
Dong vi Phong xa,
Vien Nghien cuu Hat nhan
Trung Tam Nghien
ciiu
va dieu che
Dong vj Phong xa,
Vien Nghien cim Hat nhan
Trung Tam Nghien
ctiu
va dieu che
Dong vj Phong xa,
Vien Nghien cuu Hat nhan
Trung Tam Nghien cuu va dieu che
Dong vi Phong xa,
Vien Nghien cuu Hat nhan
Trung Tam Nghien cuu va dieu che
Dong vi Phong xa,
Vien Nghien
ciiu
Hat nhan
Trung Tam Y hoc hat nhan va Dieu
tri ung budu,

Benh vien Bach Mai, Ha Noi
Vien Cong Nghe Sinh hoc,
Vien Khoa hoc Viet Nam
Benh
Vien
108,
Ha
Noi
Cac ca quan,
don
vi phoi
hap thye
hien
1.
Vien Cong nghe sinh
hoc.
Ha Noi
2.
Benh vien Bach Mai, Ha Noi
3.
BenhvienlOS,
HaNoi
^^n
MUC LUC
• •
Trang
Trang bia
Danh sach nhimg ngudi tham gia thuc hien de tai 1
Muc luc 2
Abstract 5

Tom tat 6
Md dau 7
I. Tom tat tinh hinh nghien cuu trong va ngoai nudc 8
II.
Muc tieu cua de tai 9
III.
Tom tat noi dung nghien
ctju
10
Phan I: TONG QUAN
I. Gioi thieu nhirng tien bo moi trong dieu tri mien djch phong xa RIT
11
1.1.
Cac
Idiai
niem
11
1.2.
Khang the dugc sir dung dieu tri nham dich 12
1.3.
Cac tac nhan chup hinh mdi trong RIT 18
1.4.
L/ng
dung RIT trong dieu tri NHL
18
II.
Cac thanh phan
co*
ban trong RIT 21
II.

1.
Khang nguyen CD20 vai tro va tiem nang ung dung 21
11.2.
Khang the va cong nghe tao khang the 23
11.3.
Dong vi phdng xa 28
III.
Cac phirong phap danh dau khang the voi dong vj phong xa 30
III.
1.
Dac diem chung 30
111.2.
Qua trinh danh dau phong xa
31
111.3.
Phuang phap danh dau khang the vdi
'^'l
32
111.4.
Phuang phap danh dau khang the vdi
'^'^'"Tc
va
' "in
35
IV. Nghien
cihi du-ffc
dong hoc 37
Phan II: VAT LIEU VA
PHT/ONG
PHAP NGHIEN

ClTU
38
I. Nguyen
lieu,
hoa chat, thiet bi
II.
Nghien
ciru
cac dieu kien toi
ini
danh dau khang the voi phong xa 39
II.
1.
Xac djnh ham'lugng ChT ' 39
11.2.
Xac dinh ham lugng khang the, hoat do, thdi gian phan ung 42
11.3.
Xac dinh
pH 43
11.4.
Quy trinh danh dau khang the 44
11.5.
Danh dau theo hoat do rieng 46
11.6.
Phan ung mien dich 47
11.7.
Phuong phap iodogen 48
U.S.
Danh dau khang the don chuoi 49
III.

Kiem tra chat
Iirong
san pham 49
III.
1.
Kiem tra do sach hda phdng xa 49
111.2.
Kiem tra do vd khuan 50
111.3.
Kiem tra ngi doc td vi khuan 51
111.4.
Kiem tra do dn dinh trong huyet thanh 53
111.5.
Kiem tra do dn dinh theo thdi gian bao quan 53
111.6.
Kiem tra phan bd sinh hoc tren chupt 53
III.7.
Kiem tra dao thai thudc tren thd 53
IV. Chup
hinh
phan bo thuoc phase I tren benh nhan ung
thir
NHL 54
Phan
III: KET QUA NGHIEN
CUtl
I. Ket qua danh dau khang the 56
1.1.
Ket qua kiem tra chat lugng
'^'l

danh dau 56
1.2.
Ket qua khao sat ham lugng ChT 58
1.3.
Ket qua khao sat ham lugng, hoat dp, thdi gian 61
1.4.
Kit
qua khao sat pH 62
1.5.
Ket qua dieu che san pham 63
1.6.
Ket qua dieu che san pham theo hoat dp rieng 64
1.7.
Ket qua kiem tra hoat tinh mien dich 65
1.8.
Ket qua danh dau khang the bang phuong phap iodogen 66
1.9.
Ket qua danh dau khang the dcm chudi 67
II.
Ket qua kiem tra chat
Iirong dirtfc
chat phong xa 68
II.
1.
Kiem tra dp sach hoa phdng xa 68
11.2.
Ket qua kiem tra dp vd khuan 71
11.3.
Ket qua kiem tra npi doc td vi khuan 72
11.4.

Kiem tra dp dn dinh trong huyet thanh ngudi 73
11.5.
Kiem tra dp dn dinh theo thdi gian 74
11.6.
Kiem tra phan bd sinh hpc 76
11.7.
Kiem tra dao thai sinh hpc 78
III.
Ket qua chup
hinh
phan bo thuoc phase I tren benh nhan ung
thir
80
Phan IV: KET LUAN 85
Tai lieu tham khao 86
Giai trinh cac khoan chi 87
Phu luc
1:
Tdng hgp khang the don chuoi
Phu luc 2: Kiem tra chat lugng
Phu luc 3: Lam sang
Phu luc 4: Thdng tin de tai
BANG VIET TAT
RIT Lieu phap mien dich phdng xa (radioimmunotherapy)
RJS Chup hinh nhap nhay mien dich (Radioimmunoscintigraphy)
CD Cum biet hoa (cluster differentiation)
MAb Khang the don ddng (monoclonal antibody)
NHL U lympho khdng Hogdkin (non-Hodgkin's lymphoma)
FDA
Cue

quan ly thudc va thuc pham Hoa Ky (Food and Drug Administration)
BCA Tac nhan chelat hda hai chuc nang
(bifunctional
chelating agent)
CR Dap ling toan phan (complete response)
PFS Tiep tuc sdng sdt (progression-free survival)
PR Dap ung rieng phan (partial response)
ADCC Doc tinh vdi te bao qua khang the (Antibody Dependent Cellular
Cytotoxicity)
CDC Ddc tinh vdi te bao qua bd the (Complement Dependent Cytotoxicity)
CDCC Ddc tinh vdi te bao phu thudc bd the (Complement Dependent Cellular
Cytotoxicity)
CLL Benh mau trang dang te bao lympho man tinh (Chronic Lymphocytic
Leukemia)
AML Ung thu bach cau dang tuy (Acute Myeloid Leukemia)
ALL Ung thu nguyen bao tuy cap (Acute Lymphoblastic Leukemia)
RAIT Dieu tri phdng xa mien dich
HACA Khang khang the lai d ngudi (Human Anti-Chimeric Antibody)
HAMA Khang khang the chupt (Human Anti-Murine Antibody)
HAHA Khang khang the nhan hoa d ngudi (Human
Anti-humanized
Antibody)
IL-6
Interleukin 6
Rf Retardation factor (retention factor)
NSB Phan gan khdng dac hieu (non specific binding)
EGFR Epidermal Growth Factor Receptor
HER 2 Herceptin, khang the chira ung thu
vii
HAT Hypoxanthin Aminopterin Thymidin

VEGF Vascular Endothelial Growth Factor
CDR Viing trinh dien khang nguyen
FACS phan loai te bao phat huynh quang (Fluorescence activated cell sorting)
IgG
Immunoglobulin
Sc-FvAb Khang the don chuoi (Single-Chain Fragment Variable Antibody)
cfii
Don vi do mat dp vi sinh (Colony forming unit)
ChT Chloramin T
ITLC
Sac ky lop mdng ttic thdi (Instant Thin Layer Chromatography)
FPLC Fast Protein Liquid Chromatography
STUDY ON THE RADIOLABELING OF MONOCLONAL ANTIBODY
FOR BLOOD CANCER THERAPY
Nguyen Thi Thu
Abstract
In recent years, radioimmunotherapy (RIT) has become a highly promising oncologic
therapeutic modality with established clinical efficacy, particularly in the therapy of
haematological malignancies.
Anti CD20 monoclonal antibody was labelled with
'^'l
used in the treatment of
B
cell
non Hodgkin's Lymphoma (NHL), B cell leukemia. In this study, the monoclonal antibody
Rituximab was labelled with
'^'l
using chloramin T method (ChT). The optimized ChT
concentration for the oxidation of 185 MBq of
Na'^'l

solution and
750|ag
of Rituximab was
50\igl50n\.
The reaction time was 5 minutes at room temperature. The labeling reaction has
stopped using natrimetabisulphite (SMB). Labelling efficacy was controlled by ITLC. The
reaction mixture was purified through the Sephadex G-25
PDIO
Pharmacia column. The
collected
'^'l-CD20MAb
Rituximab was filtered through a
0.20^m
milipore sterile filter. The
radiochemical labeling yield was more than 95%. Radiochemical purity of the
radiopharmaceutical after purification was more than 99%. The product has passed the test
for sterility, bacterial endotoxins, to be sufficiency invivo and invitro stable after labeling,
ready for clinical use. In the phase I trial, a radiation absorbed dose escalation and protein
mass dose escalation were undertaken.
'^'l-CD20MAb
be used on two patients with blood
cancer type NHL, has recorded immunoscintigraphy biodistribution.
The radiolabeled antibody used in clinical is safe and efficacy. It's the
radiopharmaceutical used in cancer therapy with RIT. This study has the goal of research and
develop and clinical testing. It has the potential to be a new method of treatment for RIT
domestically.
NGHIEN
ClTU
DANH DAU KHANG THE DON DONG
V6l

