DƯỢC LÍ Goodman & Gilman's The Pharmacological Basis of Therapeutics 12th, 2010

nucleotide-binding domains of PKG are expressed as a
single polypeptide, which dimerizes to form the PKG
holoenzyme.
PKG exists in two homologous forms, PKG-I and PKG-II.
PKG-I has an acetylated N terminus, is associated with the cytoplasm
and has two isoforms (Iα and Iβ) that arise from alternate
splicing. PKG-II has a myristylated N terminus, is membraneassociated
and can be localized by PKG-anchoring proteins in a
manner analogous to that known for PKA, although the docking
domains of PKA and PKG are very different structurally.
Pharmacologically important effects of elevated cyclic GMP include
modulation of platelet activation and relaxation of smooth muscle
(Rybalkin et al., 2003).
PDEs. Cyclic nucleotide phosphodiesterases form
another family of important signaling proteins whose
activities are regulated via the rate of gene transcription
as well as by second messengers (cyclic nucleotides or
Ca 2+ ) and interactions with other signaling proteins
such as β arrestin and protein kinases. PDEs hydrolyze
the cyclic 3′,5′-phosphodiester bond in cAMP and
cGMP, thereby terminating their action.
The enzymes comprise a superfamily with >50 different PDE
proteins divided into 11 subfamilies on the basis of amino acid
sequence, substrate specificity, pharmacological properties, and
allosteric regulation (Conti and Beavo, 2007). PDEs share a conserved
catalytic domain at the carboxyl terminus, as well as regulatory
domains and targeting domains that localize a given enzyme to a specific
cellular compartment. The substrate specificities of the different
PDEs include those specific for cAMP hydrolysis, cGMP hydrolysis,
and some that hydrolyze both cyclic nucleotides. PDEs (mainly PDE3
forms) are drug targets for treatment of diseases such as asthma, cardiovascular
diseases such as heart failure, atherosclerotic coronary
and peripheral arterial disease, and neurological disorders. PDE5
inhibitors (e.g., sildenafil) are used in treating chronic obstructive pulmonary
disease and erectile dysfunction. By inhibiting PDE5, these
drugs increase accumulation of cellular cGMP in the smooth muscle
of the corpus caverosum, thereby enhancing its relaxation and
improving its capacity for engorgement (Mehats et al., 2002).
Other Second Messengers
Ca 2+ . Calcium is an important messenger in all cells and
can regulate diverse responses including gene expression,
contraction, secretion, metabolism, and electrical
activity. Ca 2+ can enter the cell through Ca 2+ channels
in the plasma membrane (See “Ion Channels”, below)
or be released by hormones or growth factors from
intracellular stores. In keeping with its role as a signal,
the basal Ca 2+ level in cells is maintained in the 100 n
range by membrane Ca 2+ pumps that extrude Ca 2+ to
the extracellular space and a sarcoplasmic reticulum
(SR) Ca 2+ -ATPase (SERCA) in the membrane of the
endoplasmic reticulum (ER) that accumulates Ca 2+ into
its storage site in the ER/SR.
Hormones and growth factors release Ca 2+ from
its intracellular storage site, the ER, via a signaling
pathway that begins with activation of phospholipase C
at the plasma membrane, of which there are two primary
forms, PLCβ and PLCγ. GPCRs that couple to G q
or G i
activate PLCβ by activating the G protein α subunit
(Figure 3–8) and releasing the βγ dimer. Both the
active, G q
-GTP bound α subunit and the βγ dimer can
activate certain isoforms of PLCβ. PLCγ isoforms are
activated by tyrosine phosphorylation, including phosphorylation
by receptor and non-receptor tyrosine
kinases. For example, growth factor receptors such as
the epidermal growth factor receptor (EGFR) are receptor
tyrosine kinases (RTKs) that autophosphorylate on
tyrosine residues upon binding their cognate growth
factor. The phosphotyrosine formed on the cytoplasmic
domain of the RTK is a binding site for signaling proteins
that contain SH2 domains, such as PLCγ. Once
recruited to the RTK’s SH2 domain, PLCγ is phosphorylated/activated
by the RTK.
PLCs are cytosolic enzymes that translocate to the plasma
membrane upon receptor stimulation. When activated, they
hydrolyze a minor membrane phospholipid, phosphatidylinositol-4,
5-bisphosphate, to generate two intracellular signals, inositol-1,4,5-
trisphosphate (IP 3
) and the lipid, diacylglycerol (DAG). Both of these
molecules lead to signaling events by activating families of protein
kinases. DAG directly activates members of the protein kinase C
(PKC) family. IP 3
diffuses to the ER where it activates the IP 3
receptor
in the ER membrane causing release of stored Ca 2+ from the ER.
Release of Ca 2+ from these intracellular stores raises Ca 2+ levels in
the cytoplasm many fold within seconds and activates calmodulinsensitive
enzymes such as cyclic AMP PDE and a family of
Ca 2+ /calmodulin-sensitive protein kinases (e.g., phosphorylase
kinase, myosin light chain kinase, and CaM kinases II and IV)
(Hudmon and Schulman, 2002). Depending on the cell’s differentiated
function, the Ca 2+ /calmodulin kinases and PKC may regulate
the bulk of the downstream events in the activated cells. For example,
release of the sympathetic transmitter norepinephrine onto vascular
smooth muscle cells stimulates α adrenergic receptors,
activates the G q
-PLC-IP 3
pathway, triggers the release of Ca 2+ , and
leads to contraction by stimulating the Ca 2+ /calmodulin-sensitive
myosin light chain kinase to phosphorylate the regulatory subunit of
the contractile protein, myosin.
The IP 3
-stimulated release of Ca 2+ from the ER, its reuptake,
and the refilling of the ER pool of Ca 2+ are regulated by a novel set
of Ca 2+ channels. The IP 3
receptor is a ligand-gated Ca 2+ channel
found in high concentrations in the membrane of the ER (Patterson
et al., 2004). It is a large protein of ~2700 amino acids with three
major domains. The N-terminal region contains the IP3-binding
site, and the middle region contains a regulatory domain that can
be phosphorylated by a number of protein kinases including PKA
and PKG. The C-terminal region contains six membrane-spanning
helices that form the Ca 2+ pore. The functional channel is formed by
four subunits arranged as a tetramer. In addition to IP 3
, which
57
CHAPTER 3
PHARMACODYNAMICS: MOLECULAR MECHANISMS OF DRUG ACTION