BF211 is a synthetic molecule derived from bufalin (BF). The apoptosis-inducing effect of BF211 was stronger than that of BF while the acute toxicity of BF211 was much lower than that of BF. BF211 exhibited promising concentration-dependent anti-cancer effects in nude mice inoculated with A549 cells in vivo. The growth of A549 tumor xenografts was almost totally blocked by treatment with BF211 at 6 mg/kg. Notably, BF and BF211 exhibited differences in their binding affinity and kinetics to recombinant proteins of the α subunits of Na+/K+-ATPase. Furthermore, there was a difference in the effects of BF or BF211 on inhibiting the activity of porcine cortex Na+/K+-ATPase and in their time-dependent effects on intracellular Ca2+ levels in A549 cells. The time-dependent effects of BF or BF211 on the activation of Src, which was mediated by the Na+/K+-ATPase signalosome, in A549 cells were also different. Both BF and BF211 could induce apoptosis-related cascades, such as activation of caspase-3 and the cleavage of PARP (poly ADP-ribose polymerase) in A549 cells, in a concentration-dependent manner; however, the effects of BF211 on apoptosis-related cascades was stronger than that of BF. The results of the present study supported the importance of binding to the Na+/K+-ATPase α subunits in the mechanism of cardiac steroids and also suggested the possibility of developing new cardiac steroids with a stronger anti-cancer activity and lower toxicity as new anti-cancer agents.

Viruses are far more abundant than cellular microorganisms in the marine ecosystem. However, very few viruses have so far been isolated from marine sediments, especially hydrothermal vent sediments, hindering the understanding of the biology and ecological functions of these tiny organisms. Here, we report the isolation and characterization of a temperate bacteriophage, named PVJ1, which infects Psychrobacillus from a hydrothermal vent field in Okinawa Trough. PVJ1 belongs to the Myoviridae family of the order Caudovirales. The tailed phage possesses a 53,187 bp linear dsDNA genome, with 84 ORFs encoding structural proteins, genome replication, host lysis, etc. in a modular pattern. The phage genome is integrated into the host chromosome near the 3′-end of deoD, a gene encoding purine nucleoside phosphorylase (PNP). The phage integration does not appear to disrupt the function of PNP. The phage DNA is packaged by the headful mechanism. Release of PVJ1 from the host cell was drastically enhanced by treatment with mitomycin C. Phages encoding an MCP sharing significant similarity (≥70% identical amino acids) with that of PVJ1 are widespread in diverse environments, including marine and freshwater sediments, soils, artificial ecosystems, and animal intestines, and primarily infect Firmicutes. These results are valuable to the understanding of the lifestyle and host interactions of bacterial viruses at the bottom of the ocean.

Antibody junctional diversity in jawed vertebrates arose from RAG transposon domestication, generating CDR3‐focused V(D)J recombination with lymphocyte specificity—a constrained system limiting engineered antibody diversity. To overcome these limitations, we developed ARESEC ( A ntibody R eprogram and E xpression S ystem in E ngineered C ells), a synthetic platform that enables programmable V(D)J recombination in non‐lymphoid HEK293T cells. Through engineered recombination signal sequences (RSS) and optimized RAG1/2 expression, ARESEC enables RSS‐guided DNA recombination across all three complementarity‐determining regions (CDR1/2/3). Co‐expression of terminal deoxynucleotidyl transferase (TdT) enhanced junctional sequence diversity by 41.5%–86.3% across 3 CDRs. The platform enables native IgG production by coupling mammalian surface display with FACS‐based functional screening. Using clinical‐grade antibodies (Nivolumab and Durvalumab) as high‐affinity starting scaffolds, we achieved further efficient affinity maturation through focused CDR diversification (library complexity >10 ³ ). From these diversification libraries, we identified optimized variants including the anti‐PD‐L1 Dur1 with 2.01‐fold enhanced binding, demonstrating the system's capacity for rapid antibody optimization from minimal diversity sampling. Collectively, ARESEC establishes a synthetic platform that transcends natural V(D)J constraints, generating multi‐CDR diversification in non‐lymphoid cells to enable rapid discovery of affinity‐matured antibodies, effectively bridging immune evolution with modern antibody engineering demands.

Antibody junctional diversity in jawed vertebrates arose from RAG transposon domestication, generating CDR3-focused V(D)J recombination with lymphocyte specificity—a constrained system limiting engineered antibody diversity. To overcome these limitations, we developed ARESEC ( A ntibody R eprogram and E xpression S ystem in E ngineered C ells), a synthetic platform that enables programmable V(D)J recombination in non-lymphoid HEK293T cells. Through engineered recombination signal sequences (RSS) and optimized RAG1/2 expression, ARESEC enables RSS-guided DNA recombination across all three complementarity-determining regions (CDR1/2/3). Co-expression of terminal deoxynucleotidyl transferase (TdT) enhanced junctional sequence diversity by 41.5-86.3% across 3 CDRs. The platform enables native IgG production by coupling mammalian surface display with FACS-based functional screening. Using clinical-grade antibodies (Nivolumab and Durvalumab) as high-affinity starting scaffolds, we achieved further efficient affinity maturation through focused CDR diversification (library complexity >10 ³ ). From these diversification libraries, we identified optimized variants including the anti-PD-L1 Dur1 with 2.01-fold enhanced binding, demonstrating the system’s capacity for rapid antibody optimization from minimal diversity sampling. Collectively, ARESEC establishes a synthetic paradigm that transcends natural V(D)J constraints, generating multi-CDR diversification in non-lymphoid cells to enable rapid discovery of affinity-matured antibodies, effectively bridging immune evolution with modern antibody engineering demands.