Investigating USP42 Mutation as Underlying Cause of Familial Non-Medullary Thyroid Carcinoma.

In a family with Familial Non-Medullary Thyroid Carcinoma (FNMTC), our investigation using Whole-Exome Sequencing (WES) uncovered a novel germline USP42 mutation [p.(Gly486Arg)]. USP42 is known for regulating p53, cell cycle arrest, and apoptosis, and for being reported as overexpressed in breast and gastric cancer patients. Recently, a USP13 missense mutation was described in FNMTC, suggesting a potential involvement in thyroid cancer. Aiming to explore the USP42 mutation as an underlying cause of FNMTC, our team validated the mutation in blood and tissue samples from the family. Using immunohistochemistry, the expression of USP42, Caspase-3, and p53 was assessed. The USP42 gene was silenced in human thyroid Nthy-Ori 3-1 cells using siRNAs. Subsequently, expression, viability, and morphological assays were conducted. p53, Cyclin D1, p21, and p27 proteins were evaluated by Western blot. USP42 protein was confirmed in all family members and was found to be overexpressed in tumor samples, along with an increased expression of p53 and cleaved Caspase-3. siRNA-mediated USP42 downregulation in Nthy-Ori 3-1 cells resulted in reduced cell viability, morphological changes, and modifications in cell cycle-related proteins. Our results suggest a pivotal role of USP42 mutation in thyroid cell biology, and this finding indicates that USP42 may serve as a new putative target in FNMTC.

High-Throughput Drug Screening Revealed That Ciclopirox Olamine Can Engender Gastric Cancer Stem-like Cells.

Cancer stem cells (CSCs) are relevant therapeutic targets for cancer treatment. Still, the molecular circuits behind CSC characteristics are not fully understood. The low number of CSCs can sometimes be an obstacle to carrying out assays that explore their properties. Thus, increasing CSC numbers via small molecule-mediated cellular reprogramming appears to be a valid alternative tool. Using the SORE6-GFP reporter system embedded in gastric non-CSCs (SORE6-), we performed a high-throughput image-based drug screen with 1200 small molecules to identify compounds capable of converting SORE6- to SORE6+ (CSCs). Here, we report that the antifungal agent ciclopirox olamine (CPX), a potential candidate for drug repurposing in cancer treatment, is able to reprogram gastric non-CSCs into cancer stem-like cells via activation of SOX2 expression and increased expression of C-MYC, HIF-1α, KLF4, and HMGA1. This reprogramming depends on the CPX concentration and treatment duration. CPX can also induce cellular senescence and the metabolic shift from oxidative phosphorylation (OXPHOS) to glycolysis. We also disclose that the mechanism underlying the cellular reprogramming is similar to that of cobalt chloride (CoCl2), a hypoxia-mimetic agent.

Using a Dual CRISPR/Cas9 Approach to Gain Insight into the Role of LRP1B in Glioblastoma.

LRP1B remains one of the most altered genes in cancer, although its relevance in cancer biology is still unclear. Recent advances in gene editing techniques, particularly CRISPR/Cas9 systems, offer new opportunities to evaluate the function of large genes, such as LRP1B. Using a dual sgRNA CRISPR/Cas9 gene editing approach, this study aimed to assess the impact of disrupting LRP1B in glioblastoma cell biology. Four sgRNAs were designed for the dual targeting of two LRP1B exons (1 and 85). The U87 glioblastoma (GB) cell line was transfected with CRISPR/Cas9 PX459 vectors. To assess LRP1B-gene-induced alterations and expression, PCR, Sanger DNA sequencing, and qRT-PCR were carried out. Three clones (clones B9, E6, and H7) were further evaluated. All clones presented altered cellular morphology, increased cellular and nuclear size, and changes in ploidy. Two clones (E6 and H7) showed a significant decrease in cell growth, both in vitro and in the in vivo CAM assay. Proteomic analysis of the clones' secretome identified differentially expressed proteins that had not been previously associated with LRP1B alterations. This study demonstrates that the dual sgRNA CRISPR/Cas9 strategy can effectively edit LRP1B in GB cells, providing new insights into the impact of LRP1B deletions in GBM biology.

High VEGFA Expression Is Associated with Improved Progression-Free Survival after Bevacizumab Treatment in Recurrent Glioblastoma.

