Diphtheria toxin-based anti-human CD19 immunotoxin for targeting human CD19+ tumors.
Ontology highlight
ABSTRACT:
SUBMITTER: Zheng Q
PROVIDER: S-EPMC5527461 | biostudies-other | 2017 May
REPOSITORIES: biostudies-other
Similar Datasets
Project description:CC chemokine receptor 4 (CCR4) has attracted much attention as a promising therapeutic drug target for CCR4(+) tumor cells and Tregs. CCR4 is expressed on some tumor cells such as T-cell acute lymphoblastic leukemia (ALL), adult T-cell leukemia/lymphoma (ATLL), adult peripheral T cell lymphoma (PTCL) and cutaneous T cell lymphoma (CTCL). CCR4 is also expressed on majority of Tregs, mainly effector Tregs. In this study we have successfully developed three versions of diphtheria-toxin based anti-human CCR4 immunotoxins (monovalent, bivalent and single-chain fold-back diabody). Binding analysis by flow cytometry showed that all three versions of the anti-human CCR4 immunotoxins bound to the human CCR4(+) tumor cell line as well as CCR4(+) human PBMC. The bivalent isoform bound stronger than its monovalent counterpart and the single-chain foldback diabody isoform was the strongest among the three versions. In vitro efficacy analysis demonstrated that the bivalent isoform was 20 fold more potent in inhibiting cellular proliferation and protein synthesis in human CCR4(+) tumor cells compared to the monovalent anti-human CCR4 immunotoxin. The single-chain fold-back diabody isoform was 10 fold more potent than its bivalent counterpart and 200 fold more potent than its monovalent counterpart. The in vivo efficacy was assessed using a human CCR4(+) tumor-bearing mouse model. The immunotoxin significantly prolonged the survival of tumor-bearing NOD/SCID IL-2 receptor γ(-/-) (NSG) mice injected with human CCR4(+) acute lymphoblastic leukemia cells compared with the control group. This novel anti-human CCR4 immunotoxin is a promising drug candidate for targeting human CCR4(+) tumor cells and Tregs in vivo.
| S-EPMC5528803 | biostudies-other
Project description:Regulatory T cells (Tregs) are known to play an important role in immunoregulation and have been shown to facilitate induction of transplantation tolerance. Chemokine (C-C motif) receptor 4 (CCR4) is expressed on the surface of effector Tregs involved in controlling alloimmune and autoimmune responses. Recently we have developed a novel diphtheria-toxin based anti-human CCR4 immunotoxin for depleting CCR4+ cells in vivo. In this study, we have demonstrated that the anti-human CCR4 immunotoxin bound to porcine lymphocytes including CD4+FoxP3+ Tregs. Anti-human CCR4 immunotoxin effectively depleted CCR4+ Foxp3+ porcine Tregs in vivo. We observed depletion of up to 70-85% of the CCR4+Foxp3+ porcine Tregs in the peripheral blood and 85-91% in the lymph nodes following the anti-human CCR4 immunotoxin treatment in Massachusetts General Hospital (MGH) miniature swine. The depletion lasted for about one week with no significant reduction observed within CCR4- cell populations including CD8α+ T cells, CCR4-CD4+ T cells and B cells. In summary, anti-human CCR4 immunotoxin effectively depleted CCR4+Foxp3+ porcine Tregs in both peripheral blood and lymph nodes.
| S-EPMC5134928 | biostudies-literature
Project description:Regulatory T cells (Treg) play an important role in modulating the immune response and has attracted increasing attention in diverse fields such as cancer treatment, transplantation and autoimmune diseases. CC chemokine receptor 4 (CCR4) is expressed on the majority of Tregs, especially on effector Tregs. Recently we have developed a diphtheria-toxin based anti-human CCR4 immunotoxin for depleting CCR4(+) cells in vivo. In this study, we demonstrated that the anti-human CCR4 immunotoxin bound and depleted monkey CCR4(+) cells in vitro. We also demonstrated that the immunotoxin bound to the CCR4(+)Foxp3(+) monkey Tregs in vitro. In vivo studies performed in two naive cynomolgus monkeys revealed 78-89% CCR4(+)Foxp3(+) Treg depletion in peripheral blood lasting approximately 10 days. In lymph nodes, 89-96% CCR4(+)Foxp3(+) Tregs were depleted. No effect was observed in other cell populations including CD8(+) T cells, other CD4(+) T cells, B cells and NK cells. To our knowledge, this is the first agent that effectively depleted non-human primate (NHP) Tregs. This immunotoxin has potential to deplete effector Tregs for combined cancer treatment.
