HCT116 Colorectal Cancer Cells Research Articles

PD15, a steroidal saponin, induces apoptosis of HCT116 colorectal cancer cells via suppressing the Akt/GSK3β pathway.

PD15, a novel natural steroidal saponin extracted from the rhizomes of Paris delavayi Franchet, has demonstrated a strong cytotoxic effect against HepG2 and U87MG cells. However, its therapeutic effects on colorectal cancer (CRC) and the underlying molecular mechanisms remain unclear. MTT assay, clonogenic assay, Hoechst 33258 staining, flow cytometry, molecular docking, and western blot were used to investigate the mechanism of PD15 in HCT116 cell lines. Additionally, the anti-CRC effects of PD15 were evaluated in vivo using HCT116 xenograft models. PD15 significantly inhibited cell proliferation and induced G0/G1 phase arrest in HCT116 cells. Furthermore, PD15 upregulated cleaved Caspase 3 and 9, cleaved PARP, and Bax expression levels while downregulating Bcl-2, leading to apoptosis. Further experiments revealed that PD15 downregulated the protein expression of p-Akt and p-GSK3β, with LY294002 (a PI3K/Akt inhibitor) enhancing PD15-induced apoptosis and its effects on Akt/GSK3β-associated proteins. In addition, molecular docking demonstrated that PD15 exhibited strong binding affinity with Akt and GSK3β. Critically, PD15 inhibited CRC growth in vivo without causing apparent toxicity in mice. These findings indicate that PD15 could trigger apoptosis by suppressing the Akt/GSK3β signaling pathway in HCT116 cells.

Jujuboside B inhibits proliferation and induces apoptosis and ferroptosis in colorectal cancer cells with potential involvement of the MAPK signaling pathway.

Colorectal cancer (CRC) is one of the most prevalent and life-threatening malignancies worldwide. Jujuboside B (JUB) is a bioactive compound derived from the seeds of Ziziphus jujuba, known for its potential anticancer properties. The present study aimed to investigate the association of JUB with inhibiting the proliferation, apoptosis and ferroptosis of human CRC cells with mitogen-activated protein kinase (MAPK) pathway regulation. First, the human CRC HCT116 cell line was treated with different concentrations of JUB. Subsequently, cell viability was evaluated using MTT assay and colony formation was assessed using a colony formation assay. Flow cytometry was used to detect cell apoptosis and the levels of reactive oxygen species. Western blotting was utilized to assess the expression levels of apoptosis-related proteins, ferroptosis regulators and MAPK pathway-related proteins. In addition, biochemical assay kits were used to evaluate the levels of malondialdehyde, glutathione, total iron and ferrous iron. The results demonstrated that cell viability and colony formation were markedly decreased after JUB treatment, whilst the level of apoptosis was notably increased in a concentration-dependent manner. Using electron microscopy, cells treated with JUB exhibited typical apoptotic bodies, as well as mitochondrial swelling and cristae disruption, further demonstrating JUB-induced cell apoptosis. Western blot analysis indicated that JUB treatment markedly reduced the expression of B-cell lymphoma-2 (Bcl-2) but notably increased the expression of Bcl-2 associated X-protein and cleaved caspase-3. Additionally, JUB induced ferroptosis and inhibited the MAPK signaling pathway in CRC cells. Collectively, the findings of the present study suggest that JUB has the potential to inhibit CRC cell proliferation and induce apoptosis through regulating the MAPK pathway. Therefore, JUB may be a promising therapeutic agent for the treatment of CRC.

The Olive Oil Phenolic S-(-)-Oleocanthal Suppresses Colorectal Cancer Progression and Recurrence by Modulating SMYD2-EZH2 and c-MET Activation