DONG
VI
PHONG XA DUNG TRONG DIEU TRI
BENH UNG
THU*
MAU
Nguyen Thi Thu
Tom tat
Trong nhirng nam gan day lieu phap mien dich phdng xa (RIT) da trd thanh phucmg
thiic dieu tri ung thu day
hiia
hen vdi hieu qua lam sang ro ret dac biet la trong dieu tri cac
ung thu mau.
Khang the don ddng khang CD20 dugc danh dau vdi ddng vi phdng xa
'^'l
dimg
trong dieu tri benh u lympho khdng Hodgkin (NHL), benh ung thu te bao B. Trong nghien
ciiu
nay, khang the don ddng Rituximab dugc danh dau vdi
'^'l
bang phuong phap
Chloramin T. Ham lugng ChT tdi uu de oxy hoa 185MBq
'^'l
va
750ng
Rituximab la
50p.g/50|il.
Thdi gian phan
iing
la 3

phiit
d nhiet dp phdng. Phan ung dugc dimg bang
each
them chat khu natri metabisulphite (SMB). Hieu suat danh dau dugc kiem tra bang phuong
phap sac ky lop mdng ITLC. Hdn hgp phan
iing
dugc
Ipc
qua cdt Sephadex
G25
PDIO.
San
pham
'^'l-CD20MAb
Rituximab dugc
Ipc
qua
phii Ipc
vd triing
0,20fxm.
Hieu suat danh dau
phdng xa hon
95%.
Dp sach hda phdng xa cua san pham sau khi
Ipc
qua cot la
hem
99%. San
pham dat tieu chuan vd trung, dat tieu chuan kiem tra npi ddc td vi khuan, dn dinh invitro va
invivo cd the sir dung cho lam sang. Trong phase I thir nghiem, lieu hap thu

biic
xa va lieu
lugng protein dugc thuc hien.
'^'l-CD20MAb
Rituximab dugc sir dung lam sang tren hai
benh nhan ung thu mau dang NHL, da ghi hinh nhap nhay mien dich phan bd thudc.
Khang the danh dau phdng xa cd the sir dung tren lam sang an toan va hieu qua. Day
la dugc chat phdng xa diing trong dieu tri ung thu bang ky thuat mien dich phdng xa RIT. De
tai da dat ra muc tieu nghien ctiu dieu che va thir nghiem va cd tiem nang md dau cho hudng
dieu tri mien dich phdng xa trong nudc.
MOfDAu
Trong thap nien qua, ciing vdi su ra ddi va ngay cang hien dai hda cac thiet bi chup
hinh chan doan nhu may chup cat
Idp
dien toan SPECT va may chup cat
Idp
diing ddng vi
phdng xa phat positron PET-CT, nhieu the he thudc phdng xa mdi
cQng
da dugc nghien
ciiu
dieu che va phat trien. Su phat trien ddng bp nay da lam ddi mdi bp mat cua y hpc hat nhan
bdi chat lugng chan doan va hieu qua dieu tri. Ben canh nhiing chan doan chinh xac de phat
hien benh, viec nghien
ciiu
chiia tri benh la nhiem vu hang dau cua y hpc. Mot trong nhiing
noi bat cua y hpc hat nhan trong tri lieu la viec sir dung khang the don ddng danh dau vdi
ddng vi phdng xa dung trong dieu tri ung thu.
Ky thuat dieu tri benh bang phdng xa da ra ddi tir nhiing nam 1950 tir khi y hpc biet
dung

'•"l
trong dieu tri tuyen giap. Tir do den nay, cac dugc chat phdng xa danh dau vdi ddng
vi
' 'l
khdng ngimg nghien
ciiu
va phat trien. Dac biet la trong nhirng nam gan day, khang
the don ddng danh dau phdng xa da dugc diing trong chan doan va dieu tri lam sang. Phuong
phap chiia benh dung khang the don ddng gpi la phuong phap dieu tri nham dich. Phuong
phap dieu tri nay cdn dugc gpi la lieu phap phdng xa mien dich RIT (Radioimmunotherapy).
Nhd nhimg tien bd cua cdng nghe sinh hpc va nhiing hieu biet ve sinh hpc phan tir va sinh
hpc te bao, RIT da trd thanh lieu phap chiia tri ung thu day
hiia
hen vdi nhirng hieu qua lam
sang ro ret.
U lympho khdng Hodgkin (NHL) la benh ung thu cac lympho bao B trong mau, khi
bi ung thu, tren be mat cac lympho bao B nay bieu hien khang nguyen la, bang ky thuat sinh
hpc phan tir hien dai, cac nha khoa hpc tren the gidi da xac dinh dd la khang nguyen CD20 va
cau tnic phan
tii
ciing da xac dinh. Phan tir khang the don ddng khang CD20 (Rituximab)
dugc su dung lam sang tir nam 1997, phan tti khang the thuc hien chiic nang nham dich cua
minh bang
each
gan dac hieu len khang nguyen dich, diet te bao ung thu bang cac co che
biic
xa ion hda (khi gan vdi chat phdng xa), co che gay ddc tinh vdi te bao ung thu qua khang the
va gay ddc tinh vdi te bao ung thu qua bd the. Vdi thdi gian ban ra 8 ngay va phat tia gamma
vdi nang lugng la 364 keV, va tia beta vdi nang lugng trung binh la 192 keV,
'^'l

la ddng vi
phdng xa ly tudng cho viec chup hinh va dieu tri benh khi gan vdi phan tir khang the.
Khang the khang CD20 da dugc gan vdi ddng vi phdng xa
'^'l
bang phuong phap
danh dau phdng xa sir dung chloramin T lam chat oxy hda. Khang the danh dau dugc tinh che
va kiem tra chat lugng theo tieu chuan cua thudc phdng xa de su dung lam sang. Day la dugc
chdt phdng xa dimg trong dieu tri ung thu bang ky thuat RIT.
D6
tai da dat ra muc tieu nghien
ciiu
dieu che, thu nghiem va md dau cho hudng dieu tri mien dich phdng xa trong nudc.
I.
Tinh hinh
nghien
ciru
trong va ngoai
niro^c:
Ky thuat san xuat khang the don ddng da dugc hai nha bac hgc Cesar Milstein va
Georges Kohler de ra vao nam 1975 da dugc giai thudng Nobel vao nam
1984
bdi nhieu
iing
dung cd gia tri vugt bac trong y hpc.
Hien nay, tren the gidi vdi hon 200 Cdng ty nghien
ciiu
va san xuat khang the, da cd
23 khang the don ddng dugc FDA phe duyet cho sir dung lam sang, hang nghin loai dang
trong qua trinh nghien ciiru lam sang. Nhieu loai khang the don ddng mdi da dugc nghien ciiru
va cai tien, ddi mdi phuc vu cho cac muc dich chan doan va dieu tri trong y hpc. Nhiing ndi

bat gan day nhat la sau hon 1/4 the ky ke tir khi ra ddi, viec
sii
dung khang the dcm ddng cho
dieu tri ung thu mdi bat dau sir dung hieu qua tren lam sang. Bdi vi, trudc kia ngudi ta san
xuat khang the don ddng theo phuong phap tao ra cac khang the tren chupt, do vay khi dua
vao
CO
the con ngudi, he thdng mien dich se phat hien ra dd la chat ngoai lai, va se bi dap
ling mien dich chdng lai va dao thai khdi vat chu. Do vay ma viec dieu tri van cdn gap nhieu
han che. Trong nhirng nam
1986-1990,
cac nha khoa hpc Michael Neuberger va Greg Winter
da tao ra mot
biit
pha khi dua ra ky thuat cd ten gpi la "humanise". Nhd ky thuat nay, ddng
khang the san xuat tir chupt dugc chuyen thanh ddng khang the ngudi, thich hgp cho viec
sii
dung tren co the con ngudi va md ra trien vgng mdi trong dieu tri ung thu. Trong vai nam
gan day viec san xuat va sir dung khang the don ddng trong dieu tri ung thu la van de thdi
sir
tren toan the gidi.
Dac biet la, mdt sd ddng vj phdng xa nhu
^^""Tc,
'**Re,
'"Sm,
'"in,
^"Y,
'^'I
cd thl
gan vdi khang the don ddng, dan dudng mot