| S-EPMC4826841 | biostudies-other
Project description:Anti-CD3 immunotoxins, which induce profound but transient T-cell depletion in vivo by inhibiting eukaryotic protein synthesis in CD3+ cells, are effective reagents in large animal models of transplantation tolerance and autoimmune disease therapy. A diphtheria toxin based antiporcine CD3 recombinant immunotoxin was constructed by fusing the truncated diphtheria toxin DT390 with two identical tandem single chain variable fragments (scFv) derived from the antiporcine CD3 monoclonal antibody 898H2-6-15. The recombinant immunotoxin was expressed in a diphtheria-toxin resistant yeast Pichia pastoris strain under the control of the alcohol oxidase promoter. The secreted recombinant immunotoxin was purified sequentially with hydrophobic interaction chromatography (Butyl 650 M) followed by strong anion exchange (Poros 50 HQ). The purified antiporcine CD3 immunotoxin was tested in vivo in four animals; peripheral blood CD3+ T-cell numbers were reduced by 80% and lymph node T-cells decreased from 74% CD3+ cells pretreatment to 24% CD3+ cells remaining in the lymph node following 4 days of immunotoxin treatment. No clinical toxicity was observed in any of the experimental swine. We anticipate that this conjugate will provide an important tool for in vivo depletion of T-cells in swine transplantation models.
| S-EPMC3200572 | biostudies-literature
Project description:Programmed death-1 (PD-1), an immune checkpoint receptor, is expressed on activated lymphocytes, macrophages, and some types of tumor cells. While PD-1+ cells have been implicated in outcomes of cancer immunity, autoimmunity, and chronic infections, the exact roles of these cells in various physiological and pathological processes remain elusive. Molecules that target and deplete PD-1+ cells would be instrumental in defining the roles unambiguously. Previously, an immunotoxin has been generated for the depletion of PD-1+ cells though its usage is impeded by its low production yield. Thus, a more practical molecular tool is desired to deplete PD-1+ cells and to examine functions of these cells. We designed and generated a novel anti-PD1 diphtheria immunotoxin, termed PD-1 DIT, targeting PD-1+ cells. PD-1 DIT is comprised of two single chain variable fragments (scFv) derived from an anti-PD-1 antibody, coupled with the catalytic and translocation domains of the diphtheria toxin. PD-1 DIT was produced using a yeast expression system that has been engineered to efficiently produce protein toxins. The yield of PD-1 DIT reached 1-2 mg/L culture, which is 10 times higher than the previously reported immunotoxin. Flow cytometry and confocal microscopy analyses confirmed that PD-1 DIT specifically binds to and enters PD-1+ cells. The binding avidities between PD-1 DIT and two PD-1+ cell lines are approximately 25 nM. Moreover, PD-1 DIT demonstrated potent cytotoxicity toward PD-1+ cells, with a half maximal effective concentration (EC50 ) value of 1 nM. In vivo experiments further showed that PD-1 DIT effectively depleted PD-1+ cells and enabled mice inoculated with PD-1+ tumor cells to survive throughout the study. Our findings using PD-1 DIT revealed the critical role of pancreatic PD-1+ T cells in the development of type-1 diabetes (T1D). Additionally, we observed that PD-1 DIT treatment ameliorated relapsing-remitting experimental autoimmune encephalomyelitis (RR-EAE), a mouse model of relapsing-remitting multiple sclerosis (RR-MS). Lastly, we did not observe significant hepatotoxicity in mice treated with PD-1 DIT, which had been reported for other immunotoxins derived from the diphtheria toxin. With its remarkable selective and potent cytotoxicity toward PD-1+ cells, coupled with its high production yield, PD-1 DIT emerges as a powerful biotechnological tool for elucidating the physiological roles of PD-1+ cells. Furthermore, the potential of PD-1 DIT to be developed into a novel therapeutic agent becomes evident.
