Aryl hydrocarbon receptor (AhR), an evolutionary conserved transcription factor, is a
pleiotropic signal transductor. Thanks to its promiscuous ligand binding domain, during the
evolution of eukaryotic cells its developmental functions were integrated with biosensor functions.
Its activation by a multitude of endogenous and exogenous molecules stimulates its participation
in several pathways, some of which are linked to inflammation and breast cancer (BC). Over time,
the study of this malignancy has led to the identification of several therapeutic targets in cancer cells.
An intense area of study is dedicated to BC phenotypes lacking adequate targets. In this context, due to
its high constitutive activation in BC, AhR is currently gaining more and more attention

Guarnieri, T

English

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AhR, inflammation and breast cancerSubjects: Pharmacology & PharmacySubmitted by: Tiziana GuarnieriDefinition1. HistorySince 550 million years ago, it has constantly been expressed in animal cells, but its exact physiologicalrole remains elusive . AhR was initially identified as a xenobiotic sensor and a key regulator ofxenobiotics metabolism and persistent chemicals of concern, such as halogenated aromatic hydrocarbons(HAHs) and polycyclic aromatic hydrocarbons (PAHs). These compounds degrade very slowly in theenvironment, bio-accumulate in the food chain and are lipid soluble. Typically, they may be detected inhuman blood, adipose and breast tissue, where dichlorodiphenyldichloroethylene (DDE), the majormetabolite of DDT (dichloro-diphenyl-trichloroethane), and PCBs (polychlorinated biphenyls) are the mostprevalent contaminants. In this scenario, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is the most infamousmember of this class of environmental pollutants and one of exogenous best agonists of AhR. Consideringthe evolutionary conservation, it makes sense to hypothesize that over time, biosensor functions havebeen added to its physiological role, which to this day remains elusive. As Mulero-Navarro and FernandezSalguero recently pointed out, the role of AhR in systems homeostasis preceded its participation inxenobiotic sensing, as this function is lacking in invertebrates, where AhR homolog is a pivotal gene in thedevelopment of the nervous system, antennae, eyes and legs.2. Structure, Physiology and Target GenesIn the cytoplasm, AhR forms a macromolecular inactivating complex with two heat shock proteins 90(HSPs90), one XAP2 (hepatitis B virus X-Associated Protein 2 or AIP, Immunophilin-like Ah Receptor-interacting protein), one HSP90 co-chaperone protein named p23 and one pp60 src (Figure 1). After ligandbinding, the AhR-inactivating complex is remodeled. XAP2 is released and, soon after, the complex movesto the nuclear compartment. Here, the two HSPs90 proteins, together with p23 and pp60 src, arereplaced by AhR Nuclear Translocator (ARNT)/ hypoxia-induced factor β (HIF-1β). This heterodimer is theactive form of AhR. It regulates the transcription of target genes through binding to the xenobioticresponsive elements (XREs) in their promoter containing the sequences 5′-GCGTG-3′ or 5′-ACGTG-3′. Oncethe transcription starts, AhR separates from XRE and is exported out of the nucleus by Exportin 1 (XPO1),also known as chromosome region maintenance 1 (CRM1). In the cytoplasm, it is inactivated by 26Subiquitin–proteasome. Intriguingly, AhR self-regulates its nuclear activity as it activates the gene encodingfor Aryl hydrocarbon Receptor Repressor (AhRR). This repressor competes with AhR for the binding withARNT and forms the inactive heterodimer AhRR/ARNT. In this way, AhR lacks its partner that makes ittranscriptionally active and its nuclear activity is hindered. Another target of AhR is the TCDD-induciblePoly ADP-Ribose Polymerase (TIPARP) gene, which negatively regulates AR expression and thus AhR levels. These negative feedback mechanisms, together with the nuclear exportation and ubiquitin-proteasome-mediated degradation of AhR, regulate the transcription of AhR-responsive genes. Back in 1983, Israeland Whitlock showed that cytochrome P -450 genes (CYP450, class 1A (1 and 2)), encoding enzymaticmetalloproteins involved in endogenous and exogenous substrates transformation, are targeted from “theTCDD receptor”, later identified as the AhR. In the year 2000, Nebert group described four additional AhRtarget genes: 1. ALDH3A1 (Aldehyde dehydrogenase family 3, subfamily 1); 2. GSTA1 (Glutathione S-transferase, alpha 1); 3. NQO1, (NAD(P)H dehydrogenase quinone 1); 4. UGT1A6 (UDPglucuronosyltransferase family 1 member A6). Together with CYP1A1 AND CYP1A2 genes, they form theso-called “AhR gene battery”, which controls the cell cycle and the initiation of the apoptotic cascade.Later, other groups confirmed and extended the list of AhR-responsive genes which are involved inAhR, an environmentally sensitive transcription factor, is one of the more evolutionary conservedmolecules in living cells.