each
dac hieu den cac te bao ung thu, cac chat
phdng xa phat tia gamma
('y),
giiip cho viec phat hien chan doan va theo doi ung thu, cac chat
phdng xa phat tia beta (P) cd the tieu diet te bao ung thu.
Sir
phat trien nay ket hgp vdi su
phat trien cdng nghe may tinh, cdng nghe thong tin lam cho nganh y hpc hat nhan ngay cang
dat den dinh cao cua nen y hpc hien dai.
Khang the don ddng khang CD20 da dugc dua vao su dung lam sang de chua benh
ung thu tk bao lympho B. Day la khang the gan dac hieu len khang nguyen CD20, khang
nguyen nay bieu hien vdi mat dp cao te bao lympho B ung thu. Tren the gidi, ngudi ta da dua
khang the khang CD20 vao sir dung dieu tri lam sang
tii
nam 1997 vdi ten thucmg mai
Rituximab. Viec tao conjugate vdi ddng vi phdng xa chi nghien
ciiu
trong vai nam gan day
va chinh thiic dugc phep sir dung lam sang tir nam
2003,
dd la cac san
phdm
Bexxar,
Tositumomab.
Trung tam Nghien cim va
Di6u ch6
Ddng vi Phdng xa thudc Vien Nghien
ciiu
hat

nhan, Vien Nang lugng nguyen tir Viet Nam la don vi duy
nh4t
trong nudc hien nay dang san
xuat cac loai ddng vi phdng xa va dugc chdt phdng xa phuc vu cho viec chin doan va dieu tri
benh. Mot sd ddng vi phdng xa chi san
xuit
dugc tren
Id
phan ring hat nhan va cd gia tri sir
dung cao trong y hpc nhu '
'l,
^^""Tc,
•'^P,
'"Sm
Hien nay cac ddng vj phdng xa nay dugc
san
xudt va cung cdp thudng xuyen den cac co sd y hpc hat nhan trong nudc. Ciing vdi trang
thik
bi phuc vu cho viec san xudt va
kilm
tra chdt lugng cac dugc chdt phdng xa theo tieu
chuan QA/QC/GMP cua qudc te, Trung tam ciing dugc ddu tu mdt sd trang
thilt
bi cho san
xudt cac kit dinh lugng phdng xa mien dich
(RIA),
san xudt chdt danh ddu cac hocmon vdi
ddng vi phdng xa, danh dau cac khang the vdi dong vi phdng xa, thuc hien cac nghien
ciiu
tdi

uu qua trinh danh dau va dinh lugng.
•'
Cac nghien ciiru san xuat khang the don ddng danh ddu vdi ddng vi phdng xa hoac cac
chdt khang ung thu cho muc dich chan doan va dieu tri ung thu ciing da dugc dat ra tir nhieu
nam nay, nhung chua cd dieu kien de thuc hien. Trong cac benh ung thu mau, theo thdng ke,
ung thu mau khdng Hodgkin la dang phd bien va xay ra ca d
liia
tudi thanh thieu nien. Viec
nghien
ciiu
va san xuat cac dugc chat phdng xa khang the don ddng tai Viet Nam, phuc vu
cho chan doan va dieu tri trong nudc la viec lam cd y nghia ldn cho xa hdi, cho cac benh
nhan ung thu.
Hien nay, trong nudc cac phdng thi nghiem tai cac trudng dai hpc Y, tai Vien Cdng
Nghe Sinh Hpc Viet Nam dang thuc hien cac nghien
ciiu
ve san xuat khang the don ddng cho
chan doan va dieu tri benh ung thu. Vi vay
ITnh
vuc nghien
ciiu
danh dau khang the don ddng
trong nudc se cd dieu kien nghien
ciiu
va
iing
dung cho y hpc.
II.
Muc tieu nghien
cihi

Muc tieu cua de tai la hoan thanh quy trinh dieu che va kiem tra chat lugng dugc chat
ca khang the dcm ddng khang CD20 danh dau vdi ddng vi phdng xa
'^'l:
Hoan thanh quy trinh danh dau khang the don ddng khang CD-20 vdi ddng vi phdng
xa
'^'I.
- Hoan thanh cac quy trinh kiem tra chat lugng nhu kiem tra hieu suat danh dau, dp
sach hda phdng xa, kiem tra dp dn dinh thudc, dp vd khuan, chi nhiet td, ndi ddc td vi khuan,
cac kiem tra chat lugng sinh hpc nhu su phan bd thudc, su dao thai.
- U'ng dung lam sang chup hinh phan bd thudc tren benh nhan ung thu mau khdng
Hodgkin (NHL) phase I
III.
NQI
dung nghien
cihi:
Thuc hien 4 ngi dung chinh sau
1) Tinh che khang the don dong
Khang the don ddng don ddng khang CD20 dugc mua d dang tinh khiet, san sang
dimg trong y hpc, dugc phan tich kiem tra bang sac ky
Idng
cao ap FPLC.
2) Danh dau khang the
doti
dong vdi dong vj phong xa
''''l
Day la cdng doan quan trgng nhat de tao ra san pham chinh trong de tai, gdm cac budc
sau:
- Chpn phuong phap chloramin T (ChT): Diing chloramin T lam chdt oxy hda va natri
metabisulphit (SMB) lam chat khu.
- Khao sat hieu suat danh dau va cac yeu td anh hudng den hieu sudt danh ddu: Khao

sat cac dieu
ki?n
tdi uu de dat hieu suat danh ddu hon 90%.
- Khao sat ham lugng Chloramin T can
thilt
trong qua trinh danh ddu vdi phdng xa.
- Khao sat thdi gian phan
iing
tdi uu
dk
gdn khang
th6
vdi phdng xa.
- Ty le khang the va hoat dp phdng xa
'^'l.
- Tach hon hgp phan
iing
qua cdt
Ipc
gel Sephadex G-25: Hgp chdt danh ddu se cd
nhiSu
thanh phdn, do vay hon hgp dugc tach phan doan tinh
khilt
khang
th8
gdn vdi ddng vi
phdng xa ra khdi cac thanh phan khac. Day la cdng viec rat quan trgng trong viec tao ra san
pham tinh khiet dap
iing
yeu cau cho sir dung lam sang.

3) Kiem tra chat
Iirong
khang the danh dau phong xa
Thuc hien kiem tra chat lugng nhu dp sach hoa phdng xa, hieu suat danh dau, tinh che
san phdm danh ddu qua cdt Sephadex, kiem tra tinh dn dinh cua san pham, kiem tra chat
lugng sinh hpc dugc chdt phdng xa gdm vd trimg, chi nhiet td, phan bd sinh hpc tren ddng vat
de san phdm dat tieu chuan chat lugng cua dugc chdt phdng xa.
4)
iTng
dung chup hinh phan bo thuoc tren benh nhan ung
thir
Thuc hien phase I lam sang, sir dung che pham khang the don ddng danh dau phdng
xa tren benh nhan ung thu mau dang ung thu lympho bao B khdng Hodgkin cho muc dich
chup hinh phan bd thudc, chan doan va dieu tri benh ung thu mau khdng Hodgkin.
10
Phan I TONG QUAN TAI LIEU
I. Gidi thieu nhung tien bo moi trong dieu trj mien dich phong xa RIT
Da qua hon hai thap ky,
kl
tir khi bao cao dau tien cua khang the dugc sir dung danh
ddu vdi
^""Tc dk
chup
hirih
vi tri di can ung thu
true
trang, viec chan doan va chira benh bang
khang thl khdng ngimg phat
triln
va ddi mdi. Lieu phap dieu tri mien dich phdng xa RIT

(radioimmunotherapy) da
tiln
hda vugt bac nhd
sir
phat trien sinh hgc phan tir va cdng nghe
sinh hpc. Khang thl don ddng dugc san xuat vdi
miic
tinh khiet cao
cd
the diing tren lam
sang [1]. Ngay nay, ciing vdi cac phuong phap chiia ung thu truyen thdng, RIT gdp phan
dang kl trong viec chiia tri cac ung thu tai phat hoac khdng dap
iing
vdi hda chat. Sau day la
vai khai niem trong
linh
vuc dieu tri mien dich phdng xa RIT.
1.1.
Cac khai niem
Khang the: La cac phan tir dugc tao ra bdi he thdng mien dich cua co the, hinh
1.1
cho
thdy khang thl cd thl nhan bilt va tieu diet cac chdt ngoai lai nhu la vi khudn, virus. Khoa
hpc ngay nay cd thl san xudt ra khang thl
dl
nhan bilt nhiing bia dac hieu hoac nhirng khang
nguyen dac hieu hien dien tren cac te bao ung thu dac biet [2].
Khang nguyen
Hinh 1.1. Khang the tao ra bdi he thdng mien dich cua co the
X