Background/Objectives: Colorectal cancer (CRC) is the third most common cancer in the US and the second leading cancerassociated mortality cause. Available CRC therapies achieve modest outcomes and fail to prevent its recurrence. Epidemiological studies indicated that the Mediterranean diet rich in olive oil reduced CRC incidence. This study aimed at the identification and assessment of active anti-CRC olive phenolics. Methods: The MTT, wound-healing and colony formation assays were used to discover and assess the in vitro anti-CRC activity of olive phenolics. A nude mouse xenografting model was used to assess the in vivo CRC progression and recurrence suppressive activity of OC in pure and crude forms. OC was isolated from olive oil using liquid–liquid extractions. Results: Screening of olive phenolics for in vitro antiproliferative activity against a diverse panel of CRC cell lines identified the extra-virgin olive oil (EVOO) S-(-)-oleocanthal (OC) as the most active hit. OC showed IC50 values of 4.2, 9.8, 14.5, and 4.9 μM against HCT-116, COLO-320DM, WiDr, and SW48 CRC cells, respectively. The lysine methyltransferases SMYD2 and EZH2, along with the receptor tyrosine kinase c-MET proved aberrantly dysregulated in invasive and metastatic CRC. SMYD2 and c-MET were validated as OC molecular targets in multiple malignancies. Daily oral 10 mg/kg OC treatments over 15 days suppressed 72.5% of the KRAS mutant HCT-116-Luc cells tumors weight in male nude mice. Continued OC daily oral use after primary tumor surgical excision over an additional 40 days significantly suppressed the HCT-116-Luc locoregional tumor recurrence and totally prevented the distant tumor recurrence. The SMYD2-EZH2 expressions and c-MET activation were notably suppressed by OC treatments in vitro and in collected animal primary tumors. Conclusions: OC and olive phenolics are potential nutraceutical interventions useful for CRC control and the prevention of its relapse.

Differential effects of β-hydroxybutyrate and α-ketoglutarate on HCT-116colorectal cancer cell viability under normoxic and hypoxic low-glucose conditions: exploring the role of SRC, HIF1α, ACAT1, and SIRT2 genes.

Recent therapeutic strategies have highlighted the potential of β-hydroxybutyrate (BHB) and α-ketoglutarate (α-KG) as effective anticancer agents, particularly for colon cancer. These metabolites can modulate cellular metabolism and induce epigenetic changes, inhibiting tumor growth. Nonetheless, certain cancer cells may utilize ketone bodies, like BHB as nutrient sources under hypoxic conditions, potentially reducing treatment efficacy. Understanding these mechanisms is crucial for optimizing cancer therapies. This study evaluated the effects of BHB and α-KG on HCT-116 colorectal cancer cell viability under normoxic and low-glucose hypoxic conditions. HCT-116 cell lines were treated with different doses of BHB and α-KG in normoxic and low-glucose hypoxic conditions, and then cell viability was assessed by the MTT assay. Moreover, the mRNA expression levels of SRC, hypoxia-inducible factor 1α(HIF-1α), acetyl-CoA acetyltransferase 1 (ACAT1), and sirtuin 2 (SIRT2) genes were determined using quantitative reverse transcriptase-polymerase chain reaction (q RT-PCR). BHB significantly increased the proliferation of HCT-116 colon cancer cells under low-glucose hypoxic conditions, while α-KG maintained cell viability in normoxic conditions but not in hypoxia. BHB treatment reduced SIRT2 mRNA levels and increased ACAT1, SRC, and HIF-1α expression. Conversely, α-KG decreased ACAT1, SRC, and HIF-1α expression and increased SIRT2 levels in normoxia but could not reverse gene expression during hypoxia. Our study demonstrated that BHB and α-KG exhibited complex interactions with colon cancer cell viability under varying oxygen and glucose conditions. While BHB promoted cell proliferation in hypoxic environments, α-KG showed protective effects in normoxic conditions. This research contributed to the growing body of evidence supporting the role of metabolic modulators in cancer therapy and emphasized the importance of understanding tumor microenvironments to optimize treatment outcomes. However, the need for further research into the metabolic pathways is underscored to enhance therapeutic strategies for cancer treatment.

Mechanistic Insights into the Inhibition of Colorectal Cancer by BuShenFang through Adenomatous Polyposis Coli Expression and Wnt/β-catenin Pathway Regulation

Colorectal cancer (CRC) remains one of the leading causes of cancer-related mortality worldwide. Despite advances in chemotherapy and radiotherapy, significant side effects persist, prompting the exploration of alternative therapies such as traditional Chinese medicine (TCM). BuShenFang (BSF), a TCM formulation, is believed to exhibit anti-CRC effects, although its exact mechanism is unclear. This study aims to investigate the effects of BSF on CRC cells through the modulation of the Wnt/β-catenin pathway. The study utilized HCT116 and SW620 CRC cell lines, employing in vitro experiments including cell viability assays, colony formation, flow cytometry, and Western blot to examine the influence of BSF on cellular proliferation, apoptosis, migration, and the Wnt/β-catenin signaling pathway. Results demonstrated that BSF significantly inhibited the proliferation of CRC cells in a dose-dependent manner. The apoptotic rate was markedly increased in the 20% BSF group, while colony formation, migration, and invasion capabilities of CRC cells were notably suppressed. Furthermore, Western blot results revealed that BSF enhanced adenomatous polyposis coli (APC) expression and inhibited β-catenin, c-Myc, and Cyclin D1, key proteins in the Wnt/β-catenin pathway. In conclusion, BSF exerts anti-CRC effects by modulating the Wnt/β-catenin pathway, suggesting its potential as a complementary therapeutic agent in CRC treatment. Future studies should focus on in vivo models to validate these findings.

