Nucleic acid selection against PDL1 as a means of inhibiting immunosuppression by cancerous cells

Orlando Adrian Lara

December 3, 2013

Fall 2013

RNA N56H pool

Protein A bead based selection against PD-L1

Aptamer Stream

Aptamer selection against PD-L1 as a means of inhibiting immunosuppressive properties of cancerous cells

Abstract

            Recent studies in the field of cancer immunotherapy have shown that the Programmed death 1 receptor/Programed death ligand1 (PD-1/PD-L1) pathway contributes to the ability of cancer cells to evade the immune system. Programmed death ligand 1 (PD-L1) interacts with the surface receptor site of cytotoxic T lymphocytes PD1, causing T cell inhibition and apoptosis (Haile, S. et al. 2013).

Studies using flow cytometry show increased up regulation of PD-L1 in the tissues of patients with lymphoma, suggesting PDL1 can help promote carcinogenesis (Li, Y. et al. 2013). Iwai Y et al. (2004) demonstrated that blocking the PD-1/PD-L1 pathway—by genetic manipulations or antibody treatment— inhibited the metastasis of melanoma cells to the liver and the metastasis of colon cancer cells to the lung by enhancing the recruitment of T cells.

The use of aptamers as a means of blocking the PD-1/PD-L1 pathway is introduced as a way of promoting inhibition of immunosuppressive properties characterized by cancerous cells. Aptamers are short oligonucleotide sequences that have the ability to bind to a wide range of target molecules, including drugs, proteins, and other organic or inorganic molecules with high affinity and specificity (Ellington, A. D., 1990).

Specific Aim – Find an aptamer that will bind to PDL1. Binding an aptamer to PD-L1 has the potential to inhibit the immunosuppression of cytotoxic T cells and promote the recruitment of cytotoxic T cells for the destruction of cancer cells.

Figure 1.- An aptamer against PDL1 can be used to block the PD1/PDL1 pathway inhibiting immunosuppression of T cells and promoting tumor-immunity.

Budget

PD-L1 was obtained from R&D systems (www.rndsystems.com) and is listed under the catalog number 1019-B7-100. The protein was order as 100ug lyophilized for $329, and was reconstituted in twenty microliters of 1X PBS plus 5mM MgCl₂ for a final concentration of 100μM. The stock solution was then made into 200pmol aliquots. The cost of per round was calculated to be $32.9 for a total of 10 rounds and was received functionalized with F/C chimera.

For a link to the proposal click here


For a link to the first progress report click here

For a link to the second progress report  click here

For a link to the final report click here

References

Haile, S., Dalal, S., Clements, V., Tamada, K., & Ostrand-Rosenberg, S. (2013, August 5). Soluble CD80 Restores T Cell Activation and Overcomes Tumor Cell Programmed Death Ligand 1-Mediated Immune Suppression." J Immunol 191.5 (2013): 2829-836. Print.

Li, Y., Wang, J., & Ke, X. (2013, April 21). [Expression of PD-L1 in l... [Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2013] - PubMed - NCBI. National Center for Biotechnology Information. Retrieved September 16, 2013, from http://www.ncbi.nlm.nih.gov/pubmed/?term=Expression+of+PD- L1+in+Lymphoma+and+Its+Effect+on+Lymphoma+Proliferation+and+Chemotherapy+Resistance.

Iwai, Y., Terawaki, S., & Honjo, T. (2004, December 20). PD-1 blockade inhibits hematogenous spread of poorly immunogenic tumor cells by enhanced recruitment of effector T cells . Oxford Journals | Life Sciences & Medicine | International Immunology.

Ellington, A. D., & Szostak, J. W. (1990). In Vitro Selection Of RNA Molecules That Bind Specific Ligands. Nature, 346(6287), 818-822.