Anti-tumourigenic Effect Research Articles

Lactococcus lactis HkyuLL 10 suppresses colorectal tumourigenesis and restores gut microbiota through its generated alpha-mannosidase

ObjectiveProbiotic Lactococcus lactis is known to confer health benefits to humans. Here, we aimed to investigate the role of L. lactis in colorectal cancer (CRC).DesignL. lactis abundance was evaluated in...

Lactobacillus acidophilus suppresses non-alcoholic fatty liver disease-associated hepatocellular carcinoma through producing valeric acid

Gut probiotic depletion is associated with non-alcoholic fatty liver disease-associated hepatocellular carcinoma (NAFLD-HCC). Here, we investigated the prophylactic potential of Lactobacillus acidophilus against NAFLD-HCC. NAFLD-HCC conventional and germ-free mice were established by diethylnitrosamine (DEN) injection with feeding of high-fat high-cholesterol (HFHC) or choline-deficient high-fat (CDHF) diet. Orthotopic NAFLD-HCC allografts were established by intrahepatic injection of murine HCC cells with HFHC feeding. Metabolomic profiling was performed using liquid chromatography-mass spectrometry. Biological functions of L.acidophilus conditional medium (L.a CM) and metabolites were determined in NAFLD-HCC human cells and mouse organoids. L.acidophilus supplementation suppressed NAFLD-HCC formation in HFHC-fed DEN-treated mice. This was confirmed in orthotopic allografts and germ-free tumourigenesis mice. L.a CM inhibited the growth of NAFLD-HCC human cells and mouse organoids. The protective function of L.acidophilus was attributed to its non-protein small molecules. By metabolomic profiling, valeric acid was the top enriched metabolite in L.a CM and its upregulation was verified in liver and portal vein of L.acidophilus-treated mice. The protective function of valeric acid was demonstrated in NAFLD-HCC human cells and mouse organoids. Valeric acid significantly suppressed NAFLD-HCC formation in HFHC-fed DEN-treated mice, accompanied by improved intestinal barrier integrity. This was confirmed in another NAFLD-HCC mouse model induced by CDHF diet and DEN. Mechanistically, valeric acid bound to hepatocytic surface receptor GPR41/43 to inhibit Rho-GTPase pathway, thereby ablating NAFLD-HCC. L.acidophilus exhibits anti-tumourigenic effect in mice by secreting valeric acid. Probiotic supplementation is a potential prophylactic of NAFLD-HCC. Shown in Acknowledgments.

Peritumoural Mast Cell Density in Invasive Breast Carcinoma: Possible Antitumourigenic Effect with Potential Role for Immunotherapy

Introduction: The interaction of cancer cells and the host within the tumour microenvironment plays a major role in determining the outcome of cancers. Mast cells are part of the innate immune system that interact with tumour cells and known to have pro-tumourigenic and antitumourigenic action through their mediators. Their role in the outcome of invasive breast cancer is still not clear with studies showing both a beneficial role as well as a role in tumour progression. Aim: To assess Mast Cell Density (MCD) in tissues of invasive carcinoma of breast and compare with clinicopathological parameters to infer their biological significance. Materials and Methods: The study was a retrospective cross-sectional study carried out at the Department of Pathology, SRM Medical College Hospital & Research Centre, SRMIST, Kattankulathur, Chennai, India on 51 cases of invasive carcinoma of breast received between January 2011 and July 2013 and the study conducted between August to November 2013. Clinical parameters and histopathological findings were recorded. Mast cells were clearly demonstrated in tissue using Toluidine Blue stain at pH 2.3. The MCD within primary malignant breast tumour tissue (mastectomy specimens, n=51) was assessed using an eyepiece grid and expressed as no. of cells/per sq. mm. Statistical analysis of all parameters was performed using SPSS software (v 17.0) to analyse association of MCD with clinicopathological parameters. Results: The MCD was significantly higher in malignant breast tissue compared to non neoplastic breast tissue (p<0.0001). No significant difference in MCD was observed between tumour size groups (T1, T2, and T3) (p=0.6967). No statistically significant difference in MCD was observed between the four histological types observed (p=0.892). The majority of the tumours were of Grade 2 category (n=37), followed by Grade 3 (n=11) and Grade 1 (n=3). No significant difference in MCD was observed between the three histological grades (p=0.785). The MCD was significantly higher in patients without lymph node metastasis compared to the group with lymph node metastasis (p=0.0023*). The MCD in invasive carcinoma of breast, post-chemotherapy group was significantly higher than that in invasive carcinoma of breast pre-chemotherapy group (p=0.0064). Conclusion: The current study shows significantly increased MCD at the periphery of the carcinoma breast tissue in patients without lymph node metastasis compared to those with lymph node metastasis. Hence, a potential antitumourigenic and beneficial role of mast cells in invasive primary breast carcinoma could be inferred and may serve as one of the prognostic indicators for patient stratification for various treatment options including immunotherapy

