## 3 Metabolomics Applied to Marine Organisms in Cancer Studies
### Structure Elucidation of Bioactive Molecules from Sponges  
Marine sponges (phylum Porifera) have been largely demonstrated to be one of the richest sources of exclusive secondary metabolites with relevant bioactivity, by means of bioassays [89; 90]. Many scientists are trying to districate the complex network of causes and factors influencing the appearance of neoplastic cells [91; 92]. Metabolomics represents a powerful tool for seeking potentially new and sustainable bioactive compounds in different species of sponges. In fact, two compounds (Stylissamide A and Stylissoside A) from the marine sponge _Stylissa carteri_ sampled in the Red Sea, were extracted and analysed using LC-MS and \({}^{1}\)H-NMR. Further, these compounds were tested on two cancer cell lines. The IC\({}_{50}\) values for Stylissamide A on MCF7 (breast cancer) and on HepG2 (liver cancer) were of 21.1 \(\upmu\)M and 36.8 \(\upmu\)M, respectively, while the IC\({}_{50}\) values for Stylissoside A were of 27.5 \(\upmu\)M on MCF7 and 30.5 \(\upmu\)M on HepG2 [93]. Similar metabolomic approaches (by LC-MS) led to the isolation and characterization of two compounds from another sponge species, _Callyspongia siphonella_. The molecules were characterized by using \({}^{1}\)H-NMR and named 5-bromotrisindoline and 6-bromotrisindoline (see Figure 5 for chemical structure). In particular, 5-bromotrisindoline was effective against HT29 (colon carcinoma), OVCAR3 (ovarian carcinoma) and MM.1S (multiple myeloma) with IC\({}_{50}\) of 8, 7 and 9 \(\upmu\)M, respectively while 6-bromotrisindoline was effective against the same cancer cell lines with IC\({}_{50}\) 12.5, 9 and 11 \(\upmu\)M, respectively [94]. Some sponges living in extreme environmental conditions can also produce interesting compounds, which can be active against human cancer cell lines. This is the case of a specimen belonging to the sponge _Halicona rosea_, collected in shallow water at a hydrothermal vent site. The extract of this sponge was analysed using the MS technique. This made possible the identification of several 3-alkyl-pyridine alkaloids (3-APA) responsible for the reduction of the 70-90% of cell viability of SKBR3 breast cancer cell line [95] at the concentration of 33 \(\upmu\)g/mL in dichloromethane/methanol extracts. In particular, Cyclostelllettamine P, one out of thirteen 3-APA compounds, was characterized, for the first time, by ion mobility mass spectrometry (IMS) in time-aligned parallel (TAP) fragmentation mode. This new technique permits the separation of ionic species as they drift through a gas phase under the influence of an electric field. Then, ions isolated in this way were subjected to subsequent fragmentation in the \"trap\" region of the IMS device [96]. LC-MS led to the identification of 20 compounds that could be responsible for the cytotoxic activity of the crude extract of _Coscinoderma_ sp. on cancer cells. Crude extracts alone showed moderate activity, but were enhanced when the extract was encapsulated in liposomes. In this case, the IC\({}_{50}\) values were notably lower compared to the positive control (Doxorubicin). The tested cell lines were HepG2 (IC\({}_{50}\) = 2.2 \(\upmu\)g/mL), MCF7 (IC\({}_{50}\) = 4.1 \(\upmu\)g/mL) and CaCo2 (colon cancer, IC\({}_{50}\) = 1.7 \(\upmu\)g/mL) [97]. A new compound named Geodia taurine (see Figure 5 for chemical structure) has been isolated from the marine sponge _Geodia macandrewii_, thanks to a complex combination of two techniques for the generation of metabolomics data: UHPLC and MS. The cytotoxic activity of this compound was tested against a melanoma cancer cell line (A2058) and showed weak cytotoxic activity (IC\({}_{50}\) = 8.5 \(\upmu\)M) [98]. Similarly, a sponge coming from the deep-sea Antarctic zone belonging to the species _Lartunculia bifornis_ caught the attention of [PERSON] et al. [99]. The authors revealed, thanks to \({}^{1}\)H-NMR and MS analyses, the presence of known and unknown compounds. However, only three of them showed cytotoxic activity on the HCT-116 colon cancer cell line.