Lipid-Like Delivery Materials for Efficient siRNA Delivery - Nanotechnology for the Delivery of Therapeutic Nucleic Acids - page 162

HeLa cells was analyzed mathematically by a term that quantified
the difference between silencing with individual components and
their binary combination, the value. This value was defined such

that complete silencing achieved with a mixture of lipidoids that did
not silence on their own equaled 1, no difference between binary and
individual lipidoids equaled 0, and no silencing from binary mixtures
of lipidoids that completely silenced targets individually equaled
-1. Although many combinations did not improve targeting (

S

S = 0),

approximately 5% of the tested compounds had values greater

than 0.5, indicating synergistic delivery. Several combinations were
more effective

S

-5, silencing target gene expression
more than 75% at 20 nM. Binary potency often increased as the first
lipidoid:second lipidoid mass ratio approached 1:1 (the most evenly
mixed combination). Synergistic delivery was also seen

in vitro than 98N 12

. Two
components that silenced 20% when used independently reduced
F7 expression by 85% when combined at a relative mass ratio of
3:1.

in vivo

7.7 Next-Generation: Identifying Improved

Carriers Using Innovative Chemistry

Through a series of iterative studies, first-generation lipidoids
were optimized so that 50% reduction of a hepatic target gene
was achieved when siRNA was injected intravenously at 1 mg/kg.
Lipidoid synthesis revealed that new lipid delivery agents could be
produced without time-consuming and expensive chemical
synthesis. Utilizing three times as much lipid carrier, stable
nucleic acid lipid particles (SNALP) formulations were also shown
to silence hepatic targets in non-human primates at the same
dose.

5

Consequently, members of our lab sought to identify new
compounds that would facilitate delivery at doses less than 1 mg/
kg. The original lipoid library demonstrated that fully saturated
alkyl tails were more effective than non-alkyl tails and that lipids
attached to amines through stable amide bonds were more effective
than those connected by ester linkages.

13

To this end, members
of our lab developed a second-generation lipidoid library based
on stable chemical bonds between fully saturated alkyl tails and
amines generated by an epoxide ring-opening reaction

20

(Fig. 7.3).
Alkyl tail length was varied between 9 and 18 carbons, and amines
were selected if they were effective in previous studies or if they