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Translation and Post-Translational Modifications During Aging
Published in Alvaro Macieira-Coelho, Molecular Basis of Aging, 2017
One cytoplasmic protein that can be specifically ribosylated by at least two bacterial toxins, namely diphtheria toxin and exotoxin A, is the protein elongation factor EF-2. ADP-ribosylation of the diphthamide (modified histidine 715) residue of EF-2 results in the complete abolition of its catalytic activity.51 There is some evidence that increased ADP-ribosylation of EF-2 is correlated with cellular aging. For example, the amount of EF-2 that can be ADP-ribosylated in the presence of diphtheria toxin in cell-free extracts decreases significantly during aging of human fibroblasts in culture.67 However, no decline in the amount of ADP-ribosylatable EF-2 was observed in liver extracts prepared from calorie-restricted and freely fed Fischer 344 rats of different ages.68 Therefore, it is not clear to what extent the decline in the activity of EF-2 correlates with the decline in the rate of protein synthesis during aging.
Targeted Cytotoxicity
Published in Siegfried Matzku, Rolf A. Stahel, Antibodies in Diagnosis and Therapy, 2019
Uwe Zangemeister-Wittke, Winfried Wels
Several extremely toxic proteins of bacterial and plant origin have been employed for the preparation of chemical immunotoxins. The most commonly used are summarized in Table 4. Intoxication of cells by these proteins results from entry of the catalytic fragments into the cytoplasm of mammalian cells where they covalently modify components of the protein synthesis machinery. The bacterial toxins Pseudomonas aeruginosa exotoxin A (ETA) and diphtheria toxin (DT) secreted by toxigenic Corynebactenum diphtheriae are both synthesized as single-chain polypeptides of 59 and 67 kDa, respectively. Both toxins consist of several functional domains which were defined by crystal structure and deletion analysis (Allured et al., 1986; Hwang et al., 1987; Choe et al., 1992). Distinct domains are responsible for the binding to receptors on the surface of mammalian cells, intracellular processing, and catalytic activity. ETA and DT differ in their molecular organization, but possess identical reaction mechanisms. The catalytic domain ADP-ribosylates and inactivates eukaryotic elongation factor 2 (EF-2) which is an essential component in protein synthesis (Iglewski et al., 1977; Collier et al., 1975). This modification occurs at a post-translationally modified histidine derivative, diphthamide, which is located in the ribsomal binding site of EF-2 (described in detail by Perentesis et al., 1992). Processing of DT by extracellular proteases yields two fragments which are held together by a disulfide bond. The N-terminal A fragment of 21 kDa harbors the catalytic activity; the C-terminal B fragment of 38 kDa mediates cell binding and uptake into the cytoplasm. Due to the ability to manipulate its structure, most progress in the understanding of the mechanism of intoxication has been made with ETA. In contrast to DT the catalytic domain of ETA is located at the C-terminus (Hwang et al., 1987). After cell binding and internalization ETA is cleaved by a cellular protease and a N-terminal 28 kDa and a C-terminal 37 kDa fragment are generated. After reduction of a disulfide bond the C-terminal fragment which contains the enzymatic activity translocates to the cytoplasm (Ogata et al., 1992).
Targeting CD123 in BPDCN: an emerging field
Published in Expert Review of Hematology, 2021
Adam J DiPippo, Nathaniel R Wilson, Naveen Pemmaraju
After binding to its natural receptor (CD123 or IL3Rα), tagraxofusp is then internalized, endocytosed into the cell, and localized in endosomes where the acidic environment cleaves DT388 from its linker and is transported into the cytosol [57,58]. Here, DT388 induces ADP ribosylation of elongation factor 2 (eEF2), which has the essential function of catalyzing the translocation of RNA on the 80S ribosome. Of note, human eEF2 has a unique and conserved diphthamide residue that DT388 and other toxins (i.e. exotoxin A) can bind as a substrate and irreversibly inhibit eEF2 function [57,59]. Nonfunctional eEF2 results in cessation of cellular protein synthesis and induces cell death [19,37,57,60,61]. Additionally, tagraxofusp is not a substrate for common drug efflux pumps or P-glycoprotein, which reduces the risk of inherent cellular resistance [62, 63].
Tagraxofusp as treatment for patients with blastic plasmacytoid dendritic cell neoplasm
Published in Expert Review of Anticancer Therapy, 2020
Sophia S. Lee, Deborah McCue, Naveen Pemmaraju
Factors influencing the efficacy of tagraxofusp other than CD123 expression are largely unknown. After an investigation of tagraxofusp resistant patients, CD123 level was not associated with tagraxofusp use [40]. In Stephansky’s study (2017) with AML and BPDCN patients, decrease in expression of CD123 was again not observed during or after SL-401 in resistant BPDCN cell lines. In fact, decreased expression of DPH1 was confirmed in 6 resistant clones in further in vitro studies; and DPH1 loss was sufficient to mediate SL-401 resistance in AML and BPDCN patients [40,41]. This resistance was associated with loss of diphthamide synthesis pathway activity via DNA-methylated downregulation of DPH1 [40]. Currently, understanding the mechanism of DPH1 in tumor activity is limited and requires further investigation [40].
Biological therapy in elderly patients with acute myeloid leukemia
Published in Expert Opinion on Biological Therapy, 2023
Giulia Ciotti, Giovanni Marconi, Alessandra Sperotto, Maria B Giannini, Michele Gottardi, Giovanni Martinelli
Loss or reduction of diphthamide synthesis pathway expression due to downregulation of DPH1 and subsequent insensitivity to diphtheria toxin is been demonstrated as a mechanism of resistance to tagraxofusp that is partially mediated by altered DNA-methylation [39]. Combination with AZA may overcome this resistance pathway and enhance SL-401 effectiveness. Combination with azacytidine alone or with AZA + VEN is currently under investigation (NCT03113643) and the trial is in recruiting phase.