Inflammation and Cancer The Significance of COX-2 Dan Dixon Dept. of Biological Sciences South...
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Transcript of Inflammation and Cancer The Significance of COX-2 Dan Dixon Dept. of Biological Sciences South...
Inflammation and CancerThe Significance of COX-2
Dan Dixon
Dept. of Biological SciencesSouth Carolina Cancer Center
Inflammation and Cancer
Inflammation is a critical component of tumor progression
Cancers can arise from sites of infection, chronic irritation and inflammation.
A vital component of the tumor microenvironment are the inflammatory cells.
Inflammation promotes the neoplastic processes involving proliferation, survival, and migration.
Tumor cells have co-opted inflammatory signaling molecules and receptors.
Tumors act as wounds that fail to heal
Historical Perspective In 1863, German physician Rudolf Virchow hypothesized that certain
classes of irritants, together with the tissue injury and inflammation, promote enhanced cell proliferation.
In 1918, Yamagiwa and Ichikawa, who showed that repeated painting of coal tar onto rabbits' ears causes carcinomas.
the world’s first man-made cancer on the ears of a rabbitRudolf Virchow
Inflammation Overview In response to tissue injury, signals start and maintain a host response to
“heal” the damage.
This involves getting “activated” leukocytes (WBCs) to the proper damaged location.
Adhesion molecules and integrins with their respective receptors/ligands modulate WBC rapid adhesion, activation of gene expression, and transmigration into the tissue.
Expression of chemokines attract specific WBC populations and promote the natural progression of the inflammatory response.
The “normal” inflammatory response associated with wound healing is self-limiting. Dysregulation of key steps leads to abnormalities…
pathogenesis… neoplasia.
Monocyte
lymphocyte
neutrophil
platelets
eosinophil
Some of the Players… Neutrophils (and eosinophils): The first cells recruited to
damaged tissues and invading bactera.
Monocytes: Chemotactic factors guide monocytes to the injury migrate into the site of injury.
Macrophages: Primarily stem from differentiated monocytes; they are the main source of growth factors and cytokines. Their presence profoundly effects the surrounding microenvironment cells.
Mast cells: Release histamine, cytokines, and proteases; let the body know something is wrong.
Lymphocytes: Provide firepower for the immune response.
Platelets: They aggregate to form a fibrin clot to close the wound.
Invasive Tumor Growth: Invasive carcinomas are less organized. Neoplasia-associated angiogenesis and lymphangiogenesis produces a chaotic vascular organization of blood vessels and lymphatics where neoplastic cells interact with other cell types (mesenchymal, haematopoietic and lymphoid) and a remodelled extracellular matrix. Although the vascular network is not disrupted in the same way during neoplastic progression as it is during wounding, many reciprocal interactions occur in parallel. Neoplastic cells produce an array of cytokines and chemokines that are mitogenic and/or chemoattractants for granulocytes, mast cells, monocytes/macrophages, fibroblasts and endothelial cells. In addition, activated fibroblasts and infiltrating inflammatory cells secrete proteolytic enzymes, cytokines and chemokines, which are mitogenic for neoplastic cells, as well as endothelial cells involved in neoangiogenesis and lymphangiogenesis. These factors potentiate tumour growth, stimulate angiogenesis, induce fibroblast migration and maturation, and enable metastatic spread via engagement with either the venous or lymphatic networks.
Wound Healing: Normal tissues have a highly organized and segregated architecture. Epithelial cells sit atop a basement membrane separated from the vascularized stromal (dermis) compartment. Upon wounding or tissue assault, platelets are activated and form a haemostatic plug where they release vasoactive mediators that regulate vascular permeability, influx of serum fibrinogen, and formation of the fibrin clot. Chemotactic factors such as transforming growth factor- and platelet-derived growth factor, derived from activated platelets, initiate granulation tissue formation, activation of fibroblasts, and induction and activation of proteolytic enzymes necessary for remodelling of the extracellular matrix (for example, matrix metalloproteinases and urokinase-type plasminogen activator). In combination, granulocytes, monocytes and fibroblasts are recruited, the venous network restored, and re-epithelialization across the wound occurs. Epithelial and stromal cell types engage in a reciprocal signalling dialogue to facilitate healing. Once the wound is healed, the reciprocal signalling subsides.