Cancer-associated fibroblast senescence and its relation with tumour-infiltrating lymphocytes and PD-L1 expressions in intrahepatic cholangiocarcinoma.

Caveolin-1 (CAV1) in cancer-associated fibroblasts (CAFs) has pro- or anti-tumourigenic effect depending on the cancer type. However, its effect in intrahepatic carcinoma (ICC) remains unknown. Therefore, this study aimed to investigate the relationship between CAV1 in CAFs and tumour-infiltrating lymphocyte (TIL) numbers or PD-L1 levels in ICC patients. Consecutive ICC patients (n = 158) were enrolled in this study. The levels of CAV1 in CAFs, CD8 + TILs, Foxp3+ TILs and PD-L1 in cancer cells were analysed using immunohistochemistry. Their association with the clinicopathological factors and prognosis were evaluated. The correlation between these factors was evaluated. CAV1 upregulation in CAFs was associated with a poor overall survival (OS) (P < 0.001) and recurrence-free survival (P = 0.008). Clinicopathological factors were associated with high CA19-9 levels (P < 0.001), advanced tumour stage (P = 0.046) and lymph node metastasis (P = 0.004). CAV1 level was positively correlated with Foxp3+ TIL numbers (P = 0.01). There were no significant correlations between CAV1 levels and CD8 + TIL numbers (P = 0.80) and PD-L1 levels (P = 0.97). An increased CD8 + TIL number and decreased Foxp3+ TIL number were associated with an increased OS. In multivariate analysis, positive CAV1 expression in CAFs (P = 0.013) and decreased CD8 + TIL numbers (P = 0.021) were independent poor prognostic factors. Cellular senescence, represented by CAV1 levels, may be a marker of CAFs and a prognostic indicator of ICC through Foxp3+ TIL regulation. CAV1 expression in CAFs can be a therapeutic target for ICC.

Targeted activity of the small molecule kinase inhibitor Pz-1 towards RET and TRK kinases

We have recently described Pz-1, a benzimidazole-based type-2 RET and VEGFR2 inhibitor. Based on a kinome scan, here we show that Pz-1 is also a potent (IC50 < 1 nM) TRKA/B/C inhibitor. Pz-1 potently inhibited proliferation of human cancer cells carrying either RET- or TRKA oncoproteins (IC50 ~ 1 nM), with a negligible effect against RET- and TRKA-negative cells. By testing mutations, known to mediate resistance to other compounds, RET G810R/S, but not L730I/V, E732K, V738A and Y806N, showed some degree of resistance to Pz-1. In the case of TRKA, G595R and F589L, but not G667C, showed some degree of resistance. In xenograft models, orally administered Pz-1 almost completely inhibited RET- and TRKA-mutant tumours at 1–3 mg/kg/day but showed a reduced effect on RET/TRKA-negative cancer models. The activity, albeit reduced, on RET/TRKA-negative tumours may be justified by VEGFR2 inhibition. Tumours induced by NIH3T3 cells transfected by RET G810R and TRKA G595R featured resistance to Pz-1, demonstrating that RET or TRKA inhibition is critical for its anti-tumourigenic effect. In conclusion, Pz-1 represents a new powerful kinase inhibitor with distinct activity towards cancers induced by oncogenic RET and TRKA variants, including some mutants displaying resistance to other drugs.

Cerium oxide nanoparticles elicit antitumourigenic effect in experimental breast cancer induced by N-methyl-N-nitrosourea and benzo(a)pyrene in female Wistar rats.

