Co-transient expression of PSA-Fc and PAP-Fc fusion protein in plant as prostate cancer vaccine candidates and immune responses in mice

Abstract

Prostate speci c antigen (PSA) and prostatic acid phosphatase (PAP) have been considered as immunotherapeutic antigens for prostate cancer. The use of a single antigenic agent is unlikely to be effective in eliciting immunotherapeutic responses due to the heterogeneous and multifocal nature of prostate cancer. Thus, multiple antigens have been combined to enhance their anti-cancer effects. In the current study, PSA and PAP were fused to the crystallizable region (Fc region) of immunoglobulin G1 and tagged with KDEL, the endoplasmic reticulum (ER) retention signal motif, to generate PSA-FcK and PAP-FcK, respectively, and were transiently co-expressed in Nicotiana benthamiana. Western blot analysis con rmed the co-expression of PSA-FcK and PAP-FcK (PSA-FcK+PAP-FcK) with a 1:3 ratio in the coin ltrated plants. PSA-FcK, PAP-FcK, and PSA-FcK+PAP-FcK proteins were successfully puri ed from N. benthamiana by protein A a nity chromatography. ELISA showed that anti-PAP and anti-PSA antibodies successfully detected PAP-FcK and PSA-FcK, respectively, and both detected PSA-FcK+PAP-FcK. Surface plasmon resonance (SPR) analysis con rmed the binding a nity of the plant-derived Fc fusion proteins to FcγRI/CD64. Furthermore, we also con rmed that mice injected with PSA-FcK+PAP-FcK produced both PSA and PAP-speci c IgGs, demonstrating their immunogenicity. This study suggested that the transient plant expression system can be applied to produce the dual-antigen Fc fusion protein (PSA-FcK+PAP-FcK) for prostate cancer immunotherapy. Key MessagePAP-FcK and PSA-FcK prostate cancer antigenic proteins transiently co-expressed in plant induce their speci c humoral immune responses in mice.

Enhanced binding and inhibition of SARS‐CoV‐2 by a plant‐derived ACE2 protein containing a fused mu tailpiece

“…Additionally, two different proteins can be transiently co-expressed within a single plant system by agroinfiltration. [73] J chain, a protein that mediates IgM oligomeric assembly, facilitates the…”

Plant-Derived Anti-Human Epidermal Growth Factor Receptor 2 Antibody Suppresses Trastuzumab-Resistant Breast Cancer with Enhanced Nanoscale Binding

“…After the removal of chloroplast and cell debris using centrifugation and filtering processes, the anti-HER2 VHH-FcK proteins were purified through Protein A affinity chromatography (Amicogen, Jinju-si, Korea). The SDS-PAGE gel loaded with the purified target protein was stained with Coomassie blue staining buffer, and then the gel was destained by destaining buffer (Figure b) …”

“…The SDS-PAGE gel loaded with the purified target protein was stained with Coomassie blue staining buffer, and then the gel was destained by destaining buffer ( Figure 1 b). 51 …”

Transient Plant Expression of Recombinant Rabies Virus Glycoprotein: A strategy for Vaccine Production

“…Following cultivation, pre-cultured Agrobacterium (with an optical density reading of 0.3-0.4) was centrifuged at 3,000 rpm for 15 minutes. The resulting Agrobacterium cell pellet was then diluted in an infiltration buffer composed of 10 mM MES (pH 5.5), 10 mM MgCl 2 , and 0.1 mM acetosyringone (Oh et al, 2023). The cells, suspended in the infiltration buffer, were introduced into the leaves of 16-week-old N. benthamiana plants using a 1-mL needle syringe (Fig.…”

“…Transient expression via agro-infiltration has is becoming more preferred in the manufacturing of biologics in plants due to the cost-effectiveness and rapid protein production capabilities associated with this method (Mardanova et al, 2017;Norkunas et al, 2018). Typically, transient plant expression systems yield a higher amount of protein per unit weight of plant tissue compared to transgenic plant expression systems (Kim et al, 2021;Lim et al, 2022;Kang et al, 2023;Oh et al, 2023). However, the level of recombinant protein expression is influenced by the leaf position and the time that elapses after infiltration, as demonstrated in previous research (Yamamoto et al, 2018;Kim et al, 2021).…”