Khang the don dong: La nhirng khang the mien dich,
true
tiep chdng lai mdt yeu td
khang nguyen don ddc, hoac cac bieu vi tren phan
tii
khang nguyen [3]. Tir khi dugc san
xuat, khang the don ddng da cd nhieu iimg dung trong cac
ITnh
vuc nhu y hpc, chan nudi,
cdng nghiep Dac biet la trong chan doan va dieu tri benh, khang the don ddng khi gan vdi
chat phdng xa hoac hoa chat, sau khi tiem vao ngudi cd the phat hien sdm benh ung thu, phat
hien
sir
lan truyen ung thu, phan biet md lanh tir nhiing md ac tinh, ngan chan va loai trir
sir
phat trien cua ung thu, dieu tri cac ung thu ac tinh [4].
Dieu trj bang khang the don dong: La phuong phap dieu tri hieu qua ddi vdi te bao
ung thu nhung
miic
ddc thap nhat ddi vdi te bao binh thudng. Cach dieu tri la tiem khang the
vao tTnh mach benh nhan. Trong mau, khang the di khap noi trong co the va gan vao cac
khang nguyen dac hieu tren te bao ung thu [5]. Hoat ddng nay ciing bao ddng he thdng mien
dich
CO
the nhan biet va tieu diet cac te bao cd dinh hudng. Mac
dii
mdt vai
tl
bao binh
thudng cung bi tac ddng dac hieu gan khang the don ddng, nhung co thl se

dilu
chinh va
11
thay thl cac tl bao nay sau khi dieu tri. Cach dieu tri bang khang the don ddng la mdt dang
dilu tri mien dich thu ddng, bdi vi khang the tao ra sd lugng ldn ben ngoai co the
chii
khdng
phai do dap
iing ciia
he thdng mien dich co the [6].
Dieu tri mien djch phong xa RIT (Radioimmunotherapy) la gi? RIT la phuong
phap dieu tri ung thu gdm ca viec gan khang the don ddng vdi ddng vi phdng xa
[7].
Khi mdt
khang the don ddng gan vdi ddng vi phdng xa gpi la danh dau phdng xa. Vdi RIT, khang the
gan phdng xa se nham dich vao khang nguyen dac hieu tren te bao ung thu, phat lieu phdng
xa
true
tiep vao ung thu, lieu
biic
xa khdng chi diet te bao nd gan vao, ma cdn diet ca te bao
xung quanh gpi la hieu
iing
"crossfire" [8].
1.2.
Khang the
dirtfc sv
dung chira benh nham dich va chup hinh chan doan
Vao dau the ky 20, Pau Ehrlich, nha hoa hpc tri lieu ngudi
Diic

da dat ra ten gpi la "hat
ky dieu" de giai thich y tudng diing he mien dich
ciia
co the de nham dich dac hieu vdi cac
receptor tren vi khuan [9]. Ngay tir nhiing nam
1950
cac nha khoa hpc ngudi My nhu tien sy
David Pressman, William Bale, Irving Spar da nghien
ciiu
y tudng dinh vi ung thu cd chpn
loc bang
each
tao khang the khang cac receptor tren ung thu d loai gam nham. Hp da danh
dau phdng xa '
'l
vdi cac khang the nay va thay
dupe
ung thu bang
each
do hoat dp phdng xa
tap trung cao trong khdi u, sir dung may dem phdng xa va so sanh vdi md binh thudng [9].
Cac khang the la cac glycoprotein sinh ra bdi cac te bao B (hinh
1.2)
de chdng cac benh
viem nhiem va cac the sinh vat ngoai lai nhd he thdng tien hoa qua hang trieu nam de bao ve
ddng vat theo mdi trudng sdng [2].
Khang the
^=~^_</
s^
Te bao B

.i^d^fe,
^"^/
•Dai thu'c bao
Hinh 1.2. Dap img mien dich cua co thl
Nhu la mdt phan
ciia
he dieu hoa
miln
dich, khang thl
cQng
dugc sir dung nhu vacxin
de dieu tri, ngan chan cac viem nhiem va dugc phat
triln
dl chdng lai cac benh khac, kl ca
b?nh
ung thu. Khang the dugc md ta la phan tir cd hinh dang nhu chtr Y, hai canh tay cua chii
Y la vi tri gan cua cac
thu
the (receptor), hoac cac khang nguyen dac hieu. Dilu nay da ggi
len y nghT la cac khang
the
nay se gan vdi khang nguyen ciing loai khi dua vao trong co thl
[2].
Cai dudi cua khang the tuyen cac te bao miln dich hoac cac lymphocyte va cac bd thl,
12
cac the nay ndi tiep vdi cac khang the tuan hoan di den cac chd viem nhiem hoac
true
xuat
cac the ngoai lai [9].
Cho den nhung nam 1980, cac nghien

ciiu
diing khang the danh dau phdng xa
'^'l,
'^'l
cdn bi han che, nhung
ITnh
vuc nay da trd nen hieu qua hon nhd nhiing ddng gdp
ciia
cac nha
hoa hpc nhu Hnatowich (trudng dai hpc Massachusset) da sir dung phdi tir hoac hda chat gan
kim loai de gan khang the vdi cac ddng vi phdng xa,
each
nay tao ra nhieu kieu nhan phdng
xa gan vdi khang the de phat hien hoac dieu tri benh nhu la cac nhan phdng xa
'"in,
'"Y
[10].
Sau khi chimg minh dugc la cac khang the tao ra tir ddng vat (la khang the da ddng) cd
the phan biet cac ung thu phat trien trong chupt vdi cac cac md binh thudng, van de
thii
thach
la chuyen
iing
dung nay ddi vdi benh nhan ung thu,
tiic
la sir dung khang the khang khang
nguyen ung thu tren co the ngudi va cd the dinh vi tren cac khdi u nay.
Nam 1972, cac nha khoa hpc quan sat mot ung thu rudt d ngudi cd bieu hien khang
nguyen ung thu bieu md phdi (CEA: carcinoma embryo antigen) [9]. CEA dugc phat hien
dau tien nam 1965 bdi Phil Gold va Samuel O. Sam Freedman (trudng dai hpc McGill,

Montreal, Canada) tir dich chiet md ung thu rudt d ngudi. Cac nha khoa hpc suy luan rang
khang nguyen nay phai la mdt dich chpn loc trong qua trinh dua chat phdng xa den u rudt khi
khang the khang CEA dugc gan vdi nhan phdng xa. Vi the nam
1973,
ciing vdi F. James
Primus va Hans J. Hansen, PhD., cac nha nghien ciru tren lan dau tien thay rang, ung thu
ngudi cd the phat trien trong co the ddng vat thi nghiem, bang
each
dinh vi ung thu diing
khang the khang CEA gan phdng xa
[10,
11].
Dieu nay dua den giai doan nghien
ciiu
lam
sang, sau khi phat trien va tinh sach CEA tdi
miic
lam sang, va danh dau vdi
'^'l
(mdt ddng vi
phdng xa dimg cho con ngudi de phat hien va dieu tri benh tuyen giap), cac nha nghien
ciiu
da bat tay vao chiing minh rang ung thu trong benh nhan dugc phat hien va that
sir
dinh vi
vdi khang the gan phdng xa [9].
Cac bao cao ve sau
(1978)
cho thay, cac ung thu tao ra CEA va nhirng ung thu khac
nhu la ung thu vii, bang quang, phdi ciing san sinh ra CEA

dii
de cd the phat hien bdi cac
khang the phdng xa. Quan sat nay da cho ra ddi phuong phap sir dung khang the phdng xa de
phat hien benh, cac tac gia dat ten la Radioimmunodetection hoac Radioimmunotherapy [9].
Trong nhirng nam ke tiep, nhieu khang the don ddng mdi dugc phat hien, cac ddng vi
phdng xa khac ciing dugc dimg, cac khang the cd dang nhd hon ciing dugc sir dung td ra
thich hgp cho con ngudi. Tir khi nguyen tac dinh vi va nhin thay CEA bieu hien ung thu
dugc chiing minh, ky thuat phat hien mien djch phdng xa (radioimmunodetection) dugc
iing
dung phat hien cac ung thu va cac benh khac ngay ca benh ddng tu mau, nhdi mau co tim,
viem nhiem. Khi gan vdi
^^""Tc,
cho thay vi tri viem
nhilm,
viem tuy xuong, viem rudt thira,
da cung cap thdng tin chan doan cang nhanh hon [9].
Vao nam
1986,
lan dau tien FDA phe chuan cho sir dung 0KT3 ma ban chat la khang
the IgG2a chupt khang CD3 diing trong phdng thai ghep d nhung benh nhan ghep npi tang.
Den nam 2009, da cd 23 san pham cho phep dieu tri (bang
1.1)
va hang tram san pham dang
thvrc hien giai doan lam sang [9,
12].
13
Bang 1.1. Cac Khang thl don ddng dugc FDA phe chuan
TSn
sdn pham,
khdng

thi \k
cong ty
Orthoclone
0KT3;
muromonab-CD3
(Ortho Biotech
•
Products, L.P.)
ReoPro; abciximab
(Centocor, Inc.)
Panorex; edrecolomab
(GSK/Centocor
company)
Rituxan; rituximab
(Genentech, Inc.
vd
Biogen
Idee
Inc.);
MabThera
(F.Hofftnan-
LaRoche Ltd)
Zenapax; daclizumab
(F.Hoffman-LaRoche
Ltd)
Simulect; basiliximab
(Novartis AG)
Synagis; palivizumab
(Medlmmune
Inc. vd

Abbott Laboratories)
Remicade; infliximab
(Johnson & Johnson
and Schering Plough
Corporation)
Herceptin; trastuzumab
(Genentech, Inc. vd
F.Hoffman-LaRoche
Ltd)
Mylotarg; gemtuzumab
ozogamicin (Wyeth)
Campath; aiemtuzumab
(Genzyme Corporation
and Schering AG)
Zevalin; ibritumomab
tiuxetan (Biogen Idee
Inc.)
Humira; adalimumab
(Abbott Laboratories)
Xolair; omalizumab
(Genentech, Inc. vd
Novartis AG)
Bexxar;
'"l-
tositumomab
Bdn chat
IgG2a
chuot
Fab lai
IgG lai