The use of cerium oxide nanoparticles [CeO2 NPs] in the biomedical field has continued to gain prominencedue to its potent antioxidant property. This study was designed to assess the antitumorigenic effect of CeO2 NPs in rats administered N-methyl-N-nitrosourea [NMU] and benzo(a)pyrene (BaP). Twenty four female Wistar rats were equally assigned into four groups and treated with normal saline (control), [NMU + BaP], [NMU + BaP+CeO2 NPs], and [NMU + BaP + vincristine]. Animals were pretreated with NMU and BaP three times (age 7, 10, and 13 weeks). Thereafter, vincristine and CeO2 NPs were administered twice and three times per week, respectively, for 13 weeks. Results showed that the administration of NMU and BaP increased serum nitric oxide [NO] and myeloperoxidase [MPO] by 220% and 132%, respectively, whereas the activities of aspartate and alanine aminotransferases and level of total bilirubin remained unchanged. Furthermore, mammary inflammatory [NO and MPO] and oxidative stress (LPO) markers were increased by 37%, 19%, and 24%, respectively. Mammary superoxide dismutase, catalase, reduced glutathione, and glutathione-S-transferase were significantly decreased in [NMU + BaP]-administered rats by 165%, 146%, 35%, and 36%, respectively. Immunohistochemistry showed downregulation of Bax, p53, and caspase-3, while histology revealed the presence of malignant epithelial cells with pyknotic nuclei and high nucleocytoplasm in [NMU + BaP]-administered rats. Treatment with CeO2 NPs attenuated oxidative stress, apoptosis, and inflammation and restored the cytoarchitecture of the tissue. Overall, CeO2 NPs show an antitumourigenic effect in experimental breast cancer by targeting pathways linked to inflammation and apoptosis.

Nucleofection optimization and in vitro anti-tumourigenic effect of TRAIL-expressing human adipose-derived mesenchymal stromal cells.

BackgroundTumour homing capacity of engineered human adipose-derived mesenchymal stromal cells (ADMSCs) expressing anti-tumour agents might be the key for a much safer and yet efficient targeted tumour therapy. However, ADMSCs exhibit resistant to most gene transfection techniques and the use of highly efficient viral vectors has several disadvantages primarily concerning safety risk. Here, we optimized the use of highly efficient and safe nucleofection-based transfection using plasmid encoded for TNF-Related Apoptosis Inducing Ligand (TRAIL) into ADMSCs and investigated the potential anti-tumourigenic of TRAIL-expressing ADMSCs (ADMSCs-TRAIL) on selected cancer models in vitro.MethodsDifferent concentration of TRAIL-encoded plasmid and ADMSCs were nucleofected and the percentage of fluorescence cells were analyzed to determine the optimal condition. TRAIL protein and mRNA were validated in nucloeofected ADMSCs using ELISA and RT-PCR respectively. Evaluation of TRAIL specific death receptors were performed on both tumours (A549/lung tumour, LN18/glioblastoma and HepG2/hepatocellular carcinoma) and haematological malignant lines (REH/acute lymphocytic leukaemia, K562/chronic myelogenous leukaemia and KMS-28BM/multiple myeloma) using flow cytometry. ADMSCs-TRAIL was subsequently assessed for anti-tumourigenic properties using both proliferation assay (MTS assay) and apoptosis assay (Annexin-V / Propidium Iodide staining).ResultsNucleofection showed increased total plasmid concentration (2 μg to 8 μg) resulted in significantly higher reporter expression (11.33% to 39.7%) with slight reduction on cells viability (~10%). ADMSCs-TRAIL significantly inhibited ~50% of cell proliferation in LN18, signifying sensitivity of the cell to ADMSCs-TRAIL mediated inhibition. Inhibition of both tumour and malignant lines proliferation by ADMSCs-TRAIL conditioned medium noticed in HepG2, A549 and REH respectively, whereas K562 and KMS-28BM malignant lines exhibit resistant to ADMSCs-TRAIL mediated inhibition. Moreover, we found that native ADMSCs alone were capable of inducing apoptosis in both LN18 and HepG2 tumour lines, despite substantial increased on the percentage of apoptosis by ADMSCs-TRAIL.ConclusionADMSCs-TRAIL selectively inhibit cancer model and markedly induces apoptosis. Through investigation of the specific TRAIL death receptors expression, we saw that the receptors expression did influence the sensitivity of some but not all cancer lines to TRAIL-mediated inhibition. This study provides further insight into the anti-tumourigenic potential of ADMSCs-TRAIL on different cancer models.

