TERAPI GENE

Kiagus M Arsyad
Bagian Biologi Kedokteran dan
Andrologi
Fakultas Kedokteran UNSRI
SASARAN PEMBELAJARAN
Mahasiswa mengetahui dan memahami :
1. Macam terapi gene (Viral and Non-viral),
2. Macam virus vector untuk terapi gene :
RNA viruses (Retroviruses & Lentivirus ),
DNA viruses (Adenoviruses &
Adeno associated virus )
3. Macam penyakit yang dapat diobati dengan
terapi gene dan bukan terapi gene
4. Masa depan terapi gene
MATERI PEMBELAJARAN
1. Pendahuluan
2. Status Terapi gene saat ini
3. Pertimbangan khusus untuk Terapi
gene
4. Rencana Terapi
5. Masa Depan Terapi Gene
6. Dianosis Post natal
1. Pendahuluan
1. Pendahuluan
1. Pendahuluan
1. Pendahuluan
2. STATUS TERAPI GENE SAAT
INI

The goal of gene therapy is to add,

repair, or block the expression of
genes in the treatment of inherited,
as well as non-inherited diseases.
Somatic versus Germ line gene
therapy
Ex Vivo versus in Vivo gene therapy
 2.1. Somatic Versus Germ line

1. Somatic Treatment of Cells

All 2N Cells = Body Cells

2. Germline treatment of egg and sperm

(1N) = Gametes
Very little Germline Gene Therapy
research going on
 2.2. Ex Vivo and In Vivo Therapy

Ex Vivo In Vivo
Vector
Cell Cell

Expansion
of cells Virus/Vector

Donor/ Recipient
Recipient
Manipulated Cells
 3. PERTIMBANGAN KHUSUS
UNTUK TERAPI GENE

1. Gene Addition,
2. Gene Blocking therapies and gene
repair (antisense, ribozyme, targeted
homologous recombination),
3. Non viral methods (liposomes, Naked
DNA),
4. Viral methods (retrovirus, Adenovirus,
Adeno-associated virus, others vectors)
 3.1. NON VIRAL VECTORS

1. LIPOSOMES
1. Hollow fat molecules in solution
2. Do not pose side effect problem of viruses

2. GOLDEN BULLETS
1. Helium gun fires small gold bullets coated with
genetically altered genes
2. Used in mice cancer research
 Example of Gene Therapy
1. ADA Deficiency, Cystic Fibrosis, Hemophilia
type B
Adenosine Deaminase Deficiency (ADA) = Severe
Combined Immune Deficiency (SCID)

Autosomal Recessive Disease

SCID chosen for first human Gene Therapy

because it met certain requirements

Gene was known and cloned
Vector was available

Gene product well known
N = Normal
n = ADA Deficiency Nn X Nn
N n

N Nn

n nn

ADA Deficiency
 SCID
1. Patient has no cell-mediated immune response
and is unable to make antibodies
2. Kills quickly because of absence of immune
system
3. Previous Treatments :
Germ free environment
Bone marrow transplant
Treat phenotype with injections of Adenosine
Deaminase Enzyme mixed with Polyethylene glycol
ADA - PEG
 KONDISI NORMAL
Normal Cell

ADA gene

Deoxyadenosine Inosine
(a vector produce)

Normal T-cell function

 PADA SCID

ADA gene

Deoxyadenosine Inosine
phosphorylated

Toxic Triphosphate which kills T-cells

Immune system fails to function

 Gene Therapy Treatment of SCID

EX VIVO
1. Treatment of T cells
T-cells extracted from patient grown in tissue culture

T-cells stimulated to proliferate with IL-2

Infected with retroviruses carrying the normal ADA
gene

T-cells carrying normal gene returned to child

Disadvantages :

T-cells live for only 6 12 months in the blood
 Gene Therapy Treatment of SCID

EX VIVO
2. Treatment of Stem cells

Stem cells extracted from patient bone marrow /
umbilical cord

Infected with retroviruses carrying a RNA copy of the
normal ADA gene

Stem cells carrying normal gene returned to bone
marrow of child
Pluripotent hematopoetic stem cells produce blood
cells-including T-cells which produce normal Adenosine
Deaminase

