Abstract A053 Investigating mechanisms responsible for MAP kinase pathway resistance in RAS-altered neuroblastoma cells
Bibliographic record
Abstract
Abstract Neuroblastoma (NB) is the most common pediatric solid tumor, responsible for about 15% of childhood cancer-related mortality in the United States. Multiple therapies have been established, which include chemotherapy, radiotherapy, immunotherapy and molecularly-targeted therapy, however resistance to these therapies is the major hinderance in curing NB. RAS pathway mutations are prevalent in relapsed NB and are associated with a poorer overall survival. Therefore, comprehending the mechanisms leading to therapy resistance has become an urgent need. In this study, we attempted to identify the resistance mechanism of NB cell lines towards RAS inhibitors. RAS-altered NB cell lines are treated with trametinib, a MAP kinase pathway inhibitor to perform cell viability assays, immunoblotting and RT PCR. Copy number alteration and translocation studies are performed using spectral karyotyping (SKY) and multiplex interphase fluorescent in-situ hybridization (miFISH). We observed that unlike other RAS-altered NB cell lines (SK-N-AS, CHP-212, LA-N-6, SK-N-BE(2)-C), NB-Eb-C1, which harbors a KRAS G12D mutation shows resistance to both MRTX1133 (KRAS G12D direct inhibitor) and trametinib. Immunoblot against crucial genes regulating NB showed high expression of MYCN, the most relevant biomarker of high-risk neuroblastoma. MYCN is unfortunately undruggable hence, standard treatment for MYCN-amplified neuroblastoma patients does not include targeting MYCN itself. We therefore pursued a hypothesis-driven approach towards identifying possible mechanisms responsible for MYCN driven MEKi-resistance in RAS pathway-altered NB cells. MYCN is known to cross talk with the RAS pathway, however this mechanism is not well understood in NB. Therefore, we performed the clonal assessment of NB cell lines using miFISH and to the best of our knowledge are the first to report that, NB-Eb-C1 has a hypertriploid to hypotetraploid baseline with a low copy-number gain of MYCN. To have a profound understanding of the functionality of MYCN in NB, we have taken 3 different NB cell lines with varied MYCN status. CHP-212 (MYCN amplified-extrachromosomal), SK-N-BE(2)-C (MYCN amplified-homogenously staining region), SK-N-AS (MYCN neutral). SKY and FISH revealed that the MYCN copy-number gain in NBEB cells is the result of a translocation between chromosome 2 and 5 involving MYCN. Of note, another frequently abnormally expressed gene in NB, ALK, is co-translocated with MYCN suggesting potential involvement of both genes in imparting chemoresistance. To understand the mechanism of MYCN overexpression in NBEB we will be performing HiC to check for promoter/enhancer hijacking. Also, to better define this translocation and studying other translocations involved we are performing optical genome mapping using BioNano. Other functional experiments will be performed to look at the impact of the RAS/MAPK alteration on MYCN protein stability. We are confident that this study will contribute towards developing new combinations of targeted therapies to improve outcomes in RAS-altered NB. Citation Format: Subhra Dash, Kerstin Heselmeyer-Haddad, Stacy Stauffer, Lucas Stauffer, Danny Wangsa, Marielli E. Yohe. Investigating mechanisms responsible for MAP kinase pathway resistance in RAS-altered neuroblastoma cells [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pediatric Cancer Research; 2024 Sep 5-8; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl):Abstract nr A053.
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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 | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.003 | 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".