Glioblastoma (GB) is one of the deadliest human cancers. Many GB patients do not respond to treatment, and inevitably die within a median of 15-18 months post-diagnosis, highlighting the need for reliable biomarkers to aid clinical management and treatment evaluation. The GB microenvironment holds tremendous potential as a source of biomarkers; several proteins such as MMP-2, MMP-9, YKL40, and VEGFA have been identified as being differentially expressed in GB patient samples. Still to date, none of these proteins have been translated into relevant clinical biomarkers. This study evaluated the expression of MMP-2, MMP-9, YKL40, and VEGFA in a series of GBs and their impact on patient outcome. High levels of VEGFA expression were significantly associated with improved progression-free survival after bevacizumab treatment, thus having potential as a tissue biomarker for predicting patients' response to bevacizumab. Noteworthily, VEGFA expression was not associated with patient outcome after temozolomide treatment. To a lesser extent, YKL40 also provided significant information regarding the extent of bevacizumab treatment. This study highlights the importance of studying secretome-associated proteins as GB biomarkers and identifies VEGFA as a promising marker for predicting response to bevacizumab.

Assessing the Antitumor Potential of Variants of the Extracellular Carbohydrate Polymer from Synechocystis ΔsigF Mutant.

Cancer is a leading cause of death worldwide with a huge societal and economic impact. Clinically effective and less expensive anticancer agents derived from natural sources can help to overcome limitations and negative side effects of chemotherapy and radiotherapy. Previously, we showed that the extracellular carbohydrate polymer of a Synechocystis ΔsigF overproducing mutant displayed a strong antitumor activity towards several human tumor cell lines, by inducing high levels of apoptosis through p53 and caspase-3 activation. Here, the ΔsigF polymer was manipulated to obtain variants that were tested in a human melanoma (Mewo) cell line. Our results demonstrated that high molecular mass fractions were important for the polymer bioactivity, and that the reduction of the peptide content generated a variant with enhanced in vitro antitumor activity. This variant, and the original ΔsigF polymer, were further tested in vivo using the chick chorioallantoic membrane (CAM) assay. Both polymers significantly decreased xenografted CAM tumor growth and affected tumor morphology, by promoting less compact tumors, validating their antitumor potential in vivo. This work contributes with strategies for the design and testing tailored cyanobacterial extracellular polymers and further strengths the relevance of evaluating this type of polymers for biotechnological/biomedical applications.

Different Ability of Multidrug-Resistant and -Sensitive Counterpart Cells to Release and Capture Extracellular Vesicles.

Cancer multidrug resistance (MDR) is one of the main challenges for cancer treatment efficacy. MDR is a phenomenon by which tumor cells become resistant to several unrelated drugs. Some studies have previously described the important role of extracellular vesicles (EVs) in the dissemination of a MDR phenotype. EVs' cargo may include different players of MDR, such as microRNAS and drug-efflux pumps, which may be transferred from donor MDR cells to recipient drug-sensitive counterparts. The present work aimed to: (i) compare the ability of drug-sensitive and their MDR counterpart cells to release and capture EVs and (ii) study and relate those differences with possible distinct fate of the endocytic pathway in these counterpart cells. Our results showed that MDR cells released more EVs than their drug-sensitive counterparts and also that the drug-sensitive cells captured more EVs than their MDR counterparts. This difference in the release and capture of EVs may be associated with differences in the endocytic pathway between drug-sensitive and MDR cells. Importantly, manipulation of the recycling pathway influenced the response of drug-sensitive cells to doxorubicin treatment.

LRP1B Expression as a Putative Predictor of Response to Pegylated Liposomal Doxorubicin Treatment in Ovarian Cancer.

BACKGROUND: Pegylated liposomal doxorubicin (PLD) is among the most active therapies for recurrent/progressive ovarian cancer (OC). Low-density lipoprotein receptor-related protein 1B (LRP1B) is one of the 10 most significantly deleted genes in human cancers. It mediates endocytosis of several factors from the cellular environment including liposomes. Although the LRP1B role in cancer has not been fully disclosed, its contribution to resistance to liposomal therapies has been hypothesized. This study aimed to evaluate the impact of LRP1B protein as a possible marker of response to PLD in patients with OC. METHODS: LRP1B expression and response to PLD were analyzed in OC cell lines by qRT-PCR and PrestoBlue viability assay, respectively. LRP1B protein expression was evaluated for the first time, in tumor samples from PLD-treated patients and controls (other chemotherapies) by immunohistochemistry. Association of LRP1B staining score (determined based on intensity and percentage of positively stained cells) with clinicopathological features, response to therapy and survival outcomes was evaluated. RESULTS: OC cells with increased expression of LRP1B were more sensitive to PLD. LRP1B staining score was associated with clinicopathological features, response to therapy, and survival outcomes. Higher LRP1B levels were associated with prolonged progression-free survival. This association was more evident in patients treated with PLD and in responders to PLD. CONCLUSION: Our results support a possible role of LRP1B as a predictor of response to PLD in patients with OC.