Advantages :
Stem cells produce T-cells indefinitely
Commonly used gene
vector
 Vectors

Viral Nonviral

Associated
Adeno Retro Naked Liposo
adeno
virus virus DNA mes
virus

High efficiency High efficiency Stable, long- Extremely Increased

Infects dividing/ Insertion of term safe efficiency
nondividing small DNA expression Low compared to
cells segments High efficiency efficiency naked DNA
Transient Transient Requires Insertion of Transient
expression expression dividing cells large DNA efficiency
Immunogenic Immunogenic Insertion segments
mutagenesis 21
 Viral Vectors :
2 Types RNA and DNA viruses

RNA viruses :
Retroviral and
Lentiviruses
A retroviral genome produces
components, but no viruses
Desired gene combined with packaging
signal
Vector produced with desired gene,
reverse transcriptase, uncapable of
infection
 RNA viruses - RETROVIRUSES
1. Advantages :
1. Their envelope protein enables them to infect
dividing human cells
2. RNA copies of Human Genes incorporated into
the retro viral genome using a packaging cell

2. Disadvantages :
1. Some retroviruses cause side effects Immune
System - attacks them.
2. Inability to infect non dividing cells
 RNA virus LENTIVIRAL Vector

1. Advantages :
1. Infect dividing and non dividing cell (e.g. HIV)
2. Longest sustained expression over 6 months
3. No potent antibody response

2. Disadvantages ??
RETROVIRUS

LIFECYCLE
RNA-pol (RT) attached to
viral genomic RNA

DNA copies

Random insertions
to host genome

Producing of fresh RNAs

Translation Packaging
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/R/
 Viral Vectors :
2 Types RNA and DNA viruses

DNA viruses :
Adenoviral and
Adeno associated virus

DNA viruses ADENO associated

viruses

1. Advantages :
1. Causes no known disease
2. Do not trigger immune response
 DNA viruses - ADENOVIRAL
Cause benign Respiratory Tract Infection in Human
being = Common Cold Viruses

1. Advantages :
1. Infect dividing and non dividing cells
2. Can be used in vivo
3. Can be engineered to deliver cell killing genes to cancer cell

2. Diadvantages :
1. Short expression time
2. Elicits both cell-killing celluler response and antibody
producing humoral response
28
4. RENCANA TERAPI :
Penyakit genetik dapat diobati pada
berbagai level sesuai tahap mutasi
dari gene.
Pengobatan pada level of clinical
phenotype mengikutkan semua
intervensi medik dan bedah,
Yang penting pasien diberi edukasi
tentang penyakit genetik,
keberhasilan, komplikasi, dampak
genetik terapi, dan ketidaknyamanan
pengobatan.
 Level Intervention :
metabolik dan clinical phenotype
Penyakit Medikamento Penyakit Terapi Bedah
sa
Defisiensi
Pemberian
Obstruksi Sal Kateter
Biotinidase Biotin Kemih percutan
Gangguan Katup Vesicostomi
Pada Ibu
uretra-
Hidronefrosis
Respon Pemberian
Hernia Diafragma
Reduksi Visera
Cobalamin Cobalamin Hipoplasia dan Repair
pada ibu paru Diafragma
metylmalonic
asidiuria
Kelainan
Dexamethasone Sindrom transfusi
Dibuat Shunt

kongenital Pada anak plasenta

Adrenal kembar antara
Hiperplasia kedua anak
Mutant level
Mutant level
 Faktor lingkungan menjadi salah
satu etiologi kelainan genetik, jika
dapat diidentifikasi maka keadaan
dapat diperbaiki.
Contoh : ROKOK, dampak terhadap
paru-paru, defisiensi 1 anti
tripsin menjadi tidak berfungsi.
 Terapi Multifaktorial