IgGl
lai
IgG 1
nhan
hoa
IgGl lai
IgG 1
nhan
hoa
IgGl lai
IgGl nhan
h6a
IgG4 lai,
gan
calicheami
cin
IgG 1
nhan
hoa
IgGl
chuot,
danh dau
IgG 1
nhan
hoa
IgG 1
nhan
h6a
IgG2a,
chupt,

Khdng
nguyen
CDS
gpllt^
gpllla
va
OVPJ-
integrin
CA17-1A
CD20
CD25
CD25
RSVgpF
TNF
ERBB2
CD33
CD52
CD20
TNF
IgE
CD20
Nim
1986
1994
1995
1997
1997
1998
1998
1998

1998
2000
2001
2002
2002
2003
2003
Km
(nM)
0.83
5
-
8.0
0.3
0.1
0.96
0.1
5
0.08
10-32
14-18
0.1
0.17
1.4
sir dung trong dieu tri
Cache
hieu irng
Khoa chuc ndng te bdo T
bieu hien CD3; chong
thai loai cay ghep

Gdn thu the va doi
khdng; kim ham ket tu
tieu huyet cau
Gdn thu the vd doi khdng
Lam te bdo nhay vai hoa
cdm ung apoptosis,
ADCC va CDC
Gan thu the vd doi khdng
Gdn thu the vd doi khdng
Gdn vd trung hoa RSV;
kim ham
sy
tdi bdn vd
dung hop
Gdn phoi
tii
vd doi
khdng thy the
Ldm te bdo nhay vai hoa
trj;
kim hdm hinh thanh
mach vd tdng sinh, cdm
ung ADCC
Cam ung phd va DNA
mach kep vd ldm chet te
bdo (do calicheamicin)
Cdm ung ADCC va
CDC
Gay chet te bdo bdng
phong xa; cdm ung

apoptosis
Gdn phoi tu vd doi
khdng thy
th6;
cam ung
CDC
Gdn phoi tu vd doi
khdng thy the; gidm gidi
phong chat hieu ung vdi
dap ung di
iing
tir te bdo
bach cau ua kiem va
duOng
bdo
Gay chet
t§
bdo bdng
phong xa; cam ung
Tri benh
Phong thai loai
ghep than cap tinh
Ngdn dong mdu
do tieu huyet cau
trong phdu thuat
dong mach vdnh
Ung thu rupt ket
U lympho khong
Hodgkin vd viem
khap dang thdp

Phong thdi loai
ghep than cap
tinh
Phong thai loai
ghep than cap tinh
Chong nhiem
RSV
a
tre c6
nguy ca cao
Benh Crohn, thdp
kh6p,
vdy
n6n,
viem
lo6t
rupt ket
vd viem d6t song
cung
Ung thu vu di cdn
dang sieu bieu
hien
ERBB2
Ung thu bach cdu
dang tuy bieu hien
C'D33
Ung thu bach cdu
lypho
mdn tinh te
bdoB

U lympho khong
Hodgkin
Thdp khdp vd vay
nen
Hen suyen lau ddi
U lympho khong
Hodgkin
14
(GlaxoSmithKline)
Raptiva; efalizumab
(Genentech, Inc. vd
Serono S.A.)
Erbitux; cetuximab
(ImClone Systems Inc.
vd Bristol-Myers
Squibb Company)
Avastin; bevacizumab
(Genentech, Inc. va
F.Hoffman-LaRoche
Ltd)
Tysabri; natalizumab
(Biogen
Idee
Inc. vd
Elan Corporation,
pic)
Panitumumab
Vectibix
Ranibizumab
Lucentis

Natalizumab
Tysabri
Eculizumab Soliris
ddnh ddu
IgG 1
nhdn
h6a
IgGl lai
IgG 1
nhdn
h6a
IgG4 nhan
h6a
human
humanized
humanized
humanized
CDI
la
EGFR
VEGF
Dual dan
vj
a4-cua
a4pi-
integrin vd
a4p7-
integrin
EGFR
VEGF

2003
2004
2004
2004
2006
2006
2006
2007
3
0.2
1.1
0.3
apoptosis, ADCC vd
CDC
Gdn thy the vd doi
khdng; ngdn bach cau
gdn vai cdc te bdo khdc
Gdn thy the vd doi
khdng; kim ham te bao
tdng sinh; cam irng
apoptosis; ldm te bdo
nhay vai hoa trj va xa
tri;
kim ham hinh thdnh
mach, sy xam lan va di
cdn; cdm ung ADCC
Gdn thu the vd doi khdng
phoi tu; kim ham sy
hinh thdnh mach vd sy
tien trien cua benh di cdn

Gdn thy the va doi
khdng; kim hdm bach
cau bam dinh vai cdc thy
the doi lap ciia chung
epidermal growth factor
receptor
vascular endothelial
growth factor
T
cell
VLA4 receptor
complement system
protein
C5
Vay nen dang
mdng
Ung thu rupt ket
di cdn, ung thu
ddu vd
CO
Ung thu rupt
k6t
di cdn
Xa hoa lan toa
Colorectal cancer
Macular
degeneration
Cdc viem nhiem
(dieu trj cdc xa
cimg lien quan

den benh ty miln)
Cdc viem nhiem
paroxysmal
nocturnal
hemoglobinuria
K^:
di lyc gdn khdng nguyen
Cac kieu
siir
dung khang the trong dieu tri ung
thvt
De dieu tri ung thu, khang the dugc sir dung cd the gan ket hoac khdng gan ket. Tuy
theo
each
gan ma tao ra nhieu kieu dieu tri ung thu khac nhau. Hinh
1.4
minh hga cac
kilu sii
dung khang the trong dieu tri.
Khang the tran: La dung khang the khdng gan vdi mdt chat nao trong dieu tri ung
thu. Cac
CO
che
true
tiep tieu diet te bao ung thu khi diing khang thl
tr4n
la co
chi lie chi
tac
dung sinh hgc cua khang nguyen be mat, co che kich thich

sir chit
cd chuong trinh
ciia
tl bao
(apoptosis). Co che gian tiep la co che miln dich, dd la gay ddc tl bao phu thudc bd the
(CDCC), gay ddc tl bao phu thudc khang thl (ADCC), co
chi
gay ddc phu thudc bd thl
(CDC).
Vi du cac khang the tran dugc FDA cho phep sir dung nhu Rituximab la khang thl
khang CD20, Trastuzumab (Herceptin) la khang thl khang protein HER2, Alemtuzumab
(Campath) la khang the khang CD52, Cetuximab (Erbitux) la khang thl khang EGFR protein,
Bevacizumab (Avastin) la khang the khang protein VEGF
[13].
Nhirng diem hp che cua khang thl khdng danh dau: Tat ca cac tl bao ung thu
diu
cSn
gin vdi khang thl. Co
chi
dap
iing
miln djch cua benh nhan khdng thl khdng bi anh
15
hudng khi tuong tac bdi khang the. Cac te bao ung thu cd thl de khang vdi cac co che khang
ung thu
true
tiep ciing nhu co
chi
mien dich
ciia

khang thl [4].
1
BChdng
thl tran
Nhan phdng xa
2 KMng
the cdng hgp
Toxin
. Avidin - Biotin
3
Kh^g
thl gin nhieu budc
Tebao
ung
thir
\^r^
Bispecific
Sc-Fv-enzym
%
Prodrug Drug
Hinh
1.3. Ung dung khang the trong dieu tri mien dich
Khang the cong hop: La khang the ket hgp vdi hda chat hoac ddc chat, chat phdng
xa, liposome, cytokin, sc-Fv enzym
Tiiy
thudc vao chat gan vdi khang the ma tao ra cac
each gpi ten khac nhau nhu:
- Khang the gan vdi hda chat dieu tri lam sang gpi la khang the gan hda chat
(chemolabeled antibodies): Kieu nay hien dang sir dung nhieu tren lam sang [8].
- Khang the gan vdi phdng xa gpi la kieu khang the danh dau phdng xa: Kieu dieu tri

dimg khang the loai nay la radioimmunotherapy, vi du nhu Ibritumomab tiuxetan (Zevalin),
Tositumomab (Bexxar) hien dang dimg trong dieu tri u lympho B khdng Hodgkin. Ben canh
do,
khang the gan phdng xa ciing dugc diing de phat hien
sir
lan rdng
ciia
ung thu trong co
the nhu OncoScint (ung thu
true
trang va ung thu budng tning), ProstaScin (ung thu tuyen
tilnliet)[15].
- Khang the gan vdi ddc td gpi la ddc td mien dich (immunotoxin), chat ddc diing de
g5n
vdi khang the thudng cd ngudn gdc tir thuc vat hoac vi khuan. Vai loai ddc td vi khuan
nhu diphtherial toxin (DT) hoac pseudomonal exotoxin (PE 40), hoac ddc td tir thuc vat nhu
ricin A, saponin. Cac nghien
ciiu
lam sang da cho thay cd su co lai trong vai ung thu, nhung
nhieu van de chu yeu can giai quyet trudc khi sir dung ddc td mien dich rdng rai tren lam
sang. Chi duy nhat gemtuzumab ozogamicin (Mylotarg) la ddc td mien dich dugc FDA phe
chuan cho sir dung lam sang, chat nay chiia ddc td calicheamicin, gan vdi khang the khang
CD33,
dimg trong dieu tri ung thu bach cau
tiiy
cap (acute myelogenous leukemia AML).
16
Ddc td mien dich cdn dugc
iing
dung trong dieu tri cac ung thu lympho, u nao va cac ung thu

