Agonism at melanocortin receptor type 3 on murine macrophages inhibit neutrophil influx

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MELANOCORTIN PHARMACOLOGY
Dr Stephen J. Getting
Department of Biochemical Pharmacology William
Harvey Research Institute Charterhouse
Square Barts the London Queen Mary, University
of London
s.j.getting_at_qmul.ac.uk
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• Melanocortin
• Learning Objectives
• POMC Gene
• Melanocortin receptors
• Physiological/Pharmacological actions

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POMC gene and ACTH-derived fragments
POMC Product
-
lipotropin
b
ACTH
1-39
g
MSH
-
g
-lipotropin
ACTH
1-13

(
a-MSH)
CLIP
b- MSH
-endorphin
b
ACTH
4-10
(MEHFRWG)
ACTH
4-10
(MEHFRWG)
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Features of the POMC gene
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Amino acid sequence for peptide fragments
SYSMEHFRWGKPVKKRRPVKVYPNGAEDESA
EAFPLEF
ACTH
ACTH4-10
MEHFRWG
a-MSH
SYSMEHFRWGKPV
ß-MSH
AEKKDEGPYRMEHFRWGSPPKD
g-MSH
YVMGHFRWDRFG
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Melanocortin Receptors (MC-Rs)

• Seven transmembrane GPCRs positively coupled to
  adenylate cyclase
• Activation leads to elevation of cAMP
• Diverse physiological role by melanocortins are
  mediated via different receptors.
• To date five MC-Rs have been identified and
  cloned and are termed as MC1-5R
• MC-Rs share a sequence homology of 40 -
  60

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MC1-R-MELANOCORTIN TYPE 1 RECEPTOR
Agonist profile a-MSH gt ACTH gt g-MSH

• Identified in 1992 in Melanoma tumours
• 1996 in testis, pituitary
• 1995-1997 in cell lines of
  different lineage (macrophages, monocytes,
  endothelial, fibroblasts, Keratinocytes)
• External receptor expressed on the cellular
  cell-surface although reports of internalisation
  upon exposure to peptides.

RECEPTOR INVOVLVED IN SKIN PIGMENTATION
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MC1-R involved in skin pigmentation.
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MC2-R-MELANOCORTIN TYPE 2 RECEPTOR
Agonist profile ACTH
Expressed on the adrenal cortex (Zona fasiculata)
and adipose tissue
RECEPTOR INVOLVED IN STEROIDGENESIS
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Circadian Variation in plasma cortisol
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Endogenous corticosteroids
11b hydroxysteroid dehydrogenase
OH
1
0.5
Cortisol
Cortisone
Glucocorticoid activity
Synthetic corticosteroids
F
CH3
6
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Methylprednisolone
Dexamethasone
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Philip S Hench 1896-1965
Edward C Kendall 1886-1972
Discovered the clinical anti-inflammatory effects
of steroids
Identified, isolated, purified and synthesised
corticosteroids
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Corticosteroids are widely used in the treatment
of inflammatory and autoimmune diseases.
over 250 000 people in the United Kingdom are
taking continuous oral steroids
(Walsh LJ et al. BMJ 1996313,344-6)
Prednisone 20mg tabs
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Glucocorticoids exert effects on multiple organs
and metabolic pathways
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Unwanted effects of systemic corticosteroids
Oedema Weight gain Myopathy Glycosuria Hypertensi
on Hyperlipidaemia Thrombosis GI bleeding Peptic
ulcers Pancreatitis Colonic perforation
Glaucoma Cataracts Insomnia Depression /
Psychosis HPA axis suppression Increased
infection risk Osteoporosis Avascular / aseptic
necrosis Juvenile growth retardation
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Corticosteroid-induced peptic ulceration
Steroid -associated peptic ulcers of the stomach
and oesophagus (left) are usually small,
superficial and multiple.
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Corticosteroid-induced glaucoma
Photo of normal optic nerve on left and optic
nerve with glaucoma damage on the right (note the
central white cupping or depression representing
nerve fiber  loss)
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response to stress
stressor
physiological 'tone'
Glucocorticoid hormones prevent overzealous
rebound responses to stress.
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Cushings Syndrome

• Caused by too much ACTH which results in
  overproduction of cortisol
• Caused by benign pituitary adenoma (70 of
  endogenous cases)
• Lung tumours (15 of endogenous cases)
• Benign and malignant adrenal tumours (15 of
  endogenous cases)

The most common cause is exogenous ingestion of
glucorticoids.
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Symptoms of Cushing's syndrome Hypertension Ather
osclerosis Congestive heart failure Oedema Menstru
al irregularities Psychological
disturbances Osteoporosis Increased infections
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The classical moon face associated with
corticosteroid use
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Buffalo hump in a 34yr old woman taking
corticosteroids
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MC3-R-MELANOCORTIN TYPE 3 RECEPTOR
Agonist profile g-MSH ACTH gt a-MSH

• Identified in 1993 in Brain, Gut and Placenta
• 1996 in Heart
• 1999 in leucocytes (macrophages)
• Found to have a wide distribution within the
  brain, including the Hippocampus, thalamus and
  midbrain.
• It has been reported that it may function as an
  auto-receptor, regulating the release of MSH
  peptides from POMC neurones.

