The dysadherin/FAK axis promotes individual cell migration in colon cancer.

Dysregulation of cancer cell motility is a key driver of invasion and metastasis. High dysadherin expression in cancer cells is correlated with invasion and metastasis. Here, we found the molecular mechanism by which dysadherin regulates the migration and invasion of colon cancer (CC). Comprehensive analysis using single-cell RNA sequencing data from CC patients revealed that high dysadherin expression in cells is linked to cell migration-related gene signatures. We confirmed that the deletion of dysadherin in tumor cells hindered local invasion and distant migration using in vivo tumor models. In this context, by performing cell morphological analysis, we found that aberrant cell migration resulted from impaired actin dynamics, focal adhesion turnover and protrusive structure formation upon dysadherin expression. Mechanistically, the activation of focal adhesion kinase (FAK) was observed in dysadherin-enriched cells. The dysadherin/FAK axis enhanced cell migration and invasion by activating the FAK downstream cascade, which includes the Rho family of small GTPases. Overall, this study illuminates the role of dysadherin in modulating cancer cell migration by forcing actin dynamics and protrusive structure formation via FAK signaling, indicating that targeting dysadherin may be a potential therapeutic strategy for CC patients.

EMMPRIN promotes spheroid organization and metastatic formation: comparison between monolayers and spheroids of CT26 colon carcinoma cells.

Background: In vitro studies often use two-dimensional (2D) monolayers, but 3D cell organization, such as in spheroids, better mimics the complexity of solid tumors. To metastasize, cancer cells undergo the process of epithelial-to-mesenchymal transition (EMT) to become more invasive and pro-angiogenic, with expression of both epithelial and mesenchymal markers. Aims: We asked whether EMMPRIN/CD147 contributes to the formation of the 3D spheroid structure, and whether spheroids, which are often used to study proliferation and drug resistance, could better model the EMT process and the metastatic properties of cells, and improve our understanding of the role of EMMPRIN in them. Methods: We used the parental mouse CT26 colon carcinoma (CT26-WT) cells, and infected them with a lentivirus vector to knock down EMMPRIN expression (CT26-KD cells), or with an empty lentivirus vector (CT26-NC) that served as a negative control. In some cases, we repeated the experiments with the 4T1 or LLC cell lines. We compared the magnitude of change between CT26-KD and CT26-WT/NC cells in different metastatic properties in cells seeded as monolayers or as spheroids formed by the scaffold-free liquid overlay method. Results: We show that reduced EMMPRIN expression changed the morphology of cells and their spatial organization in both 2D and 3D models. The 3D models more clearly demonstrated how reduced EMMPRIN expression inhibited proliferation and the angiogenic potential, while it enhanced drug resistance, invasiveness, and EMT status, and moreover it enhanced cell dormancy and prevented CT26-KD cells from forming metastatic-like lesions when seeded on basement membrane extract (BME). Most interestingly, this approach enabled us to identify that EMMPRIN and miR-146a-5p form a negative feedback loop, thus identifying a key mechanism for EMMPRIN activities. These results underline EMMPRIN role as a gatekeeper that prevents dormancy, and suggest that EMMPRIN links EMT characteristics to the process of spheroid formation. Conclusions: Thus, 3D models can help identify mechanisms by which EMMPRIN facilitates tumor and metastasis progression, which might render EMMPRIN as a promising target for anti-metastatic tumor therapy.

Identification of extracellular matrix-related biomarkers in colon adenocarcinoma by bioinformatics and experimental validation.