khac [16].
Cai tien dieu tri bang khang the don ddng: Hieu qua
ciia
dieu tri cdn nhieu han che bdi
khang the san xuat tir tap doan te bao lai
ciia
chupt. Sau thdi gian dieu tri, he thdng mien dich
cua
CO
the phat hien khang the la chat ngoai lai va tieu diet ngay khi dua vao co the. Vi ly do
nay ma cac nha khoa hpc da gan gen mang doan hoat tinh
ciia
khang the chupt vdi gen mang
khang the ngudi. Viec san xuat gen khang the chupt - ngudi gpi la khang thl don ddng kham
"chimeric" hoac "humanized" [17]. Khang the mdi tao ra nay gidng nhu khang the ngudi, vi
vay khdng bi he thdng mien dich co the phat hien. Cach dieu tri mdi hon niia la diing mdt
doan khang the thay vi diing ca phan tir. Cac doan khang the nhd nay cd the di den ung thu
tdt hon, vi vay viec chira tri se hieu qua hon.
Khang the gan nhieu
bu'd'c
(multistep targeting): La phuong phap mdi dat ra nhim
khac phuc su
ehiu
lieu cua nhiing te bao lanh binh thudng trong qua trinh luu thdng cua
khang the gan phdng xa, ddng thdi ciing lam gia tang hieu qua dieu tri. Cd nhieu phuong
phap de tien hudng dich, dd la diing khang the gan vdi biotin va streptavidine (Ab/SA). Hgp
chat nay tiem vao cd kha nang dinh vi len te bao ung thu. Sau khi tich liiy cue dai, Ab/SA di
vao vi tri khdi u. Tiep theo la cho lieu
^"Y-DOTA-Biotin.
^°Y-DOTA-biotin

nay cd kich
thudc nhd nen xam nhap,
Ipt
vao khdi u nhanh chdng, tai dd nd gan vdi phiic SA-Ab, phat
lilu
phdng xa den ung thu. Phan tir DOTA biotin khdng phdng xa nhanh chdng dao thai ra
khdi mau va qua nudc tieu ra ngoai. Phuong phap nay hieu qua va giam thdi gian
ehiu
lieu
phdng xa cua co quan lanh [8].
E3l4u
trj ba
bu'cS-c tl6n
hu-6-no Ctfch
(pretargeting)
I
\^
Bu'ffc
I:
Tiem Bio-IgG
BifoclliTiemStAv,
dao
thai StAv-IgG
ra
ktioi
mau
BLro-c
III:
Tiem
Biotini

deinh diu
phong
xa
Hoat do phong xa loc qua
th^n Vci th^i
ra
kh6i cff thS
i^B^"
Hinh
1.4. Khang the tien hudng dich
Mdt phuong phap
tiln
hudng dich khac nira la dimg enzym gin khang thl dac hieu,
sau khi tiem dua thudc tien than (prodrug) chua hoat hoa vao, khi vao dich, thudc tiln than
dugc enzym hoat hoa, chuyin thanh thudc, tieu diet ung thu. Phuong phap nay gpi la lieu
phap khang the
kit
hgp enzym va thudc tiln than chdng ung thu ADEPT (antibody-directed
enzyme prodrug therapy) [9].
17
L3.
Cac tac nhan chup hinh moi trong dieu trj RIT
Tir khi dugc gan vdi cac chat phdng xa, cac khang the, va ca nhirng doan khang the,
khang cac khang nguyen khac nhau lien quan
din
cac loai benh tat da dugc phat triln dl hien
hinh dac biet cac loai benh nay
[17,18].
Cac benh diing khang thl danh diu phdng xa dl chup
hinh nhu ung thu

true
trang (satumomab pendetic, arcitumomab), ung thu
tuyin
tiln liet
(capromab, pendetide), u lympho khdng Hodgkin, ung thu gan, nhdi mau co tim (imciromab
penlefate) nhiem triing va viem (sulesomab, fanolesomab), nhiem trimg dac hieu va tic mach
mau. Dong vi phdng xa phd
biin
la
^'""Tc,
'"in,
'^''l,
^^Ga,
^^Cu
va
'^F
la tac nhan cho chup
hinh PET va phat triln phuong phap immunoPET
(hinhl.5),
se cho ra nhiing hinh anh cd dp
phan giai cai tiln dang kl khi diing vdi ddng vj
ciia
PET. Steven M. Larson MD, trung tam
ung thu Memorial Sloan - Kettering (My) la ngudi diu tien dua ra phuong phap nhim dich
cac ung thu diing khang thl danh diu vdi ddng vi phdng xa phat positron dimg
I
[9].
Hinh
1.5. May PET va chup hinh immunoPET
1.4.

Ung dung RIT trong dieu trj benh u lymphoma khong Hodgkin.
Benh u lympho khdng Hodgkin la dang ung thu cac te bao lympho B trong he mien
dich. Cimg vdi benh Hodgkin, la cac benh ve cac hach bach huyet, benh khdng Hodgkin la
b$nh
ciing gidng nhu benh ung thu bach cau dang nguyen bao lympho cap Acute
Lymphobllastic Leukaemia (ALL), cac te bao trong he bach huyet khiem khuyet va phat triln
vd dp xam chiem tuy xuong va ngan chan hoat ddng san xuat cac te bao quan trpng khac
trong mau, gay thieu mau, dau xuong va nhiem triing Day la mdt loai benh phd
biin,
xay
ra theo mpi
liia
tudi. Cac te bao ung thu nay mang nhiing dau hieu dac trung bat thudng, dd
la khang nguyen. Khang nguyen CD20 dac biet xuat hien 50.000 - 200.000 vi tri tren phan
tii
lympho bao B ac tinh (hinh 1.6). Do vay, khang the don ddng khang CD20 la cdng cu ly
tudng de nhan biet nhirng khang nguyen tren nhirng te bao ac tinh dd va tieu diet.
Khai niem ve dieu tri nham dich da dugc cac nha khoa hgc dua ra tir nhieu nam trudc.
Vao
nam
1953,
Pressman va Komgold da chiing minh rang cac khang the cd the nham dich
dSc
hieu len te bao ung thu [9]. Tuy vay, phai tdi nam
1975,
khi Kohler va Milstein nhan giai
thudng Nobel cho cdng trinh nghien ciiru
ciia
hg ve ky thuat tao te bao lai hybridoma, viec san
xuit khang the don ddng nham dich khang nguyen dac hieu mdi bat dau di vao hien thuc [9].

18
Nam
1979,
Nadler va cdng
sir
da dieu tri benh nhan u lymphd ac tinh dau tien bang khang the
don
ddng
[19].
Hinh 1.6. Te bao B ung thu cd bieu
hien khang nguyen CD20
Sir
dap ling dieu tri phu thudc vao nhieu yeu td nhu the tich khdi u, tdc do phat trien,
muc bieu hien khang nguyen, lieu hap thu
biic
xa vao khdi u, dp nhay cua tia
biic
xa Vi
vay, cin chup hinh RIT vdi
'^'l
cho NHL. Ndi chung, viec dieu tri ung thu dugc che ngu bdi
3 hinh thiic la giai phiu,
biic
xa va hoa tri. Neu ung thu dugc phat hien sdm khi nd dugc
dinh vi, giai phau hoac
hire
xa, cd the
dii
de kiem soat va loai trir triet de benh. Dieu tri
biic

xa
diln
hinh gom chilu xa ngoai len viing dac biet
ciia
co the cd khdi u, hoac "hat gidng"
biic
xa se phat tia trong viing ap sat ung thu (brachytherapy) dang tiec la ung thu thudng dugc
phat hien sau khi nd da di can khdi viing ung thu gdc den cac md va co quan khac. Dieu tri
bing hoa chat la phuong phap dieu tri khdi dau, khang the dugc dimg de gan vdi thudc khang
ung thu, hoac chat phdng xa, de di sudt co the va phan phdi lieu phdng xa den cac te bao ung
thu.
Trong 30 nam qua, ti le mac phai benh NHL ngay cang tang d My, trd thanh mot ung
thu diing hang
thii
5 va nguy co tir vong diing hang
thii
sau d My. Vdi gan 270.000 ngudi
My hien nay, udc tinh cd 60.000 ca mdi dugc chan doan va gan mdt nira trong sd dd dugc
dilu tri. Lymphocyte binh thudng va ac tinh bieu hien cac receptor khac nhau, chiing la dich
cho cac khang the don ddng. Gerald DeNardo, MD, Sally DeNardo, MD, trudng dai hpc
California, Davis, la nhimg ngudi dau tien da chiing minh viec tri lieu thanh cdng NHL vdi
khang the phdng xa [5]. Oliver Press, da tien phong trong viec diing phdng xa lieu cao
ciia
'^'l
gin khang the khang CD20, de dieu tri NHL. Ong va cdng su da chiing minh tinh kha thi
va thuan lgi cua phdi hgp giiia RIT va hda tri. Thuc ra la dieu tri hudng dich NHL dimg
khang thl khang
CD
19,
CD20 chupt (khdng gan phdng xa) da dugc nghien

