With the advancement of genetic engineering, monoclonal antibodies (mAbs) have made far-reaching progress in the treatment of various human diseases. However, due to the high cost of production, the increasing demands for antibody-based therapies have not been fully met. Currently, mAb-derived alternatives, such as antigen-binding fragments (Fab), single-chain variable fragments, bispecifics, nanobodies, and conjugated mAbs have emerged as promising new therapeutic modalities. They can be readily prepared in bacterial systems with well-established fermentation technology and ease of manipulation, leading to the reduction of overall cost. This review aims to shed light on the strategies to improve the expression, purification, and yield of Fab fragments in Escherichia coli expression systems, as well as current advances in the applications of Fab fragments.

Activating transcription factor 5 (ATF5) belongs to the activating transcription factor/cyclic adenosine monophosphate (cAMP) response element-binding protein family of basic region leucine zipper transcription factors. ATF5 plays an important role in cell stress regulation and is involved in cell differentiation and survival, as well as centrosome maintenance and development. Accumulating evidence demonstrates that ATF5 plays an oncogenic role in cancer by regulating gene expressions involved in tumorigenesis and tumor survival. Recent studies have indicated that ATF5 may also modify the gene expressions involved in other diseases. This review explores in detail the regulation of ATF5 expression and signaling pathways and elucidates the role of ATF5 in cancer biology. Furthermore, an overview of putative therapeutic strategies that can be used for restoring aberrant ATF5 activity in different cancer types is provided.

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Abstract Aim NIH3T3 cell line with expression of human receptor for advanced glycation end-products (hRAGE) transduced with lentivirus vectors was used to analyze affinity, biological activity, and/or molecular mechanisms of molecules targeting the hRAGE pathway. Method The DNA fragment coding for hRAGE gene was integrated into the genome of NIH3T3 cells using lentivirus transduction. Cells expressing hRAGE were selected with puromycin, and the level of hRAGE expression was analyzed by Western blot. To establish a stable cell line, colonies of hRAGE-expressing cells were generated, and the level of RAGE expression in each engineered cell line was analyzed within 20 generations. Flow cytometry assay was used to verify affinity of anti-hRAGE antibody binding to hRAGE on the surface of engineered cells. The engineered NIH3T3 cell line was applied to assess effects of anti-hRAGE blocking antibody on amyloid β-induced cells apoptosis by CCK-8 assay. Results The engineered NIH3T3 cell line (hRAGE-NIH3T3) could stably express human RAGE. Commercial anti-RAGE polyclonal antibody could recognize and bind to human RAGE on the surface of hRAGE-NIH3T3 but not original NIH3T3 cells. In addition, hRAGE-NIH3T3 was more sensitive to RAGE pathway-dependent stimulation. Our data show that the hRAGE-NIH3T3 cell line established is an excellent tool in the study of RAGE-targeting molecules based on the cellular level, biological function, and RAGE-mediated molecular mechanisms.