(57a) Polymeric Mechanical Amplifiers of Tumor Cell Mechanotransduction and Cell Death | AIChE

(57a) Polymeric Mechanical Amplifiers of Tumor Cell Mechanotransduction and Cell Death

Authors 

Mitchell, M. J. - Presenter, Massachusetts Institute of Technology
Langer, R. - Presenter, Massachusetts Institute of Technology

Polymeric Mechanical Amplifiers of Tumor Cell Mechanotransduction and
Cell Death

Michael J. Mitchell1, Robert Langer1

1Chemical Engineering and David H. Koch Institute for
Integrative Cancer Research, MIT, Cambridge MA 02139



Introduction: It has become evident that tumor cells are
responsive to mechanical forces in vivo,
and prove critical to tumor cell proliferation and death. Recent work has shown
that tumor cells exposed to fluid shear forces increase receptor-mediated
signaling. We hypothesized that biocompatible, polymeric micro- and nanoparticles
conjugated to the tumor cell surface via free amine coupling act as mechanical
amplifiers in presence of fluid shear forces to increase mechanotransduction,
and can be exploited to increase the efficacy of therapeutic ligands.

Materials and Methods: Polymeric particles ranging from 100 nm–6 μm
in size were conjugated to free amines on tumor cells via NHS crosslinker chemistry (Fig.
1A
). Nondegradable polystyrene and degradable (PLGA, PCL) particles bound
to tumor cells were assessed using flow cytometry, brightfield,
and confocal fluorescence microscopy. A cone-and-plate viscometer was used to
apply a fluid shear force (2.0 dyn/cm2) to tumor cell suspensions
and to amplify the force exerted by polymeric particles on tumor cells (Fig.1B). Tumor cells (COLO 205, PC-3)
were treated with 1 μg/mL of a TNF-related apoptosis-inducing ligand
(TRAIL) to assess amplified mechanotransduction and receptor-mediated apoptosis
in the presence of polymeric particles. An annexin-V
apoptosis assay was used to characterize the mode of cell death. Caspase colorimetric assays and inhibitors (Z-VAD-FMK) were
utilized to assess caspase-dependence in the mechanotransduction
response.

Figure 1: (A)
Conjugation of polymer amplifiers to tumor cell surface via NHS crosslinker chemistry. (B) Polymeric particles amplify force exerted on the tumor cell
surface under fluid forces to increase mechanotransduction and therapeutic
efficacy. (C) Confocal and brightfield images of 1 μm polymeric particles
conjugated to the surface of tumor cells. Scale bars: 10 μm. (D) Particle-functionalized tumor cell
viability after TRAIL treatment in the presence of shear force.*P<0.05**P< 0.01. NS: not significant. (E,F) Annexin-V
apoptosis assays of non-functionalized and functionalized tumor cells treated
with TRAIL in the presence of shear forces. Lower left/right and upper
left/right quadrants denote viable, early apoptotic, late apoptotic, and
necrotic cells, respectively.  

Results and Discussion: NHS crosslinker chemistry was successfully used to conjugate
polymeric particles to the tumor cell surface (Fig. 1C). In the presence
of fluid shear forces, it was found that polymeric particles act to mechanical
amplify tumor cell mechanotransduction, as evidenced by increased
receptor-mediated apoptosis and decreased tumor cell viability in the presence
of the therapeutic ligand TRAIL (Fig. 1D). Additionally, amplification
of TRAIL-mediated apoptosis was increased with particles of increasing size (Fig.
1D
), demonstrating that increasing the force exerted on the cell surface
with larger particles amplified the therapeutic response. Annexin-V apoptosis
assays showed the addition of conjugated polymeric particles to the cell
surfaces nearly doubled tumor cell apoptosis in the presence of TRAIL under
shear forces (Fig.1E,F), and inhibition assays
revealed the response to be caspase-dependent
apoptosis.

Conclusions: 
These
data demonstrate that polymeric particles, both degradable and non-degradable,
act as mechanical amplifiers of tumor mechanotransduction in the presence of
shear forces, and are exploited to increase therapeutic efficacy. Clinically,
this approach shows that increased mechanical force applied to target tumor
cells can increase sensitivity to therapeutic ligands.

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