[1]1different steps of cell lifecycle and metabolism. These include also Phase III transporters, as the Proteins 2and 3 associated to multidrug resistance (MRP2-MRP3), the organic anionic and cationic transportproteins (OATP and OCTP), the breast cancer resistance proteins (BCRP) . More recently, it has beensuggested that AhR participates in pluripotency and stemness regulation through a possible link withtransposable elements. Alternative, “unorthodox” signaling has been described in the last few years, as ithas been observed that some AhR-responsive genes do not contain the XREs. This is the case forplasminogen activator inhibitor-1 (PAI-1), encoding a fibrinolysis inhibitor which has been connected toinflammatory endothelial fibrosis. The PAI-1 promoter is not a classical XRE, but is responsive to TCDD,one of the more potent exogenous agonists of AhR binding. In fact, Wright group described anonconsensus XRE (NC-XRE) in PAI-1 promoter which interacts with AhR alone when it is associated toKruppel-like factor 6 (KLF6). AhR dimerizes with the KLF family member KLF6 and binds to a novelnonconsensus XRE (NC-XRE) in the promoter of target genes after TCDD exposition. NC-XRE and XRE haveno sequence homology and interact with different proteins, this suggesting that AhR also has differenttargets. In particular, they demonstrated that the complex AhR/KLF6 is of pivotal importance in thecontrol of cell cycle, as it regulates the expression of p21Cip1, or, cyclin-dependent kinase inhibitor 1A(CDKN1A), which inhibits the cell cycle and regulates cell metastasis by switching between invasion andproliferation. Considering these data, it is reasonable to hypothesize that originally AhR had a checkpointrole in the cellular metabolism and that, over time, AhR has acquired the ability to bind to a multitude ofmolecules, both exogenous and endogenous. Over time, it has been demonstrated that, due to itspromiscuous binding site, AhR is responsive not only to exogenous, but also to a variety of naturalmolecules, among which some derivatives of arachidonic acid as prostaglandins and leukotrienes,lipotoxin A 4 and 7-ketocholesterol , the hemoglobin catabolites bilirubin and biliverdin, the tryptophanproducts kynurenine (Kyn), tryptamine and 6-formylindolo[3,2-b] carbazole (FICZ), some indolemetabolites derived from diet and from host bacteria metabolism, such as Indole-3-acetic acid (IAA) andIndol [3,2-b]carbazole (ICZ) (Figure 2). The list of possible AhR ligands is constantly growing and some ofthese compounds are now included into the category of selective AhR modulators (SAhRMs), that bindAhR with low to medium affinity. Interestingly, they can behave as an agonist, antagonist or a mixedagonist/antagonist, depending on the metabolic set-up of the organism with which they come intocontact. Most information about the role of AhR in organ and system physiology comes from multiplesystemic abnormalities described in AhR /AhR  mice. Obviously, these rodents are not sensitive to TCDDand are also smaller in size than AhR /AhR  mice. They exhibit a spectrum of anomalies that includesreduced fertility, portal tract fibrosis, hepatocyte microvesicular fatty metamorphosis, cardiachypertrophy, epidermal hyperplasia, T cell deficiency in the spleen and altered circadian rhythms .Figure 1. The Aryl hydrocarbon receptor (AhR) pathway. This transcription factor usually resides in thecytoplasm, where it associates with some inactivating molecules: two HSPs90s, one XAP2 or AIP, oneHSP90 co-chaperone protein named p23, one pp60(c-src). Once it binds a ligand, XAP2 leaves thecomplex that moves to the nucleus. Here, the two HSP-90s, p23 and pp60-src drop AhR that binds toARNT. This new complex is now ready to