Backgrounds: Extracellular matrix (ECM) is an important component of tumor microenvironment, and its abnormal expression promotes tumor formation, progression and metastasis. Methods: Weighted gene co-expression network analysis (WGCNA) was used to identify ECM-related hub genes based on The Cancer Genome Atlas (TCGA) colon adenocarcinoma (COAD) data. COAD clinical samples were used to verify the expression of potential biomarkers in tumor tissues, and siRNA was used to explore the role of potential biomarkers in cell proliferation and epithelial-mesenchymal transition (EMT). Results: Three potential biomarkers (LEP, NGF and PCOLCE2) related to prognosis of COAD patients were identified and used to construct ERGPI. Immunohistochemical analysis of clinical samples showed that the three potential biomarkers were highly expressed in tumor tissues of COAD patients. Knockdown of LEP, NGF or PCOLCE2 inhibited COAD cell proliferation and EMT. Dictamnine inhibited tumor cell growth by binding to these three potential biomarkers based on molecular docking and transplanted tumor model. Conclusion: The three biomarkers can provide new ideas for the diagnosis and targeted therapy of COAD patients.

Selective but not pan-CDK inhibition abrogates 5-FU-driven tissue factor upregulation in colon cancer.

Thromboembolic events are complications in cancer patients and hypercoagulability has been linked to the tissue factor (TF) pathway, making this an attractive target. Here, we investigated the effects of chemotherapeutics and CDK inhibitors (CDKI) abemaciclib/palbociclib (CDK4/6), THZ-1 (CDK7/12/13), and dinaciclib (CDK1/2/5/9) alone and in combination regimens on TF abundance and coagulation. The human colorectal cancer (CRC) cell line HROC173 was treated with 5-FU or gemcitabine to stimulate TF expression. TF+ cells were sorted, recultured, and re-analyzed. The effect of treatment alone or in combination was assessed by functional assays. Low-dose chemotherapy induced a hypercoagulable state and significantly upregulated TF, even after reculture without treatment. Cells exhibited characteristics of epithelial-mesenchymal transition, including high expression of vimentin and mucin. Dinaciclib and THZ-1 also upregulated TF, while abemaciclib and palbociclib downregulated it. Similar results were observed in coagulation assays. The same anticoagulant activity of abemaciclib was seen after incubation with peripheral immune cells from healthy donors and CRC patients. Abemaciclib reversed 5-FU-induced TF upregulation and prolonged clotting times in second-line treatment. Effects were independent of cytotoxicity, senescence, and p27kip1 induction. TF-antibody blocking experiments confirmed the importance of TF in plasma coagulation, with Factor XII playing a minor role. Short-term abemaciclib counteracts 5-FU-induced hypercoagulation and eventually even prevents thromboembolic events.

Berberis vulgaris L. Root Extract as a Multi-Target Chemopreventive Agent against Colon Cancer Causing Apoptosis in Human Colon Adenocarcinoma Cell Lines.

Berberis vulgaris L. (Berberidaceae) is a shrub that has been widely used in European folk medicine as an anti-inflammatory and antimicrobial agent. The purpose of our study was to elucidate the mechanisms of the chemopreventive action of the plant's methanolic root extract (BVR) against colon cancer cells. Studies were conducted in human colon adenocarcinoma cell lines (LS180 and HT-29) and control colon epithelial CCD841 CoN cells. According to the MTT assay, after 48 h of cell exposure, the IC50 values were as follows: 4.3, 46.1, and 50.2 µg/mL for the LS180, HT-29, and CCD841 CoN cells, respectively, showing the greater sensitivity of the cancer cells to BVR. The Cell Death Detection ELISAPLUS kit demonstrated that BVR induced programmed cell death only against HT-29 cells. Nuclear double staining revealed the great proapoptotic BVR properties in HT-29 cells and subtle effect in LS180 cells. RT-qPCR with the relative quantification method showed significant changes in the expression of genes related to apoptosis in both the LS180 and HT-29 cells. The genes BCL2L1 (126.86-421.43%), BCL2L2 (240-286.02%), CASP3 (177.19-247.83%), and CASP9 (157.99-243.75%) had a significantly elevated expression, while BCL2 (25-52.03%) had a reduced expression compared to the untreated control. Furthermore, in a panel of antioxidant tests, BVR showed positive effects (63.93 ± 0.01, 122.92 ± 0.01, and 220.29 ± 0.02 mg Trolox equivalents (TE)/g in the DPPH•, ABTS•+, and ORAC assays, respectively). In the lipoxygenase (LOX) inhibition test, BVR revealed 62.60 ± 0.87% of enzyme inhibition. The chemical composition of BVR was determined using a UHPLC-UV-CAD-MS/MS analysis and confirmed the presence of several known alkaloids, including berberine, as well as other alkaloids and two derivatives of hydroxycinnamic acid (ferulic and sinapic acid hexosides). The results are very promising and encourage the use of BVR as a comprehensive chemopreventive agent (anti-inflammatory, antioxidant, and pro-apoptotic) in colorectal cancer, and were widely discussed alongside data from the literature.