ciiu
tir hon 20
nSm
nay, nhung tir khi phat trien ky thuat khang the khang CD20 kham (chimeric) giiia ngudi
va chupt, gpi la rituximab, da tao ra mdt cudc
each
mang trong dieu tri NHL va lam nen tang
cho nhung nghien
ciiu
xa hon trong viec diing khang the don ddng de dieu tri cac ung thu
khac [5].
Dieu tri NHL diing Rituximab cho thay su thuyen giam mdt
each
tdng the trong mdt
mia sd benh nhan NHL cd dp ac thap (indolent) trong phac dd phdi hgp dieu tri vdi hda chat,
nd cd the cai thien hieu qua hda tri, ket qua la ung thu bien mat hoan toan trong nhieu benh
19
nhan NHL. Dac biet hip dan la khdng cd cac ddc tinh nhu vdi cac thudc khang ung thu
thudng thay nhu su iirc che (suppression) cac te bao
mau.
Id
mieng va rung tdc [9].
Cac nha nghien
ciiu
nhu Thomas Witzig, Gregory Wiseman, Christine White la
nhung ngudi da cd cdng trong nghien
ciiu
lam sang cho thay nhiing uu diem cua viec dieu tri
khang the danh diu phdng xa khi so sanh vdi khang thl khdng danh dau phdng xa, va dugc
FDA cho phep sir dung lam sang san pham dilu tri ung thu ten la

^''V-ibritumomab
tiuxetan.
Cimg vdi phac do phdi hgp dilu tri rituximab vdi
^°Y-ibritumomab
tiuxetan, mdt khang the
khang CD20 ban chit chupt gin vdi ddng vi phdng xa kim loai
^"Y
da dugc dieu che. San
pham nay da thuong mai tir nam 2002 dimg dilu tri NHL
[51].
Mdt san pham khac diing de
dieu tri phdng xa mien dich dugc phep sir dung lam sang tir nam 2003 la khang the khang
CD20 gan
'•"l,
ten la
'^'
I-tositumomab
diing trong dieu tri NHL. Mac
dii
khang the nay la
khang the ed ban chat chupt, ca hai san phim deu cho thay budc cai tien dang ke de tieu diet
ung thu khi so sanh vdi khang the khdng gan phdng xa (bang
1.2)
[9].
Bang 1.2. Cac khang the danh dau phdng xa dang dugc nghien
ciiu
dieu tri
STT
01
02

03
04
05
06
07
08
09
10
09
11
12
13
14
16
17
18
19
Tac nhan
Y-ibritumomab tiuxetan
I-tositumomab
^"Y-epratuzumab
anti-CD22 IgG
^'^Bi-HuM195
anti-CD33 IgG
'^'l-BC8
IgG
^"Y
anti -Tac IgG
^VHU3S193
'"LU-J591

IgG
^V-PAM4
IgG
i3ij/90Y.cc49.deitaCH2
'25l/I31i.A33
^''Y-T84.66
anti-CEA IgG
'^'l-chTNT
'^'l-Hepama-1
IgG
'^'l-cG150IgG
'^'l
-8H9 IgG
'*^Re-bivatuzumab
'"l-425IgG
'^'l -81C6
anti-tenascin;
^"At-
81C6;^°Y-BC-2andBC4
Dieu tri
U lympho khdng Hodgkin (NHL)
U lympho khdng Hodgkin (NHL)
U lympho khdng Hodgkin (NHL)
Benh bach cau (leukemia)
Benh bach cau (leukemia)
T-cell
lymphomas; non-Hodgkin's and
Hodgkin's lymphomas
Ung thu budng triing (ovarian cancer)
Ung thu tuyen tien liet (prostate cancer)

Ung thu tuyen tuy (pancreatic cancer)
Ung thu rudt ket (colorectal cancer)
Ung thu rudt ket (colorectal cancer)
Cac ung thu cd khang nguyen CEA
(CEA producing cancers)
Ung thu phdi (lung cancer)
Ung thu gan (hepatocellular cancer)
Ung thu than (renal cancer)
CNS or leptomeningeal cancer
Ung thu dau va cd (head and neck cancer)
Ung thu nao (brain cancers) (gliomas)
U than kinh dem (glicomas)
20
Bang
1.2
tdm tit nhilu loai khang thl danh diu phdng xa trong nghien
ciiu
lam sang,
cho thay rang RIT ddng vai trd tich cue trong dieu tri da dang ung thu. Viec
lira
chpn chat
danh dau thich hgp cho dilu tri nhim dich la mdt thach thiic chu yeu, trong khi
chgii lira
nhan
phdng xa thich hgp, phuong phap danh diu thich hgp va ngay ca dang khang the sir dung
trong ky thuat nhim dich la tit ca nhiing thay ddi cin xem xet can than
tiiy
theo tinh trang
lam sang.
Ngoai ra, cac phucmg phap cai tiln de gia tang lieu

biic
xa chpn
Ipc
den khdi u can
dugc tinh den. Mdt phuong phap mdi cd hieu qua ro ret la khu tni phdng xa vao khu vuc ung
thu nhu la viing ung thu hoac trong khdi u. That vay, ca hai quan diem nay da cho thay nhirng
ket qua lam sang dang khich le khi phat lilu phdng xa danh dau khang the den cac viing
phlu
thuat trong dieu tri ung thu nao va ung thu
vii
da dugc md ta bdi Pietro Riva, Giovanni
Paganelli, Vien Nghien
ciiu
ung thu Chau Au, Milan, Italy. Hoac la dua phdng xa true tiep
vao khoang bung (Ruby M. Meredith, MD, PhD, trudng dai hpc Alabama, Birmingham) [9].
II.
Cac thanh phan co ban trong RIT
Cd ba thanh phan co ban trong RIT, dd la khang nguyen, khang the va ddng vi phdng
xa. Sau day la cac tinh chat va dac diem cua ba thanh phan dimg trong dieu tri RIT:
II.l. Khang nguyen CD20, vai tro va tiem nang
irng
dung
He mien dich thu dugc la mdt co che phdng ve vd ciing hieu qua tdn tai trong hau het
cac loai ddng vat cd xuong sdng bac cao. He thong nay tao ra cac dap
iing
bao ve nhanh, dac
hieu cao, khang lai mam benh trong mdi trudng sdng. De cd the tao ra dap
iing
khang lai bat
cii

vi sinh vat nao, dau tien he mien djch can vi sinh vat dd. Day la diem ma he mien djch thu
dugc khac vdi cac he thdng khac trong co the, vdi kha nang nhan biet cac loai phan tir khac
nhau tao thanh vi khuan. Cac thanh phan dugc nhan ra nay gpi la khang nguyen. Cac co chat,
hoac cu the hon la cac phan tir dugc nhan ra bdi mdt hoac nhieu thanh td
ciia
dap
iing
mien
djch thu dugc gpi la khang nguyen [2].
He mien djch dugc tien hda de nhan ra tat ca cac phan tir ngoai lai khdng phai
ciia
vat
chu, ve mat nguyen ly, cac khang nguyen bao ham mdt phd rdng bao
triim
tat ca cac nhdm
phan tir hiru co, tir cac hgp chat hda hpc don gian nhat cho tdi cac dai phan tir phiic tap nhat.
Khang nguyen tren te bao ung thu phan bd khdng ddng nhat, bieu hien vdi mat dp
cao,
cd the gan dac hieu vdi khang the don ddng, khdng thay ddi hoac dieu chinh sau khi gan
vdi khang nguyen, khang nguyen dich khong luu thdng trong he tuan hoan hoac rcri ra he tuan
hoan. Sau day la vai dac trung
ciia
khang nguyen CD20.
II.l.l.
Dac
trung
cau
true
CD20,
cdn cd ten khac la