interact with XRE of geneslike CYP1A1, CYP1A2, CYP1B1 and AHRR, usually containing the sequences 5’-GCGTG-3′ or 5′-ACGTG-3′.AhR also targets NC-XREs, which bind AhR, while not containing the canonical consensus sequence. Just[1]− −+ +[2]before the transcription starts, AhR moves from XREs, is translated out of the nucleus by XPO1 (notshown) and is degraded in the cytoplasm by 26S ubiquitin–proteasome. AhRR = Aryl HydrocarbonReceptor Repressor; AIP = Immunophilin-like Ah Receptor-interacting protein); ARNT = Aryl HydrocarbonNuclear Translocator; CYP1A1 = Cytochrome P450 Family 1 Subfamily A Member 1; CYP1A2 =Cytochrome P450 Family 1 Subfamily A Member 2; CYP1B1 = Cytochrome P450 Family 1 Subfamily BMember 1; HSPs90 = Heat Shock Proteins 90; NC-XRE = Nonconsensus Response Element; p23 =Proteolytically Resistant 23-kDa protein; pp60(c-src) = Proto-oncogene tyrosine-protein kinase 60(Sarcoma); XAP2 = Hepatitis B virus X-Associated Protein 2; XPO1 = Exportin 1; XRE = XenobioticResponse Element. Figure 2. Endogenous and exogenous ligands of AhR. Endogenous ligands come from anabolism(kynurenine, tryptamine, β-Naphtoflavone, FICZ, prostaglandins, leukotrienes) and catabolism (bilirubin,biliverdin) of individuals and their commensal flora (microbiota: Indole-3-Carbinole; Indole-3-Acetic acid;Indol [3,2-b]carbazole; 7-ketocholesterol). Exogenous ligands can be manmade products which have aheterogeneous nature and uses (DDE, DDT, PCBs, TCDD, BaP, DMBA, Omeprazole Tranilast). Severalexogenous ligands of natural origin are contained in plants and vegetable foods (Flavonoids, Resveratrol,Luteolin, Quercetin, Epigallocatechin, Genistein, Daidzein). DDE = dichlorodiphenyldichloroethylene; DDT= dichloro-diphenyl-trichloroethane; PCBs = polychlorinated biphenyls; TCDD = 2,3,7,8-tetrachlorodibenzo-p-dioxin; BaP = Benzo [a] pyrene; DMBA = 7,12-Dymethylbenz-[α]-anthracene.3. AhR and InflammationFrom 2000 onwards, various groups have reported a connection between AhR and inflammatoryphenomena in different experimental models. In 2002, Hennig group showed that in endothelial cells, thePCB-mediated activation of AhR promotes inflammatory atherosclerotic phenomena, passing through thetranscriptional activation of NF-B and the increase in IL6 production. NF-kB/AhR interactions ininflammation were also described in detail by Tian and colleagues and, in the same period, an interestingreview by Dalton was focused on the connection between AhR, inflammatory signaling and oxidativestress. Over the years, AhR’s pathway has been associated with various inflammatory markers, includingcyclooxygenase-2 (COX2), tumor necrosis factor apha (TNFalpha),matrix metalloproteinases (MMPs), early growth response 1 (EGR1), prostaglandin E 2 (PGE2), microsomalPGE2 synthase (mPGE2S), NF-kB component RelB, RelA, inducible nitric oxide synthase (iNOs), interleukin8 (IL8)  .4. AhR and Breast CancerMore generally, inflammation is an ascertained risk factor for the onset of a plethora of pathologies,among which cancer. Here, a frequently emerging role in the modulation of the inflammatory state isemerging for AhR .The connection between inflamed phenotype and neoplasia is now evident in BC. BC[3][4]is the most common invasive malignancy among women in the industrialized world. It is a complex,multifactorial and extremely heterogeneous disease. Genetic factors account for up to 10% of all BCs.Among inherited mutations, BRCA1 e BRCA2 (BReast CAncer) suppressor genes are the most involved inBC susceptibility. BRCA1 interacts both at transcriptional and post-transcriptional level with estrogenspathway, in order to limit their positive eects on proliferation of mammary tissues.The lack of this control is a well-known risk factor for TNBC occurrence and this can occur due to BRCA1epigenetic mutations, among which AhR-instigated hypermethylation .This type of BC aects women inall age groups. It is a highly aggressive neoplasia whose cells, do not express ER, PR and HER2. Therefore,it has fewer options for targeted treatment, as hormone therapy and HER2 drugs cannot be used. For thisreason, its outcome is not favorable. Over the last 10 years, the urgency to identify alternativetherapeutic options for TNBC has fueled the selection and the screening of selected molecular targets,some of them belonging to the AhR pathway. Goode and colleagues proposed that in TNBC