Network-based elucidation of colon cancer drug resistance mechanisms by phosphoproteomic time-series analysis.

Aberrant signaling pathway activity is a hallmark of tumorigenesis and progression, which has guided targeted inhibitor design for over 30 years. Yet, adaptive resistance mechanisms, induced by rapid, context-specific signaling network rewiring, continue to challenge therapeutic efficacy. Leveraging progress in proteomic technologies and network-based methodologies, we introduce Virtual Enrichment-based Signaling Protein-activity Analysis (VESPA)-an algorithm designed to elucidate mechanisms of cell response and adaptation to drug perturbations-and use it to analyze 7-point phosphoproteomic time series from colorectal cancer cells treated with clinically-relevant inhibitors and control media. Interrogating tumor-specific enzyme/substrate interactions accurately infers kinase and phosphatase activity, based on their substrate phosphorylation state, effectively accounting for signal crosstalk and sparse phosphoproteome coverage. The analysis elucidates time-dependent signaling pathway response to each drug perturbation and, more importantly, cell adaptive response and rewiring, experimentally confirmed by CRISPR knock-out assays, suggesting broad applicability to cancer and other diseases.

Impact of the thyroid hormone T3 and its nuclear receptor TRα1 on colon cancer stem cell phenotypes and response to chemotherapies.

Colorectal cancers (CRCs) are highly heterogeneous and show a hierarchical organization, with cancer stem cells (CSCs) responsible for tumor development, maintenance, and drug resistance. Our previous studies showed the importance of thyroid hormone-dependent signaling on intestinal tumor development and progression through action on stem cells. These results have a translational value, given that the thyroid hormone nuclear receptor TRα1 is upregulated in human CRCs, including in the molecular subtypes associated with CSC features. We used an established spheroid model generated from the human colon adenocarcinoma cell line Caco2 to study the effects of T3 and TRα1 on spheroid formation, growth, and response to conventional chemotherapies. Our results show that T3 treatment and/or increased TRα1 expression in spheroids impaired the response to FOLFIRI and conferred a survival advantage. This was achieved by stimulating drug detoxification pathways and increasing ALDH1A1-expressing cells, including CSCs, within spheroids. These results suggest that clinical evaluation of the thyroid axis and assessing TRα1 levels in CRCs could help to select optimal therapeutic regimens for patients with CRC. Proposed mechanism of action of T3/TRα1 in colon cancer spheroids. In the control condition, TRα1 participates in maintaining homeostatic cell conditions. The presence of T3 in the culture medium activates TRα1 action on target genes, including the drug efflux pumps ABCG2 and ABCB1. In the case of chemotherapy FOLFIRI, the increased expression of ABC transcripts and proteins induced by T3 treatment is responsible for the augmented efflux of 5-FU and Irinotecan from the cancer cells. Taken together, these mechanisms contribute to the decreased efficacy of the chemotherapy and allow cells to escape the treatment. Created with BioRender.com .

Effects of down-regulated carbonic anhydrase 8 on cell survival and glucose metabolism in human colorectal cancer cell lines.

