Investigating tissue penetration and bystander effect of the hypoxia targeted anticancer drug SN36506

Abstract

Hypoxia is a hallmark of solid tumours that is associated with malignant progression and therapy resistance. Hypoxia-activated prodrugs (HAPs) exploit hypoxia as a basis of tumour selectivity for therapeutic gain. SN36506 is a second-generation analogue of PR-104, designed specifically to circumvent the off-target systemic toxicities associated with aldoketo reductase 1C3 (AKR1C3) activation. Previous investigations have shown that the anti-tumour activity of HAPs is dependent on their ability to diffuse through tumour tissues to its hypoxic-activating cells (extravascular transport), and for the released metabolites to diffuse through-out the tumour to potentiate cell killing (bystander effect). This thesis investigates the pharmacological properties of SN36506 in tumour tissues using spheroid co-cultures and a well-validated in silico spatially-resolved pharmacokinetic/pharmacodynamic (SR-PK/PD) modelling approach. A pro(drug) transport model was developed from experimental determinations of cellular uptake, metabolism and diffusion with flux of SN36506 measured across multicellular layer (MCL) cultures by mass spectrometry. The fitted parameters were then validated by comparing experimental cell killing in spheroid co-cultures to model simulations. MCL transport studies indicate that the high affinity cellular uptake and bioreductive metabolism of SN36506 slows its extravascular transport across tumour tissues. However, the active metabolites (SN36506H, SN36506M and SN39751) are capable of diffusing from their hypoxic activating cells through a tissue-like cell density into the media reservoirs of the diffusion chambers by means of a bystander effect. Spheroid co-culture experiments confirmed a substantial bystander effect, with the effector released from the metabolically-competent ‘activator’ cells capable of sensitising the metabolically-incompetent ‘target’ cells measured by clonogenic assay. Spheroid regrowth delay reinforced the bystander effect with growth inhibition proportional to the fraction of activators in the co-cultures. The fitted parameters for the pro(drug) transport model were used to simulate bystander killing in good agreement with clonogenic assays but the spheroid regrowth model requires further refinement. Overall, this thesis couples experimental and computational modelling approaches to investigate tissue pharmacological properties of SN36506. Demonstration of a marked bystander effect, similar in magnitude to PR104A, together with its previously demonstrated lack of AKR1C3 activation, indicates further investigation is warranted to determine the therapeutic potential of SN36506 in patients with solid malignancies.