Aptamer-targeted inhibition of mTOR in T cells enhances antitumor immunity Alexey Berezhnoy, Iris Castro, Agata Levay, Thomas R. Malek, and Eli Gilboa J Clin Invest. 2014;124(1):188–197
Background (siRNA) siRNA (small interfering RNA) - ability to inhibit target genes Aptamer - oligonucleic acid molecules that bind to a target molecule. Aptamer-siRNA conjugate - delivery vehicle composed of a targeting part (aptamer) and a therapeutic part (siRNA).
Background (T Cells) Memory T cells – T cells that have encountered and responded to their specific antigen. They can recognize invaders and at a 2 nd encounter can reproduce to mount a faster and stronger immune response than the 1 st time.
Background Elevated levels of mTOR promote the accumulation of short-lived effectors rather than memory cells. Pharmacological agents can have undesirable effects due to their broad range
Aim of the study Aptamer-targeted siRNA inhibition of mTOR in CD8+ T cells enhances a vaccine-induced memory and an antitumor immunity that is superior to pharmacological treatment.
Methods siRNA specific to raptor (a component of mTORC1) Aptamer that binds to 4-1BB, a molecule that is expressed on CD8+ T cells Test if siRNA inhibits mTORC1 activity. Test if mTOR inhibition causes formation of memory CD8+ T cell Test if siRNA can enhance vaccine- induced memory responses
Methods Goal: 4-1BB-raptor is capable of inhibiting mTOR activity OT-I cells transferred into mice, and mice vaccinated with OVA peptide and then treated with: rapamycin 4-1BB aptamer-GFP siRNA (4-1BB-GFP) 4-1BB aptamer-raptor siRNA (4-1BB-raptor) mTORC activity measured
Results The 4-1BB-raptor conjugate inhibited the activity of mTORC1, but not mTORC2, Rapamycin inhibited both mTOR complexes. The target specificity of raptor siRNA inhibition is shown
Methods Goal: mTOR inhibition leads to the generation of enhanced memory CD8+ T cell responses OT-I cells transferred into mice, mice vaccinated with OVA peptide and then treated with: rapamycin 4-1BB aptamer-GFP siRNA (4-1BB-GFP) 4-1BB aptamer-raptor siRNA (4-1BB-raptor) Cells were analyzed by flow cytometry.
Cells treated with rapamycin or 4-1BB-raptor have greater CD62Lhi CD44hi or CD62LhiCD127hi phenotype. Phenotypes show an enhanced potential to develop into memory cells.
Methods Goal: 4-1BB-raptor can enhance vaccine-induced memory responses Mice were vaccinated with B16 melanoma cells (GVAX) and then treated with: Rapamycin 4-1BB aptamer-GFP siRNA conjugate (4-1BB-GFP) 4-1BB aptamer-raptor siRNA conjugate (4-1BB-raptor) Then, on day 50, the mice were challenged with parental B16 tumor cells and survival was determined
Results Treatment with 4-1BB-raptor enhanced GVAX-induced antitumor immunity. Rapamycin is less effective at promoting memory responses and has less antitumor immunity
Author’s Interpretation siRNA inhibition of mTORC1 enhances vaccine-induced immunity. Aptamer-targeted siRNA superior to rapamycin in terms of protective antitumor immunity in mice. Rapamycin inhibits mTORC1 and mTORC2. Since mTORC2 is in cell survival, and glucose homeostasis, inhibition could be harmful Cell targeting reduces dose of siRNA needed, reducing the risk of nonspecific immune activation
Critique Side-effects of inhibition of mTOR on other cellular processes was not examined Enhancement of memory T-cells not directly measured Mechanism of the immune response in the enhancement of antitumor immunity still needs to be investigated 4-11B expression is upregulated on cells other than CD8+ so more specific targeting receptors will improve the specificity Currently, only in mouse model, no human studies conducted so therapeutic applications not completely confirmed
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