Surface display of glycosylated Tyrosinase related protein-2 (TRP-2) tumour antigen on Lactococcus lactis

Abstract

BackgroundThe exploitation of the surface display system of food and commensal lactic acid bacteria (LAB) for bacterial, viral, or protozoan antigen delivery has received strong interest recently. The Generally Regarded as Safe (GRAS) status of the Lactococcus lactis coupled with a non-recombinant strategy of in-trans surface display, provide a safe platform for therapeutic drug and vaccine development. However, production of therapeutic proteins fused with cell-wall anchoring motifs is predominantly limited to prokaryotic expression systems. This presents a major disadvantage in the surface display system particularly when glycosylation has been recently identified to significantly enhance epitope presentation. In this study, the glycosylated murine Tyrosinase related protein-2 (TRP-2) with the ability to anchor onto the L. lactis cell wall was produced in suspension adapted Chinese Hamster Ovary (CHO-S) cells by expressing TRP-2 fused with cell wall anchoring LysM motif (cA) at the C-terminus.ResultsA total amount of 33 μg of partially purified TRP-2-cA from ~6.0 g in wet weight of CHO-S cells was purified by His-tag affinity chromatography. The purified TRP-2-cA protein was shown to be N-glycosylated and successfully anchored to the L. lactis cell wall.ConclusionsThus cell surface presentation of glycosylated mammalian antigens may now permit development of novel and inexpensive vaccine platforms.

A review on Lactococcus lactis: from food to factory

“…More importantly, this enables the lactococcal cells to carry heterologous proteins without being genetically modified, a method which have also been demonstrated with Newcastle disease virus hemagglutinin-neuraminidase (HN) protein for specific targeting of breast cancer cells [33]. In addition, eukaryotic proteins which require post-translational modifications can also be expressed in eukaryotic hosts, and subsequently attached to L. lactis for delivery [34]. A variation of this method uses GEM (gram-positive enhancer matrix) particles which are killed non-recombinant lactococcal cells devoid of most intact cell wall components and intracellular materials.…”

“…These include a vaccine against human papilloma virus type-16 induced tumours where L. lactis surface displaying the E7 antigen whist secreting IL-12 was shown to provide full prophylactic protection in immunized mice and was also able to induce regression of palpable tumours in tumour-induced mice [101]. Other cancer antigens expressed using L. lactis includes glycosylated tyrosinase related protein-2 (TRP-2) tumour antigen against melanoma (although this has not gone to animal trials) [34] and carcinoembryonic antigen (CEA) against colon cancer in mice [102]. The latter showed successful induction of immune response in mice as indicated by higher levels of CEA-specific secretory IgA compared to controls.…”

Targeting Melanoma Hypoxia with the Food-Grade Lactic Acid Bacterium Lactococcus Lactis

“…Finally, the strategy proposed in this study has a therapeutic potential, by engineering L. lactis to express and deliver molecules of health interest directly in the hypoxic microenvironment of solid tumors. L. lactis has been used as a vehicle to deliver therapeutic molecules into the human body, including cytokines/ligands (e.g., IL-10, IL-22, RANKL, TGF-β1) [53][54][55], tumor-associated antigens (TAAs) (e.g., HPV-16 E7, TRP-2) [56,57], antioxidant enzymes [58], and protease inhibitors [59].…”

Molecular dynamics simulation and experimental study of the surface-display of SPA protein via Lpp-OmpA system for screening of IgG

“…Display of heterologous proteins on the bacterial surface has been demonstrated as a multi-strategy approach to develop an e cient vaccine for S. aureus development (Kalyanasundram et al, 2015), screening of antibody libraries (Cavallari, 2017), development of whole-cell bioadsorbents (Tafakori et al, 2012), and biosensors (Furst et al, 2017). Chimeric protein system of the Lpp′-OmpA is used as an anchor and loads heterologous proteins onto the Gram-negative bacterial surface (Yang et al, Georgiou et al, 1996).…”