LC-MS/MS-based proteomics and metabolomics of HCT-116 colorectal cancer cells: A potential anticancer activity of atorvastatin

Colorectal cancer (CRC) is the third most prevalent tumor in men, the second most common in women, and the fourth leading cause of mortality worldwide. Statins reduce cholesterol levels by hampering the function of 3-hydroxy-3-methyl-glutaryl-CoA reductase enzymes in cholesterol synthesis. Strains have shown anticancer effects against CRC. However, statins’ anticancer mechanism is yet unknown. Liquid chromatography–mass spectrometry (LC–MS)-based untargeted metabolomics and proteomics were employed to study statins’ effects on CRC using the CRC cell line HCT-116. These approaches were utilized to identify potential underlying metabolic pathways and proteins altered in atorvastatin (a statin)-treated HCT-116 cells. Compared to the control, atorvastatin significantly altered numerous metabolites in HCT-116 cells, including a reduction in the levels of decanoylcarnitine and octanoyl-L-carnitine and in the biosynthesis and metabolism of amino acids like alanine and citrate cycle. Proteomic study showed that atorvastatin-treated HCT-116 cells expressed 127 proteins differently from controls. Novel findings were among them, such as centromere-associated protein E, cytochrome c oxidase subunit 6A1 mitochondrial, and hyaluronan synthase 1. The findings indicate that atorvastatin may have potential anticancer characteristics and highlight the essential role metabolomics and proteomics to understand complex metabolic pathways and proteins relevant to cancer and develop novel treatment targets.

Development of 1,2,3-triazole hybrids as multi-faced anticancer agents co-targeting EGFR/mTOR pathway and tubulin depolymerization.

Novel 1,2,3-triazole hybrids bearing various substituents have been synthesized as potential anticancer agents. Ligand-based approach has been adopted to design these compounds relying on the hybridization of 1,2,3-triazole with α,β-unsaturated carbonyl, 5- and 6-membered heterocyclic scaffolds. All synthesized members were investigated for their cytotoxic potency against nine types comprising 60 panels of human cancerous cells by the US National Cancer Institute: Development Therapeutic Program (US_NCI_DTP). Among the tested members, 4b, 4e, and 4h showed prominent cytotoxic effects (> 80% growth inhibition: GI) on a wide panel of tested cancer cell lines, mainly melanoma and colorectal cancer redeeming their selection for five dose testing. Presenting low nanomolar GI50 concentrations, two representative potent anticancer compounds 4b and 4e were subjected to cytotoxicity testing on colon normal cell (FHC) to investigate their safety window and they showed less toxicity to normal cells at the concentration required to produce anticancer effect. Furthermore, 4b and 4e were exposed to additional mechanistic studies in colorectal cancer cell HCT-116 suggesting multifaceted mechanisms of action. A study into the effects of cytotoxic chemicals 4b and 4e on cell cycle progression regulation showed triggered the arrest of cell cycles during the G1 and S phases. Moreover, 4b and 4e caused cell death mainly through apoptosis the thing that has been reinforced by the elevated Bax: Bcl2 ratio, as well as concentrations of caspases 3 and 9 within HCT-116. Further, both compounds showed prominent inhibition profiles against tubulin polymerization as well as EGFR catalytic activity reaching down to low-digit micromolar and sub-micromolar concentrations, respectively, as compared to positive reference controls. Compounds' impacts on gene expression of cancer-associated and EGFR-downstream signaling markers including TNFα, IL-6, and mTOR, were explored in HCT-116 highlighted significant downregulations versus the untreated cells. Docking studies demonstrated the specific fit of 4b and 4e into EGFR and the colchicine binding site of tubulin.

Targeting the Hippo and Rap1 signaling pathways: the anti-proliferative effects of curcumin in colorectal cancer cell lines.

