In silico studies of green fluorescent protein-tagged Hsp27-HPV16 E7 fusion protein and evaluation of cytokine secretion from antigen-presenting cells exposed to the fusion protein-carrying tumor-derived exosomes
Bibliographic record
Abstract
Objective: Exosome (Exo)-based therapies have attracted considerable interest due to their potential as carriers for therapeutic molecules and their capacity to elicit anti-tumor immune responses. The objective of this study was to engineer TC-1 tumor cell line-derived exosomes with GFP-tagged heat shock protein (Hsp) 27-human papillomavirus (HPV)16 E7 fusion protein and evaluation of cytokine secretion from antigen-presenting cells (APCs: macrophages and dendritic cells) exposed to the engineered exosomes in vitro.Methods: In this study, different in silico methods were employed to evaluate the Hsp27-E7 and Hsp27-E7-GFP fusion proteins as potential vaccine candidates. Regarding to the in silico data, the Hsp27-E7-GFP fusion gene was subcloned into pCDH lentiviral vector for production of lentivirions harboring the Hsp27-E7-GFP fusion protein in eukaryotic cells. Subsequently, the TC-1 tumor cells were transduced with these lentivirions to isolate the engineered exosomes (i.e., Exo-Hsp27-E7-GFP) using the ExoQuick-TCTM kit and their characterization using physicochemical methods. Finally, the secretion of key cytokines (IFN-γ, TNF-α, and IL-10) was evaluated through incubation of antigen-presenting cells (APCs) with the engineered Exo-Hsp27-E7-GFP using enzyme-linked immunosorbent assay (ELISA).Results: Our in silico data showed that both the Hsp27-E7 and Hsp27-E7-GFP constructs were soluble and non-allergenic, and exhibited strong interaction with TLR4. Indeed, the linkage of GFP did not affect the physicochemical properties, and interaction of Hsp27-E7 with the immune receptors. Moreover, western blot analysis confirmed the presence of Hsp27-E7-GFP fusion protein in the isolated exosomes. The Exo-Hsp27-E7-GFP could significantly enhance the secretion of TNF-α and IL-10 from APCs compared to Exo and Exo-GFP, as well.Conclusions: These engineered vesicles derived from tumor cells demonstrate the capacity to induce effective immunity, suggesting their potential as a promising strategy in the development of cell-free vaccine candidates.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.003 | 0.001 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 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 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
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 teacher head, 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".