Baci, D. | bioRxiv Science

bioRxiv2021

A polyphenol-rich extract of Olive Mill Wastewater Enhances cancer chemotherapy effects while mitigating cardiac toxicity.

BackgroundCardiovascular toxicities still remain one of the most undesirable side effects in cancer patients receiving chemotherapy, and cardiotoxicity has been detected associated with many therapeutic regimens. A number of mechanisms are reported for these effects, some of which are related to inflammation, oxygen radical generation, mitochondrial damage. Extra-virgin olive oil (EVOO) is rich in cancer preventive polyphenols endowed with anti-inflammatory, antioxidant activities which could exert protective effects on the heart cells. One very interesting derivative of EVOO preparation is represented by purified extract form waste waters. Here, we investigated the anti-cancer activity when combined with chemotherapeutics as well as potential cardioprotective activities of a polyphenol-rich extract from waste product of the EVOO, named A009. Methods and ResultsMice bearing prostate cancer (PCa) xenografts were treated with cisplatin with and without A009. Tumor cell growth was reduced by cis and by A009 and further hindered by the combination. The effects of the A009 extract on cardiovascular toxicities was investigated in vivo. Hearts of mice were analyzed, and the mitochondria were studied by transmission electron microscopy. A protection activity by A009 was observed. To confirm the in vivo data obtained with cisplatin therapy, tumor cell lines and rat cardiomyocytes were treated with cisplatin in vitro with and without A009. A009 enhanced cisplatin and 5FU reduced cancer cell growth while did not further affect co-treated rat cardiomyocytes. Another frequently used chemotherapeutic agent 5-fluorouracil (5FU), was also tested in this assay a similar effects were observed. The cardioprotective effects of the A009 extract towards 5 FU chemotherapy were further investigated in a second system of in vitro cultures, on cardiomyocytes freshly isolated from mice pups. These cells were treated with 5-fluorouracil and A009. Wastewater extract mitigated toxicity of the fluorpyrimidine. ConclusionsIn vivo, we found synergisms of A009 and cisplatin in prostate cancer treatment. Hearts of mice xenografted with PCa cell lines and receiving co-treatments of A009 extracts along with cisplatin had reduced mitochondria damage compared to chemotherapy alone, indicating a cardioprotective role. A009 in vitro was additive to cisplatin and 5FU to reduce cancer cell growth while did not further affect rat cardiomyocytes cell cultures treated with cisplatin and 5FU. The A009 extract also rescued the proliferation rate of neonatal murine cardiomyocytes treated with 5-Fluorouracil. Our study demonstrates that the polyphenol rich purified A009 extracts enhances the effect of chemotherapy in vitro and in vivo but mitigates effects on heart and heart cells. It could therefore represent a potential candidate for cardiovascular prevention in patients undergoing cancer chemotherapy.

cancer biology↗

bioRxiv2020

Prostate cancer peripheral blood NK cells show enhanced CD9, CD49a, CXCR4, CXCL8, MMP-9 production, and secrete monocyte-recruiting and polarizing factors