• Invasion: Macrophages secrete a variety of proteases to breakdown the basement membrane around areas of proliferating tumor cells thereby prompting their escape into the surrounding stromal tissue.
• Angiogenesis: In areas of tumor hypoxia, macrophages cooperate with tumor cells to induce a vascular supply for the area by up-regulating a number of angiogenic growth factors. These proangiogenic factors stimulate vascular endothelial cells in neighboring areas to migrate into new vessels for the tumor.
• Immunosuppression: Macrophages secrete factors that suppress the anti-tumor functions of innate immune system.
• Metastasis: Macrophages associated with tumor vessels secretes factors that guide tumor cells toward blood vessels where they then escape into the circulation. In the stromal compartment, macrophages secrete growth factors to stimulate tumor cell growth and motility.
The Roles of Tumor-Associated Macrophages in Tumor Progression
Chronic Inflammation is a Risk Factor for Cancer
Ulcerative colitis (UC) AdenocarcinomaChronic
InflammationDysplasia
• Patients with UC have a 5 to 7-fold greater risk of getting colon cancer.• UC persisting for 35-40 years increases the risk 20-35%.• Colon cancer associated with IBD has the worst prognosis. • Management with anti-inflammatory agents reduce incidence of cancer.
Ulcerative Colitis and Increased Risk of Colorectal Cancer Over Time
Eaden JA, et al. Gut. 2001;48:526. Overall prevalence of CRC in any UC patient is 3.7%
Time from diagnosis (years)
UC
0 5 10 15 20 25 30
5
10
15
20
25
1
Cum
ulat
ive
prob
abili
ty (
%)
Inflammatory Syndromes and Their Associated Cancers
Inflammatory bowel disease Colorectal cancer
Gastric intestinal metaplasia Gastric cancer
Barrett’s esophagus Esophageal cancer
Chronic hepatitis Hepatocellular carcinoma
Chronic pancreatitis Pancreatic cancer
Oral leukoplasia Head/neck cancer
Atypical adenomatous hyperplasia Non-small cell lung cancer
Ductal carcinoma in situ Breast cancer
Prostatic intraepithelial neoplasia Prostate cancer
Bladder dysplasia Bladder cancer
Cervical dysplasia Cervical cancer
Actinic keratoses Skin cancer
Chronic Inflammation Promotes Tumor Development
mutation
initiated epitheliumepithelial cells
Promotion
benign
Inflammatory cells express factors that stimulate cell
growth and progression
carcinoma
Malignant conversion
Chronic activation promotes continued inflammation, angiogenesis, and ECM remodeling
Insulted stromal cells recruit activated
inflammatory cells
Activated monocyte/macrophages
Anti-Inflammatory Drugs as Chemotherapeutic Agents
Aspirin and NSAIDs: Aspirin and NSAIDs inhibit prostaglandin synthesis. Aspirin inhibits platelet aggregation. Flurbiprofin has anti-metastatic effects. NSAIDs promote apoptosis, interfere with cell-cycle progression, and stimulate immune surveillance.
Anti-TNFAntibodies: Effective in controlling IBD. Regulates TNF activity which promotes cytokine, chemokine, adhesions, MMPs, and pro-angiogenic activities.
Adhesion Molecule Receptor Antagonists: Cancer cells and tumors contain mucins and ligands for cell adhesion molecules. Blocking adhesion may alter metastasis.
MMP Inhibitors: MMPs can promote tumor growth and also attenuate growth.
Cyclooxygenase and Arachidonic Acid Metabolism
• Arachidonic acid (AA) is a 20-carbon polyunsaturated fatty acid.
• AA is liberated from the membrane glycerophospholipids by phospholipases.
• 3 Major pathways convert AA into biologically active eicosanoids.
• Cyclooxygenase (COX) catalyses a key step in the formation of prostaglandins.
