AcroPrep Protein Purification Plates Facilitate Testing of Novel Cancer Vaccine Strategy

Many ongoing efforts to develop anti-cancer vaccines have, to some extent, been underwhelming. Vaccines targeting tumor cell surface proteins are often subject to immune tolerance, lowering the ultimate effectiveness of the therapy. Personalized cancer therapies have met with notably greater success, but these treatments cannot be applied universally.

In this publication,^[1] the authors sought to develop a more effective cancer vaccine strategy by targeting antigen-presenting dendritic cells (DCs) and invariant natural killer T-cells (iNKT cells). Anti-inflammatory DCs are particularly apt at promoting a potent anti-tumor response in CD8 T cells. INKT cells promote the maturation of pro-inflammatory DCs, indirectly promoting CD4 and CD8 T cell activation.  Both fusion proteins that the group developed and tested are based on protein sequences that are conserved from person-to-person, so either one could potentially form the basis of anti-cancer vaccine that could be applied universally.

In the first phase of their study, the research group made use of the immune system-related cell surface molecule known as CD1d as their fusion protein partner; CD1d is expressed by several types of immune cell, including iNKT cells and all DC subsets.  The authors designed a fusion protein consisting of the CD1d molecule fused with anti-HER2 protein; HER2 being a marker well-known for its expression on the surface of breast cancer cells. This fusion protein was used in combination with a therapeutic cancer vaccine, which has previously shown promising results in clinical studies. For the second phase of their study, the team tested an alternative fusion protein targeted primarily to a unique subpopulation of anti-inflammatory DCs that provoke a strong CD8 T cell activation.

To test the new fusion proteins in a mouse model, the scientists first needed the purify the fusion proteins to remove any endotoxin contamination. Endotoxins released from bacterial cells during sample preparation can skew research results by triggering an immune response. 

The authors used Pall’s AcroPrep™ Advance 96-Well Filter Plates With Mustang® Q/S Ion Exchange Membrane to carry out protein purification based on Mustang Q anion exchange chromatography.

The Mustang Q anion exchange membrane is designed to withstand high flow rates for rapid purification, with a high binding capacity for plasmid DNA, viral particles, and negatively charged proteins and molecules such as endotoxins.

The AcroPrep 96-well filter plates used in this study are ideal in small-scale protein purification applications. In addition to plates equipped with the Mustang Q anion exchange membrane, Pall also supply these plates with Mustang S cation exchange membrane. These plates can be used either manually or within an automated system and are available with 350 µL and 1 mL well volumes. Below is additional data demonstrating the high protein binding capacity of these membranes:

Each step in the purification process requires optimization in order to maximize yield and purity of the target protein. Read our Application Note: Efficient Multi-Well Protein Purification for more information on the use of filter plates to optimize protein purification and protocols describing a typical purification strategy for each of the Mustang membranes..

Following protein purification, the authors investigated the anti-tumor effects of the two new fusion proteins in mice bearing grafted human-derived HER2 tumor cells. Both fusion protein strategies promoted a potent CD8 T cell response associated with tumor regression and tumor growth delay. The CD1d fusion protein also elicited potent iNKT cell activation. Based on these results, the authors plan to continue their research into this promising new cancer fighting strategy.

Learn more about how AcroPrep Advance filtration products support a wide range of biomedical research applications.

Reference:

1. Corgnac S., et al. Recombinant fusion proteins for targeting dendritic cell subsets in therapeutic cancer vaccine. Methods in Enzymology. 632: 521-543. 2020.