## 4 Discussion
PRDX1 was highly expressed and is the predominant peroxiredoxin isoform in cells. According to the Human Protein Atlas (www.proteinatlas.org), comparing A549 cells with several other cell lines show that these cells rank along those with lower catalase levels. In addition, the levels of PRDX2 were undetected in A549 cells compared with other cell lines. Similar results in terms of gene expression were observed in studies with lung cancer. Some authors compared the antioxidant levels between tumor and tumor-free lung tissue. The lung tumor tissue showed lower levels of catalase expression than healthy tissue, while similar levels of GPX1 were detected in both cancerous and non-cancerous tissue. In addition, by performing the immunohistochemical localization of catalase, the results revealed lower or no expression of this enzyme in tumor cells [55]. [PERSON] et al. analyzed the expression of several PRDXs in multiple cell lines, and the absence of expression of PRDX2 was also found in A549 cells [56]. Overall, our analysis, along with the information reported elsewhere, pointed out the low gene expression level of these key enzymes, particularly catalase, in NSCLC cell lines, suggesting their propensity to changes in redox balance, thus anticipating the usefulness of the MnPs-based redox modulators, in the context of this type of cancer. Cancer cells use high levels of ROS to their own advantage for proliferation. Yet, the balance is delicate, and depending upon the levels of endogenous antioxidants, a tiny amount of external ROS may drive cancer cells to death. We first evaluated the cytotoxic impact of MnTnHex in both NSCLC cell lines using complementary methodologies. MnTnHex, as a single drug, exhibited a concentration-dependent decrease in terms of cell viability, revealing quite low IC\({}_{50}\) values, in some cases reaching the sub-micromolar range. This fact, we believe, is of high relevance since it demonstrates the ability of this compound to act as a promising stand-alone drug in NSCLC. It should be emphasized that our previously published results with MnTnHex using cells of non-tumor origin clearly revealed opposite results. Indeed, this compound had already been assessed in V79 lung fibroblasts (MTT reduction assay, 24-h incubation) [33] as well as in renal cells (Vero cells) (CV assay, 48 h-incubation) [57]. In both cases, MnTnHex at concentrations up to 25 \(\upmu\)M showed no evidence of cytotoxicity. Conversely, in our previous studies, MnTnHex already exhibited high cytotoxic effects in renal cancer cells (786-O) [29], which is in line with the herein observed results with NSCLC cells and demonstrates this important feature in different invasive cancer cells. While there are some experimental differences in the protocols used, it is clear that MnTnHex has differential effects in malignant vs. non-tumor cells. The gene expression data concerning catalase, and PRDX2, suggest that both cell lines are under high oxidative stress. Even without a cisplatin-derived source of H2O2, Mn porphyrins alone employ enough high levels of H2O2 to inflict oxidative damage to numerous enzymes, inactivating them and suppressing thus their activity [25; 28]. Such results are supported by data showing that the IC\({}_{50}\) values for MnTnHex in the present study are lower in both cell lines (between 0.7 \(\upmu\)M and 2.1 \(\upmu\)M depending on the type of assay and cell) than for cisplatin alone, indicating its potency in enhancing cellular oxidative stress beyond repair, and in turn death. By combining MnTnHex with cisplatin, we further aimed to study a different aspect of the potential usefulness of this compound in NSCLC. That is its ability to increase the efficacy of chemotherapy treatment. Our group has been studying the sensitivity of cisplatin in different cell lines. Regarding A549 cells, these cells are more sensitive to cisplatin than H1975 cells having IC\({}_{50}\) values up to 2.8 \(\upmu\)M (Table S2).