To study the NSCLC evolutionary genomic landscape between primary and metastatic sites and the dynamics of intratumour heterogeneity over time combined with detailed clinical, histopathological and cancer phenotypic annotation for each patient, in order to significantly improve the outcomes of NSCLC patients (e.g. reduce their chance of recurrence and improve survival).

The importance of intratumour genetic and functional heterogeneity is increasingly recognised as a driver of cancer progression and survival outcome. Understanding how tumour clonal heterogeneity impacts upon therapeutic outcome, however, is still an area of unmet clinical and scientific need. TRACERx (TRAcking non-small cell lung Cancer Evolution through therapy [Rx]), a prospective study of patients with primary non-small cell lung cancer (NSCLC), aims to define the evolutionary trajectories of lung cancer in both space and time through multiregion and longitudinal tumour sampling and genetic analysis. By following cancers from diagnosis to relapse, tracking the evolutionary trajectories of tumours in relation to therapeutic interventions, and determining the impact of clonal heterogeneity on clinical outcomes, TRACERx may help to identify novel therapeutic targets for NSCLC and may also serve as a model applicable to other cancer types.

Inclusion Criteria:

• Written Informed consent
• Patients ≥18 years of age, with early stage IIA-IIIB disease (according to TNM 8th edition) who are eligible for primary surgery. Patients with a radiological staging of IB (N0) who could be upstaged to IIA-IIIB following surgery (due to the presence of possible nodal involvement on the pre-operative scan) may also be included, but will be withdrawn if post-surgical staging remains IB.

Histopathologically confirmed NSCLC, or a strong suspicion of cancer on lung imaging necessitating surgery (e.g. diagnosis determined from frozen section in theatre)

• Primary surgery in keeping with NICE guidelines planned (see section 9.3)
• Agreement to be followed up at a TRACERx site
• Performance status 0 or 1
• Minimum tumour diameter at least 15mm to allow for sampling of at least two tumour regions (if 15mm, a high likelihood of nodal involvement on pre-operative imaging required to meet eligibility according to stage, i.e. T1N1-3)

Exclusion Criteria:

• Any other* malignancy diagnosed or relapsed at any time, which is currently being treated (including by hormonal therapy).

• Any other* current malignancy or malignancy diagnosed or relapsed within the past 3 years**.

  *Exceptions are: non-melanomatous skin cancer, stage 0 melanoma in situ, and in situ cervical cancer

  **An exception will be made for malignancies diagnosed or relapsed more than 2, but less than 3, years ago only if a pre-operative biopsy of the lung lesion has confirmed a diagnosis of NSCLC.

• Psychological condition that would preclude informed consent

• Treatment with neo-adjuvant therapy for current lung malignancy deemed necessary

• Post-surgery staging is not IIA-IIIB

• Known Human Immunodeficiency Virus (HIV), Hepatitis B Virus (HBV), Hepatitis C Virus (HCV) or syphilis infection.

• Sufficient tissue, i.e. a minimum of two tumour regions, is unlikely to be obtained for the study based on pre-operative imaging

Stage I-IIIA NSCLC eligible for primary surgery