VB6-845: recombinant engineering of an Ep-CAM-specific Fab antibody for optimal delivery of de-immunized bouganin (de-bouganin)
Bibliographic record
Abstract
Proc Amer Assoc Cancer Res, Volume 46, 2005 683 Tumor-targeting immunotoxins are composed of the variable regions of an antibody linked to a bacterial, fungal or plant toxin. Although several trials have shown promising clinical benefits, the induction of a human anti-toxin antibody response (HATA) is still a major limitation to achieving therapeutic success. At Viventia, VB5-845, a Fab version of an anti-EpCAM scFv antibody, was genetically linked to a de-immunized form of bouganin (de-bouganin), a potent, plant-derived, type I ribosome-inactivating protein (RIP), to create the antibody-toxin construct VB6-845. To determine the optimal antibody-de-bouganin orientation, several forms of a dicistronic expression unit were generated, expressed and tested for potency. In each case, the dicistronic unit was cloned into the pING3302 vector under the control of the arabinose-inducible araBAD promoter and transformed in E104 E.coli . Upon induction, the presence of the PelB leader sequence directed the secretion of the Fab-de-bouganin fusion protein into the culture supernatant. Two of several constructs tested comprised a pelB leader sequence adjacent to either the VH-CH or VL-CL domains linked to de-bouganin by a protease-sensitive linker. Immediately following was the second unit comprising either, pelB-VL-CL or the pelB-VH-CH domain as well as an N-terminal histidine affinity tag. For constructs where de-bouganin was re-positioned to the N-terminal end, Western-blot analysis showed no detectable product and only C-terminal linked de-bouganin yielded an intact soluble protein. Similarly, the addition of a second de-bouganin domain in the molecule with the optimal orientation did not yield product. A competition assay with Proxinium™, a scFv version of VB6-845, but containing a variant of Pseudomonas exotoxin A, demonstrated that the Ep-CAM specificity of VB6-845 was unaltered when engineered into a Fab format. In addition, cell-free and MTS assays demonstrated that de-bouganin retained its potency when conjugated to the Fab fragment. Using Ep-CAM positive cell lines, CAL-27 and OVCAR-3, the IC50 of VB6-845 was 1 to 2 nM. The development of VB6-845, comprising a tumor-targeting antibody fragment linked to de-immunized bouganin should permit repeat systemic administration of this drug and hence offer a greater clinical benefit.
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".