CRC has the third-highest cancer incidence and death. Many human cancers, including colorectal cancer, are connected to abnormal signaling pathway gene expression. Many human malignancies include Hippo and Rap1 signaling. This research examined curcumin's therapeutic effects on colorectal cancer cell lines' Hippo and Rap1 signaling pathway genes. The role of the above signaling pathways is considered in colorectal cancer development. No research has examined curcumin's influence on key genes in these pathways; thus, this work ismeant to uncover its more precise mechanism. First, the gene expression omnibus database is queried to discover GSE8671, a dataset that contains differentially expressed genes associated in CRC formation. DAVID was used to discover the corporation of these genes and signaling pathways (Hippo and Rap1), and the cancer genome atlas (TCGA) database was utilized to select genes and assess their expression and biomarker potential. MTT, apoptosis, and quantitative PCR were used to assess whether curcumin is therapeutic for colorectal cancer cell lines. An in-silico analysis identified the dysregulation of several critical genes AXIN2, MYC, TEAD4, MET, LPAR1, and ADCY9 in colorectal cancer, highlighting their involvement in the Hippo and Rap1 signaling pathways. Experimental assessments, including MTT assays, apoptosis assays, and quantitative PCR (qPCR) analysis, demonstrated that the targeted modulation of these genes effectively inhibits cancer cell proliferation. Specifically, treatment with curcumin resulted in a significant reduction in cell viability in HT-29 and HCT-116 colorectal cancer cell lines, thereby facilitating apoptotic cell death. Furthermore, curcumin administration was associated with the upregulation of LPAR1 and ADCY9 gene expression, while concurrently downregulating AXIN2, MYC, TEAD4, and MET in both cell lines. This study reveals compelling evidence of curcumin's potent anticancer properties, highlighting its transformative influence on the Hippo and Rap1 signaling pathways within colorectal cancer cells. These findings not only underscore curcumin's potential as a therapeutic agent but also pave the way for innovative strategies in the fight against colorectal cancer.

Folic acid-targeted β-lactoglobulin nanocarriers for enhanced delivery of 5-fluorouracil and sodium butyrate in colorectal cancer treatment.

Colorectal cancer (CRC) remains a significant public health concern, emphasizing the need for innovative therapeutic strategies to improve patient outcomes. This study aimed to develop a highly efficient nanocarrier for targeted drug delivery, enhancing drug efficacy while minimizing concentrations and limiting adverse effects. We synthesized protein-based β-lactoglobulin (βlg) nanoparticles (NPs), loaded with 5-fluorouracil (5-FU) and sodium butyrate (NaB), and further functionalized with folic acid (FA) for specific targeting of folate receptor-positive CRC cells. The βlg-5-FU-NaB-FA nanoplatforms exhibited a well-defined size of 208 nm with a narrow size distribution (PDI ≈ 0.5). Zeta potential measurements showed a value of -11.4 mV, indicating stability and suitability for drug delivery. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) confirmed the nanocarrier's spherical morphology and efficient distribution. Drug release profiles demonstrated that the NPs released more drugs at neutral to alkaline pH levels, attributed to pectin's ionization properties. The efficacy of the prepared βlg-5-FU-NaB-FA nanoplatforms was investigated on HCT116 and Caco2 CRC cells, along with the normal cell line CRL-1831. The βlg-5-FU-NaB-FA nanoplatforms exhibited remarkable cytotoxicity against both HCT116 and Caco2 CRC cells compared to free drugs, highlighting the efficacy of targeted delivery in folate receptor-positive cells. These NPs induce cell apoptosis and cell cycle arrest more effectively than free drugs, demonstrating their potential for targeted cancer therapy. Furthermore, a decrease in the expression of crucial genes involved in the Wnt signaling pathway was observed, which offers a valuable understanding of their underlying mechanism. Collectively, our results suggest that the FA-targeted βlg nanocarriers represent a promising platform for the efficient and targeted delivery of 5-FU and NaB in folate receptor-positive CRC. This novel nanocarrier holds the potential to enhance therapeutic outcomes while minimizing side effects, providing a new avenue for the treatment of CRC.

The Role of Indoxyl Sulfate in Exacerbating Colorectal Cancer During Chronic Kidney Disease Progression: Insights into the Akt/β-Catenin/c-Myc and AhR/c-Myc Pathways in HCT-116 Colorectal Cancer Cells.

