patient: 6-1 tissue: non tumor lesion of liver biopsy agent: peretinoin time: 8 weeks agent dose: 300mg clincal outcome: Recurrence of HCC
Treatment protocol
Peretinoin, a member of the acyclic retinoid family, is expected to be an effective chemo preventive drug for HCC.The patients had undergone curative surgical resection or radiofrequency ablation (RFA). They received 300 mg/day or 600 mg/day of peretinoin for 8 weeks and were followed up for 88 weeks with 600 mg/day of peretinoin. Hepatic gene expression profiling obtained from non tumor lesion of these patients prior to administering peretinoin and 8 weeks after starting peretinoin treatment
Extracted molecule
total RNA
Extraction protocol
Total RNA was isolated from liver biopsy samples using an RNA extraction kit (Micro RNA Extraction Kit, Stratagene, La Jolla, CA, USA). Aliquots of total RNA (5 ug) were subjected to amplification with antisense RNA (aRNA) using a Message AmpTM aRNA kit (Ambion, Austin, TX, USA) as recommended by the manufacturer
Label
Cy5
Label protocol
As a reference for each microarray analysis, aRNA samples prepared from the normal liver tissue from one of the patients were used. Test RNA samples fluorescently labeled with cyanine (Cy) 5 and reference RNA labeled with Cy3 were used for microarray hybridization
Peretinoin, a member of the acyclic retinoid family, is expected to be an effective chemo preventive drug for HCC.The patients had undergone curative surgical resection or radiofrequency ablation (RFA). They received 300 mg/day or 600 mg/day of peretinoin for 8 weeks and were followed up for 88 weeks with 600 mg/day of peretinoin. Hepatic gene expression profiling obtained from non tumor lesion of these patients prior to administering peretinoin and 8 weeks after starting peretinoin treatment
Extracted molecule
total RNA
Extraction protocol
Total RNA was isolated from liver biopsy samples using an RNA extraction kit (Micro RNA Extraction Kit, Stratagene, La Jolla, CA, USA). Aliquots of total RNA (5 ug) were subjected to amplification with antisense RNA (aRNA) using a Message AmpTM aRNA kit (Ambion, Austin, TX, USA) as recommended by the manufacturer
Label
Cy3
Label protocol
As a reference for each microarray analysis, aRNA samples prepared from the normal liver tissue from one of the patients were used. Test RNA samples fluorescently labeled with cyanine (Cy) 5 and reference RNA labeled with Cy3 were used for microarray hybridization
Hybridization protocol
Reference papers; Honda M, Yamashita T, Ueda T, Takatori H, Nishino R, Kaneko S. Hepatology. 2006 Nov;44(5):1122-38, Honda M, Kaneko S, Kawai H, Shirota Y, Kobayashi K. Gastroenterology. 2001 Mar;120(4):955-66.
Scan protocol
Quantitative assessment of the signals on the slides was carried out by scanning on a ScanArray 5000 (General Scanning, Watertown, MA, USA) followed by image analysis using GenePix Pro 4.1(Axon Instruments, Union City, CA, USA)
Description
300mgRecurrence
Data processing
Because the levels of expression of some housekeeping genes were also changed, especially in advanced stages of liver disease, for the normalization of data we averaged intensities of all spots obtained with Cy3 and Cy5 in each of the 16 rectangles and adjusted intensity of each corrected DNA spot as the average intensity ratio Cy5/Cy3 = 1.0. This global normalization of intensity provided smaller variance of CyS/Cy3 ratio and almost equivalent results to normalization using the housekeeping genes. Reference papers; Honda M, Yamashita T, Ueda T, Takatori H, Nishino R, Kaneko S. Hepatology. 2006 Nov;44(5):1122-38, Honda M, Kaneko S, Kawai H, Shirota Y, Kobayashi K. Gastroenterology. 2001 Mar;120(4):955-66.