LRP1B: A Giant Lost in Cancer Translation.

Low-density lipoprotein receptor-related protein 1B (LRP1B) is a giant member of the LDLR protein family, which includes several structurally homologous cell surface receptors with a wide range of biological functions from cargo transport to cell signaling. LRP1B is among the most altered genes in human cancer overall. Found frequently inactivated by several genetic and epigenetic mechanisms, it has mostly been regarded as a putative tumor suppressor. Still, limitations in LRP1B studies exist, in particular associated with its huge size. Therefore, LRP1B expression and function in cancer remains to be fully unveiled. This review addresses the current understanding of LRP1B and the studies that shed a light on the LRP1B structure and ligands. It goes further in presenting increasing knowledge brought by technical and methodological advances that allow to better manipulate LRP1B expression in cells and to more thoroughly explore its expression and mutation status. New evidence is pushing towards the increased relevance of LRP1B in cancer as a potential target or translational prognosis and response to therapy biomarker.

Predictive Biomarkers and Patient Outcome in Platinum-Resistant (PLD-Treated) Ovarian Cancer.

Identification of predictive biomarkers for ovarian cancer (OC) treatment, particularly in the platinum-resistant/refractory setting, is highly relevant for clinical management. E-cadherin, vimentin, and osteopontin (OPN) are proteins associated with tumor microenvironment (TME) remodelling that play key roles in cancer. This study aimed to evaluate the association between the staining patterns of these proteins with survival outcomes in a series of OC patients, namely in patients with platinum-resistant/refractory disease. Low E-cadherin expression and high vimentin expression in all patient groups (as well as for E-cadherin in the platinum-resistant arm) were significantly associated with longer overall survival (OS). Low cytoplasmic OPN expression (and cytoplasmic and membrane OPN in the platinum-resistant arm) were significantly associated with longer OS. In patients that responded to treatment (pegylated liposomal doxorubicin (PLD) or other), low cytoplasmic OPN expression was also associated with longer progression-free survival (PFS). In the other hand, high nuclear OPN-c expression in patients that respond to treatment was associated with longer OS and longer PFS. Longer PFS was also associated with high expression of both nuclear and cytoplasm OPN-c, in platinum-resistant patients and in those that responded to PLD. Our study indicates that the expression of E-cadherin, vimentin, and OPN may have prognostic implications. Nuclear OPN-c and cytoplasm OPN expression are putative predictive markers in platinum-resistant (PLD treated) ovarian cancer patients.

Deep Sequencing Analysis Reveals Distinctive Non-Coding RNAs When Comparing Tumor Multidrug-Resistant Cells and Extracellular Vesicles with Drug-Sensitive Counterparts.

Multidrug resistance (MDR) is one of the main limitations of cancer treatment. The overexpression of drug-efflux pumps, such as P-glycoprotein (P-gp), is a major cause of MDR. Importantly, different studies have shown that extracellular vesicles (EVs) participate in the communication between MDR cells and drug-sensitive counterparts, promoting dissemination of the MDR phenotype. In the present work, we aimed to identify RNA species present in MDR cells and in EVs released by those cells, which may be associated with the MDR phenotype. The RNA content from two pairs (leukemia and lung cancer) of MDR (P-gp overexpressing) cells and their drug-sensitive counterparts, as well as from their EVs, was analyzed by deep sequencing. Our results showed distinctive transcripts for MDR cells and their EVs, when compared with their drug-sensitive counterparts. Remarkably, two pseudogenes (a novel pseudogene and RNA 5.8S ribosomal pseudogene 2) were found to be increased in EVs released by MDR cells in both leukemia and lung cancer models. Moreover, six miRs (miR-204-5p, miR-139-5p, miR-29c-5p, miR-551b-3p, miR-29b-2-5p, and miR-204-3p) exhibited altered levels in lung cancer MDR cells and their EVs. This study provides insights into the contribution of EVs to MDR.

Characterization and antitumor activity of the extracellular carbohydrate polymer from the cyanobacterium Synechocystis ΔsigF mutant.

