MicroRNAs as Engineering Targets: Pathway Manipulation to Impact Bioprocess Phenotypes - MicroRNAs as Tools in Biopharmaceutical Production - page 77

of IL-1 transduction pathway and reduced the secretion of IL-6 and IL-8, thereby
participating in the negative feedback loop which suppresses the imbalance of the
senescence -associated secretory phenotype activity (Bhaumik et al. 2009 ).
The engineering of protein secretory pathway via manipulation of miRNA levels
is feasible for industrial scale applications in CHO cell cultures, especially consid-
ering that homologs for miR-9 and miR-146a/b have been reported in CHO cells
(Hackl et al. 2011 ; Johnson et al. 2011 ). This research is in its early stages and
more studies are needed to identify the effects of manipulating specific genes on
the recombinant protein secretion rates, and to identify the CHO-specific miRNAs
involved in targeting those genes.

5.3 Global Modification of MicroRNA Expression to Improve

CHO Cells Bioprocesses

Global modification of miRNA expression levels is another strategy to improve CHO
cell performance. The fact that mature miRNA expression levels in tumors are usu-
ally lower than their levels in healthy tissues (Lu et al. 2005 ), and over-expression of
miRNAs frequently prevents transformation of cells to cancer-like phenotype, indi-
cates their possible anti-oncogenic role (Imam et al. 2010 ; Lee et al. 2010 ). Therefore,
the global down-regulation of miRNA expression may improve proliferative capac-
ity of the cells and eliminate the stress response activation pathways (Barron et al.
2011a ).
Modification of miRNA expression levels by genetic or epigenetic means, and
by varying the levels of the miRNA biosynthesis enzymes and degradation factors,
should be considered for CHO cell engineering (Barron et al. 2011a ). An example
of genetic regulation of miRNA profiles is the effect of the oncogenic transcription
factor, c-Myc, which down-regulates miRNA expression levels by direct association
with promoters of pri-microRNAs (Chang et al. 2008 ). Another study showed the
opposite effect, where c-Myc activates the pro-oncogenic miR 17-92 cluster by direct
binding to this cluster's promoter (O'Donnell et al. 2005 ). Other transcription factors,
such as Hif-1alpha, NF-

B and p53 were also shown to affect expression of miRNA
clusters by interaction with their promoters (Sun et al. 2010 ).
Epigenetic regulation of a single miRNA or an miRNA cluster can be initiated
by DNA methylation and histone modifications; it was shown by the activation of
epigenetically silenced miR-127 in human cancer cell lines (Saito et al. 2006 ). It is
also possible to alter miRNA biogenesis by knocking down the miRNA processing
enzymes Dicer and Drosha, or by over-expression of degradation factors such as
XRN2 (Martello et al. 2010 ).
The global modification of miRNA expression needs to be carefully investigated
to avoid potential harmful effects on cell homeostasis. For example, even though
knockout of Dicer and Drosha can be implemented to globally down-regulate miRNA
expression, it can negatively affect cell-cycle progression and mitosis in some cell
lines (Martello et al. 2010 ). More research needs to be conducted to clarify the effects
of global miRNA alterations on cellular functions in CHO cells.

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