Bi/CNS 150
Lecture 29.5
Friday, December 7, 2012
“Inside-out” Actions for Psychiatric Drugs
Henry Lester
1
How do psychiatric drugs work?
1. Statement of the problem:
Antispychotics, Antidepressants, Bipolar drugs
2. Lessons from nAChRS;
3. Pharmacokinetics
4. Detailed hypotheses:
Antipsychotic drugs
SSRI Antidepressant drugs
“Fast” NMDA blocker antidepressants
5. Tests of “inside-out” mechanisms for psychiatric drugs
Psychiatric drugs bind to classical targets within early exocytotic pathways:
Therapeutic effects
Biological Psychiatry, Dec 2012
Henry A. Lester, Julie M. Miwa, and Rahul Srinivasan
2
Eroom’s law applies especially to neural drugs
Scannell, Nature Revs Drug Disc. 2012
3
Contemporary ideas about psychiatric drugs
have emphasized binding to
the classical targets at synapses. . .
“Inside-out” mechanisms emphasize binding to
the same classical targets, but within the
endoplasmic reticulum and cis-Golgi
Some psychiatric drugs, their targets,
logP values, and half lives
antipschizophrenic
recreational /
abused / addictive
chlorpromazine
(Thorazine)
dopamine D2 receptor, GPCR
logP 5.2, 16-30 hr
antidepressant
ketamine
(“special K”)
NMDA glutamate receptor
logP 2.2, 3-5 hr
nicotine
acetylcholine receptor
logP 1.2, 0.5 -2 hr
clozapine (Clozaril)
5-HT2A serotonin receptor, GPCR
logP 3.2, 8-12 hr
fluoxetine
(Prozac)
serotonin transporter
logP 3.4, 24-72 hr
logP = log (solubility in octanol / water)
5
Like most drugs, nicotine is a weak base.
Its neutral form passes through 6 plasma membranes in ~ 20 s
Alveolar epithelium
Brain capillary
Endothelial
cells
Lungs
Astrocyte
End-feet
Blood
CSF
H+
logP = 1.1 = log (solubility in octanol / water)
6
“Inside-out” Drug Action by Nicotine at α4β2 nAChRs
Na+
Classical Pathway:
Channel activation
& desensitization
nAChR
Plasma
membrane
Ca2+
Clathrin
Secretory
vesicle
Early
endosome
Golgi
COPI
Nicotine in CSF
ATF6
Lysosome
Golgi
complex
COPII vesicle
Sec 13/31
COPI
ATF6
Pharmacological
Chaperoning→
upregulation
COPII
Sec24
Sec23
Endoplasmic
reticulum
Sar1
PERK
IRE1 nAChR
M3-M4 loop
Unfolded protein response
→ Do neurons survive
Despite stressors?
ATF4
eIF2α
+
XBP1
H+
UPRE
BiP
ER
Nucleus
IRE1
PERK
7
Three possible results of nicotine-nAChR binding in the endoplasmic reticulum
1. Agonist binding eventually favors stable,
high-affinity states (a “chaperone”)
unbound
agonist
2. Nicotine binding at subunit interface favors
assembled nAChRs (a “matchmaker”)
Bound states with
increasing affinity
nicotine
20 sec
Free Energy
106
channels
“closed”
AC
“activated”
Highest
affinity
?
“desensitized”
Reaction Coordinate
3. Nicotine may displace lynx, directing nAChRs toward cholesterol-poor domains (an “escort”)
nicotine
lynx
8
The three arms of the ER stress / unfolded protein response pathway
R. L. Wiseman, C. M. Haynes, D. Ron Cell 2010
9
Inside-out Pharmacology of Nicotine Effects at α4β2 nAChRs
During chronic exposure to nicotine, α4β2 nAChRs are selectively upregulated.
Now we’re assessing gene expression in identified neurons chronically
exposed to nicotine.
Pharmacological chaperoning is necessary but not sufficient for upregulation.
Upregulation proceeds similarly in clonal cells, rodent brains, and smokers’ brains.
Other sequelae of chaperoning: changed stoichiometry, reduced ER stress and
reduced UPR.
Inside-out pharmacology is a powerful concept for nearly all CNS drugs:
They are all membrane-permeant weak bases.
10
The discovery criteria for psychiatric drugs lead to excellent intracellular chaperoning
1. High bioavailability implies high membrane permeation
All psychiatric drugs have logP > 2
2. Good stability in the body implies simple or little enzymatic breakdown.
Half-life is ~ 1 day.
3. Good selectivity, few off-target effects imply high-affinity binding to the target
Kd < 1 μM, often ~ 10 nM
a. “Chaperoning”:
(i) Transporter ligands are
organic substrates
ions,
or antagonists,
They favor two major binding states, “inward” vs “outward”.
