Biomedical Engineering Reference
In-Depth Information
7 Which Genes Should Be Delivered to Create a Biological
Pacemaker?
Classical pharmacology is not only about the active substance, but also about the
formulation as a whole. This is not different in gene therapy. It is necessary to select
the most appropriate viral vector system produced at the best quality as discussed
above, but, within this vector, the specificity of transcription is of equal importance.
Instead of a constitutive promoter which has been used in most of the previous gene
therapy approaches, conditional gene expression may be preferred. Cardiac specific
promoters will be useful for cell-type-specific expression [16]. Some cardiac
promoters are well described, e.g., the myosin light chain 2v (MLC-2v) promoter
(ventricular specific) [14, 32, 45], the Į-myosin heavy chain promoter (atrial and
ventricular specific) [14], and the atrial natriuretic factor (ANF) promoter (atrial
specific) [49]. However, at present, constitutive promoters are suitable for
experimental purposes.
About the active substance, the therapeutical gene, we can be more specific. In our
opinion, HCN channels are the most promising candidates to deliver. This is because
they incorporate autonomic modulation by direct allosteric interaction with cAMP
and they are only activated upon hyperpolarization. The latter results in channel
inactivity during the action potential plateau phase. The knock-down of the inward
rectifier potassium current, in contrast, is known to prolong action potentials and may
cause excess prolongation of repolarization resulting in torsade de pointes, while
oscillations in resting membrane potential may also constitute inappropriate
membrane depolarizations and cardiac arrhythmias.
If the delivery of a HCN channel is considered, HCN2 and HCN4 are primary
candidates. HCN4 is highly expressed in the SA-node and a mutation in this channel
is associated with familial sinus bradycardia [29]. Activation of HCN2, on the other
hand, may have the advantage of being much faster (i.e., having a shorter response
time) [44]. Single channel analysis has shown that
I
f
in human atrial myocytes closely
resemble characteristics of HCN4 or HCN2 + HCN4 [28].
A bio-engineered designer channel could be the ultimate transgene [1] and some
interesting results in this research area are now becoming available. The group of
Marbán described the conversion of a depolarization-activated K-channel into a
hyperpolarization-activated channel, which also became permeable to Na. With
multiple mutations, this yielded a mutant channel with a pacemaker-current
phenotype [17]. The group of Rosen described improved pacemaker function using
the E324A point mutation in the S4-S5 linker (linker of the fourth and fifth
transmembrane segments) of murine HCN2 (mHCN2). This mHCN2-E324A channel
induces faster and more positive pacemaker current activation compared to normal
mHCN2. This bioengineered channel was already tested in vivo, when it was applied
in the left bundle branch of four dogs using Ad vectors. Treated dogs showed faster
rhythms with stronger catecholamine sensitivity than mHCN2 and controls [5]. If
these mutations eventually lead to precise rate modulation, issues on the regulation of
gene expression might be irrelevant.
