154P - Alternative splicing in non-small cell lung cancer evolution

Background

Alternative splicing allows a single gene to encode different transcripts and has been shown to occur frequently in cancer. Alternative splicing may be important for tumour evolution and immune evasion as it can result in a more diverse proteome and regulate expression levels. However, alternative splicing’s role in non-small cell lung cancer (NSCLC) evolution remains unclear.

Methods

We developed our own bioinformatics pipeline that extracts splice junctions from sequencing data and investigated alternative splicing in the multiregional TRACERx421 cohort, consisting RNAseq data from 947 primary NSCLC tumour regions from 354 patients and 96 tumour-adjacent normal samples. In 14 patients, we validated our pipeline using an orthogonal method that detects MET exon 14 skipping and found a 100% concordance rate.

Results

Using our pipeline, we found similar levels of total splicing between tumour and normal samples. Most splicing events were ubiquitous: 86% and 92% of splicing events in LUAD and LUSC tumours respectively occurred in all regions of the given tumour. Interestingly, the fraction of non-ubiquitous splicing events per tumour was highly correlated with the mean intratumor expression distance, a measure of transcriptional diversity. It also explained an additional 7.37% of intratumor heterogeneity, adding to the impact of other selected genomic and clinical features described in Martínez-Ruiz et al. 2023. Moreover, we found non-transcript-disrupting RB1 exon 21 skipping in four tumours, occurring at high transcript frequency, that could disrupt RB1-E2F binding and consequently increase cell proliferation.

Conclusions

Overall, we assessed alternative splicing’s role in NSCLC tumour evolution. Going forward, we plan to explore the impact of splicing on neoantigen expression to evaluate the potential role of this process in tumour-immune evasion.

Editorial acknowledgement

Clinical trial identification

Legal entity responsible for the study

The authors.

Funding

CRUK, Rosetrees Trust.

Disclosure

C. Swanton: Financial Interests, Personal, Invited Speaker, Activity took place in 2016: Pfizer, Celgene; Financial Interests, Personal, Invited Speaker, October 26th 2020: Novartis; Financial Interests, Personal, Invited Speaker: Roche/Ventana, BMS, AstraZeneca, MSD, Illumina, GSK; Financial Interests, Personal, Advisory Board, Ad Board - November 12th, 2020: Amgen; Financial Interests, Personal, Advisory Board, Current - since 2018: Genentech; Financial Interests, Personal, Advisory Board: Sarah Canon Research Institute; Financial Interests, Personal, Advisory Board, Joined October 2020. Also have stock options: Bicycle Therapeutics; Financial Interests, Personal, Other, Consultancy: Medicxi; Financial Interests, Personal, Advisory Board, Member of the Science Advisory Board. Also had stock options until June 2021: GRAIL; Financial Interests, Personal, Other, Consultancy agreement: Roche Innovation Centre Shanghai; Financial Interests, Personal, Advisory Board, 29 November - 1 December 2022: Novartis; Financial Interests, Personal, Invited Speaker, Oncology Collective - 2nd Nov - 4 Nov 2022 - Atlanta, USA: Roche; Financial Interests, Personal, Advisory Board, ctDNA advisory Board - 24th March 2023: AstraZeneca; Financial Interests, Personal, Invited Speaker, Pfizer Oncology 'Leading the revolution for the future: Pfizer; Financial Interests, Personal, Advisory Board, Scientific Advisory Board and Stock options from September 2023: Relay Therapeutics; Financial Interests, Personal, Advisory Board, Member of the Scientific Advisory Board: SAGA Diagnostics; Financial Interests, Personal, Full or part-time Employment, Chief Clinician since October 2017: Cancer Research UK; Financial Interests, Personal, Ownership Interest, Co-Founder of Achilles Therapeutics. Also, have stock options in this company: Achilles Therapeutics; Financial Interests, Personal, Stocks/Shares, Stocks owned until June 2021: GRAIL, Apogen Biotechnologies; Financial Interests, Personal, Stocks/Shares: Epic Biosciences, Bicycle Therapeutics; Financial Interests, Personal, Stocks/Shares, Stock options: Relay Therapeutics; Financial Interests, Institutional, Research Grant, Funded RUBICON grant - October 2018 - April 2021: Bristol Myers Squibb; Financial Interests, Institutional, Research Grant, Collaboration in minimal residual disease sequencing technologies: Archer Dx Inc; Financial Interests, Institutional, Research Grant: Pfizer, Boehringer Ingelheim; Financial Interests, Institutional, Invited Speaker, Chief Investigator for the MeRmaiD 1and 2 clinical trials and chair of the steering committee: AstraZeneca; Financial Interests, Institutional, Research Grant, Research grant from Oct 2019 - July 2023 - Genetics of CIN and SCNAs for Targeted Discovery (SCEPTRE): Ono Pharmaceutical; Financial Interests, Institutional, Research Grant, Research Grants from 2015: Roche; Financial Interests, Personal, Other, Co-chief investigator: NHS-Galleri Clinical Trial; Financial Interests, Institutional, Research Grant, from October 2022: Personalis; Non-Financial Interests, Personal, Principal Investigator, Chief Investigator for MeRmaiD 1and 2 clinical trials: AstraZeneca; Non-Financial Interests, Personal, Member of Board of Directors, From 2019-2022: AACR; Non-Financial Interests, Personal, Other, Board of Directors: AACR; Non-Financial Interests, Personal, Advisory Role, EACR Advisory Council member: EACR. N. McGranahan: Financial Interests, Institutional, Stocks/Shares: Achilles Therapeutics. All other authors have declared no conflicts of interest.