Cyanobacterial extracellular carbohydrate polymers are particularly attractive for biotechnological applications. Previously, we determined the monosaccharidic composition of the polymer of a Synechocystis ΔsigF overproducing mutant. Here, we further characterized this polymer, demonstrated that it is possible to recover it in high yields, and successfully use it for biomedical research. This amorphous polymer is formed by a mesh of fibrils/lamellar structures with high porosity, is constituted by high molecular mass fractions, is highly sulfated and displays low viscosity, even in highly concentrated aqueous solutions. FTIR analysis confirmed the presence of several functional groups. We demonstrated that the ΔsigF polymer has strong biological activity, decreasing the viability of melanoma, thyroid and ovary carcinoma cells by inducing high levels of apoptosis, through p53 and caspase-3 activation. Therefore, the ΔsigF Synechocystis mutant is a promising platform for the sustainable production of biological active carbohydrate polymer(s) with the desired characteristics for biomedical applications.

The Antitumor Activity of a Lead Thioxanthone is Associated with Alterations in Cholesterol Localization.

The search for novel anticancer small molecules and strategies remains a challenge. Our previous studies have identified TXA1 (1-{[2-(diethylamino)ethyl]amino}-4-propoxy-9H- thioxanthen-9-one) as a hit compound, with in vitro antitumor potential by modulating autophagy and apoptosis in human tumor cell lines. In the present study, the mechanism of action and antitumor potential of the soluble salt of this molecule (TXA1.HCl) was further investigated using in vitro and mouse xenograft tumor models of NSCLC. Our results showed that TXA1.HCl affected steroid biosynthesis, increased RagD expression, and caused abnormal cellular cholesterol localization. In addition, TXA1.HCl treatment presented no toxicity to nude mice and significantly reduced the growth of human NSCLC cells xenografts in mice. Overall, this work provides new insights into the mechanism of action of TXA1, which may be relevant for the development of anticancer therapeutic strategies, which target cholesterol transport.

OPNa Overexpression Is Associated with Matrix Calcification in Thyroid Cancer Cell Lines.

Osteopontin (OPN) spliced variants (OPN-SV: OPNa, OPNb, and OPNc) are aberrantly expressed in tumors and frequently associated with cancer progression. This holds true for papillary thyroid carcinoma (PTC), which is the most common type of thyroid cancer (TC). PTC often presents with desmoplasia and dystrophic calcification, including psammoma bodies (PB). This work aimed to investigate total OPN (tOPN) and OPN-SV expression and their association with the presence of PB in the PTC classical variants (cPTC), as well as the involvement of OPN-SV in matrix calcification of TC cell lines. We found that cPTC samples presenting PB showed higher OPN expression levels. In TC cell lines, OPNa overexpression promotes higher matrix calcification and collagen synthesis when compared to that of clones overexpressing OPNb or OPNc. In response to OPN knockdown, calcification was inhibited, paralleled with the downregulation of calcification markers. In conclusion, our data evidenced that OPN expression is associated with the presence of PB in cPTC samples. Among the OPN-SV, OPNa is the main contributor to matrix calcification in tested TC cells, providing clues to a better understanding on the biology and ethiopathogenesis of the calcification process in TC cells.

Liposomal therapies in oncology: does one size fit all?

Liposomal therapies opened the chapter of nanomedicine, in 1995, with the approval of liposomal doxorubicin (Doxil®) for the treatment of numerous types of cancer. For the first time, liposomes permitted the employment of potent chemotherapeutic agents with improved pharmacokinetic and pharmacodynamic profiles, with less undesired side effects. Liposomal therapies allow the drug encapsulation and more selective delivery in the tumor bed, particularly due to the enhanced permeation and retention effect. These unique characteristics explain why liposomal therapies are being increasingly considered as alternatives in cancer therapy and represent an important research field with the recent approval for clinical use and emergent formulations in clinical trials. Even so, the response rate of liposomal therapies varies between 5 and 71%, and they are still indistinguishably given to every patient. As already well-demonstrated for conventional chemotherapies and targeted therapies, there is also the need for predictive biomarkers that allow a better use of liposomal drugs, with higher patient quality of life. Our aim in this review was to address the approved liposomal therapies and to summarize the information concerning possible predictive markers of response in their various clinical applications, to personalize each patient treatment and maximize its efficacy.

Synthesis and Evaluation of the Tumor Cell Growth Inhibitory Potential of New Putative HSP90 Inhibitors.

