The effect of TSPP-mediated photodynamic therapy and Parecoxib in experimental tumours

Abstract

Aims

The study investigated the effects of the combined treatment Parecoxib (Pcox) and 5,10,15,20-tetra-sulphonato-phenyl-porphyrin(TSPP)-mediated photodynamic therapy on Walker 256 carcinosarcoma.

Main methods

Five groups of male Wistar rats were used: the control group, treated with TSPP, group 2, irradiated 24 h thereafter, group 3, treated with Pcox and irradiated 24 h thereafter, groups 4 and 5 treated with combined therapies, TSPP and Pcox before irradiation, and Pcox 24 h after TSPP and irradiation respectively. Tumour inflammation, growth and non-growth factors, apoptosis/necrosis rate and oxidative/nitrosative stress markers were investigated.

Key findings

Malondialdehyde levels and cyclooxygenase (COX)-2 expression increased significantly in the group treated with Pcox after TSPP-PDT when compared with TSPP + IR group (p < 0.05, p < 0.001 respectively), in correlation with a decrease in glutathione levels (p < 0.05). The quantification of apoptosis, based on the TUNEL-assay, and necrosis rate revealed an increase of apoptotic/necrotic index in the same group (p < 0.05). On the other hand, Pcox administered before irradiation showed a significant increase in both vascular endothelial growth factor (VEGF) and COX-2 levels (p < 0.05) and in nitric oxide production (p < 0.01), when compared with the control group.

Significance

The administration of Pcox after TSPP-mediated PDT showed promising antitumoural effects, leading to an increase in oxidative and nitrosative stress as well as apoptosis/necrosis rate in tumour tissue. These results show that combined regimens that involve selective COX-2 inhibitors administration after irradiation may improve the therapeutic effectiveness of PDT.

Introduction

Treating cancer has always been one of medicine's most ambitious goals, with a plethora of worldwide researchers struggling to find the cure. Nonetheless, in spite of the improved techniques and advances in surgery, radio- and chemotherapy, all these methods still involve a considerable degree of invasiveness and significant side-effects. It is thus natural that all the attempts to match this goal should be considered and even embraced, provided they are efficient. This is also the case of photodynamic therapy (PDT) (Brown, 2012).

PDT is a relatively novel treatment for cancer such as head and neck, esophageal, lung, bladder, gastric and colorectal cancer, cholangiocarcinoma, as well as certain non-malignant, mainly dermatological diseases and skin rejuvenation. The therapy involves the administration of a photosensitizing agent (photosensitizer, PS) and specific activation by light in its absorption spectrum (Buytaert et al., 2007). This results in a sequence of photochemical and photobiological processes that cause irreversible photodamage to tumour tissues. Specifically, PDT-induced oxidative stress targets and damages various cellular macromolecules such as proteins, lipids and nucleic acids. Among these targets, the peroxidation of lipids is particularly damaging since the formation of lipoperoxidation products leads to a facile propagation of free radicals and to membrane disintegration (Agostinis et al., 2011). Besides the cancer cells, tumour microenvironment and vasculature are also injured, without affecting the survival of the adjacent normal tissue. Therefore, in order to achieve optimal tumour damage and avoid any sort of tumour resistance, it is vital that the triad of direct tumour cell photodamage, destruction of tumour vasculature and activation of innate immunity mechanisms is respected (Baldea and Filip, 2012).

PDT also induces the expression of angiogenic and survival factors including vascular endothelial growth factor (VEGF), cyclooxygenase (COX)-2, matrix metalloproteinases (MMPs) and nitric oxide (NO) (Ferrario et al., 2000). Recent studies found that MMPs are indirectly involved in angiogenesis by cleaving extracellular matrix (ECM)-bound VEGF (Lee et al., 2005). Besides this, PDT-linked inflammation is an essential component of PDT outcome. It has been shown that eliciting or inhibiting immune mechanisms through interplay of inflammatory and anti-inflammatory mediators, respectively, is PS and light dosage-dependent (Agostinis et al., 2011).

Based upon the basic mechanism and in an attempt to enhance PDT, researchers designed novel modalities to improve its outcome. Several studies have shown that highly selective COX-2 inhibitors are good candidates as adjuvants (Dannenberg et al., 2001, Ferrario et al., 2005). They specifically target the inducible isoform of cyclooxygenase, which is involved in inflammation and tumour angiogenesis, and consequently in tumour cell survival. TSPP, 5,10,15,20-tetra-sulfo-phenyl-porphyrin, is an alternative to the already used PSs, whose specific aggregation characteristics explain the yield of considerable quantities of reactive oxygen species in a high aggregated state (Ion, 1997). Both in vitro and in vivo studies proved the efficiency of this synthetic PS with minimal side effects (Clichici et al., 2010).

Given these premises, the study aims to assess comparatively the effects of Parecoxib (Pcox) on TSPP-mediated PDT and to analyse its effects in two different therapeutic regimens, using Walker 256 subcutaneous carcinosarcoma as an experimental model. Inflammation, targeted growth and non-growth factors, apoptosis and necrosis index and oxidative/nitrosative stress markers and their interaction following PDT and Pcox administration will be investigated.