Carbonic anhydrase 8 (CA8) is a member of the α-carbonic anhydrase family but does not catalyze the reversible hydration of carbon dioxide. In the present study, we examined the effects of CA8 on two human colon cancer cell lines, SW480 and SW620, by suppressing CA8 expression through shRNA knockdown. Our results showed that knockdown of CA8 decreased cell growth and cell mobility in SW620 cells, but not in SW480 cells. In addition, downregulated CA8 resulted in a significant decrease of glucose uptake in both SW480 and SW620 cells. Interestingly, stable downregulation of CA8 decreased phosphofructokinase-1 expression but increased glucose transporter 3 (GLUT3) levels in SW620 cells. However, transient downregulation of CA8 fails to up-regulate GLUT3 expression, indicating that the increased GLUT3 observed in SW620-shCA8 cells is a compensatory effect. In addition, the interaction between CA8 and GLUT3 was evidenced by pull-down and IP assays. On the other hand, we showed that metformin, a first-line drug for type II diabetes patients, significantly inhibited cell migration of SW620 cells, depending on the expressions of CA8 and focal adhesion kinase. Taken together, our data demonstrate that when compared to primary colon cancer SW480 cells, metastatic colon cancer SW620 cells respond differently to downregulated CA8, indicating that CA8 in more aggressive cancer cells may play a more important role in controlling cell survival and metformin response. CA8 may affect glucose metabolism- and cell invasion-related molecules in colon cancer, suggesting that CA8 may be a potential target in future cancer therapy.

LncRNA ANRIL Promotes Glucose Metabolism and Proliferation of Colon Cancer in a High-Glucose Environment and is Associated with Worse Outcome in Diabetic Colon Cancer Patients.

BACKGROUND: The potential involvement of type 2 diabetes mellitus (T2DM) as a risk factor for colon cancer (CC) has been previously reported. Epigenetic changes, such as deregulation of long non-coding RNA (lncRNA) and microRNA (miR), have been linked to the advancement of CC; however, the effects of high glucose levels on their deregulation and, in turn, colon cancer remain unexplored. METHODS: Fifty patients had a dual diagnosis of CC and T2DM, and 60 patients with CC without diabetes mellitus were included in the study. qRT-PCR was used to examine the expression of lncRNA ANRIL and miR-186-5p in tissue samples. ANRIL, miR-186-5p, and their downstream target genes HIF-1α, PFK, HK, Bcl-2, and Bax were also determined in CC cell lines under various glucose conditions. Glucose uptake, lactate production and cells proliferation were estimated in CC cell lines. RESULTS: A significant upregulation of ANRIL expression levels (p<0.001) and a significant downregulation of miR-186-5p expression (p<0.001) in diabetic colon cancer specimens compared to those in non-diabetic colon cancer group were observed. MiR-186-5p expression levels were inversely correlated with ANRIL expression levels, blood glucose levels and HbA1c%. Concerning in vitro model, a significant upregulation of ANRIL, downregulation of miR-186-5p, upregulation of HIF-1α, glycolytic enzymes and activation of antiapoptotic pathway was detected in higher glucose concentrations than lower one. There was a significant increase of glucose uptake, lactate accumulation and proliferation of the Caco2 and SW620 cell lines in a dose dependent manner of glucose concentrations. Moreover, a significant positive correlation between glucose uptake and ANRIL expression was shown. CONCLUSIONS: A high-glucose environment can increase the tumor-promoting effect of ANRIL. ANRIL can promote glucose metabolism and colon cancer proliferation by downregulating miR-186-5p with subsequent upregulation of glycolysis enzymes expression and inhibition of apoptosis.

Upregulation of EMR1 (ADGRE1) by Tumor-Associated Macrophages Promotes Colon Cancer Progression by Activating the JAK2/STAT1,3 Signaling Pathway in Tumor Cells.

