Sains Malaysiana 46(8)(2017): 1289–1297

http://dx.doi.org/10.17576/jsm-2017-4608-15

 

A Gain of Function p53 Gene Mutant Promotes Growth Suppression in Human Liver Cancer Cells

(Pemulihan Fungsi Mutan Gen p53 Menggalakkan Penindasan Pertumbuhan Sel Kanser Hati Manusia)

 

NOR ADZIMAH JOHDI*, SITI NURMI NASIR & RAHMAN JAMAL

 

UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Jalan Yaacob Latif, Bandar Tun Razak, 56000 Cheras, Kuala Lumpur, Federal Territory, Malaysia

 

Diserahkan: 18 Ogos 2016/Diterima: 11 Januari 2017

 

ABSTRACT

Primary liver cancer is one of the most common cancer in the world with highest cancer mortality rate. The most common type of primary liver cancer is hepatocellular carcinoma (HCC). There are many risk factors for liver cancer and currently available treatments for HCC are largely inadequate. Gene mutation and dysfunction of p53 are common and is recognized as an important molecular event in hepatocarcinogenesis. Therefore, replacement of the aberrant p53 gene is an attractive approach in the treatment of HCC providing an alternative treatment for primary HCC. In this study, we assessed whether the transfection with wild-type p53 gene is able to restore the pro-apoptotic effects and evaluate the feasibility of gene therapy in fixing a faulty p53 molecule. We established a non-viral cationic lipid-based p53 gene delivery into two human HCC cell lines namely HLF and PLC/PRF/5 cells. Both cell lines have mutations in the p53 gene. We compared the results with the normal liver cell line, WRL68, that constitutively expresses the wild-type p53 gene. In this study, the introduction of wild-type p53 gene into HLF and PLC/PRF/5 cells resulted in an increased of p53 gene expression, protein expression and cells growth inhibition shown in MTS reduction cell viability assay, FITC-Annexin V and PI apoptosis assay, western blot and caspase activity assay. In summary, the study provides a promising therapeutic approach for p53 gene delivery into HCC patients. The p53 gene delivery can be instituted together with chemotherapy as a combination treatment to induce apoptosis.

 

Keywords: Apoptosis; cell lines; hepatocellular carcinoma; p53 gene; transfection

 

ABSTRAK

Kanser hati utama adalah salah satu kanser umum di dunia dengan kadar kematian kanser tertinggi. Jenis yang paling biasa untuk kanser hati utama adalah karsinoma hepatosel (HCC). Terdapat banyak faktor risiko untuk kanser hati utama dan rawatan untuk HCC sebahagian besarnya tidak mencukupi. Mutasi gen dan kehilangan fungsi gen p53 adalah faktor yang paling biasa dan gen p53 diiktiraf sebagai antara rantaian molekul yang penting dalam proses pembentukan kanser hati. Oleh itu, pemulihan mutan gen p53 yang defektif adalah satu pendekatan yang menarik dalam rawatan HCC sebagai rawatan alternatif untuk HCC utama. Dalam kajian ini, penilaian dilakukan sama ada transfeksi gen p53 normal ke atas titisan sel HCC mampu mengembalikan kesan pro-apoptotik dan potensi terapi gen dalam pemulihan molekul p53 yang defektif. Ini melibatkan transfeksi gen p53 normal menggunakan teknik lipid kationik kepada dua jenis titisan sel HCC manusia iaitu HLF dan PLC/PRF/5. Kedua-dua jenis titisan sel ini mempunyai mutasi dalam gen p53. Keputusan kajian dibandingkan dengan titisan sel hati normal, WRL68, yang mengekspres gen p53 normal. Transfeksi gen p53 normal ke dalam titisan sel HLF dan PLC/PRF/5 menyebabkan peningkatan ekspresi gen p53, pertambahan ekspresi protein p53 dan perencatan pertumbuhan titisan sel-sel yang ditunjukkan dalam assai MTS sel pengurangan daya maju, assai apoptosis FITC-Annexin V & PI, pemendapan barat dan aktiviti caspase. Secara ringkasnya, kajian ini menyediakan pendekatan terapeutik yang boleh digunakan untuk menyalurkan gen p53 ke dalam sel pesakit HCC. Penyampaian gen p53 boleh dimulakan bersama dengan kemoterapi sebagai rawatan kombinasi untuk mendorong aktiviti apoptosis.

 

Kata kunci: Apoptosis; gen p53; hepatosel karsinoma; titisan sel; transfeksi

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*Pengarang untuk surat-menyurat; email: adzimah@ppukm.ukm.edu.my