Quantifying EpCAM heterogeneity of circulation-tumor-cells (CTCs) from small cell lung cancer (SCLC) patients.

H.Sorotsky, M.Aparanthi, D.Z.Wang, F.McFadden, S.N.Popescu, R.M.Mohamadi, M.Pereira, J.Weiss, D.Patel, S.Majeed, M.Cabanero, A.G.Sacher, P.A.Bradbury, N.B.Leighl, F.A.Shepherd, M.S.Tsao, G.Lui, S.O.Kelly, B.H.Lok

Objectives: To investigate the effect of pazopanib on different CTCs subpopulations in patients with recurrent SCLC and evaluate their clinical relevance.

We kindly thank the researchers at University of Toronto for this collaboration, and for sharing the results obtained with their system.

Methods

Methods: Blood samples from 20 SCLC pts were processed through the MagRC platform. Magnetic nanoparticles conjugated with anti-EpCAM antibodies were incubated with whole blood samples then introduced into the MagRC device where CTCs are sorted by differently sized nickel micromagnets within microfluidic channels. Captured CTCs are ranked into 8 zones that correlate with EpCAM expression levels (zone 1 = highest to 8 = lowest). For 8 pts, all samples were processed at a 1mL/hr flow rate (fr), and for 12 pts, a 0.5mL/hr fr was also studied; 66% of all chips were processed at a 1ml/hr fr and 34% at a 0.5ml/hr fr. The average zone for each chip was compared to the flow rate, age, and stage (extensive-stage (ES) vs limited-stage (LS)). The differences were tested using the Wald test within the linear mixed effects model.

Materials

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Results

Before treatment, CTCs could be detected in 50% of patients by CellSearch; phenotypic characterization of CTCs demonstrated that 50%, 46.6% and 27.6% of patients had CD45-/TTF1+, CD45-/CD56+ and TTF-1+/CD56+ CTCs, respectively. Additionally, 59% of CTCs were TTF-1+/VEGFR2+ and 53% CK+/VEGFR2+. One pazopanib cycle resulted to a significant decrease of the number of CTCs (CellSearch: p=0.043) and CK+/VEGFR2+ cells (p=0.027). At the time of PD, both the total number of CTCs (p=0.027) and the number of the different subpopulations were significantly increased compared to post-1st cycle values; this increased CTCs number was associated with a significant increase of TTF-1+/VEGFR2+ (p=0.028) and CK+/VEGFR2+ CTCs (p=0.018). In multivariate analysis, only the number of CTCs as assessed by CellSearch after one treatment cycle was significantly associated with OS (HR: 0.21; p=0.005).

Conclusions: Pazopanib has a significant effect on different subpopulations of CTCs in patients with relapsed SCLC; the detection of VEGFR2+ CTCs during treatment could be a surrogate marker associated with resistance to pazopanib.

Keywords: CD56; CTCs; CellSearch; Immunofluorescence; Pazopanib; SCLC; TTF-1; VEGFR2.