Investigation of strategies to enhance the delivery of oncolytic vaccinia virus

Vaccinia Virus (VV) is an attractive vector for oncolytic virotherapy given its potential for tumour-selective tropism, efficient cell-to-cell spread, rapid replication in cancer cells, large transgene coding capacity and stimulation of anti-tumour immunity. It has a well-defined safety profile and is being assessed in late￾stage clinical trials.
Systemic delivery of VV has shown limited clinical success in patients due to a knockdown in infectivity, as a result of rapid immune-mediated neutralisation, and poor penetration into tumours. These factors have often restricted its route of delivery to intratumoural (IT) injection. For VV to become applicable across a wider range of patients/cancer indications, amenability to systemic delivery is required. This may be achieved using strategies that modulate VV by genetic or chemical means and/or that alter the physiology of target tumours. Chemical stealthing has not previously been applied to enhance the pharmacokinetics (PK) of enveloped viruses such as VV. Ultrasound (US)-mediated cavitation has been previously shown to effectively enhance the delivery of therapeutics, including VV, to solid tumours.
In this thesis, US-mediated cavitation was applied to DLD and CAPAN-2 xenografts after IT administration of VV with the aim of producing more homogenous intratumoural infection. The limited success observed in the IT context provided motivation to instead enhance the stability of VV for improved IV administration. Flow virometry demonstrated that VV could be effectively modified using an amphiphilic polymer (to create PCVV). This PCVV showed reduced natural tropism, a 20% decrease in anti-VV antibody binding and could be used as a platform for the attachment of an antibody to permit retargeting of infection to the tumour associated antigen mucin-1 (MUC1).
PK analysis of VV, PCVV and aMUC1-PCVV showed circulation kinetics for PCVV and aMUC1-PCVV were enhanced at two different doses (approximately 5-fold higher circulating dose at 5 minutes vs VV). The hypothesis that the application of US-mediated cavitation during this 5 minute window could be used to enhance extravasation of PCVV and aMUC1-PCVV into and throughout tumours, was explored. When samples were administered IV and US-mediated cavitation was applied to CAPAN-2 xenografts, increased infection (as assessed by transgene expression) was observed. The impact of cavitation on aMUC1-PCVV transgene expression was highest (a mean increase of 530-fold compared to non US exposed tumours). Tumour growth studies showed that combining aMUC1-PCVV, SSPs and US enabled substantial and significant tumour regression (-73%).
The work presented in this thesis brings to light the challenges of achieving effective and uniform delivery to tumours after both IT and IV administration of VV and contributes to the necessary progress VV needs to make if it is going to achieve its full clinical potential.

Keywords: polymer coating
cellular targeting
Flow virometry
poxvirus
vaccinia virus
pancreatic cancer
oncolytic virotherapy
viruses
ultrasound
capan-2
cholesterol-PEG
cavitation
mucin-1
Advisor: Carlisle, R