COOH
Arachidonic Acid
Cycloxygenase Lipoxygenase Epoxygenase
Prostaglandins (PGs)Thromboxanes (TXs)
Leukotrienes (LTs)Lipoxins (LXs)
Epoxy Acids (EETs)Dihydroxy Acids
PGE2, PGF, and PGI2
RELAX VASCULARSMOOTH MUSCLE
PGE2 and PGI2
INCREASERENAL BLOOD FLOW
PGE2 and PGI2
RELAX BRONCHIALSMOOTH MUSCLE;
PGF CONTRACTS IT
PGE2 and PGF
CONTRACT UTERINESMOOTH MUSCLE;PGI2 RELAXES IT
PGE2 and PGI2
PROTECTGASTRIC MUCOSA
TxA2 PROMOTESPLATELET AGGREGATION;
PGI2 INHIBITS IT
Prostaglandin Regulate Physiological Functions
Prostaglandins are biologically active
phospholipid molecules that regulate many physiological
functions
Proper balance of prostaglandins are critical for normal
homeostasis
Prostaglandin Signaling Mechanisms
Prostaglandins control cellular function through
G-coupled membrane receptors and nuclear
receptors
Increased PGE2 Promotes Many Facets of Tumorigenesis
VascularAngiogenesis
ImmuneSuppression
ReducedApoptosis
ProliferationMotility
VEGF
PGE2
MetastasisInvasion
MMP-2 MMP-9 BCL-2
PI3-KActivation
IL-10 IL-12
Multiple COX Enzymes Exist in Cells
COX-1: Constitutively present
COX-2: Inducible isoform
“COX-3”: Splice variant of COX-1
Feature COX-1 COX-2
Expression Constitutive Inducible
Protein Size 72 kDa 72/74 kDa
Gene Size 22 kb 8.3 kb
mRNA size 2.7 kb 4.5 kb; contains multiple AU-rich elements in 3’UTR
Localization ER, nuclear envelope ER, nuclear envelope
Cell & Tissue Expression Platelets, stomach, kidney, Expressed in most cells or tissues after stimulation withcolon, most tissues cytokines, growth factors, or tumor promoters
Gene Regulation Constitutive low-level expression Immediate-early response gene, rapidly transcribed,mRNA is rapidly degraded, translation is controlled
Differing Functions of COX-1 & COX-2
Arachidonic Acid
COX-1(constitutive)
Homeostasis• Stomach/GI protection• Platelet aggregation• Renal blood flow
COX-2(inducible)
Pathophysiology• Inflammation, Pain• Fever• Cancer• Morbus Alzheimer• Ischemia (CNS)
Evidence of a COX-2 Dependent Role in NeoplasiaEpidemiological Studies
Decreased risk of CRC-associated deaths in aspirin users. The NSAID sulindac decreases the size and number of polyps (FAP). Prostaglandin levels are increased in CR tumors. Overexpression of COX-2 detected in adenomas and adenocarcinomas.
Animal Studies Min mice and AOM-treated rats have elevated COX-2 levels. Sulindac and other NSAIDs attenuate intestinal tumor and xenografted cancer cell growth in mice.
Cellular Studies Overexpression of COX-2 in epithelial cells results in: Decreased apoptosis Angiogenesis (increased VEFG, FGF, PDGF… expression) Metastatic potential (increased adhesion and MMP expression)
Genetic Model Mice defective in COX-2 have a dramatic reduction (86%) in colorectal polyp formation.
Increased COX-2 = Increased Prostaglandins
COX-2-derived PGs promote cell growth, proliferation, and angiogenic gene expression in cells composing a tumor
Genetic “Removal” of
COX-2 Suppresses
Tumorigenesis
COX-2Genotype
COX-2 gene dosage effect on polyp number
COX-2
COX-2COX-2
Tot
al P
olyp
Num
ber/
Mou
se
(100%)
(34%)
(14%)
Involvement of COX-2 in the Progression of Colon Cancer
NormalEpithelium
CarcinomaEarly Adenoma Late Adenoma
APCmutation
TGFß IIRhMSH
K-rasmutation
p53mutation
- COX-2 +/- COX-2 ++ COX-2 +++ COX-2
Loss of COX-2 gene regulation
COX-2 overexpression
Increased PG levels
NSAIDs: A History of COX Inhibition• BC: Ancient Greeks and Romans used salicylate extracts derived from willow
leaves as analgesics and antipyretics.• Middle Ages: Medicinal herb gardens featured salicylate containing wintergreen
and meadowsweet plants.
