Evaluation of antitumor and immunomodulatory properties of Indicaxanthin from Opuntia Ficus Indica (L. Mill) fruit

Cancer is a growing health problem around the world and according to estimates from the International Agency for Research on Cancer (IARC), 14.1 million new cancer cases and 8.2 million cancer deaths worldwide have been reported in 2012 (Ferlay et al., 2015). By 2030, the global burden is expected to grow to 21.7 million new cancer cases and 13 million cancer deaths simply due to the growth and aging of the population. Indicaxanthin ((2S)-2,3-dihydro-4-[2-[(2S)-2a-carboxypyrrolidin-1- yl]ethenyl]pyridine-2a,6-dicarboxylic acid), a betalain pigment from cactus pear fruit, has been the object of sound experimental work over the latest years. As many phytochemicals, indicaxanthin is a redox-active compound and has been shown to act as antioxidant in a number of in vitro studies (Allegra et al., 2005; Turco Liveri et al., 2009). Interestingly, thanks to its charged portions, ionizable groups and lipophilic moieties, it is amphiphilic at physiological pH (Turco Liveri et al., 2009) and has been demonstrated to interact with cell membranes (Tesoriere et al., 2006; Turco Liveri et al., 2009). This feature is critical to allow bioactive compounds to interact with cells and to initiate signaling events. In this regard, indicaxanthin has been showed to modulate specific redox-dependent signaling pathways involved in macrophage activation and apoptosis, epithelial and endothelial dysfunction in vitro (Allegra et al., 2014; Tesoriere et al., 2015). Remarkably, and in contrast with the majority of dietary phytochemicals, indicaxanthin is highly bioavailable (Tesoriere et al., 2004). The molecule has been shown to cross unaltered intestinal epithelial cell in vitro being absorbed through paracellular junctions (Tesoriere et al., 2013). In line with that, indicaxanthin has been found in human plasma at a 7 μM peak concentration 3 h after the ingestion of four cactus pear fruits containing 28 mg of the pigment (Tesoriere et al., 2004). Moreover, its amphiphilicity allows it to cross the blood-brain-barrier and localize within the CNS (Allegra et al., 2015). Finally, thanks to its bioavailability and redox-modulating properties, indicaxanthin exerts significant pharmacological effects in vivo. Indeed, oral administration of the PhC at nutritionally-relevant doses (2 μmol/kg) generates, in rats, a plasma peak concentration of 0.2 μM able to exert strong anti-inflammatory effects in an in vivo model of acute inflammation (Allegra et al., 2014). The causative link between inflammation and melanoma has accurately been explored in the recent years (Bald et al., 2014; Meyer et al., 2011; Reinhardt et al., 2017; Soudja et al., 2010). Experiments in mice revealed that UV-induced skin inflammatory responses can cause the reactive proliferation and migration of melanocytes (Zaidi et al., 2011). More recently, it has been shown that reciprocal interactions between melanoma and immune cells in a pro-inflammatory microenvironment provide a source of phenotypic heterogeneity that drives therapy resistance and metastasis (Bald et al., 2014; Landsberg et al., 2012). In keeping this perspective, we decided to investigate the effects of Indicaxanthin against human melanoma cell proliferation and in a model of cutaneous melanoma. We here demonstrate that indicaxanthin induces apoptosis of human melanoma cells through the inhibition of the NF-κB pathway and the downstream anti-apoptotic signaling events in vitro and these effects were paralleled in vivo in a murine model of melanoma. Finally, preliminary data on six healthy volunteers, showed that indicaxanthin is able to modulate TNF- and Il-6 production in a whole blood ex vivo model. Furthermore, the phytochemical induces an increase in the phagocytosis of 5 different Gram-negative pathogens on whole blood assay, without exerting antimicrobial effects on them. Interestingly, preliminary data on 4 of the 6 volunteers showed that the observed effects maybe attributed to the modulation of LTB4 levels, strictly correlated to the activation of immune cells.