Abstract 3732: Targeted photodynamic therapy enhances the therapeutic efficacy of combination therapy (PDT and chemotherapy) on chemoresistant melanoma cells

Abstract

Abstract Cutaneous melanoma remains refractory to current therapies such as chemotherapy. Failure of treatment has been attributed to enhanced self-renewal capacity and the over-expression of ABC transporters by therapy resistant populations. A need therefore exists to identify and evaluate new approaches. Alternative combinatorial approaches comprising chemotherapeutics, photodynamic therapy (PDT) and targeted-PDT have been proposed to overcome resistance. Objective This study set to investigate the efficacy of novel (targeted-PDT-IR700) and standard combinatorial effect of PDT and chemotherapy (Hypericin-PDT and DTIC) as an adjunctive combination therapy treatment to decrease cellular chemoresistance in melanoma. This was assessed in an in vitro model, to show differences in killing of melanoma cells. Methodology Single chain variable fragments (scFv) of panitumumab were cloned into a transient eukaryotic SNAP expression vector co-expressing cytosolic proteins and transfected into HEK293T cells. Purified recombinant protein was confirmed by SDS-PAGE and Western blot analyses. Benzylguanine (BG) modified IR700 was irreversibly conjugated in a 1:1 stochiometric ratio to the 1711(scFv)-SNAP using the self-labelling property of the suicide enzyme SNAP-tag. Binding activity and cytotoxicity was documented on EGFR expressing melanoma cells using flow cytometry, confocal microscopy and XTT assays. Cytotoxic activity, self-renewal capacity and ABC transporters expression (ABCB5 and ABCG2) after the combination therapy (HYP-PDT and DTIC) was determined using XTT, clonogenic assays and flow cytometry. Results Binding and internalization of purified 1711(scFv)-SNAP conjugated to BG-modified Alexa 488 or IR700 was successfully confirmed by confocal microscopy and flow cytometry. Cell viability showed cell specific cytotoxicity of the photosensitizer labeled fusion proteins upon light induction, at nanomolar concentrations which were lower than those used for the non-targeted combinatorial approach (HYP-PDT and DTIC). Moreover, DTIC resistant melanoma cells were less sensitive to therapeutic treatment than their non-resistant counterpart. This correlated with an increased self-renewal capacity and ABC transporter expression in DTIC resistant melanoma in comparison to their non-resistant counterpart. This study describes a proof of concept of selective targeting and elimination of EGFR expressing melanoma cells by targeted photoimmunotherapy using a panitumumab 1711-scFv fused to SNAP tag and conjugated to near infrared photosensitizer IR700 which improve therapeutic efficacy with lower concentration and reduced sides effect. This work highlights the efficacy of targeted PDT and suggest that it is highly applicable to improve the current clinical arsenal of therapies. Note: This abstract was not presented at the meeting. Citation Format: Lester M. Davids, Fleury Nsole Biteghe, Eden Padayachee, Stefan Barth. Targeted photodynamic therapy enhances the therapeutic efficacy of combination therapy (PDT and chemotherapy) on chemoresistant melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3732.