BI,
la mdt khang nguyen dac hieu don ddng te bao lympho
B,
bieu hien tren te bao lympho B d mdt sd giai doan nhat djnh. CD20 la mdt protein khdng
glycosyl hoa cd khdi lugng phan tir khoang 33-35 kDa. Gene ma hoa CD20 nam tren nhiem
sac the sd
11,
canh dai, locus
12-13.
Cau tnic va vi tri ciia CD20 dugc md ta trong hinh
1.7:
21
Viiig
xuyen
dng
CD40
QCXJUOC
. Ngoai lio
Ldp phosphol^iid
'
kep
-Baotutfng
viaig noi bdo
Hinh
1.7: Cau tnic va vi tri
ciia
CD20 tren mang te bao
CD20 la mdt protein cd 4 viing xoan xuyen mang, ca 2 dau tan
NH2
va COOH deu nam

trong bao tuong. Hai vdng ngoai bao
ciia
CD20 rat khac nhau ve kich thudc. Vdng nhd hau
nhu khdng
Id
ra khdi mang, vdng ldn cd dp dai khoang 46 axit amin, tir axit amin sd 140 den
axit amin sd 185. Tren vdng ldn cd cau
disulfid
giira axit amin sd 167 va 183. Vdng nay chiia
da sd cac epitope cua cac khang the don ddng [52].
11.1.2.
Chirc nang
Den nay, chiic nang
ciia
CD20 van chua dugc biet mdt
each
rd rang. Tren nhirng con
chupt cd te bao B khdng bieu hien CD20 khdng thay cd
sir
suy giam chiic nang te bao B ro
ret. Tuy nhien, mdt sd nghien
ciiu
da chiing minh CD20 cd vai trd trung gian trong nhieu hoat
ddng sinh hpc cua te bao B thdng qua
sir
gan khang the nhu:
Ddi vdi
sir
di chuyen
ciia

ddng canxi vao te bao: su chuyen vi tri va gan dac hieu CD20
vao cac "mang" lipid sau khi CD20 gan khang the la dieu kien tien quyet ddi vdi khdi dau su
dan truyen tin hieu va/hoac
sir
tao thanh cac kenh canxi.
Ddi vdi chu ki te bao: CD20 cd vai trd quan trpng trong dieu hoa su phat trien chu ki te
bao (cell cycle progression), it nhat la d te bao B binh thudng. Bang chiing la cac khang the
don ddng hudng dich CD20 deu gay thay ddi chu ki te bao, lam day nhanh su chuyen te bao
B tir pha
GO
sang Gl, cac khang the don ddng cimg nhdm vdi
BI
lai
lic
che su chuyen te bao
B tu pha
Gl
sang pha S/G2 va pha M.
11.1.3.
Tiem nang
irng
dung cua khang nguyen CD20
Tai My, moi nam cd khoang 55.000 nguoi dugc chan doan mic benh u lympho khdng
Hodgkin, trong dd 65% khdng dap
iing
vdi dieu tri. Benh nay hien diing
thii
6 trong cac
nguyen nhan gay chet do ung thu. Khoang 85% den 90% NHL cd ngudn gdc do tl bao B va
bieu hien cac chi thi khang nguyen be mat te bao B. Ngudi mac benh nay cd thdi gian sdng

trung binh
8-10
nam [20]. Tai Viet Nam, theo tai lieu tai Hdi Nghj Ung thu nam 2008 td chiic
tai benh vien Ung budu thanh phd Hd Chi Minh, benh NHL la mdt trong
10
loai ung thu
thudng gap, xuit do chuin theo tudi la 4,6/100.000 (nam) va 33,2/100.000 (nir). Ghi nhan
nhirng benh ung thu hay gap d tre em tai 5 tinh thanh giai doan 2001-2004, NHL diing hang
thiir hai d Ha Ndi (15%), Hai Phdng (15,4%), Thai Nguyen (14,3%), Thira Thien Hue
(16,4%),
diing
thii
ba d Cin Tho (20,9%). Tudi trung binh la 9, dinh cao nhit
7-10
tudi, dinh
tilp
theo la cudi tudi thieu nien. Ti le
nam:nii
la 3:1
[51].
22
U lympho loai nay cd thl chuyin tdi
miic
do nang hon ve mat md hgc (histologic) va
sau dd rat khd dieu trj. Benh phat triln cham va thudng thuyen giam trong nhieu nam, nhung
cudi ciing se gay tir vong cho ngudi benh.
CD20 la mdt phan tir dich
tilm
nang trong y hpc bdi khang nguyen nay chi bieu hien
tren te bao B;

miic
bieu hien cao (khoang
100.000
phan
tir/tl
bao); khdng bien dang hay ddng
hda khi gan vdi khang thl; va thudng khdng phat hien d dang hda tan [21].
6
chupt binh
thudng va chupt chuyin gen CD20
ciia
ngudi
diu
cho thiy CD20 la dich hieu qua cho
sir
lam
giam te bao B. Ngoai ra, do CD20 khdng
bilu
hien trong cac te bao gdc tao mau, khdng de
dang hi ddng hda va khdng bj tach rdi khi gin vdi khang the danh dau phdng xa nen la mdt
dich hap dan cho lieu phap miln djch phdng xa [3].
II.2.
Khang the va cong nghe tao khang the:
Khang the sir dung trong RIT trudc
hit
phai dap
iing
tinh dac hieu, va cd ai luc cao
vdi antigen, trpng lugng phan tir nhd khdng thay ddi va cd dap
iing

mien djch nhu HAMA,
HAHA, HACA
II.2.1.
Cau triic:
Cau tnic chung cua khang the gdm hai chuoi nhe va hai chuoi nang ddng nhat, gan
vdi nhau nhd cac lien ket disulfide. Cd 5 nhdm globulin mien djch (IgG,
IgA,
IgM,
IgD
va
IgE) vdi trinh tu va sd mien trong cac viing hang djnh tren chudi nang khac nhau. Cd hai
ddng phan chudi nang la
(k
va
K).
Globulin mien dich G (IgG) la loai khang the dac tnmg
trong huyet thanh binh thudng va dugc nghien
ciiu
phd bien nhat (hinh 1.8). Globulin mien
djch cau thanh tir cac don vj gap nep gdm 100 axit amin, mdi don vj cd cau tnic ddc lap
gidng nhau gpi la kieu gap nep globulin mien djch. Mien dau N
ciia
mdi polypeptide (tren
chudi nang va nhe), gpi la vimg bien ddi (V), rat khac nhau. Cac mien cdn lai cd trinh tu
gidng nhau gpi la viing bat bien (C). So sanh trinh tu viing V cho thay tinh kha bien khdng
phan bd deu ma tap trung vao ba viing gpi la cac viing sieu bien [2].
Mofv <ry»t
(b)
(«)
Hinh 1.8. Cau tnic

ciia
IgG. (a) So dd
ciia
IgG tieu bilu. Cac chudi L va H dugc thl
hien la cac ddng ke dam, vdi cau disulfide ndi phan tir lien
kit
cac gdc Cys (S-S). (b) Ciu
tnic lap the cua IgG.
23
11.2.2.
Ai
lu-c
(affinity) va do bam (avidity)
Khi khang the don ddng dugc tao ra bdi ky thuat te bao lai cd hai vj tri gan khang
nguyen va huyet thanh chiia cac khang the da ddng (cd kha nang gan vdi nhieu yeu td trinh
dien khang nguyen), khang the cd the da trj trong phan
iing
vdi khang nguyen.
Khi trong mdt khang nguyen cd nhieu vj tri vdi khang the gidng nhau (dai phan tir
hoac vi sinh vat) phan
iing
vdi khang the da tri, ai luc gan tang manh vi tat ca cac lien ket
khang nguyen-khang the phai bi pha vo ddng thdi trudc khi khang nguyen va khang the tach
ra. Do dd, nang lugng gan
ciia
khang nguyen da tri vdi cac vj tri gan khang nguyen nam tren
khang the da tri se ldn hon tdng ai luc cua mdi vi tri gan vdi mdt khang nguyen. Luc gan giiia
mot khang the da tri va khang nguyen da trj gpi la dp bam (avidity), de phan biet vdi ai
lire.
Dp bam cua mdt khang the vdi khang nguyen cua nd dugc xac djnh bdi tdng tat ca

cac tuong tac ddc lap xay ra giira cac vj tri gan khang nguyen nam khang the va cac yeu td
trinh dien nam tren khang nguyen. Dp bam
ciia
mdt khang the vdi khang nguyen cua nd phu
thudc ldn vao ai luc cua cac vj tri gan rieng biet vdi cac yeu td trinh dien tren khang nguyen.
Nd ldn hon tdng cua cac ai luc nay neu ca hai vj tri gan khang nguyen cua mdt khang the gan
vdi khang nguyen. Do dd, cho
dii
moi vj tri gan khang nguyen cd ai luc thap (nhu IgM mdi
tao ra trong dap
iing
mien djch), thi cac khang the van cd chiic nang trong he mien djch.
11.2.3.
Ban chat va
irng
dung cua khang the khang CD20 Rituximab
Rituximab la khang the khang CD20 dau tien dugc FDA phe chuan vao cudi nam
1997 su dung cho ngudi. Rituximab dugc tao thanh bang ky thuat di truyen, dung hgp vimg
bien ddi chudi nang va nhe
ciia
chupt vdi vimg hang djnh
IgGl/K
cua ngudi (hinh 1.9).
Khang the lai tao thanh cd kha nang tieu diet cac te bao B in vitro va lam giam lugng te bao
B trong mau ngoai vi in vivo. Su giam te bao B trong
tiiy
xuong,
lach
va cac hach lympho
chua dugc nghien cim ky [23].

I I
=
Vung
bien
3Si
cua chuot
••
Vung
hixig dinh CU4 ngujcyi
= Vung hing
3mh
Fc cua
ngu^
Vi
=
Vung
bien doi chuoi nhe
v
= Vung
bien
d6i
chuoi nang
Cl =
Vung hing dinh chuoi nhe
Cy =
Vung hing dinh chuoi nang
Hinh
1.9. Cau tnic cua khang the Rituximab.
FDA phe chuan Rituximab phan ldn dua tren cac thir nghiem ddi vdi u lympho te bao
B khdng Hodgkin tai phat nhe. Ty le dap

iing
trong nhdm khd chiia nay la khoang 50% khi
chi
sii
dung Rituximab. Sau dd, tren 380.000 trudng hgp da dugc kiem chimg cho thay dp an
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