ABSTRACTBackground Natural killer (NK) cells are effector lymphocytes of the innate immunity. Two major NK cell subsets are mostly present in the peripheral blood (pNKs): the cytotoxic CD56dimCD16+ NK cell subset (90-95% of pNKs), and the low cytotoxic, highly cytokine-producing CD56brightCD16-/low NK cell subset (5-10% of pNKs). It has been demonstrated that NK cells in peripheral blood of patients with several tumors are altered. We have shown that in NSCLC and colon cancer, tumor associated circulating NK (pTA-NK) and tumor infiltrating NK (TI-NK) are skewed towards the CD56brightCD16-/low phenotype. We have detected the production of pro-inflammatory and pro-angiogenic cytokines and chemokines. Other groups are reporting similar observations. There is still a lack of knowledge concerning the phenotype of pNK cells in prostate cancer (PCa). Here, we phenotypically and functionally characterized peripheral blood NK (pNK) from PCa patients (PCa pTA-NKs) and investigated their production of soluble factors, with endothelial cells and macrophage stimulatory action.Methods NK cell subset distribution was investigated in the peripheral blood of PCa patients, by multicolor flow cytometry (FC) for surface antigens expression. Protein arrays were performed to characterize the secretome on FACS-sorted pNK cells. Secreted products from FACS-sorted PCa TA-NKs were used to characterize their production of pro-inflammatory molecules. Secreted products from FACS-sorted PCa pTA-NKs were also used to stimulate endothelial cells and monocytes and macrophages, determining their ability to recruit and polarize them. Alterations of endothelial cells and monocytes, following exposure to secreted products from FACS-sorted PCa pTA-NKs, was assessed by RT-PCR. To confirm these observations, secreted products from 3 different PCa (PC-3, DU-145, LNCaP) cell lines were used to assess their effects on human NK cell polarization, by multicolor flow cytometry.Results Circulating NK cells from prostate cancer patients have been studied before, mostly for their impaired lytic functions. However, here we are the first to report that circulating pNK cells from PCa patients acquire a CD56brightCD9+CD49a+CXCR4+ phenotype with pro-inflammatory properties. We observed a similar polarization of heathy-donor derived pNK cells exposed to secreted products of three different PCa cell lines. Increased production of CXCL8, CXCR4, MMP-9, pro-inflammatory and reduced production of TNFα, IFNγ and Granzyme-B was detected. PCa TA-NKs released factors able to support angiogenesis in vitro and increased the expression of CXCL8, ICAM-1 and VCAM-1 mRNA in endothelial cells, confirming a pro-inflammatory signature. Secretome analysis revealed the ability of PCa pTA-NKs to release pro-angiogenic cytokines/chemokines involved in monocyte recruitment and M2-like polarization. In experimental setting, secreted products from PCa pTA-NKs can recruit THP-1 monocyte and polarize THP-1-differentiated macrophage towards CD206/Arginase1/IL-10/CXCL8-expressing M2-like/TAMs.Conclusions Our results show that PCa pTA-NKs are effector cells able to produce pro-inflammatory angiogenesis factors able to stimulate endothelial cells, attract monocytes and polarize macrophage to an M2-like type. Our data provides a rationale for the possible use of pNK profiling in clinical studies on PCaGraphical Abstract: Representative cartoon illustrating the pro-angiogenic features of PCa pTA-NKs.A) direct effects of PCa pTA-NKs in supporting angiogenesis by interacting with endothelial cells. B) Proposed model for PCa pTA-NK pro-angiogenic activities via macrophage recruitment and polarization.Competing Interest StatementThe authors have declared no competing interest.ABBREVIATIONSADCCAntibody Dependent Cellular CytotoxicityADKAdenocarcinomaANGAngiogeninANGIOP1Angiopoietin 1ANOVAAnalysis of VarianceCCLChemokine Ligand (C-C motif)cDNAcomplementary DNACMConditioned MediaCRPCCastration Resistant Prostate CancerCXCLChemokine Ligand (C-X-C motif)DNAM-1DNAX accessory molecule 1dNK cellsdecidual Natural Killer cellsEBMEndothelial cell Basal MediumEGMEndothelial cell Growth MediumFACSFluorescence-Activated Cell SortingFBSFetal Bovine SerumFCFlow CytometryFSCForward ScatterGAPDHGlyceraldehyde-3-Phosphate DehydrogenaseGM-CSFGranulocyte-Macrophage Colony-Stimulating FactorHBVHepatitis B VirusHCHealthy ControlHCVHepatitis C VirusHIVHuman Immunodeficiency VirusHUVECHuman Umbilical Vein Endothelial CellICAMInter Cellular Adhesion Molecule-1IFN-γ = Interferon γIHCImmuno Histo ChemistryIL-Interleukin-I-TACInterferon-inducible T-cell Alpha ChemoattractantmAbsmonoclonal AntibodiesMCP-1/CCL2Monocyte Chemoattractant Protein-1MMPsMatrix Metallo ProteinasesMNCsMono Nuclear CellsNKNatural KillerNKG2DNatural Killer receptor Group 2 DNSCLCNon-Small Cell Lung CancerPAIPlasminogen Activator InhibitorPBPeripheral Blood cellsPBMCsperipheral Blood Mononuclear CellsPCaProstate CancerPD-L1Programmed Death receptor Ligand 1PGE2Prostaglandin E2PlGFPlacental Growth FactorPMAPhorbol Merystate AcetateP/SPenicillin / StreptomycinPTA-NKsProstate Tumor Associated Natural Killer cellsRANTESRegulated upon Activation, Normal T Cell Expressed and Presumably SecretedSSCSide ScatterSTATSignal Transducer and Activator of TranscriptionTAMsTumor Associated MacrophagesTGFβTransforming Growth Factor-βTIMPTissue Inhibitor of Metallo-ProteinaseTMETumor MicroenvironmentTNF αTumor Necrosis Factor-αTregT Regulatory CellsuPARurokinase-type Plasminogen Activator ReceptorVCAMVascular Cellular Adhesion Molecule-1VEGFVascular Endothelial Growth FactorVEGFRVascular Endothelial Growth Factor ReceptorView Full Text

cancer biology↗