| S-EPMC10443333 | biostudies-literature
Project description:Cancer remains one of the leading causes of death worldwide. Conventional therapeutic strategies usually offer limited specificity, resulting in severe side effects and toxicity to normal tissues. Targeted cancer therapy, on the other hand, can improve the therapeutic potential of anti-cancer agents and decrease unwanted side effects. Targeted applications of cytolethal bacterial toxins have been found to be especially useful for the specific eradication of cancer cells. Targeting is either mediated by peptides or by protein-targeting moieties, such as antibodies, antibody fragments, cell-penetrating peptides (CPPs), growth factors, or cytokines. Together with a toxin domain, these molecules are more commonly referred to as immunotoxins. Targeting can also be achieved through gene delivery and cell-specific expression of a toxin. Of the available cytolethal toxins, diphtheria toxin (DT) is one of the most frequently used for these strategies. Of the many DT-based therapeutic strategies investigated to date, two immunotoxins, OntakTM and TagraxofuspTM, have gained FDA approval for clinical application. Despite some success with immunotoxins, suicide-gene therapy strategies, whereby controlled tumor-specific expression of DT is used for the eradication of malignant cells, are gaining prominence. The first part of this review focuses on DT-based immunotoxins, and it then discusses recent developments in tumor-specific expression of DT.
| S-EPMC6813239 | biostudies-literature
Project description:To establish an inducible liver injury mouse model and transplant human hepatocytes to obtain liver-humanized mice.We crossed three mouse strains, including albumin (Alb)-cre transgenic mice, inducible diphtheria toxin receptor (DTR) transgenic mice and severe combined immune deficient (SCID)-beige mice, to create Alb-cre/DTR/SCID-beige (ADSB) mice, which coincidentally harbor Alb-cre and DTR transgenes and are immunodeficient. As the Cre expression is driven by the liver-specific promoter Alb (encoding ALB), the DTR stop signal flanked by two loxP sites can be deleted in the ADSB mice, resulting in DTR expression in the liver. ADSB mice aged 8-10 wk were injected intraperitoneally (i.p.) with diphtheria toxin (DT) and liver damage was assessed by serum alanine aminotransferase (ALT) level. Two days later, mouse livers were sampled for histological analysis, and human hepatocytes were transplanted into the livers on the same day. A human ALB enzyme-linked immunosorbent assay was performed 7, 14, 21 and 28 d after transplantation. Human CD68 immunohistochemistry was performed 30 and 90 d after transplantation.We crossed Alb-cre with DTR and SCID-beige mice to obtain ADSB mice. These mice were found to have liver damage 4 d after i.p. injection of 2.5 ng/g bodyweight DT. Bodyweight began to decrease on day 2, increased on day 7, and was lowest on day 4 (range, 10.5%-13.4%). Serum ALT activity began to increase on day 2 and reached a peak value of 289.7 ± 16.2 IU/mL on day 4, then returned to background values on day 7. After transplantation of human liver cells, peripheral blood human ALB level was 1580 ± 454.8 ng/mL (range, 750.2-3064.9 ng/mL) after 28 d and Kupffer cells were present in the liver at 30 d in ADSB mice.Human hepatocytes were successfully repopulated in the livers of ADSB mice. The inducible mouse model of humanized liver in ADSB mice may have functional applications, such as hepatocyte transplantation, hepatic regeneration and drug metabolism.
| S-EPMC5526763 | biostudies-other
Project description:Epidermal growth factor receptor (EGFR) is often overexpressed in head and neck squamous cell carcinoma (HNSCC) and represents a top candidate for targeted HNSCC therapy. However, the clinical effectiveness of current Food and Drug Administration (FDA)-approved drugs targeting EGFR is moderate, and the overall survival rate for HNSCC patients remains low. Therefore, more effective treatments are urgently needed. In this study, we generated a novel diphtheria toxin-based bivalent human epidermal growth factor fusion toxin (bi-EGF-IT) to treat EGFR-expressing HNSCC. Bi-EGF-IT was tested for in vitro binding affinity, cytotoxicity, and specificity using 14 human EGFR-expressing HNSCC cell lines and three human EGFR-negative cancer cell lines. Bi-EGF-IT had increased binding affinity for EGFR-expressing HNSCC compared with the monovalent version (mono-EGF-IT), and both versions specifically depleted EGFR-positive HNSCC, but not EGFR-negative cell lines, in vitro. Bi-EGF-IT exhibited a comparable potency to that of the FDA-approved EGFR inhibitor, erlotinib, for inhibiting HNSCC tumor growth in vivo using both subcutaneous and orthotopic HNSCC xenograft mouse models. When tested in an experimental metastasis model, survival was significantly longer in the bi-EGF-IT treatment group than the erlotinib treatment group, with a significantly reduced number of metastases compared with mono-EGF-IT. In addition, in vivo off-target toxicities were significantly reduced in the bi-EGF-IT treatment group compared with the mono-EGF-IT group. These results demonstrate that bi-EGF-IT is more effective and markedly less toxic at inhibiting primary HNSCC tumor growth and metastasis than mono-EGF-IT and erlotinib. Thus, the novel bi-EGF-IT is a promising drug candidate for further development.