Previously, we reported that epidermal growth factor-like module-containing mucin-like hormone receptor-like 1 (EMR1/ADGRE1) is abnormally expressed in colon cancer (CC) and is a risk factor for lymph node metastasis (LNM) and poor recurrence-free survival in patients with abundant tumor-associated macrophages (TAMs). However, the signaling pathways associated with EMR1 expression in CC progression remain unclear. In this study, we aimed to explore the role of EMR1 and its signaling interactions with macrophages in CC progression. Spatial transcriptomics of pT3 microsatellite unstable CC tissues revealed heightened Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling in EMR1-HL CC with LNM compared to EMR1-N CC without LNM. Through in vitro coculture of CC cells with macrophages, EMR1 expression by CC cells was found to be induced by TAMs, ultimately interacting with upregulated JAK/STAT signaling, increasing cell proliferation, migration, and motility, and reducing apoptosis. JAK2/STAT3 inhibition decreased the levels of EMR1, JAK2, STAT1, and STAT3, significantly impeded the proliferation, migration, and mobility of cells, and increased the apoptosis of EMR1+ CC cells compared to their EMR1KO counterparts. Overall, TAMs-induced EMR1 upregulation in CC cells may promote LNM and CC progression via JAK2/STAT1,3 signaling upregulation. This study provides further insights into the molecular mechanisms involving macrophages and intracellular EMR1 expression in CC progression, suggesting its clinical significance and offering potential interventions to enhance patient outcomes.

Raman Imaging-A Valuable Tool for Tracking Fatty Acid Metabolism-Normal and Cancer Human Colon Single-Cell Study.

Altered metabolism of lipids is a key factor in many diseases including cancer. Therefore, investigations into the impact of unsaturated and saturated fatty acids (FAs) on human body homeostasis are crucial for understanding the development of lifestyle diseases. In this paper, we focus on the impact of palmitic (PA), linoleic (LA), and eicosapentaenoic (EPA) acids on human colon normal (CCD-18 Co) and cancer (Caco-2) single cells using Raman imaging and spectroscopy. The label-free nature of Raman imaging allowed us to evaluate FAs dynamics without modifying endogenous cellular metabolism. Thanks to the ability of Raman imaging to visualize single-cell substructures, we have analyzed the changes in chemical composition of endoplasmic reticulum (ER), mitochondria, lipid droplets (LDs), and nucleus upon FA supplementation. Analysis of Raman band intensity ratios typical for lipids, proteins, and nucleic acids (I1656/I1444, I1444/I1256, I1444/I750, I1304/I1256) proved that, using Raman mapping, we can observe the metabolic pathways of FAs in ER, which is responsible for the uptake of exogenous FAs, de novo synthesis, elongation, and desaturation of FAs, in mitochondria responsible for energy production via FA oxidation, in LDs specialized in cellular fat storage, and in the nucleus, where FAs are transported via fatty-acid-binding proteins, biomarkers of human colon cancerogenesis. Analysis for membranes showed that the uptake of FAs effectively changed the chemical composition of this organelle, and the strongest effect was noticed for LA. The spectroscopy studies have been completed using XTT tests, which showed that the addition of LA or EPA for Caco-2 cells decreases their viability with a stronger effect observed for LA and the opposite effect observed for PA. For normal cells, CCD-18 Co supplementation using LA or EPA stimulated cells for growing, while PA had the opposite impact.

Feasibility study of multimodal imaging for redox status and glucose metabolism in tumor.