Medikamentosa
Pembedahan
Contoh;
Pada DM, terapi insulin menjadi terapi

pilihan .
Pada kelainan katup jantung

bawaan,labiopalatoschizis, stenosis
pylorus, pembedahan adalah drug of
choice
 Efek terapi insulin secara intensif pada 3
jenis komplikasi DM tipe 1/ rata rata 100
pasien /th

Terapi Terapi % Resiko

Konvensiona Intensi Drop Out
l f
Retinopati 4,7 1,2 76

Albuminuri 3,4 2,2 34

a
Neuro Pati 9,8 3,1 69
 Penyakit Gen tunggal ditandai dengan
defisiensi.
Terapi dapat sukses jika struktur
biokimiawi diketahui, tetapi pemulihan
kesehatan secara normal dapat gagal.
Penelitian Hayes dan partner : Angka
harapan hidup meningkat, 15% setelah
terapi, setelah kelainan monogenik
diterapi dari 65 kasus kelainan bawaan.
 Gen Yang tidak dikenali,
patogenesis belum dipahami,
Mutan adalah salah satu penyebab
penyakit genetik yang tidak
dikenal patofisiologi dan kelainan
biokimiawi ada 75 % dari kelainan
genetik
Pada PKU (Fenil Keton Uria), bagaimana
mekanisme tingginya kadar fenilalanin dapat
merusak fungsi otak belum dipahami.

Pada Duchenne Muscular Dystrophy (DMD),

dan Cystic Fibrosis, panyakit yang
mempengaruhi fungsi protein masih banyak
pengetahuan dasar yang belum diketahui
 Penyakit gangguan metabolisme
galaktosa, suatu kelainan autosom
resesif,tidak sempurnanya enzim
Galaktosa-1 Fosfat Uridiltransferase
GALT
Galaktosa-1 fosfat UDPG
Bayi dgn galaktosemia sering lahir
normal, tapi diawal perkembangan dpt
tjd sirosis hepatis dan katarak, minggu
pertama setelah pemberian susu.
 Terapi Galaktosemia dengan diet susu
lengkap dapat berhasil, pada wanita
ada hambatan karena adanya toksik
galaktosa mengakibatkan kelaianan
pada saluran telurnya.
Penyakit cistinosis, ditandai akumulasi
sistein pada lisosom dan adanya defek
pada pelepasan sistein, dapat
mengakibatkan gangguan fungsi ginjal
 Pengobatan dapat dilakukan
tapi
efek samping baru timbul
kemudian

Contoh pada hemofilia, ada bentukan

anti bodi yang dapat menembus protein,
pada proses pembekuan, sehingga zat
besi yang hasilkan dapat berlebihan
akibatnya penanganan menjadi lebih
sulit.

Pada pasien yang menerima cangkok

ginjal, dapat terjadi hipotyroidisme
 Pengobatan dapat dilakukan tapi
efek samping baru timbul kemudian

Kelainan islet sel pankreas Diabetes,

dan gangguan syaraf.
Penderita Retinoblastoma, setelah
berhasil diterapi tumornya, dalam
perkembangannya dapat timbul
Osteosarcoma, diduga adanya mutasi
gen.
Pada prinsipnya terapi gene dapat
memperpanjang usia tapi ada dampaknya
kelainan organ ditempat lain bisa terjadi
 5. MASA DEPAN TERAPI
GENE :
Gen therapy has not yet to become a
routine and is only beginning to
emerge from strictly experimental
stage of development.
Most gene therapy trials to date have
focused more on investigating
potential feasibility and safety rather
than having been designed to induce
benefit.
 5. MASA DEPAN TERAPI
GENE :
In fact, there are finishing few trails if
any- that have occurred convincing
evidence of clinical benefit to the
patients who have been treated.
On the other hand, there is no reason
to doubt that technological barriers to
its success will ultimately surmounted.
Tugas Mandiri :
1. Buatlah Pedigree dari keluarga anda
dari anda sampai 3 generasi ke atas ,
2. Catatlah penyakit penyakit yang
dijumpai pada keluarga ayah dan Ibu
3. Penyakit mana yang termasuk
penyakit keturunan,
4. Buatlah artikel salah satu penyakit
tersebut dan pengobatannya saat ini