(ii) GPCR ligands (see next slide):
agonists
antagonists
allosteric modulators
“inverse” agonists
b. “Matchmaking”:
(i) Neurotransmitter transporters must homodimerize before leaving the ER
(ii) GPCRs homo- and heterodimerize,
in some cases required for ER export,
in some cases favored by ligands
11
Pharmacological chaperoning of GPCRs
receptor
mutant /WT
drug class
reference
adenosine A1
mutant
agonists; antagonists
(Malaga-Dieguez et al., 2010)
dopamine D4
both
transported dopamine;
quinpirole; antagonists
gonadotropinreleasing hormone
mutant
antagonists
(Van Craenenbroeck et al.,
2005)
(Conn and Ulloa-Aguirre,
2011)
histamine H2
both
agonist, inverse agonist
(Alewijnse et al., 2000)
opsin
mutant
--
(Noorwez et al., 2008)
δ-opioid
mutant
antagonist
(Leskela et al., 2012)
μ-opioid
mutant
agonists, antagonists
(Chaipatikul et al., 2003)
mutant
antagonist
(Fan et al., 2005)
both
both
both
both
antagonist, inverse agonist
antagonist
antagonist
antagonists
(Tao, 2010)
(Hawtin, 2006)
(Robert et al., 2005)
(Wuller et al., 2004)
melanin conc.
hormone
melanocortin-4
vasopressin V1a
vasopressin V1b/V3
vasopressin V2
12
Two mechanisms for gene activation downstream from antipsychotic drugs
Most papers suggest . . .
βarrestin
Intracellular
messenger
bg
Enzyme
or
channel
ATF6
Drug+
in CSF
Golgi
H+
Neutral
permeant
drug
ATF6,
CREBH
Endoplasmic
reticulum
kinase
+
+
cascade
IRE1
PERK
+
a
Golgi
+
ATF4
p-eIF2α
Transcription
factors
XBP1
Nucleus
Transcription
factors
UPRE
+
+
+
In CSF
+
Drug+
B. Intracellular pharmacological
chaperoning of GPCR,
and downstream effects
+
A. Inhibition of plasma membrane GPCR ,
and downstream effects
We suggest . . .
H+
ER
BiP
IRE1
PERK
Nucleus
“Nearly” cell-autonomous actions of SSRI antidepressant treatment
Kellermann group
14
Adult Neurogenesis
Inside-out actions would occur here
Other diagrams
Samuels & Hen, Eur J. Neurosci, 2011
15
Gene activation is too brief to account for the “therapeutic lag”
Axonal transport provides a natural delay in the
“inside-out” mechanism.
Speed: ~ 1 mm / day.
Mouse hippocampus
Suggests that equivalent effects would require
briefer delays in animals with shorter axons
Marks et al, 1985
Days of nicotine infusion
Dendritically localized events
16
How does acute ketamine produce antidepressant effects within 2 hr?
(1) involve BDNF synthesis & release,
(3) require protein synthesis,
Monteggia & Duman groups suggest . . .
The effects
(2) occur in the dendrites,
(4) do not require gene activation.
We suggest . . .
Outside-in
NMDA Receptor
Inside-out
Ca2+
BDNF
secretion
BDNF
secretion
Decreased
Ca2+ flux
Dendritic
Golgi
Escorting
kinases↓
COPII
+
+
pPERK↓
+
+
BDNF↑
+
Dendritic
ER
BDNF↑
p-eIF2α↓
BDNF
mRNA
BDNF
mRNA
NMDA Receptor
+
H+
ER
BiP
IRE1
PERK
17
“Acid trapping” of nicotine might
1. keep nAChRs
desensitized until
they are exocytosed;
2. serve as a
reservoir for nicotine
nAChR
Cell
membrane
pH
nic+
nicCSF
5.2
100
6.0
30
6.3
20
6.5
10
6.7
3
7.2
1
7.2
1
Clathrin
Secretory
vesicle
Early
endosome
COPI
Lysosome
Golgi
complex
Nicotine in CSF
COPI
COPII
Endoplasmic
reticulum
nAChR
&
See detailed calculations
for antipsychotics:
Tischbirek et al,
Neuron 2012
What knowledge do we need next?
As usual, we need cell biology & biochemistry
1. Reconstituted, cell-free systems for ER exit and retrieval
2. Better real-time markers for compartmentalized receptors and transporters
a. Imaging mass spectrometry
b. Plasma membrane binding only? Possible with impermeant derivatives
c. ER binding only? More challenging, especially for antagonists.
3. Better measurements of pathway-specific gene activation (RNA-Seq)
4. Analyze newly synthesized proteins
19
Three concepts used in describing complex diseases such a schizophrenia
Polygenic
the disease occurs only if several genotypes are present together
Genetically Multifactorial
several distinct genes (or sets of genotypes) can independently cause the disease
Partially penetrant
nongenetic or epigenetic factors are required, or the disease is inherently stochastic
Genetically
Multifactorial
Polygenic
Partially
Penetrant
20
Contemporary ideas about psychiatric diseases
have emphasized synaptic and signaling deficits . . .
“Inside-out” mechanisms emphasize that
~30% of a cell’s proteins enter the ER,
and additional nuclear and cytoplasmic proteins
control their synthesis & trafficking.
GABAergic “chandelier cell”
in human cerebral cortex has
many large axon terminals . . .
. . . and plentiful somatic ER
Ch
axon
Ch
Chterminals
terminals
Pyramidal
Cells
~ 100 μm
DeFelipe, Brain (1999) 122, 1807 (Cajal Institute, Madrid)
Jones, J. Comp. Neurol., 1984 22
Bi/CNS 150
End of Lecture 29.5
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