Abstract 3993: New paradigms for the modulatory actions of p38δ MAPK restorative expression in esophageal squamous cancer cell growth, migration and response to chemotherapy: A possible future therapeutic target.

Abstract Background: Esophageal cancer is an aggressive tumour which responds poorly to both chemotherapy and radiation therapy and has a poor prognosis. Approximately half of patients diagnosed with localized esophageal cancer die of metastatic disease within the first 2 years following tumour resection. Thus, a greater understanding of the biology of esophageal cancer is needed in order to identify novel therapeutic targets. Among these targets p38 MAPK isoforms are becoming increasingly important for a variety of cellular functions. A better understanding of the role(s) of p38 MAPKs may provide useful therapeutic tools for the management of human cancers. Methods: We analysed p38 MAPK isoform expression in both cancer cell lines and corresponding human normal and tumour tissue including esophageal, liver, lung, prostate, colon, renal and pancreatic. We observed that over 75% of both cell lines and human tumour tissue samples fail to express the specific isoform p38δ MAPK. To evaluate the role(s) of p38δ and active p-p38δ MAPK in esophageal cancer progression we developed a series of constructs: p38δ MAPK (pcDNA3-FLAG-p38δ MAPK), active p-p38δ MAPK (pcDNA3-MKK6b(E)-(Gly-Glu)5-FLAG-p38δ) and inactive p38δDN MAPK (pcDNA3-MKK6b(E)-(Gly-Glu)5-FLAG-p38δDN). The active p-p38δ plasmid was generated by removing the MKK6b(E) stop codon in pcDNA3-MKK6b(E). Secondly, FLAG-p38δ was amplified with a 5′(Gly-Glu)5 linker. Linear blunt-ended pcDNA3-MKK6b(E) and blunt ended (Gly-Glu)5-FLAG-p38δ were ligated. The dominant negative p-p38δ plasmid (p-p38δDN) was generated from pcDNA3-MKK6b(E)-(Gly-Glu)5-FLAG-p38δ by replacing Thr180 with Ala and Tyr182 with Phe. Stable esophageal squamous cancer cell lines expressing these plasmids were generated. Results: Re-introduction of p38δ or p-p38δ proved to be (a) anti-proliferative, (b) anti-migratory and (c) pro-apoptotic. We observed a time-dependent decrease in proliferation in esophageal cancer cells stably transfected with p38δ MAPK. This anti-proliferative effect was exacerbated with cells transfected with p-p38δ MAPK. Using a Boyden chamber and a wound healing assay we observed a decrease in cellular migration and invasion with cells transfected with p38δ or p-p38δ MAPK. Finally using Annexin V-FITC/PI double labeled flow cytometry we observed that p38δ and p-p38δ MAPK confers a greater sensitivity of esophageal cancer to certain chemotherapeutic drugs but resistance to others. Conclusion: We now provide data for the anti-tumourigenic effect of p38δ MAPK in esophageal cancer. Our research may provide a new potential target for the treatment of esophageal cancer and metastases. Interestingly, our findings are also applicable to other cancer types namely renal, prostate and pancreatic cancer which also lack p38δ MAPK isoform expression. Citation Format: Carol O'Callaghan, Liam Fanning, Aileen Houston, Orla P. Barry. New paradigms for the modulatory actions of p38δ MAPK restorative expression in esophageal squamous cancer cell growth, migration and response to chemotherapy: A possible future therapeutic target. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3993. doi:10.1158/1538-7445.AM2013-3993