• 1763: Edward Stone reported on use of willow bark powder as an anti-inflammatory.
• 1853: Von Gerhardt synthesizes a crude form of aspirin (acetylsalicyclic acid).
• 1860: Felix Hoffman working for Bayer synthesizes pure aspirin.
• 1949: The NSAID Phenylbutazone introduced.
• 1963: Indomethacin introduced.
• 1971: Vane and Piper demonstrated NSAIDs inhibit prostaglandin production.
• 1974: Ibuprofen introduced.
• 1976: Miyamoto et al purified the COX-1 enzyme.
• 1989: Simmons et al (and others) identified the COX-2 enzyme.
• 1999: The COXIBs Celebrex (celecoxib) and Vioxx (rofecoxib) introduced.
Aspirin and NSAIDs• NSAIDs have pain-relieving (analgesic) effects as well as effects of
reducing inflammation and fever.
• NSAIDs are non-selective COX-1 and COX-2 enzymatic inhibitors.
• Aspirin is an irreversible inhibitor of COX; Other NSAIDs are reversible.
• Aspirin protects against CV disease; NSAIDs are chemopreventive.
• Chronic NSAID use can lead to GI ulcers and renal problems.
Long-Term Aspirin and NSAID Use Reduces The Risk of Developing Colorectal Cancer
0 0.5 1 1.5 2
Relative Risk of Colorectal Cancer
Kune 1988
Rosenberg 1991
Suh 1993, men
Suh 1993, women
Peleg 1994
Muscat 1994, men
Muscat 1994, women
Muller 1994
Reeves 1996
Bansal 1996
LaVecchia 1997
Rosenberg 1998
Coogen 2000A
Coogen 2000B
Friedman 1998
adapted from Thun et al, JNCI, 2002
NSAIDs• Non Steroidal Anti-Inflammatory Drugs (NSAIDs) are the oldest and most widely used drugs in human history.
• NSAIDs have pain-relieving (analgesic) effects as well as effects of reducing inflammation and fever.
• NSAIDs are non-selective COX-1 and COX-2 enzymatic inhibitors.
• Chronic NSAID use reduces CRC risk ~50% and reduce polyp size and number.
• Extended NSAID use can lead to GI ulcers and renal problems.
Generic Name Brand Names
Made by several companies
Motrin®, Advil®, Motrin IB®
Naprosyn®, Aleve®
Relafen®
Aspirin
Ibuprofen
Naproxen
Nabumetone
COXIBs
• COX-2 inhibitors (COXIBs) are a special category of NSAIDs that target only COX-2 enzyme.
• Because they do not block the actions of the COX-1 enzyme, COXIBs generally have the side-effects of traditional NSAIDs.
• COXIBs reduce CRC polyp burden by 31% in colon cancer patients.
• COX-2 inhibitors also don’t offer the same kind of protection against heart disease.
Purpose-designed selective inhibitors of COX-2: the COXIBs.
Generic Name Brand Names
Celebrex®
Vioxx®
Celecoxib
Rofecoxib
Valdecoxib Bextra
Although rofecoxib (Vioxx) has been withdrawn, celecoxib (Celebrex) and valdecoxib (Bextra) remain on the US market.
Lumiracoxib and etoricoxib remain under consideration by the FDA.
Grosser, T. et al. J. Clin. Invest. 2006;116:4-15
Schematic depiction of the structural differences between the substrate-binding channels of COX-1 and COX-2 that allowed
the design of selective inhibitors
Study Drug Results
Arthritis StudiesVIGOR Vioxx
CLASS Celebrex
Vioxx associated w/higher risk of CV events
No difference in CV events
Colon Cancer StudiesAPPROVe
APC
PreSAP
Vioxx
Celebrex
Celebrex
2-fold increase risk of CV-events.Vioxx removed from market.
2.5-fold increase risk of CV-events.FDA recommends limited use of Celebrex and Bextra.
No difference in CV events
COXIBs are not without their problems…Summary of relevant trials examining CV events associated with extended COXIB use
The APPROVe study compared Vioxx vs. placebo in the prevention of colon cancer
Confirmed Cardiovascular Events with Extended Vioxx Use
After 18 months increases CV events were detected and led to removal of Vioxx from the market.