Understanding the tumor redox status is important for efficient cancer treatment. Here, we noninvasively detected changes in the redox environment of tumors before and after cancer treatment in the same individuals using a novel compact and portable electron paramagnetic resonance imaging (EPRI) device and compared the results with glycolytic information obtained through autoradiography using 2-deoxy-2-[18F]fluoro-d-glucose ([18F]FDG). Human colon cancer HCT116 xenografts were used in the mice. We used 3-carbamoyl-PROXYL (3CP) as a paramagnetic and redox status probe for the EPRI of tumors. The first EPRI was followed by the intraperitoneal administration of buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, or X-ray irradiation of the tumor. A second EPRI was performed on the following day. Autoradiography was performed after the second EPRI. After imaging, the tumor sections were evaluated by histological analysis and the amount of reducing substances in the tumor was measured. BSO treatment and X-ray irradiation significantly decreased the rate of 3CP reduction in tumors. Redox maps of tumors obtained from EPRI can be compared with tissue sections of approximately the same cross section. BSO treatment reduced glutathione levels in tumors, whereas X-ray irradiation did not alter the levels of any of the reducing substances. Comparison of the redox map with the autoradiography of [18F]FDG revealed that regions with high reducing power in the tumor were active in glucose metabolism; however, this correlation disappeared after X-ray irradiation. These results suggest that the novel compact and portable EPRI device is suitable for multimodal imaging, which can be used to study tumor redox status and therapeutic efficacy in cancer, and for combined analysis with other imaging modalities.

Evaluation of the anticancer effects exerted by 5-fluorouracil and heme oxygenase-1 inhibitor hybrids in HTC116 colorectal cancer cells.

Colon cancer remains a clinical challenge in industrialised countries. Its treatment with 5-Flurouracil (5-FU) develops many side effects and resistance. Thus, several strategies have been undertaken so far, including the use of drug cocktails and polypharmacology. Heme oxygenase-1 (HO-1) is an emerging molecular target in the treatment of various cancers. We recently demonstrated that a combination of HO-1 inhibitors with 5-FU and the corresponding hybrids SI1/17, SI1/20, and SI1/22, possessed anticancer activity against prostate and lung cancer cells. In this work, we evaluated these hybrids in a model of colon cancer and found that SI1/22 and the respective combo have greater potency than 5-FU. Particularly, compounds inhibit HO-1 activity in cell lysates, increase ROS and the expression of HO-1, SOD, and Nrf2. Moreover, we observed a decrease of pro-caspase and an increase in cleaved PARP-1 and p62, suggesting apoptotic and autophagic cell death and potential application of these drugs as anticancer agents.

Coculture with Colon-26 cancer cells decreases the protein synthesis rate and shifts energy metabolism toward glycolysis dominance in C2C12 myotubes.

Cancer cachexia is the result of complex interorgan interactions initiated by cancer cells and changes in patient behavior such as decreased physical activity and energy intake. Therefore, it is crucial to distinguish between the direct and indirect effects of cancer cells on muscle mass regulation and bioenergetics to identify novel therapeutic targets. In this study, we investigated the direct effects of Colon-26 cancer cells on the molecular regulating machinery of muscle mass and its bioenergetics using a coculture system with C2C12 myotubes. Our results demonstrated that coculture with Colon-26 cells induced myotube atrophy and reduced skeletal muscle protein synthesis and its regulating mechanistic target of rapamycin complex 1 signal transduction. However, we did not observe any activating effects on protein degradation pathways including ubiquitin-proteasome and autophagy-lysosome systems. From a bioenergetic perspective, coculture with Colon-26 cells decreased the complex I-driven, but not complex II-driven, mitochondrial ATP production capacity, while increasing glycolytic enzyme activity and glycolytic metabolites, suggesting a shift in energy metabolism toward glycolysis dominance. Gene expression profiling by RNA sequencing showed that the increased activity of glycolytic enzymes was consistent with changes in gene expression. However, the decreased ATP production capacity of mitochondria was not in line with the gene expression. The potential direct interaction between cancer cells and skeletal muscle cells revealed in this study may contribute to a better fundamental understanding of the complex pathophysiology of cancer cachexia.NEW & NOTEWORTHY We explored the potential direct interplay between colon cancer cells (Colon-26) and skeletal muscle cells (C2C12 myotubes) employing a noncontact coculture experimental model. Our findings reveal that coculturing with Colon-26 cells substantially impairs the protein synthesis rate, concurrently instigating a metabolic shift toward glycolytic dominance in C2C12 myotubes. This research unveils critical insights into the intricate cellular cross talk underpinning the complex pathophysiology of cancer cachexia.

