Aberrant activation of Wnt signaling results in unregulated accumulation of cytosolic β-catenin, which subsequently enters the nucleus and promotes transcription of genes that contribute to cellular proliferation and malignancy. Here, we sought to eliminate pathogenic β-catenin from the cytosol using designer ubiquibodies (uAbs), chimeric proteins composed of an E3 ubiquitin ligase and a target-binding domain that redirect intracellular proteins to the proteasome for degradation. To accelerate uAb development, we leveraged a protein language model–driven algorithm called SaLT&PepPr to computationally design “guide” peptides with affinity for β-catenin, which were subsequently fused to the catalytic domain of a human E3 called carboxyl terminus of Hsp70-interacting protein. Expression of the resulting peptide-guided uAbs in colorectal cancer cells led to the identification of several designs that greatly reduced the abnormally stable pool of free β-catenin in the cytosol and nucleus while preserving the normal membrane–associated subpopulation. This selective knockdown of pathogenic β-catenin suppressed Wnt/β-catenin signaling and impaired tumor cell survival and proliferation. Furthermore, one of the best degraders selectively decreased cytosolic but not membrane-associated β-catenin levels in livers of BALB/c mice following delivery as a lipid nanoparticle–encapsulated mRNA. Collectively, these findings reveal the unique ability of uAbs to selectively deplete abnormal proteins in vitro and in vivo and open the door to peptide-programmable biologic modulators of other disease-causing proteins.
Tianzheng Ye, A. Alamgir, C. Robertus et al.· Science Advances· 0 citations
Dengue virus (DENV) remains a major global health burden, with four antigenically distinct serotypes (DENV-1–4) posing a significant challenge for vaccine development. Dengue non-structural protein 1 (NS1) has been associated with additional protection and reduced disease severity, supporting its inclusion in vaccine design. In this study, we designed a consensus NS1 (cNS1) antigen by integrating sequence elements from all four DENV serotypes (78–89% amino acid identity) to enhance cross-serotype antigenic coverage. The cNS1 sequence was encoded as a nucleoside-modified mRNA and formulated in lipid nanoparticles (mRNA–LNPs). Immunization of BALB/c mice with a low dose (0.2 µg) of cNS1 mRNA–LNP induced broadly reactive NS1-specific IgG responses that recognized NS1 proteins from all four serotypes. In addition, the vaccine elicited interferon-γ (IFN-γ)–producing T cell responses against peptide pools derived from multiple DENV serotypes, indicating the activation of cross-reactive cellular immunity. While broad immune recognition was achieved, this was accompanied by lower serotype-specific response magnitudes as a trade-off. In conclusion, the cNS1 mRNA vaccine induces cross-serotype humoral and cellular immune responses in mice, highlighting the potential of consensus antigen design to broaden immune recognition of DENV NS1. These findings support the further development of NS1-based immunogens as complementary components of next-generation dengue vaccines aimed at achieving broad and effective protection.
Kittipan Tharakhet, E. Prompetchara, Chirayus Khawsang et al.· PLoS ONE· 0 citations
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