Epidemiological studies suggest an increased risk of colorectal cancer (CRC) aggravation in patients with chronic kidney disease (CKD). Our previous study demonstrated that indoxyl sulfate, a uremic toxin whose concentration increases with CKD progression, exacerbates CRC through activation of the AhR and Akt pathways. Consequently, indoxyl sulfate has been proposed to be a significant link between CKD progression and CRC aggravation. The present study aimed to investigate the roles of c-Myc and β-Catenin, which are hypothesized to be downstream factors of indoxyl sulfate-induced AhR and Akt activation, in CRC cell proliferation and EGF sensitivity in HCT-116 CRC cells. Indoxyl sulfate significantly induced CRC cell proliferation at concentrations exceeding 62.5 µM, a process suppressed by the c-Myc inhibitor 10058-F4. Indoxyl sulfate activated the Akt/β-Catenin/c-Myc pathway as evidenced by the Akt inhibitor MK2206, which decreased both β-Catenin and c-Myc protein levels, and the β-Catenin inhibitor XAV-939, which reduced c-Myc protein levels. The AhR antagonist CH223191 also inhibited c-Myc upregulation, indicating involvement of the AhR/c-Myc pathway. MK2206 partially attenuated the indoxyl sulfate-induced AhR transcriptional activity, suggesting that Akt activation influences the AhR/c-Myc pathway. MK2206, CH223191, and 10058-F4 suppressed the increase in EGFR protein levels induced by indoxyl sulfate, indicating that the Akt/β-Catenin/c-Myc and AhR/c-Myc pathways enhance the sensitivity of HCT-116 CRC cells to EGF. These findings indicate that the elevation of indoxyl sulfate levels in the blood, due to CKD progression, could worsen CRC by promoting the proliferation of CRC cells and enhancing EGF signaling. Therefore, indoxyl sulfate could potentially serve as a therapeutic target for CRC aggravation in patients with CKD.

Overexpression of lncRNA LINC00294 Induces Cell Cycle Arrest and Apoptosis in Colorectal Cancer by Regulating the miR‐499a‐5p/LARP4B Axis

ABSTRACTIncreasing long noncoding RNAs (lncRNAs) have been found to participate in regulating the progression of colorectal cancer (CRC), which is a common gastrointestinal malignancy. Here, the specific role and mechanisms of lncRNA LINC00294 were investigated in CRC. The expression levels of LINC00294, miR‐499a‐5p, and La‐related protein 4B (LARP4B) in CRC cells (HCT116 and SW620) and tissues were assessed by RT‐qPCR. The viability, proliferation, cell cycle, and apoptosis of HCT116 and SW620 cells were determined by CCK‐8, EdU, and flow cytometry assays. Protein levels of cell cycle and cell apoptosis markers were measured by western blot analysis. FISH assay was performed to evaluate the subcellular localization of LINC00294 in CRC cells. Luciferase reporter assay, RNA pull‐down assay, as well as RIP assay verified the interactions between miR‐499a‐5p and LINC00294 (or LARP4B). The xenograft model was established in mice to investigate the function of LINC00294 in vivo. LINC00294 presented a decreased expression level in CRC cells and tissues. LINC00294 overexpression suppressed the cell proliferative capacity, promoted cell cycle arrest, and induced cell apoptosis in CRC HCT116 and SW620 cells. LINC00294 interacted with miR‐499a‐5p, and miR‐499a‐5p targeted LARP4B. MiR‐499a‐5p was upregulated while LARP4B was downregulated in CRC cells. In rescue assays, LARP4B knockdown reversed the inhibition of LINC00294 overexpression on malignant phenotypes of HCT116 and SW620 cells. In the xenograft model, LINC00294 overexpression inhibited tumor growth in vivo. LINC00294 exerted an antitumor function in CRC by forming a LINC00294/miR‐499a‐5p/LARP4B regulatory network.

Creation of a Model Line of Tumor Cells with Inducable Expression of Adenoviral E1A to Study Its Antiproliferative and Cytotoxic Properties In Vitro and In Vivo

Over the past decades, gene therapy based on the adenoviral E1A has proven its benefit against a number of tumor diseases, both in animal models and in clinical studies. It has been shown that in addition to its own antiproliferative activity, E1A also has the ability to enhance the cytotoxic effect of some anticancer drugs. The use of E1A in combination therapy can solve a number of problems in clinical oncology, among which the most pressing is the problem of drug-resistance of tumor cells. This work describes the establishment of a cell model based on human colorectal cancer cells HCT116 and cisplatin-resistant HCT116/C cells with doxycycline-inducible expression of adenoviral E1A. We have shown the concentration-dependent and time-dependent dynamics of E1A expression upon doxycycline treatment, and shown the antiproliferative effect of adenoviral E1A in the HCT116-E1A and HCT116/C-E1A cells in vitro in experiments assessing viability in MTT and clonogenic activity tests and in vivo in xenograft mouse models. Thus, as a result of our work, a model was created to explore the antiproliferative and sensitizing properties of E1A in platinum-sensitive and platinum-resistant colorectal cancer cells and to search for new approaches to anticancer therapy both in vitro and in vivo. The resulting cell line is a convenient model for selecting the most promising combinations of cytostatic drugs with E1A-based gene therapy.