NXPH4 can be used as a biomarker for pan-cancer and promotes colon cancer progression.

NXPH4 promotes cancer proliferation and invasion. However, its specific role and mechanism in cancer remain unclear. Transcriptome and clinical data for pan-cancer were derived from the TCGA database. K-M survival curve and univariate Cox were used for prognostic analysis. CIBERSORT and TIMER algorithms were employed to calculate immune cell infiltration. Gene set enrichment analysis (GSEA) was employed for investigating the function of NXPH4. Western blot verified differential expression of NXPH4 in colon cancer. Functional assays (CCK-8, plate clonogenicity assay, wound healing assay, and Transwell assay) confirmed the impact of NXPH4 on proliferation, invasion, and migration of colon cancer cells. Dysregulation of NXPH4 in pan-cancer suggests its potential as a diagnostic and prognostic marker for certain cancers, including colon and liver cancer. High expression of NXPH4 in pan-cancer might be associated with the increase in copy number and hypomethylation. NXPH4 expression in pan-cancer is substantially linked to immune cell infiltration in the immune microenvironment. NXPH4 expression is associated with the susceptibility to immunotherapy and chemotherapy. Western blot further confirmed the higher expression of NXPH4 in colon cancer. Knockdown of NXPH4 significantly suppresses proliferation, invasion, and migration of colon cancer cell lines HT-29 and HCT116, as validated by functional assays.

Colon cancer inhibitory properties of Caulerpa lentillifera polysaccharide and its molecular mechanisms based on three-dimensional cell culture model.

Caulerpa lentillifera is rich in polysaccharides, and its polysaccharides show a significant effect in different biological activities including anti-cancer activity. As an edible algae-derived polysaccharide, exploring the role of colon cancer can better develop the application from a dietary therapy perspective. However, more in-depth studies of C. lentillifera polysaccharide on anti-colon cancer activity and mechanism are needed. In this study, we found that Caulerpa lentillifera polysaccharides (CLP) showed potential anti-colon cancer effect on human colon cancer cell HT29 in monolayer (IC50 = 1.954 mg/mL) and spheroid (IC50 = 0.402 mg/mL). Transcriptomics and metabolomics analyses revealed that CLP had an inhibitory effect on HT29 3D spheroid cells by activating aminoacyl-tRNA biosynthesis as well as arginine and proline metabolism pathways. Furthermore, the anti-colon cancer effects of CLP were confirmed through other human colon cancer cell HCT116 and LoVo in monolayer cells (IC50 = 1.890 mg/mL and 1.437 mg/mL, respectively) and 3D spheroid cells (IC50 = 0.344 mg/mL and 0.975 mg/mL, respectively), and three patient-derived organoids with IC50 values of 6.333-8.780 mg/mL. This study provided basic data for the potential application of CLP in adjuvant therapeutic food for colon cancer on multiple levels, while further investigation of detailed mechanism in vivo was still required.

H2S-driven chemotherapy and mild photothermal therapy induced mitochondrial reprogramming to promote cuproptosis.

The elevated level of hydrogen sulfide (H2S) in colon cancer hinders complete cure with a single therapy. However, excessive H2S also offers a treatment target. A multifunctional cascade bioreactor based on the H2S-responsive mesoporous Cu2Cl(OH)3-loaded hypoxic prodrug tirapazamine (TPZ), in which the outer layer was coated with hyaluronic acid (HA) to form TPZ@Cu2Cl(OH)3-HA (TCuH) nanoparticles (NPs), demonstrated a synergistic antitumor effect through combining the H2S-driven cuproptosis and mild photothermal therapy. The HA coating endowed the NPs with targeting delivery to enhance drug accumulation in the tumor tissue. The presence of both the high level of H2S and the near-infrared II (NIR II) irradiation achieved the in situ generation of photothermic agent copper sulfide (Cu9S8) from the TCuH, followed with the release of TPZ. The depletion of H2S stimulated consumption of oxygen, resulting in hypoxic state and mitochondrial reprogramming. The hypoxic state activated prodrug TPZ to activated TPZ (TPZ-ed) for chemotherapy in turn. Furthermore, the exacerbated hypoxia inhibited the synthesis of adenosine triphosphate, decreasing expression of heat shock proteins and subsequently improving the photothermal therapy. The enriched Cu2+ induced not only cuproptosis by promoting lipoacylated dihydrolipoamide S-acetyltransferase (DLAT) heteromerization but also performed chemodynamic therapy though catalyzing H2O2 to produce highly toxic hydroxyl radicals ·OH. Therefore, the nanoparticles TCuH offer a versatile platform to exert copper-related synergistic antitumor therapy.