Receptor proposed to be involved in modulating
inflammation
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MC4-R-MELANOCORTIN TYPE 4 RECEPTOR
Agonist profile a-MSH ACTH gt g-MSH

• Expressed in multiple sites within the brain
  (cotex, thalamus, hypothalamus, brain stem)
• Not detected in any other organ in human,
  although one study has proposed message on
  adipose tissue.
• Distribution much wider than MC3-R

RECEPTOR INVOLVED IN CONTROLLING FOOD INTAKE
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MC5-R-MELANOCORTIN TYPE 5 RECEPTOR
Agonist profile a-MSH gt ACTH gt g-MSH

• MC5-R is ubiquitously expressed in many
  peripheral tissues including adrenal glands, fat
  cells, kidneys, lung , liver etc
• It has been proposed that the seborrhoea observed
  in Parkinsons
• Disease has been attributed to an increased
  activity of MSH and that receptor blocking might
  lead to a treatment.

RECEPTOR INVOLVED IN SEBORRHOEA
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Cardinal signs of inflammation
The accumulation and subsequent activation of
leukocytes are central events in the pathogenesis
of virtually all forms of inflammation
Inflammation Basic Principles and Clinical
Correlates 2nd Ed. Gallin JI, Goldstein IM and
Snyderman R, 1992
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detachment

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Experimental Murine Colitis
                                                
 

• Reduces Faecal Blood
• Inhibits weight loss
• Reduces TNF-a and Nitric Oxide
• In lower colon

Rajora, N. et al., Peptides 1997, 18, 381-385
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Endotoxin-induced intestinal inflammation

• reducing neutrophil migration
• reduce severity of the lesions macroscopically
  and microscopically in the distal ileum
• San, T. et al., Peptides 2001, 22(12),
  2077-2082.

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Mesenteric I/R Injury

• a-MSH reduced depressed intestinal transit
• reduced myeloperoxidase activity in ileal
• cytoplasmic extracts
• reduced NF-kB activation in ileal nuclear
• extracts following I/R
• Hassoun, H.T. et al., Am. J. Physiol. 2002,
  282, G1059-1068

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Lung Inflammation

• a-MSH inhibits eosinophil migration
• and mucus secretion
• (Rapp et al., J. Immunol, 2003, 171(1) 353-359.)

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The Gout by James Gilray, 1799. Gout depicted
as an evil demon attacking a toe.
By Royal Authority by George Cruickshank. A
gout sufferer helped onto his horse.
2.2 million cases of gout in the USA equivalent
to 160 American males 37 million working days
are lost in the USA every year to gout.
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GOUT-CLINICAL PICTURES
After years of gouty attacks, patients develop a
chronic arthritis resulting in bone and cartilage
destruction and deformity. Uric acid crystals
deposit within and surrounding the joint causing
a chronic destructive inflammatory process.
X-rays characteristically show punched out
erosions.
X-ray showing soft tissue swelling and erosion
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ACTH is effective in the clinical treatment of
gout with an efficacy over and above that
obtainable with conventional glucocorticoid
therapy Ritter et al., J. Rheumatol, 1994
696-699
Must be another mechanism Action besides reflex
stimulation Of the HPA axis
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in vivo Inflammation
Time (h)
-0.5 0 2
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Agonists and Antagonists
PMN accumulation
MSU crystals (3 mg in 0.5 ml PBS)
Chemokine levels by ELISAs

Getting et al., 1999 J. Immunol, 162 7446-7453
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MELANOCORTIN AGONISTS INHIBIT PMN RECRUITMENT
EFFECT BLOCKED BY MC3-R ANTAGONISTS
Getting et al., 1999 J. Immunol, 162
7446-7453
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MC3-R mRNA AND FUNCTIONALITY
Getting et al., 1999 J. Immunol, 162
7446-7453
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Injection of MSU crystals (1 mg in 50 ml)
2-72 h knee joints were examined and inflammatory
exudates collected and analysed for 1
Arthritic Score 2 Size of knee joint 3
PMN accumulation by light microscopy 4
Cytokine contents by specific ELISAs
Getting et al., 2002 Arthritis Rheumatism,
46 2765-2775
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MC3-R AGONISTS INHIBIT JOINT INFLAMMATIONEFFECT
BLOCKED BY ANTAGONIST
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REPEATED ADMINISTRATION OF MSU CRYSTALS
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Lipton et al News physiol. Sci, 15,192-195.
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Diverse Physiological Roles Mediated By MC-Rs

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Effects of melanocortins
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