MP16-06 EXPLOITING NOVEL THERAPEUTIC TARGETS TO BLOCK METASTASIS OF CLEAR CELL RENAL CELL CARCINOMA (RCC)
Bibliographic record
Abstract
You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology I (MP16)1 Apr 2019MP16-06 EXPLOITING NOVEL THERAPEUTIC TARGETS TO BLOCK METASTASIS OF CLEAR CELL RENAL CELL CARCINOMA (RCC) Jan Rudzinski*, Natasha Govindasamy, Konstantin Stoletov, Adrian Fairey, and John Lewis Jan Rudzinski*Jan Rudzinski* More articles by this author , Natasha GovindasamyNatasha Govindasamy More articles by this author , Konstantin StoletovKonstantin Stoletov More articles by this author , Adrian FaireyAdrian Fairey More articles by this author , and John LewisJohn Lewis More articles by this author View All Author Informationhttps://doi.org/10.1097/01.JU.0000555342.90934.fcAboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVES: Metastatic RCC is incurable, and the current therapeutic paradigm is centered around controlling the disease burden to prolong overall survival. We conducted a whole human genome short hairpin RNA (shRNA) screen on human squamous cell carcinoma (Hep3) cells to identify a panel of novel functional genes that are required for productive cell motility and successful metastatic dissemination. One such novel protein target encodes for chromosome 14 open reading frame 142 (C14orf42), which has been demonstrated to be up regulated in metastatic clear cell RCC. The objective of our study was to characterize the impact of C14orf142 on clear cell RCC in vitro motility, invasion, and in vivo vascular extravasation. METHODS: Benign proximal convoluted tubule cells (PCT) and clear cell RCC cell lines 786-0 (derived from renal tissue) were obtained from American Type Culture Collection (ATCC). Targeted genomic editing to knockout (KO) C14orf142 was achieved with CRISPR-Cas9 system. Successful protein knockdown was validated using western blot analysis. To measure impact of gene KO on in vitro invasion we conducted the FITC-gelatin degradation assay. To measure the combined effect of invasion and productive cell migration in vitro we utilized the modified Boyden chamber assay coated with 0.1% gelatin. To study cancer cell vascular extravasation in vivo, 786-0 cells were injected IV into fertilized avian embryos. For statistical analysis, t-test was used to evaluate differences between groups with p value of ≤0.05 accepted as statistically significant. RESULTS: The baseline expression of C14orf142 was significantly higher in 786-0 (1.078AUD±0.11) compared to PCT(0.12AUD±0.04) (p≤0.05). The CRISPR-Cas9 targeted genomic editing resulted in generation of 786-0 clones with complete KO of C14orf142. The FITC-gelatin degradation assay demonstrated significant difference in gelatin degradation between 786-0 scramble and 786-0 CRISPR KO clones (100% vs 27%±5.56%, respectively, p≤0.05). The modified Boyden chamber assay also showed significant difference in productive cell migration between 786-0 scramble and 786-0 CRISPR KO clones (100% vs 42±3.46%, respectively, p≤0.05). Compared to 786-0 scramble clones, 786-0 CRISPR KO clones demonstrated significant reduction in extravasation on the avian embryo model (100% vs 46.33±8.37%, respectively, p≤0.05). CONCLUSIONS: Up regulation of novel target protein, C14orf142, in 786-0 clear cell RCC cells may play an important role in cancer cell invasion, productive cell migration, and vascular extravasation. Source of Funding: Kidney Cancer Research Network (KCRN); Kidney Cancer Canada (KCC); Canadian Institute of Health Research (CIHR) Edmonton, Canada© 2019 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 201Issue Supplement 4April 2019Page: e207-e208 Advertisement Copyright & Permissions© 2019 by American Urological Association Education and Research, Inc.MetricsAuthor Information Jan Rudzinski* More articles by this author Natasha Govindasamy More articles by this author Konstantin Stoletov More articles by this author Adrian Fairey More articles by this author John Lewis More articles by this author Expand All Advertisement PDF downloadLoading ...
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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 | 0.001 | 0.001 |
| Insufficient payload (model declined to judge) | 0.009 | 0.002 |
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".