The Best MPhil Thesis in the Faculties of Dentistry, Engineering, Medicine and Science (University of Hong Kong), Li Ka Shing Prize / published_or_final_version / Pathology / Master / Master of Philosophy

Simple Summary Gene therapy is a novel approach to treat diseases by introducing corrective genetic information into target cells. Adeno-associated virus vectors are the most frequently applied gene delivery tools for in vivo gene therapy and are also studied as part of innovative anticancer strategies. Here, we report on the natural preference of AAV2 vectors for hepatocellular carcinoma (HCC) compared to nonmalignant liver cells in mice and human tissue. This preference in transduction is due to the improved intracellular processing of AAV2 vectors in HCC, resulting in significantly more vector genomes serving as templates for transcription in the cell nucleus. Based on this natural tropism for HCC, novel therapeutic strategies can be designed or existing therapeutic approaches can be strengthened as they currently result in only a minor improvement of the poor prognosis for most liver cancer patients. Abstract Although therapeutic options are gradually improving, the overall prognosis for patients with hepatocellular carcinoma (HCC) is still poor. Gene therapy-based strategies are developed to complement the therapeutic armamentarium, both in early and late-stage disease. For efficient delivery of transgenes with antitumor activity, vectors demonstrating preferred tumor tropism are required. Here, we report on the natural tropism of adeno-associated virus (AAV) serotype 2 vectors for HCC. When applied intravenously in transgenic HCC mouse models, similar amounts of vectors were detected in the liver and liver tumor tissue. In contrast, transduction efficiency, as indicated by the level of transgene product, was moderate in the liver but was elevated up to 19-fold in mouse tumor tissue. Preferred transduction of HCC compared to hepatocytes was confirmed in precision-cut liver slices from human patient samples. Our mechanistic studies revealed that this preference is due to the improved intracellular processing of AAV2 vectors in HCC, resulting, for example, in nearly 4-fold more AAV vector episomes that serve as templates for gene transcription. Given this background, AAV2 vectors ought to be considered to strengthen current—or develop novel—strategies for treating HCC.

Hu, Baoguang. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 121-143). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Hepatocellular carcinoma (HCC) is the third commonest cancer worldwide. However HCC is considered to be highly resistant to chemotherapy. Gene therapies aimed to regulate Bd-2 proteins may sensitize HCC cells to chemotherapy. Studies have demonstrated that Bid/tBid are crucial in hepatocyte apoptosis. Bid also plays important roles in the development and chemotherapeutic sensitivity of HCC. The objective of this study is to test effect of Ad/AFPtBid and chemotherapeutic agents in combination on an orthotopic HCC model. / In conclusion, (1) Ad/AFPtBid can specifically target and effectively suppress the AFP-producing HCC. (2) Ad/AFPtBid can significantly sensitize HCC to 5-FU, their combination can significantly increase the anti-tumor effectiveness. (3) Ad/AFPtBid shows little toxicity in vivo. (4) The complementary effect of tBid and 5-FU on different phases of the cell cycle may explain the better therapeutic result if both are used to treat HCC. (5) The elucidation of phase specific effect of tBid points to a possible therapeutic option that combines tBid with different phase specific agents to treat HCC. / It is well established that many apoptosis inducers act in a cell cycle-specific fashion. This leads us to hypothesize that tBid might have phase specific effect. So, we tested the susceptibility of Hep3B cells at 00/01, S or G2/M phases to tBid. The results revealed that tBid significantly reduced Hep3B cells in G0/G1 phase, increased cells in G2/M phase. On the contrary, 5-FU arrested Hep3B cells in G0/G1 phase, and significantly reduced cells in G2/M phase. The levels of cell cycle-related proteins were altered in line with the result of the cell cycle. This suggests Hep3B cells in G0/G1 phase may be more susceptible to tBid. The complementary effects tBid and 5-FU on different phases of the cell cycle may explain the better therapeutic result if both are used to treat HCC. / The mice bearing orthotopic HCC tumors were treated with Ad/AFPtBid alone or in combination with 5-FU/Dox. Serum AFP levels were measured to mornitor tumor progression. The mice were killed four weeks after treatment. Liver tissues were subjected to immunohistochemical staining of proliferation cell nuclear antigen (PCNA) and TUNEL staining. Another batch of mice was observed for survival rate over a six month period. In addition, possible side effects of Ad/AFPtBid were tested in BALB/c mice. Results demonstrated that Ad/AFPtBid significantly inhibited Hep3B tumor growth. The combination of Ad/AFPtBid with 5-FU was more effective in tumor regression than either agent alone. However, the combination of Ad/AFPtBid with Dox treatment failed to demonstrated better effect than Dox treatment alone because the mice that received Dox exhibited serious weight loss. Tumor tissues from Ad/AFPtBid alone or combination treatment groups showed a decrease in cells positive for PCNA, and an increase in apoptosis by TUNEL staining, indicating that Ad/AFPtBid induced tumor regression through its pro-apoptotic effect. Inflammatory cell infiltration was also increased. Furthermore, Ad/AFPtBid did not suppress the hepatic tumor formed by non-AFP producing SK-HEP-1 or DLD-1. Finally, Ad/AFPtBid and 5-FU in combination results in better survival rate. No acute toxic effect of Ad/AFPtBid was observed. / Ma, Shihong. / "December 2009." / Adviser: CHEN Gong George. / Source: Dissertation Abstracts International, Volume: 72-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 114-138). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. Ann Arbor, MI : ProQuest Information and Learning Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.