Cannabidiol as potential anticancer drug

Over the past years, several lines of evidence support an antitumourigenic effect of cannabinoids including Δ(9)-tetrahydrocannabinol (Δ(9)-THC), synthetic agonists, endocannabinoids and endocannabinoid transport or degradation inhibitors. Indeed, cannabinoids possess anti-proliferative and pro-apoptotic effects and they are known to interfere with tumour neovascularization, cancer cell migration, adhesion, invasion and metastasization. However, the clinical use of Δ(9)-THC and additional cannabinoid agonists is often limited by their unwanted psychoactive side effects, and for this reason interest in non-psychoactive cannabinoid compounds with structural affinity for Δ(9)-THC, such as cannabidiol (CBD), has substantially increased in recent years. The present review will focus on the efficacy of CBD in the modulation of different steps of tumourigenesis in several types of cancer and highlights the importance of exploring CBD/CBD analogues as alternative therapeutic agents.

Rapamycin blocks hepatoblastoma growth in vitro and in vivo implicating new treatment options in high-risk patients

Activation of the protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signalling pathway plays a central role in the formation of hepatoblastoma (HB), the most common liver cancer in childhood. Blocking this pathway with specific mTOR inhibitors such as the immunosuppressant rapamycin is being currently tested for a variety of cancers. Here, we report that rapamycin treatment induced a significant dose-dependent inhibition of cell viability and promoted apoptosis in HB cells in vitro. Moreover, rapamycin inhibited AKT/mTOR signalling by dephosphorylation of the downstream target p70S6 kinase (p70S6K). Most importantly, treating subcutaneous HUH6 xenograft tumour bearing mice orally with 5mg/kg/day rapamycin for three weeks resulted in a striking reduction of tumour growth, as evidenced by reduced volume and weight, and moderately lowered tumour-specific alpha-fetoprotein (AFP) serum levels. The anti-tumourigenic effect was primarily ascribed to a significantly reduced proliferation rate upon p70S6K dephosphorylation, as microvascular density of rapamycin-treated compared to vehicle-treated tumours stayed grossly unchanged. Of uttermost clinical importance, we found no evidence for a feedback-loop activation of AKT in vivo. In conclusion, we demonstrate that rapamycin effectively inhibits HB growth both in vitro and in vivo by blocking AKT/mTOR signalling at the level of p70S6K and that rapamycin should be considered to treat HB patients especially those to be indicated for liver transplantation to benefit from its anti-tumourigenic and immunosuppressive properties.

Do antidiabetic medications play a specific role in differentiated thyroid cancer compared to other cancer types?

The risk for differentiated thyroid cancer, like for many other types of cancer, is increased in obese individuals and people with intermediate hyperglycaemia. The incidence of all cancers, with the exception of thyroid cancer, is also increased in type 2 diabetes mellitus patients. The review compares the prevalence of thyroid carcinoma and other cancers in obese, people with intermediate hyperglycaemia and patients with diabetes and summarizes mode of action and anti-tumourigenic effect of common antidiabetic medications. The over-expression of dipeptidyl peptidase IV in the tumours, not seen in the other cancer types, is suggested as a potential reason for the unique situation in thyroid cancer.

IFN-γ combined with targeting of XIAP leads to increased apoptosis-sensitisation of TRAIL resistant pancreatic carcinoma cells

The tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) is a specific and potent inducer of apoptosis in cancer cells, but the resistance of many tumour cells to TRAIL still represents a major hurdle for the clinical treatment of tumours with TRAIL. As apoptosis is regulated by the balance of activities of several anti-apoptotic factors and pro-apoptotic factors, we analysed the relative contribution of the two sides and found that down-regulation of Bcl-x L and in particular XIAP, but not c-Flip, sensitised the TRAIL resistant pancreatic cancer cell line Panc-1. A combination of both XIAP and Bcl-x L knock-downs showed no substantial added benefit indicating that both act in the same pathway. Notably, the degree of sensitisation by silencing of anti-apoptotic genes was further elevated by concomitantly increasing the pro-apoptotic potential in Panc-1 cells through over-expression of TRAIL-R1 or IFN-γ-mediated increases in caspase-8 levels. Similar sensitisation effects were obtained for another TRAIL-resistant pancreatic tumour cell line, AsPC-1. Our findings demonstrate that modulation of the balance between anti- and pro-apoptotic pathways from both sides by inhibition of apoptosis-antagonists and stimulation of pro-apoptotic factors provides the best way to enhance the anti-tumourigenic effect of TRAIL.