| S-EPMC8024719 | biostudies-literature
Project description:The TNF-like weak inducer of apoptosis (TWEAK; TNFSF12) receptor Fn14 (TNFRSF12A) is expressed at low levels in normal tissues but frequently highly expressed in a wide range of tumor types such as lung, melanoma, and breast, and therefore it is a potentially unique therapeutic target for these diverse tumor types. We have generated a recombinant protein containing a humanized, dimeric single-chain anti-fibroblast growth factor-inducible 14-kDa protein (Fn14) antibody fused to recombinant gelonin toxin as a potential therapeutic agent (designated hSGZ). The hSGZ immunotoxin is a highly potent and selective agent that kills Fn14-positive (Fn14(+)) tumor cells in vitro. Treatment of cells expressing the MDR protein MDR1 (ABCB1B) showed no cross-resistance to hSGZ. Induced overexpression of Fn14 levels in MCF7 cells through HER2 (ERBB2) signaling translated to an improved therapeutic index of hSGZ treatment. In combination with trastuzumab, hSGZ showed an additive or synergistic cytotoxic effect on HER2(+)/Fn14(+) breast cancer cell lines. Also, hSGZ treatment inhibited Erb3/Akt signaling in HER2-overexpressing breast cancer cells. Pharmacokinetic studies in mice revealed that hSGZ exhibited a biexponential clearance from plasma with a rapid initial clearance (t1/2? = 1.26 hours) followed by a seven-fold longer plasma half-life (t1/2? = 7.29 hours). At 24, 48, and 72 hours after injection, uptake of the hSGZ into tumors was 5.1, 4.8, and 4.7%ID/g, with a tumor-to-muscle ratio of 5.6, 6.2, and 9.0, respectively. Therapeutic efficacy studies showed significant tumor inhibition effects using an MDA-MB-231/Luc breast cancer xenograft model. Our findings show that hSGZ is an effective anticancer agent and a potential candidate for clinical studies.
| S-EPMC3805367 | biostudies-literature
Project description:Hepatocellular carcinoma (HCC) is a major global malignancy, responsible for >90% of primary liver cancers. Currently available therapeutic options have poor performances due to the highly heterogeneous nature of the tumor cells; recurrence is highly probable, and some patients develop resistances to the therapies. Accordingly, the development of a novel therapy is essential. We assessed gene therapy for HCC using a diphtheria toxin fragment A (DTA) gene-expressing plasmid, utilizing a non-viral hydrodynamics-based procedure. The antitumor effect of DTA expression in HCC cell lines (and alpha-fetoprotein (AFP) promoter selectivity) is assessed in vitro by examining HCC cell growth. Moreover, the effect and safety of the AFP promoter-selective DTA expression was examined in vivo using an HCC mice model established by the hydrodynamic gene delivery of the yes-associated protein (YAP)-expressing plasmid. The protein synthesis in DTA transfected cells is inhibited by the disappearance of tdTomato and GFP expression co-transfected upon the delivery of the DTA plasmid; the HCC cell growth is inhibited by the expression of DTA in HCC cells in an AFP promoter-selective manner. A significant inhibition of HCC occurrence and the suppression of the tumor marker of AFP and des-gamma-carboxy prothrombin can be seen in mice groups treated with hydrodynamic gene delivery of DTA, both 0 and 2 months after the YAP gene delivery. These results suggest that DTA gene therapy is effective for HCC.
| S-EPMC7072394 | biostudies-literature