tumors andcell lines, AhR expression is directly proportional to tumoral aggressive behavior, both in vivo and in vitro.In MDA-MB231, a TNBC cell line, they observed the attenuation of the malignant phenotype after AHRknockdown. Considering these premises, AhR inhibition could be a promising therapeutic target in TNBC.This result can be obtained in vivo by the administration of selected antagonists, among which genistein.The dietary intake of this isoflavone had been shown to  nhibit AhR interaction with BRCA1 exon 1 in micemammary tissue and AhR-driven hypermethylation of CpG in BRCA1 gene. These results are consistentwith those obtained by the administration of the synthetic alpha-naphthoflavone and the natural flavonoidGalangin in MCF7 cells. The antiproliferative properties of these molecules have long been known and aresupported by data identifying them as AhR modulators in TNBC. Interestingly, galangin and the naturalisoflavone genistein are also known for their anti-inflammatory properties . 5. Conclusions In conclusion, it appears evident that there is a clear link between AhR pathway, inflammation and BC,particularly triple negative (Figure 3). Since inflammation is one of the factors that favors and supportstumor transformation through several factors, among which AhR, it is reasonable to include AhR amongthe targets of a combined anti-inflammatory/antitumoral therapy. [5][6][7] Fig. 3. AhR pathway interactions in cancer breast cells. AhR is usually overexpressed in cancerbreast cells. Here, it  correlates with several markers of:  1.  proliferation (NF-κB, IL6, c-Myc, BRCA1); 2.DNA repair (BRCA genes); 3. cell migration (kynurenine) 4. metastatic behavior (MMPs and PLAU); 5.immune system (Th17 and Treg cells); 6. inflammation (NF-κB, IL6, Kyn).  BRCA = BReast CAncer gene;Kyn = Kynurenine; c-Myc = Avian myelocytomatosis virus oncogene cellular homolog; IL-6 = interleukin6;MMP = matrix metalloproteinase; NF-kB =  nuclear factor kappa-light-chain-enhancer of activated Bcells; PLAU = Plasminogen Activator, Urokinase (gene); ↑ = increase; ↓ = decrease Retrieved from https://encyclopedia.pub/2089References1. 12. Marlowe, J.L.; Puga, A.; Aryl hydrocarbon receptor, cell cycle regulation, toxicity, and tumorigenesis.. J. CellBiochem. 2005, 96, 1174, 10.1002/jcb.20656.2. 18. Esser, C.; Rannug, A.; The aryl hydrocarbon receptor in barrier organ physiology, immunology, and toxicology.Pharmacol. Rev. 2015, 67, 259, 10.1124/pr.114.009001.3. Drew Neavin; Duan Liu; Balmiki Ray; Richard M. Weinshilboum; The Role of the Aryl Hydrocarbon Receptor (AHR) inImmune and Inflammatory Diseases. International Journal of Molecular Sciences 2018, 19, 3851,10.3390/ijms19123851.4. Tiziana Guarnieri; Provvidenza Maria Abruzzo; Alessandra Bolotta; More than a cell biosensor: aryl hydrocarbonreceptor at the intersection of physiology and inflammation. American Journal of Physiology-Cell Physiology 2020,318, C1078-C1082, 10.1152/ajpcell.00493.2019.5. Donato F. Romagnolo; Andreas J. Papoutsis; Christina Laukaitis; Ornella I. Selmin; Constitutive expression of AhR andBRCA-1 promoter CpG hypermethylation as biomarkers of ERα-negative breast tumorigenesis. BMC Cancer 2015, 15,1-11, 10.1186/s12885-015-2044-9.6. Gennifer D. Goode; Billy R. Ballard; H. Charles Manning; Michael L. Freeman; Yibin Kang; Sakina E. Eltom; Knockdownof aberrantly upregulated aryl hydrocarbon receptor reduces tumor growth and metastasis of MDA-MB-231 humanbreast cancer cell line.. International Journal of Cancer 2013, 133, 2769-80, 10.1002/ijc.28297.7. Donato F. Romagnolo; Kevin D. Daniels; Jonathan T. Grunwald; Stephan A. Ramos; Catherine R. Propper; Ornella I.Selmin; Epigenetics of breast cancer: Modifying role of environmental and bioactive food compounds.. MolecularNutrition & Food Research 2016, 60, 1310-29, 10.1002/mnfr.201501063.8. Donato F. Romagnolo; Kevin D. Daniels; Jonathan T. Grunwald; Stephan A. Ramos; Catherine R. Propper; Ornella I.Selmin; Epigenetics of breast cancer: Modifying role of environmental and bioactive food compounds.. MolecularNutrition & Food Research 2016, 60, 1310-29, 10.1002/mnfr.201501063.KeywordsAryl Hydrocarbon Receptor;Cytochrome P450 Family;TCDD

AhR, inflammation and breast cancer

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AhR, inflammation and breast cancer

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