Betulinic acid induces apoptosis and inhibits hedgehog signalling in rhabdomyosarcoma

Background:Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in childhood with the ability to resist apoptosis by the activation of survival promoting and anti-apoptotic proteins.Methods:Efficacy of the apoptosis-inducing agent betulinic acid (BA) was determined in RMS cell cultures and in vivo by measuring cell viability, survival, apoptosis, hedgehog signalling activity, and neovascularisation.Results:Betulinic acid had a strong cytotoxic effect on RMS cells in a dose-dependent manner. The BA treatment caused a massive induction of apoptosis mediated by the intrinsic mitochondrial pathway, which could be inhibited by the broad-range caspase inhibitor zVAD.fmk. Exposure of hedgehog-activated RMS-13 cells to BA resulted in a strong decrease in GLI1, GLI2, PTCH1, and IGF2 expression as well as hedgehog-responsive luciferase activity. Intraperitoneal injection of 20 mg BA per kg per day significantly retarded growth of RMS-13 xenografts in association with markedly higher counts of apoptotic cells and down-regulation of GLI1 expression compared with control tumours, while leaving microvascular density, cell proliferation, and myogenic differentiation unaffected.Conclusion:Our data show that induction of apoptosis and inhibition of hedgehog signalling are important features of the anti-tumourigenic effect of BA in RMS and advices this compound for the use in a multimodal therapy of this highly aggressive paediatric tumour.

Endogenous neural precursor cells display extensive tropism for experimental glioblastomas and improve survival in an animal model

Gliomas are the major tumour entity of the central nervous system. The term glioma implies that these cells may originate from glial cells, although this has not been proven. Recent evidence indicates an interaction and interrelationship of glioma cells with stem cells. Neural stem cells generate all neurons, astrocytes and oligodendrocytes of the developing brain. They persist through adulthood in two specialized stem cell niches. A number of recent studies suggest that altered proliferation and differentiation of neural stem and precursor cells could be the primary source for glioma. Previously, it was demonstrated that cultured, immortalised neural precursors display tropism for gliomas, and that such precursors exert an anti-tumourigenic effect on glioma.

Evidence that prostate gonadotropin‐releasing hormone receptors mediate an anti‐tumourigenic response to analogue therapy in hormone refractory prostate cancer

Gonadotropin-releasing hormone analogue (GnRHa) therapy is an established method of androgen withdrawal in the treatment of prostate cancer. The present study investigated if the expression of prostate GnRH receptors (GnRHRs) might influence the response to GnRHa. GnRHR protein expression was first studied in a panel of prostate cancer cell lines. In androgen-dependent cells, GnRHR expression was unchanged following acute or chronic androgen withdrawal. In these cells, GnRHa significantly inhibited androgen-induced cell proliferation (p = 0.01). In contrast, GnRHa was unable to further suppress basal levels of cell proliferation induced by androgen withdrawal. In androgen-independent prostate cancer cells, variable levels of GnRHR expression were observed. In these cells, GnRHa treatment blocked cell proliferation (p = 0.001) and invasion (up to 70%) induced by fibroblast growth factor stimulation. Crucially, this effect was only evident in cells that expressed high levels of the GnRHR. GnRHa treatment also significantly inhibited the ability of these cells to recover from a cytotoxic insult (50% inhibition). The clinical significance of prostate GnRHR was tested by immunohistochemistry in a preliminary cohort of patients treated with GnRHa or surgical castration. There was no association between GnRHR expression and pathological grade, clinical stage, time to PSA nadir (p = 0.82) (n = 35) or progression to hormone refractory disease (p = 0.22) (n = 21), irrespective of the treatment method. GnRHa therapy in the presence of high GnRHR expression however, was found to be associated with longer disease-specific survival (mean survival 85 months, p = 0.002). In contrast, high GnRHR expression was not associated with survival among surgically castrated patients (mean survival 50 months, p = 0.7). Taken together, these data support the notion of a functional interaction between GnRHa and the GnRHR, which results in an anti-tumourigenic effect on prostate cancer cells. Findings from this report have direct implications for the use of GnRHR as a novel therapeutic target in hormone refractory prostate cancer.