Signature Development and Validation Protocol for an Epigenetic Assay in Diagnosing Lung Cancer

This clinical testing research study outlines the validation process for an epigenetic assay targeting host peripheral blood cell and the associated host DNA methylation signatures designed to diagnose lung cancer. The overall protocol process will involve three distinct stages representing three patient cohorts with up to 250 subjects per cohort across 5 populations of patients. Cohort one will act as signature development phase; cohort two will act as signature finalization phase; and cohort three will act as a validation cohort phase. Each cohort will include the following patient populations with up to 50 patients per population:

1. Lung cancer; No chemotherapy. Surgical resection or stereotactic body radiation therapy (SBRT) only
2. Lung cancer; With chemotherapy. All stages including remission
3. At risk group (Lung cancer screening population)
4. Control group with mixed comorbid disease EXCLUDING tobacco use of greater than 10 pack years (PY) or chronic obstructive pulmonary disease (COPD)
5. Healthy controls with no comorbid disease

• Epigenetic Signature Assay (Other): Up to 15 ml of blood will be collected from each patient at various time points throughout their 5 years of participation. DNA extraction, bisulfite conversion and analysis of epigenetic markers through PCR or next-generation sequencing will be performed. An epigenetic signature assay will then be identified.

Primary outcomes

• Identification of tumor-associated host methylation signature [5 years]: Genome-wide methylation profile of whole blood from lung cancer patients, pre-cancer patients, patients undergoing therapy, and control subjects.
• Technology development [5 years]: The investigators will develop array-based assays using whole-blood samples, focused on disease-specific methylation sites to provide early diagnosis, prognosis, and therapeutic efficacy prediction.
• Technology validation [1 year]: The investigators will validate identified blood-based circulating methylation signatures in patients with undiagnosed pulmonary nodules.

Secondary outcomes

• EORTC QLQ-C30 Questionnaire [5 years]: The European Organization for Research and Treatment of Cancer core quality of life questionnaire, (EORTC QLQ-C30) is a cancer-specific quality of life instrument designed to assess health-related quality of life of cancer patients and is intended to be supplemented by disease-specific questionnaires, such as the lung cancer specific module, the EORTC QLQ-LC13. The EORTC QLQ-C30 incorporates nine multi-item scales including five functional scales (Physical, Role, Cognitive, Emotional and Social Functioning); three symptom scales (Fatigue, Pain and Nausea/Vomiting); and a Global Health Status/QoL scale. Six single item scales are also included (Dyspnea, Insomnia, Appetite Loss, Constipation, Diarrhea and Financial Difficulties). All of the scales and single-item measures range in score from 0 to 100. A higher score represents a higher level of symptom burden.
• EORTC QLQ-LC13 Questionnaire [5 years]: The European Organization for Research and Treatment of Cancer quality of life questionnaire module on Lung Cancer, EORTC QLQ-LC13, is a supplementary questionnaire meant to be used in conjunction with the full QLQ-C30. The QLQ-LC13 incorporates one multi-item scale to assess dyspnea, and a series of single items assessing pain, coughing, sore mouth, dysphagia, peripheral neuropathy, alopecia, and hemoptysis. All of the scales and single-item measures range in score from 0 to 100. A higher score represents a higher level of symptom burden.
• SF-36v2 Questionnaire [5 years]: The short-form of 36 questions (SF-36v2) is a multipurpose, 36-item survey that measures eight domains of health: physical functioning, role limitations due to physical health, bodily pain, general health perceptions, vitality, social functioning, role limitations due to emotional problems, and mental health. It yields scale scores for each of these eight health domains, and two summary measures of physical and mental health. Higher scores indicate better health status.

Sex: All, ages: 18 Years to any, accepts healthy volunteers

Inclusion Criteria:

* 18 years old or older
* Patient at University of Maryland Baltimore Washington Medical Center
* Willing and able to consent to study procedures listed in the protocol
* Ability to speak and understand English

Exclusion Criteria:

* Younger than 18 years old
* Patient not cared for at University of Maryland Baltimore Washington Medical Center
* Unable to consent to study procedures listed in the protocol
* Unable to speak or understand English

• Results publication: Li Y, Fan Z, Meng Y, Liu S, Zhan H. Blood-based DNA methylation signatures in cancer: A systematic review. Biochim Biophys Acta Mol Basis Dis. 2023 Jan 1;1869(1):166583. doi: 10.1016/j.bbadis.2022.166583. Epub 2022 Oct 18. PMID 36270476
• Results publication: Terp SK, Stoico MP, Dybkaer K, Pedersen IS. Early diagnosis of ovarian cancer based on methylation profiles in peripheral blood cell-free DNA: a systematic review. Clin Epigenetics. 2023 Feb 14;15(1):24. doi: 10.1186/s13148-023-01440-w. PMID 36788585
• Results publication: Ibrahim J, Peeters M, Van Camp G, Op de Beeck K. Methylation biomarkers for early cancer detection and diagnosis: Current and future perspectives. Eur J Cancer. 2023 Jan;178:91-113. doi: 10.1016/j.ejca.2022.10.015. Epub 2022 Oct 27. PMID 36427394
• Results publication: Guo Y, Yin J, Dai Y, Guan Y, Chen P, Chen Y, Huang C, Lu YJ, Zhang L, Song D. A Novel CpG Methylation Risk Indicator for Predicting Prognosis in Bladder Cancer. Front Cell Dev Biol. 2021 Sep 1;9:642650. doi: 10.3389/fcell.2021.642650. eCollection 2021. PMID 34540821
• Results publication: Xie Y, Li P, Sun D, Qi Q, Ma S, Zhao Y, Zhang S, Wang T, Wang J, Li S, Gong T, Xu H, Xiong M, Li G, You C, Luo Z, Li J, Wang C, Du L. DNA Methylation-Based Testing in Peripheral Blood Mononuclear Cells Enables Accurate and Early Detection of Colorectal Cancer. Cancer Res. 2023 Nov 1;83(21):3636-3649. doi: 10.1158/0008-5472.CAN-22-3402. PMID 37602818
• Results publication: Wang T, Li P, Qi Q, Zhang S, Xie Y, Wang J, Liu S, Ma S, Li S, Gong T, Xu H, Xiong M, Li G, You C, Luo Z, Li J, Du L, Wang C. A multiplex blood-based assay targeting DNA methylation in PBMCs enables early detection of breast cancer. Nat Commun. 2023 Aug 7;14(1):4724. doi: 10.1038/s41467-023-40389-5. PMID 37550304