Targeting viral proteins using insulin-like growth factor 2-fused lysosomal targeting chimera platform to combat viral infection.

Dual Targeting Engineered Binding Proteins Block SARS-CoV-2 Infection and Complement Activation.

The rapid evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and associated complement overactivation challenge current antiviral strategies that mainly target viral entry. This study aimed to develop dual targeting engineered binding proteins capable of simultaneously blocking viral infection and complement activation. Two proteins, SBP10 and SBP16, were engineered by integrating an ACE2-mimicking peptide with a mannose-binding lectin (MBL) domain. Binding affinity and antiviral activity were evaluated using biochemical and functional assays, including inhibition of S protein-ACE2 interaction, neutralization of multiple SARS-CoV-2 variants, and assessment of lectin pathway-mediated complement activation. Both SBP10 and SBP16 bound the spike protein with low-nanomolar affinity and effectively blocked its interaction with ACE2. The proteins exhibited broad-spectrum neutralizing activity against several variants, including Alpha, Beta, Delta, and Omicron. Moreover, they significantly suppressed spike-induced activation of the lectin complement pathway. In vivo experiments further demonstrated that treatment with SBP10 or SBP16 markedly reduced spike protein-induced lung injury. In conclusion, SBP10 and SBP16 function as dual targeting engineered binding proteins that inhibit viral entry while attenuating complement-mediated inflammation, highlighting a promising therapeutic strategy for controlling SARS-CoV-2 infection and its associated immune dysregulation.

A receptor-Fc based SFV replicon Trim-Away platform for targeted viral protein degradation.

BACKGROUND Developing effective antiviral strategies is urgently needed during global viral pandemics. Traditional approaches, including small-molecule inhibitors, neutralizing antibodies, and RNA interference (RNAi), often face challenges such as drug resistance, limited specificity, and inefficient delivery. These limitations highlight the pressing need for innovative strategies focused on the targeted degradation of viral proteins. METHODS We developed an optimized Trim-Away system employing a receptor-Fc fusion protein strategy. This system integrates the E3 ubiquitin ligase TRIM21 with engineered receptor-Fc proteins to ensure highly specific recognition and intracellular degradation. A key innovation is the use of the Semliki Forest virus (SFV) self-amplifying replicon (pSFV). This platform enables sustained and robust expression of the Trim-Away components. Furthermore, this plasmid-based delivery eliminates the need for protein purification, thereby streamlining the process and improving delivery efficiency. RESULTS The system effectively degrades diverse viral targets. Specifically, it successfully degraded the spike proteins of both wild-type SARS-CoV-2 and its Omicron variant. It also targeted adeno-associated virus (AAV) capsid proteins. In vivo assays further confirmed that the self-amplifying replicon markedly reduces AAV-encoded luciferase expression. These data demonstrate that the system maintains high potency even at low dosages. CONCLUSIONS Our findings demonstrate that the pSFV-driven Trim-Away system is a powerful tool for viral protein degradation. The receptor-Fc strategy provides a significant advantage against rapidly mutating viruses. This study establishes a versatile and adaptable platform for future antiviral intervention.

TROP2-targeting chimeras (TRTACs) for tumor-selective membrane protein degradation and enhanced drug delivery

Extracellular targeted protein degradation (eTPD) systems typically utilize lysosome-targeting receptors (LTRs) to mediate internalization and lysosomal degradation of extracellular and membrane proteins. While multiple LTRs have been discovered, there remains a compelling need to seek for new LTRs, particularly those with clear clinical relevance, to expand the therapeutic potential of eTPD. Here we report trophoblast cell surface antigen-2 (TROP2), a clinically validated tumor-associated antigen, as a promising tumor-selective LTR. We engineer TROP2-targeting chimeras (TRTACs) by genetically fusing a TROP2-binding nanobody to nanobodies against specific target proteins. We show that TRTACs can induce tumor cell-selective degradation of diverse membrane proteins, including epithelial growth factor receptor (EGFR), human epithelial growth factor receptor 2 (HER2), and programmed death-ligand 1 (PD-L1). The EGFR-targeted TRTAC significantly inhibits tumor cell proliferation and shows potent antitumor activity in vivo. We further design TRTAC-drug conjugates (TRTAC-DCs) by attaching cytotoxic payloads to TRTACs, enabling targeted protein degradation together with enhanced drug delivery. TRTAC-DCs show significantly enhanced activity against HER2- and EGFR-positive tumors both in vitro and in vivo, with minimal toxicity observed in normal tissues. These findings establish TROP2 as a robust LTR and provide a versatile eTPD platform with profound translational potential for tumor treatment.

Targeted Degradation of Capsid by Host-Hijacking PROTAC Overcomes Drug Resistance.

The escalating prevalence of HIV-1 drug-resistant variants and the toxicity limitations of conventional antiretroviral therapies necessitate therapeutic strategies with novel mechanisms of action. This study focuses on HIV-1 capsid (CA), an essential replication-related viral protein. We developed CA-targeted proteolysis-targeting chimera (PROTAC) degraders by conjugating PF74-derived CA ligand IIA-4 with VHL E3 ligase ligand. Among these, VHL-3 exhibited potent anti-HIV-1 activity in MT-4 cells (EC50 = 3.0 ± 1.5 nM), a 300-fold improvement over PF74. Mechanistic studies confirmed VHL-3 dose- and time-dependently reduced CA levels in HEK293T cells (early stage, DC50 = 812 nM; late stage, DC50 = 252 nM) via a proteasome-driven pathway. Notably, it effectively degraded clinically relevant CA-resistant mutants (N74D, K70R). This work pioneers the development of CA-targeted degraders, providing a framework for next-generation anti-HIV therapies with high potency and resistance barriers.

ALOX15 orchestrates mitochondrial antiviral immunity and serves as a host target for anti-influenza therapy

Intracellular Delivery of Antibody-Based Therapeutics for Targeted Oncogenic c-Myc Degradation in Mammalian Cells and Mice.

Targeting the intrinsically disordered oncoprotein c-Myc remains challenging due to its lack of druggable pockets, hindering small-molecule inhibitor development for decades. Antibody-based strategies utilizing TRIM21-mediated targeted protein degradation (TPD), such as TRIM-Away, offer a promising alternative for endogenous c-Myc degradation but are limited by ineffective intracellular antibody delivery and uneven endogenous TRIM21 levels in various cancer cells. To overcome these, herein we disclose an acid-responsive nanoplatform using amorphous carbonated calcium phosphate nanoparticles (ACCP NPs) for effective intracellular co-delivery of TRIM21 and antibodies. Under mild biomimetic conditions, TRIM21-antibody complexes were effectively mineralized and encapsulated in the formed nanoparticles. Following endocytic uptake, these acid-sensitive ACCP NPs disassembled in endo/lysosomes, facilitating subsequent endo/lysosomal escape and cytosolic cargo release, leading to eventual intracellular TPD. In vitro, mineralized TRIM21-antibody complexes from ACCP NPs effectively degraded various intracellular targets, particularly c-Myc, across diverse cell types, including those that are TRIM21-deficient. Specifically, m-T21-c-Myc Ab nanoparticle degraded c-Myc via the ubiquitin-proteasome system and concurrently reduced MAX levels, synergistically inhibiting c-Myc transcriptional activity. In vivo, m-T21-c-Myc Ab significantly suppressed tumor growth without major organ toxicity. This work establishes ACCP NP-based TPD as a versatile and efficient platform for TRIM21/antibody-mediated degradation and a promising strategy for targeting other "undruggable" proteins.