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Tailoring In Vivo Cytotoxicity Assays to Study Immunodominance in Tumor-specific CD8+ T Cell Responses
Published on: May 6, 2019
Foxp3 inhibitory peptide encapsulated in a novel CD25-targeted nanoliposome promotes efficient tumor regression in
Alejandro Serrano1,2, Noelia Casares2,3, Iñaki F Trocóniz1,2
1Department of Pharmaceutical Sciences, School of Pharmacy and Nutrition, University of Navarra, Pamplona, Spain.
Abstract:
P60, a Foxp3 inhibitory peptide, can hinder the regulatory T cell (Treg) activity and impair tumor proliferation. However, low systemic stability and poor specificity have led to daily dosing to achieve therapeutic effect. Therefore, this study aims to improve P60 stability and specific delivery through its encapsulation in liposomes targeting CD25, constitutively expressed in Tregs. P60 liposomes formulated with DSPE-PEG750 or DSPE-PEG2000 were incubated with DSPE-PEG2000-Maleimide micelles conjugated to Fab' fragments of anti-CD25 to develop two targeted formulations or immunoliposomes (IL): IL-P602000 (DSPE-PEG2000 only) and IL-P60750 (combining DSPE-PEG750 and DSPE-PEG2000). P60 encapsulation efficiency was 50%-60% irrespective of PEG chain length. Treg uptake was 2.5 and 14 times higher for IL-PEG750 compared with IL-PEG2000 and non-targeted liposomes, respectively, in in-vitro assays. In fact, IL-P60750 allowed CD8+ T cells ex-vivo proliferation in presence of Treg at doses 10-20 times lower than for free P60. Antitumor response of P60 and IL-P60750 in monotherapy and combined with anti-PD-1 was evaluated in MC38 and LLCOVA tumor bearing mice. In MC38 model, IL-P60750 monotherapy induced total tumor regression in 40% of mice reaching 100% for anti-PD-1 combination. This effect was associated with a significant increase in activated CD8+ T cells in tumors. Notably, IL-P60750 also inhibited human Treg in ex-vivo assay, showing the translational capability of this formulation. In conclusion, IL-P60750 formulated with different PEG chain lengths, has demonstrated antitumor efficacy by selective inhibition of Treg activity and enhances the effect of anti-PD1. Altogether, this novel IL represents a promising nanoplatform for cancer immunotherapies.
Insights
This study developed targeted immunoliposomes (IL-P60_750) to improve the stability and delivery of P60, a Foxp3 inhibitory peptide, for cancer immunotherapy. The novel formulation demonstrated significant antitumor efficacy by selectively inhibiting regulatory T cell activity and enhancing anti-PD-1 therapy.
Area of Science:
- Immunology
- Nanotechnology
- Cancer Research
Background:
- Regulatory T cells (Tregs) suppress anti-tumor immunity.
- P60 peptide inhibits Treg activity but has poor stability and specificity.
- Current P60 therapy requires frequent dosing due to limitations.
Purpose of the Study:
- To enhance P60 stability and achieve targeted delivery to Tregs.
- To develop P60-loaded liposomes (immunoliposomes, IL) targeting CD25 on Tregs.
- To evaluate the anti-tumor efficacy of the novel immunoliposome formulation.
Main Methods:
- Formulated P60-loaded liposomes with varying DSPE-PEG chain lengths.
- Conjugated anti-CD25 Fab' fragments to liposomes to create targeted immunoliposomes (IL-P60_750 and IL-P60_2000).
- Assessed Treg uptake, P60 encapsulation efficiency, T cell proliferation, and in-vivo anti-tumor response in mouse models.
Main Results:
- P60 encapsulation efficiency was 50%-60% regardless of PEG length.
- IL-P60_750 showed significantly higher Treg uptake compared to IL-P60_2000 and non-targeted liposomes.
- IL-P60_750 monotherapy induced complete tumor regression in 40% of mice and enhanced anti-PD-1 combination therapy efficacy.
Conclusions:
- IL-P60_750 formulation demonstrates improved stability and targeted delivery of P60.
- This targeted approach selectively inhibits Treg activity, enhancing anti-tumor immune responses.
- IL-P60_750 represents a promising nanoplatform for advancing cancer immunotherapy.
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