The Use of Three-Dimensional DNA Fluorescent In Situ Hybridization (3D DNA FISH) for the Detection of Anaplastic Lymphoma Kinase (ALK) in Non-Small Cell Lung Cancer (NSCLC) Circulating Tumor Cells

Abstract

Tumor tissue biopsy is often limited for non-small cell lung cancer (NSCLC) patients and alternative sources of tumoral information are desirable to determine molecular alterations such as anaplastic lymphoma kinase (ALK) rearrangements. Circulating tumor cells (CTCs) are an appealing component of liquid biopsies, which can be sampled serially over the course of treatment. In this study, we enrolled a cohort of ALK-positive (n = 8) and ALK-negative (n = 12) NSCLC patients, enriched for CTCs using spiral microfluidic technology and performed DNA fluorescent in situ hybridization (FISH) for ALK. CTCs were identified in 12/20 NSCLC patients ranging from 1 to 26 CTCs/7.5 mL blood. Our study revealed that 3D imaging of CTCs for ALK translocations captured a well-defined separation of 3′ and 5′ signals indicative of ALK translocations and overlapping 3′/5′ signal was easily resolved by imaging through the nuclear volume. This study provides proof-of-principle for the use of 3D DNA FISH in the determination of CTC ALK translocations in NSCLC.

Heterogeneous Tumor-Immune Microenvironments between Primary and Metastatic Tumors in a Patient with ALK Rearrangement-Positive Large Cell Neuroendocrine Carcinoma

“…The enumeration and characterization of CTCs provided a minimally invasive and, therefore, repeatable, method despite being present in extremely low numbers, enabling the sampling of tumor cells from peripheral blood and monitoring PD-L1 expression on tumor cells over time. In addition, EML4-ALK rearrangements have been reported to be found in CTCs [ 56 ]. Thus, combining CTC and peripheral immune subset analyses may make it possible to longitudinally evaluate serial human specimens during treatment (at pre-treatment, early-on-treatment, and progression time points), which allow for deep analysis to unveil potential mechanisms of therapeutic resistance [ 16 , 50 , 57 , 58 ].…”

Epidermal Growth Factor Receptor (EGFR)-Mutated Non-Small-Cell Lung Cancer (NSCLC)

“…Molecular characterization of CTCs has revealed that genomic changes of the tumor can be readily assessed via CTCs. This was recently highlighted by a number of studies identifying translocations/mutations in anaplastic lymphoma kinase (ALK) and EGFR in CTCs [ 74 , 75 , 76 , 77 , 78 ]. For EGFR mutations, the sensitivity of the assay increased with higher numbers of detected CTCs with concordance between EGFR mutation status in the tumor and CTC samples [ 77 ].…”

Genomic Instability in Circulating Tumor Cells

“…In clinical practice, CTCs represent a real-time, minimally invasive alternative source of tumor material. CTCs can be used to assess the landscape of primary and metastatic tumor in the body over time by sampling the blood serially over the course of treatment [ 147 ]. CTCs information can be used to monitor disease progression and to predict treatment response.…”

“…CTCs information can be used to monitor disease progression and to predict treatment response. The analysis of CTCs can also be used to select patients for targeted therapies [ 77 , 147 ].…”

“…A number of studies have reported the presence of ALK-translocated CTCs in blood from NSCLC patients [ 80 , 147 , 148 ]. Provencio et al, 2017, used CTCs for the dynamic monitoring of NSCLC patients with ALK rearrangement, as an example [ 80 ].…”