BACKGROUND: Heat shock protein 90 (HSP90) is a well-known target for cancer therapy. In a previous work, some of us have reported a series of 3-aryl-naphtho[2,3-d]isoxazole-4,9-diones as inhibitors of HSP90. METHODS: In the present work, various compounds with new chromenopyridinone and thiochromenopyridinone scaffolds were synthesized as potential HSP90 inhibitors. Their binding affinity to HSP90 was studied in vitro. Selected compounds (5 and 8) were further studied in various tumor cell lines regarding their potential to cause cell growth inhibition, alter the cell cycle profile, inhibit proliferation, and induce apoptosis. Their effect on HSP90 client protein levels was also confirmed in two cell lines. Finally, the antitumor activity of compound 8 was studied in A431 squamous cell carcinoma xenografts in nude mice. RESULTS: Our results indicated that treatment with compounds 5 and 8 decreased the proliferation of tumor cell lines and compound 8 induced apoptosis. In addition, these two compounds were able to downregulate selected proteins known as "clients" of HSP90. Finally, treatment of xenografted mice with compound 5 resulted in a considerable dose-dependent inhibition of tumor growth. CONCLUSIONS: Our results show that two new compounds with a chromenopyridinone and thiochromenopyridinone scaffold are promising putative HSP90 inhibitors causing tumor cell growth inhibition.

Identification of the metabolic alterations associated with the multidrug resistant phenotype in cancer and their intercellular transfer mediated by extracellular vesicles.

Multidrug resistance (MDR) is a serious obstacle to efficient cancer treatment. Overexpression of P-glycoprotein (P-gp) plays a significant role in MDR. Recent studies proved that targeting cellular metabolism could sensitize MDR cells. In addition, metabolic alterations could affect the extracellular vesicles (EVs) cargo and release. This study aimed to: i) identify metabolic alterations in P-gp overexpressing cells that could be involved in the development of MDR and, ii) identify a potential role for the EVs in the acquisition of the MDR. Two different pairs of MDR and their drug-sensitive counterpart cancer cell lines were used. Our results showed that MDR (P-gp overexpressing) cells have a different metabolic profile from their drug-sensitive counterparts, demonstrating decreases in the pentose phosphate pathway and oxidative phosphorylation rate; increases in glutathione metabolism and glycolysis; and alterations in the methionine/S-adenosylmethionine pathway. Remarkably, EVs from MDR cells were capable of stimulating a metabolic switch in the drug-sensitive cancer cells, towards a MDR phenotype. In conclusion, obtained results contribute to the growing knowledge about metabolic alterations in MDR cells and the role of EVs in the intercellular transfer of MDR. The specific metabolic alterations identified in this study may be further developed as targets for overcoming MDR.

Suppression of spindly delays mitotic exit and exacerbates cell death response of cancer cells treated with low doses of paclitaxel.

Microtubule-targeting agents (MTAs) are used extensively for the treatment of diverse types of cancer. They block cancer cells in mitosis through the activation of the spindle assembly checkpoint (SAC), the surveillance mechanism that ensures accurate chromosome segregation at the onset of anaphase. However, the cytotoxic activity of MTAs is limited by premature mitotic exit (mitotic slippage) due to SAC silencing. Here we have explored the dual role of the protein Spindly in chromosome attachments and SAC silencing to analyze the consequences of its depletion on the viability of tumor cells treated with clinically relevant doses of paclitaxel. As expected, siRNA-mediated Spindly suppression induced chromosome misalignment and accumulation of cells in mitosis. Remarkably, these cells were more sensitive to low-doses of paclitaxel. Sensitization was due to an increase in the length of mitotic arrest and high frequency of multinucleated cells, both correlated with an exacerbated post-mitotic cell death response as determined by cell fate profiling. Thus, by affecting both SAC silencing and chromosome attachment, Spindly targeting offers a double-edged sword that potentiates tumor cell killing by clinically relevant doses of paclitaxel, providing a rationale for combination chemotherapy against cancer.

A novel curcumin derivative which inhibits P-glycoprotein, arrests cell cycle and induces apoptosis in multidrug resistance cells.