Design, Synthesis, and Biological Evaluation of Novel Urea-Containing Carnosic Acid Derivatives with Anticancer Activity.

A series of novel carnosic acid 1 derivatives incorporating urea moieties at the C-20 position was synthesized and evaluated for their antiproliferative activity against the HCT116 colorectal cancer cell line. Most derivatives demonstrated enhanced antiproliferative activity compared to that of carnosic acid 1. The most promising derivatives were tested in other colorectal cancer cell lines (SW480, SW620, and Caco-2), melanoma (A375), and pancreatic cancer (MiaPaca-2). Derivative 14 consistently demonstrated the highest activity across all tested cancer cell lines, showing selectivity for cancer cells over normal cells. Further investigation of the mechanism of action in SW480 cells revealed that compound 14 induced cell cycle arrest at the G0/G1 phase by downregulating CDK4 and CDK6. Molecular docking studies revealed that compound 14 established several interactions with key residues in the active site of CDK6. Additionally, compound 14 also reduced ROS production. In summary, our results strongly indicate that compound 14 has potential as a lead compound in the development of innovative anticancer drugs.

Green hydrothermal synthesis of gallic acid carbon dots: characterization and cytotoxic effects on colorectal cancer cell line

Colorectal cancer (CRC) remains a leading cause of cancer-related deaths worldwide, necessitating the development of novel therapeutic approaches. Carbon dots (CDs) have emerged as promising nanoparticles for biomedical applications due to their unique properties. Gallic acid (GA), an anticancer agent, is effective against various tumor types. This study explores the potential of gallic acid-derived carbon dots (GA-CDs) as an innovative anticancer agent against HCT-116 CRC cells, focusing on apoptosis signaling pathways. GA-CDs were synthesized using a one-pot hydrothermal method. Characterization was conducted using transmission electron microscopy (TEM), Fourier transform infrared (FT-IR) spectroscopy, and ultraviolet-visible (UV-vis) absorption spectroscopy. The cytotoxicity of GA and GA-CDs on HCT-116 cells was evaluated using the MTT assay at various concentrations over 24 and 48 h. Cellular uptake was assessed via fluorescence microscopy, and apoptosis was analyzed using acridine orange/propidium iodide (AO/PI) staining. Total RNA extraction followed by complementary DNA (cDNA) synthesis via reverse transcription-PCR was performed, and real time-PCR (Q-PCR) was conducted to examine the expression of apoptosis-related genes includingCaspase-3,Bax, andBcl-2. Characterization confirmed the successful synthesis of spherical GA-CDs. GA-CDs exhibited dose- and time-dependent cytotoxicity, with IC50 values of 88.55 μg ml-1for GA-CDs and 192.2 μg ml-1for GA after 24 h. Fluorescence microscopy confirmed the efficient uptake of GA-CDs by HCT-116 cells. AO/PI staining showed a significant increase in apoptotic cell numbers after treatment with GA-CDs. Q-PCR analysis revealed overexpression ofCaspase-3 andBaxgenes in GA-CD-treated cells, though no significant changes were observed in the expression ofBcl-2 or theBax/Bcl-2 ratio. GA-CDs demonstrated potent anticancer properties by inducing apoptosis and reducing cell viability in HCT-116 cells. These findings suggest the potential of GA-CDs as a novel therapeutic agent for CRC treatment, warranting further investigation into their mechanism of action andin vivoefficacy.

Combining in silico and in vitro approaches for understanding the mechanism of action of the galactomannan extracted from Cassia grandis seeds against colorectal cancer

This study aimed to investigate the antitumor activity of galactomannan extracted from Cassia grandis seeds (GCg) against colorectal cancer cells using both experimental and computational approaches. Galactomannan was extracted from C. grandis seeds and prepared into solutions with varying concentrations. The cytotoxicity of these solutions was tested on HT-29 and HCT-116 colorectal cancer cell lines using the MTT assay. Additionally, computational evaluations, including molecular docking and molecular dynamics simulations, were performed to explore the potential binding interactions of GCg with cyclin-dependent kinase 2 (CDK2). The experimental results demonstrated that GCg significantly inhibited the proliferation of HT-29 cells, especially at concentrations of 5 mg/mL. On the other hand, no concentration inhibited >30 % of HCT-116 cells. Computational analysis revealed that GCg could bind to the ATP-binding site of CDK2, promoting the inactive DFG-out conformation, similar to the known inhibitor K03861. This interaction suggests a mechanism through which GCg may exert its anticancer effects. GCg exhibits significant cytotoxic activity against HT-29 colorectal adenocarcinoma cells, likely through the inhibition of CDK2; however, its efficacy against HCT-116 cells is limited, possibly due to structural differences in the molecular targets. To the best of the authors' knowledge, no studies have explored the applications of GCg in cancers, particularly colorectal ones. Further studies are needed to explore the antimetastatic effects and potential clinical applications of GCg in colorectal cancer treatment.

