Abstract 6472: Clonal lineage and somatic hypermutation analysis of chronic lymphocytic leukemia by long-amplicon IGH chain sequencing
Bibliographic record
Abstract
Abstract Background: Current next-generation sequencing (NGS) approaches for analyzing SHM commonly rely on multiplex primers targeting the framework 1 (FR1) or leader region of the IGH variable gene in combination with joining gene primers to amplify rearranged IGH chains from gDNA template. Limitations include the potential for joining gene mutations to interfere with primer binding and an inability to evaluate isotype. Here we present a method for translational research investigations of IGH chain SHM employing multiplex FR1 and isotype (constant gene) specific primers (Oncomine IGH-LR assay primers) to amplify IGH chains from RNA template. We evaluated performance by comparing SHM values obtained from NGS of RNA from 54 CLL samples amplified using Oncomine IGH-LR primers to values obtained by Sanger sequencing or NGS of RNA amplified using FR1 or leader region/J gene primers. Methods: IGH chains from 54 CLL samples derived from two separate sequencing sites (Site 1: 24 samples, Site 2: 30 samples) were amplified from peripheral blood using Oncomine IGH-LR assay followed by sequencing via the Ion Gene Studio S5. Clonotyping, clonal lineage identification and somatic hypermutation analysis was performed by Ion Reporter via comparison to the IMGT reference database. Oncomine IGH-LR assay SHM values were compared to those obtained via NGS-based sequencing utilizing FR1/J gene primers (Site 1) or Sanger sequencing utilizing IGH-leader or FR1 and joining gene primers (Site 2). Results: IGHV SHM values were highly concordant between NGS approaches (Spearman cor =.957, Site 1) and between Sanger sequencing and NGS approaches (Spearman cor = .849, Site 2). Sequence data obtained using Oncomine IGH-LR assay enables more in-depth clonal lineage analysis, including evaluation of isotype representation and subclonal evolution. Conclusions: These results support the robustness and reliability of multiplex FR1 and constant gene based IGH chain amplification for the translational research characterization of somatic hypermutation in CLL and other B cell neoplasms. Citation Format: Jayde Chang, Zadie Davis, Graeme Quest, Harriet Feilloter, Michelle Toro, Geoffrey Lowman, Loni Pickle, Fiona Hyland, Timothy Looney. Clonal lineage and somatic hypermutation analysis of chronic lymphocytic leukemia by long-amplicon IGH chain sequencing [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6472.
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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 | 0.001 | 0.001 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.001 | 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) | 0.002 | 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".