Cancer multidrug resistance (MDR) is a major limitation to the success of cancer treatment and is highly associated with the overexpression of drug efflux pumps such as P-glycoprotein (P-gp). In order to achieve more effective chemotherapeutic treatments, it is important to develop P-gp inhibitors to block/decrease its activity. Curcumin (1) is a secondary metabolite isolated from the turmeric of Curcuma longa L.. Diverse biological activities have been identified for this compound, particularly, MDR modulation in various cancer cell models. However, curcumin (1) has low chemical stability, which severely limits its application. In order to improve stability and P-gp inhibitory effect, two potential more stable curcumin derivatives were synthesized as building blocks, followed by several curcumin derivatives. These compounds were then analyzed in terms of antitumor and anti-P-gp activity, in two MDR and sensitive tumor lines (from chronic myeloid leukemia and non-small cell lung cancer). We identified from a series of curcumin derivatives a novel curcumin derivative (1,7-bis(3-methoxy-4-(prop-2-yn-1-yloxy)phenyl)hepta-1,6-diene-3,5-dione, 10) with more potent antitumor and anti-P-gp activity than curcumin (1). This compound (10) was shown to promote cell cycle arrest (at the G2/M phase) and induce apoptosis in the MDR chronic myeloid leukemia cell line. Therefore it is a really interesting P-gp inhibitor due to its ability to inhibit both P-gp function and expression.

Modulation of Autophagy by a Thioxanthone Decreases the Viability of Melanoma Cells.

(1) Background: Our previous studies unveiled the hit thioxanthone TXA1 as an inhibitor of P-glycoprotein (drug efflux pump) and of human tumor cells growth, namely of melanoma cells. Since TXA1 is structurally similar to lucanthone (an autophagy inhibitor and apoptosis inducer) and to N10-substituted phenoxazines (isosteres of thioxanthones, and autophagy inducers), this study aimed at further assessing its cytotoxic mechanism and evaluating its potential as an autophagy modulator in A375-C5 melanoma cells; (2) Methods: Flow cytometry with propidium iodide (PI) for cell cycle profile analysis; Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, flow cytometry with Annexin V/PI labeling and Western blot for apoptosis analysis were conducted. A pharmacophore approach was used for mapping TXA1 onto pharmacophores for autophagy induction. Autophagy analyses included transmission electron microscopy for visualization of autophagic structures, fluorescence microscopy for observation of monodansylcadaverine (MDC) staining, pattern of LC3 expression in the cells and acridine orange staining, and Western blot for autophagic proteins expression; (3) Results: TXA1 induced autophagy of melanoma cells at the GI50 concentration (3.6 µM) and apoptosis at twice that concentration. Following treatment with TXA1, autophagic structures were observed, together with the accumulation of autophagosomes and the formation of autophagolysosomes. An increase in LC3-II levels was also observed, which was reverted by 3-methyladenine (3-MA) (an early stage autophagy-inhibitor) but further increased by E-64d/pepstatin (late-stage autophagy inhibitors). Finally, 3-MA also reverted the effect of TXA1 in cellular viability; (4) Conclusion: TXA1 decreases the viability of melanoma cells by modulation of autophagy and may, therefore, serve as a lead compound for the development of autophagy modulators with antitumor activity.

An Aqueous Extract of Tuberaria lignosa Inhibits Cell Growth, Alters the Cell Cycle Profile, and Induces Apoptosis of NCI-H460 Tumor Cells.

Tuberaria lignosa (Sweet) Samp. is found in European regions, and has antioxidant properties due to its composition in ascorbic acid and phenolic compounds. Given its traditional use and antioxidant properties, the tumor cell growth inhibitory potential of aqueous extracts from T. lignosa (prepared by infusion and decoction) was investigated in three human tumor cell lines: MCF-7 (breast adenocarcinoma), NCI-H460 (non-small cell lung cancer), and HCT-15 (human colorectal adenocarcinoma). Both extracts inhibited the growth of these cell lines; the most potent one being the T. lignosa extract obtained by infusion in the NCI-H460 cells (GI50 of approximately 50 µg/mL). Further assays were carried out with this extract in NCI-H460 cells. At 100 µg/mL or 150 µg/mL it caused an increase in the percentage of cells in the G0/G1 phase and a decrease of cells in S phase of the cell cycle. Additionally, these concentrations caused an increase in the percentage of apoptotic cells. In agreement, a decrease in total poly (ADP-ribose) polymerase (PARP) and pro-caspase 3 levels was found. In conclusion, the T. lignosa extract obtained by infusion was more potent in NCI-H460 cells, altering the cell cycle progression and inducing apoptosis. This work highlights the importance of T. lignosa as a source of bioactive compounds with tumor cell growth inhibitory potential.