ONC206, an imipridone derivative, demonstrates anti-colorectal cancer activity against stem/progenitor cells in 3D cell cultures and in patient-derived organoids.

Colorectal cancer (CRC) remains one of the most frequently diagnosed and life-threatening malignancies worldwide. CRC's high recurrence rates and drug resistance have been correlated with a subpopulation of dormant slowly dividing cells termed CRC stem cells (CCSCs). Consequently, there is a pressing need to identify novel therapeutics that can effectively and specifically target CCSCs. Imipridones are promising structurally related anticancer molecules that showed efficacy in several solid and hematological preclinical models and phase I/II/III clinical trials. This study mainly aimed to assess the potential anticancer effects of ONC206, an imipridone derivative, on CRC three-dimensional in vitro culture systems using HCT116 and HT29 cells. Importantly, the study aimed at using CRC patient-derived organoids (PDOs) to test the potential therapeutic effect of ONC206. Two-dimensional cell proliferation, viability, migration, and invasion assays were used to assess the effects of ONC206 on two colorectal cancer cell lines, HCT116 and HT29, in vitro. Immunofluorescence imaging, flow cytometry, and western blot analysis were also performed to investigate the mechanism of action of this drug. Sphere formation assay and CRC PDOs were employed to evaluate the effect of ONC206 on CRC cells in a 3D setting and specifically its potency in targeting the CRC stem/progenitor subpopulation of cells. Our results showed that ONC206 was more potent than its parental moleculeONC201 in inhibiting the proliferation and viability of HCT116 and HT29 cells. Moreover, ONC206 significantly reduced the migration and invasion indices of CRC cells. These effects were accompanied by an increase in reactive oxygen species (ROS) production, sub-G1 phase accumulation, and apoptosis in HCT116 and HT29 cells. Furthermore, ONC206 significantly inhibited the 3D colonospheres growth and self-renewal ability of CCSCs more potently than ONC201, which was associated with a decrease in the expression of CSC-related markers. Lastly, ONC206 significantly reduced the growth of organoids derived from CRC patients. Collectively, our findings demonstrate that ONC206 is an effective anticancer molecule capable of targeting CCSCs, which may represent a novel therapeutic strategy that can overcome CRC resistance and recurrence.

4-(Pyrazolyl)benzenesulfonamide Ureas as Carbonic Anhydrases Inhibitors and Hypoxia-Mediated Chemo-Sensitizing Agents in Colorectal Cancer Cells.

Hypoxia in tumors contributes to chemotherapy resistance, worsened by acidosis driven by carbonic anhydrases (hCA IX and XII). Targeting these enzymes can mitigate acidosis, thus enhancing tumor sensitivity to cytotoxic drugs. Herein, novel 4-(pyrazolyl)benzenesulfonamide ureas (SH7a-t) were developed and evaluated for their inhibitory activity against hCA IX and XII. They showed promising results (hCA IX: KI = 15.9-67.6 nM, hCA XII: KI = 16.7-65.7 nM). Particularly, SH7s demonstrated outstanding activity (KIs = 15.9 nM for hCA IX and 55.2 nM for hCA XII) and minimal off-target kinase inhibition over a panel of 258 kinases. In NCI anticancer screening, SH7s exhibited broad-spectrum activity with an effective growth inhibition full panel GI50 (MG-MID) value of 3.5 μM and a subpanel GI50 (MG-MID) range of 2.4-6.3 μM. Furthermore, SH7s enhanced the efficacy of Taxol and 5-fluorouracil in cotreatment regimens under hypoxic conditions in HCT-116 colorectal cancer cells, indicating its potential as a promising anticancer agent.