Comparative evaluation of hesperetin-loaded graphene oxide nanosheets (Hsp-GO) as a drug delivery system for colon cancer: synthesis and anticancer efficiency assessment.

BACKGROUND: Graphene oxide nanosheets (GONS) are recognized for their role in enhancing drug delivery and effectiveness in cancer treatment. With colon cancer being a prevalent global issue and the significant side effects associated with chemotherapy, the primary treatment for colon cancer alongside surgery, there is a critical need for novel therapeutic strategies to support patients in combating this disease. Hesperetin (HSP), a natural compound found in specific fruits, exhibits anti-cancer properties. The aim of this study is to investigate the effect of GONS on the LS174t colon cancer cell line. METHODS: In this study, an anti-cancer nano-drug was synthesized by creating a hesperetin-graphene oxide nanocomposite (Hsp-GO), which was subsequently evaluated for its efficacy through in vitro cell toxicity assays. Three systems were investigated: HSP, GONS, and HSP-loaded GONS, to determine their cytotoxic and pro-apoptotic impacts on the LS174t colon cancer cell line, along with assessing the expression of BAX and BCL2. The morphology and properties of both GO and Hsp-GO were examined using scanning electron microscopy (SEM), X-ray diffraction, and Fourier transform infrared spectroscopy (FTIR). RESULTS: The Hsp-GO nanocomposite displayed potent cytotoxic and pro-apoptotic effects on LS174t colon cancer cells, outperforming individual treatments with HSP or GONS. Cell viability assays showed a significant decrease in cell viability with Hsp-GO treatment. Analysis of BAX and BCL2 expression revealed elevated BAX and reduced BCL2 levels in Hsp-GO treated cells, indicating enhanced apoptotic activity. Morphological analysis confirmed successful Hsp-GO synthesis, while structural integrity was supported by X-ray diffraction and FTIR analyses. CONCLUSIONS: These study highlight the potential of Hsp-GO as a promising anti-cancer nano-drug for colon cancer therapy.

Mitochondrial genome transfer drives metabolic reprogramming in adjacent colonic epithelial cells promoting TGFß1-mediated tumor progression.

Although nontumor components play an essential role in colon cancer (CC) progression, the intercellular communication between CC cells and adjacent colonic epithelial cells (CECs) remains poorly understood. Here, we show that intact mitochondrial genome (mitochondrial DNA, mtDNA) is enriched in serum extracellular vesicles (EVs) from CC patients and positively correlated with tumor stage. Intriguingly, circular mtDNA transferred via tumor cell-derived EVs (EV-mtDNA) enhances mitochondrial respiration and reactive oxygen species (ROS) production in CECs. Moreover, the EV-mtDNA increases TGFß1 expression in CECs, which in turn promotes tumor progression. Mechanistically, the intercellular mtDNA transfer activates the mitochondrial respiratory chain to induce the ROS-driven RelA nuclear translocation in CECs, thereby transcriptionally regulating TGFß1 expression and promoting tumor progression via the TGFß/Smad pathway. Hence, this study highlights EV-mtDNA as a major driver of paracrine metabolic crosstalk between CC cells and adjacent CECs, possibly identifying it as a potential biomarker and therapeutic target for CC.