In vitro characterization of oncolytic virus in combination with STAT3 inhibition for improved therapy against pancreatic ductal adenocarcinoma

Master's Thesis , 2017 , 52 Pages

Biology - Diseases, Health, Nutrition

Since there is accumulating evidence that STAT3 is constitutively activated in PDAC and important for the progression and maintenance of this disease, the goal of this thesis was to establish a PDAC model in the lab to further investigate and characterize if the synergistic effect of VSV with STAT3 inhibition would also be effective on this cancer type. Aiming to unravel as many characteristics as in vitro studies allow, this study utilized not just 2D and 3D systems, but also cocultural approaches on both molecular and morphological levels.

Pancreatic ductal adenocarcinoma (PDAC) is considered as a very severe diagnosis because of the poor survival rates and the aggressive nature. Further investigations show the high complexity of the mutational cascades in PDAC initiation and progression. The importance of the tumor microenvironment is clarified, but as different studies led to paradoxic results in targeting the stroma, it gives us evidence that it is just poorly understood yet. Given the limitations of current treatments and the failure of various novel approaches, a necessity of further investigating new therapeutic options is unquestionable.

Inhaltsverzeichnis (Table of Contents)

• I. Introduction
• 1.1 Pancreatic cancer
• 1.1.1 PDAC development and progression
• 1.1.2 Microenvironment
• 1.1.3 Current treatment options and their limitations
• 1.2 Oncolytic viruses
• 1.2.1 Vesicular stomatitis virus
• 1.3 Signal transducer and activator of transcription 3
• 1.4 Aim of this thesis
• II. Material
• 2.1 Cell lines
• 2.2 Reagents
• 2.3 Antibodies
• 2.4 Consumables
• 2.5 Appliances
• III. Methods
• 3.1 Cell culture
• 3.1.1 3D cell culture systems
• 3.1.2 Co-culture systems
• 3.2 Viability assay
• 3.3 Tissue culture infection dose
• 3.4 Western blotting
• 3.5 Flow cytometric analysis
• 3.6 Confocal microscopy
• 3.7 Luciferase assay
• 3.7.1 Plasmid DNA preparation
• 3.7.2 Transfection
• 3.7.3 Interferon induction and response
• 3.8 Scratch assay
• IV. Results
• 4.1 PDAC cells are susceptible to rVSV-GFP-mediated oncolysis in vitro
• 4.2 S3I-201 causes reduction of PDAC cell viability in vitro
• 4.3 Combination therapy enhances PDAC killing and safety
• 4.4 Combinational therapy in the microenvironment
• 4.5 Effects of S3I-201 and VSV in co-culture
• 4.6 Combination therapy in 3D
• 4.7 Inhibition of migration
• V. Discussion

Zielsetzung und Themenschwerpunkte (Objectives and Key Themes)

This thesis aims to investigate the effect of combining oncolytic vesicular stomatitis virus (VSV) with STAT3 inhibition as a novel treatment option for pancreatic ductal adenocarcinoma (PDAC). The study focuses on characterizing the cytotoxic effects of this combinational therapy in vitro, utilizing both 2D and 3D cell culture systems, as well as co-culture approaches.

• The oncolytic potential of VSV against PDAC cells
• The cytotoxic effects of STAT3 inhibition on PDAC cells
• The synergistic effects of combining VSV with STAT3 inhibition on PDAC cells
• The impact of the combinational therapy on the tumor microenvironment, particularly cancer-associated fibroblasts
• The ability of the combinational therapy to modulate the migratory and proliferative potential of both PDAC cells and cancer-associated fibroblasts

Zusammenfassung der Kapitel (Chapter Summaries)

• I. Introduction: This chapter provides an overview of pancreatic cancer, including its development, progression, and microenvironment. It also discusses current treatment options and their limitations, highlighting the need for novel therapies. The introduction introduces oncolytic viruses and their mechanisms of action, specifically focusing on vesicular stomatitis virus (VSV). The chapter concludes with a detailed explanation of STAT3, its role in cancerogenesis, and the potential of STAT3 inhibitors as therapeutic targets.
• II. Material: This chapter details the materials used in the study, including cell lines, reagents, antibodies, consumables, and appliances.
• III. Methods: This chapter describes the methods used in the study, including cell culture techniques, viability assays, viral titer determination, Western blotting, flow cytometry, confocal microscopy, luciferase assays, and scratch assays.
• IV. Results: This chapter presents the results of the study, demonstrating the cytotoxic effects of VSV and STAT3 inhibition, both individually and in combination, on PDAC cells. The chapter also examines the effects of the combinational therapy on the tumor microenvironment and the migratory/proliferative potential of both PDAC cells and fibroblasts.

Schlüsselwörter (Keywords)

Pancreatic ductal adenocarcinoma, oncolytic virus, vesicular stomatitis virus, STAT3 inhibition, tumor microenvironment, cancer-associated fibroblasts, apoptosis, migration, proliferation.

• College: LMU Munich
• Author: Simon Dedic (Author)
• Publication Year: 2017
• Pages: 52
• Catalog Number: V420553
• ISBN (eBook): 9783668714854
• ISBN (Book): 9783668714861
• Language: English
• Tags: stat3