Bismuth oxide nanoparticles inhibit HCT116 colorectal cancer cells by inducing apoptosis, cell cycle arrest and ROS production

Nanoparticles (NPs) have recently gained traction for anticancer use. Colorectal cancer (CRC) is one of the most common cancer types in the world, but only a few studies have previously reported the anticancer action of Bi2O3 NPs towards CRC, and the underlying mechanisms are not well understood. In this study, 1 mg/ml Bi2O3 NPs showed 60 % and 8 % inhibition in HCT116 and HT-29 cells, respectively. In the HCT116 spheroid, Bi2O3 NPs showed reduced inhibition of 27 % at 1 mg/ml due to the more stringent tissue architecture. Both CRC cells showed successful internalization of Bi2O3 NPs through flow cytometry and ICP-MS. The NPs mainly killed HCT116 cells by inducing late-stage apoptosis (∼17 %). In addition, Bi2O3 NPs also induced S and G2/M cell cycle arrest (∼4 % and ∼10 %) by targeting CDK2 protein in HCT116 cells. In HT-29 cells, Bi2O3 NPs did not trigger apoptosis but induced ∼22 % G2/M cell cycle arrest. Bi2O3 NPs also induced ROS production, and potently inhibited the cell migration. 300 μg/ml Bi2O3 NPs also exhibited anti-angiogenic action in the angiogenesis assay. In conclusion, the present study highlights the potential anticancer effect of Bi2O3 NPs towards CRC, and further investigation is warranted to improve the biocompatibility, targeted selectivity, and tumor-penetrating capacity.

TMEM206 Contributes to Cancer Hallmark Functions in Colorectal Cancer Cells and Is Regulated by p53 in a p21-Dependent Manner.

Acid-induced ion flux plays a role in pathologies where tissue acidification is prevalent, including cancer. In 2019, TMEM206 was identified as the molecular component of acid-induced chloride flux. Localizing to the plasma membrane, TMEM206 contributes to cellular processes like acid-induced cell death. Since over 50% of human cancers carry loss of function mutations in the p53 gene, we aimed to analyze how TMEM206 is regulated by p53 and its role in cancer hallmark function and acid-induced cell death in HCT116 colorectal cancer (CRC) cells. We generated p53-deficient HCT116 cells and assessed TMEM206-mediated Cl- currents and transcriptional regulation using the patch-clamp and a dual-luciferase reporter assay, respectively. To investigate the contribution of TMEM206 to cancer hallmark functions, we performed migration and metabolic activity assays. The role of TMEM206 in p53-mediated acid-induced cell death was assessed with cell death assays. The TMEM206 mRNA level was significantly elevated in human primary CRC tumors. TMEM206 knockout increased acid-induced cell death and reduced proliferation and migration, indicating a role for TMEM206 in these cancer hallmark functions. Furthermore, we observed increased TMEM206 mRNA levels and currents in HCT116 p53 knockout cells. This phenotype can be rescued by transient overexpression of p53 but not by overexpression of dysfunctional p53. In addition, our data suggest that TMEM206 may mediate cancer hallmark functions within p53-associated pathways. TMEM206 promoter activity is not altered by p53 overexpression. Conversely, knockout of p21, a major target gene of p53, increased TMEM206-mediated currents, suggesting expression control of TMEM206 by p21 downstream signaling. Our results show that in colorectal cancer cells, TMEM206 expression is elevated, contributes to cancer hallmark functions, and its regulation is dependent on p53 through a p21-dependent mechanism.

Synthesis, In Silico Prediction, and In Vitro Evaluation of Anti-tumor Activities of Novel 4'-Hydroxybiphenyl-4-carboxylic Acid Derivatives as EGFR Allosteric Site Inhibitors.

Allosteric inhibition of EGFR tyrosine kinase (TK) is currently among the most attractive approaches for designing and developing anti-cancer drugs to avoid chemoresistance exhibited by clinically approved ATP-competitive inhibitors. The current work aimed to synthesize new biphenyl-containing derivatives that were predicted to act as EGFR TK allosteric site inhibitors based on molecular docking studies. A new series of 4'-hydroxybiphenyl-4-carboxylic acid derivatives, including hydrazine-1-carbothioamide (S3-S6) and 1,2,4-triazole (S7-S10) derivatives, were synthesized and characterized using IR, 1HNMR, 13CNMR, and HR-mass spectroscopy. Compound S4 had a relatively high pharmacophore-fit score, indicating that it may have biological activity similar to the EGFR allosteric inhibitor reference, and it scored a relatively low ΔG against EGFR TK allosteric site, indicating a high likelihood of drug-receptor complex formation. Compound S4 was cytotoxic to the three cancer cell lines tested, particularly HCT-116 colorectal cancer cells, with an IC50 value comparable to Erlotinib. Compound S4 induced the intrinsic apoptotic pathway in HCT-116 cells by arresting them in the G2/M phase. All of the new derivatives, including S4, met the in silico requirements for EGFR allosteric inhibitory activity. Compound S4 is a promising EGFR tyrosine kinase allosteric inhibitor that warrants further research.