Circulating Tumor Cell Detection and Capture by Photoacoustic Flow Cytometry in Vivo and ex Vivo

Abstract

Despite progress in detecting circulating tumor cells (CTCs), existing assays still have low sensitivity (1–10 CTC/mL) due to the small volume of blood samples (5–10 mL). Consequently, they can miss up to 103–104 CTCs, resulting in the development of barely treatable metastasis. Here we analyze a new concept of in vivo CTC detection with enhanced sensitivity (up to 102–103 times) by the examination of the entire blood volume in vivo (5 L in adults). We focus on in vivo photoacoustic (PA) flow cytometry (PAFC) of CTCs using label-free or targeted detection, photoswitchable nanoparticles with ultrasharp PA resonances, magnetic trapping with fiber-magnetic-PA probes, optical clearance, real-time spectral identification, nonlinear signal amplification, and the integration with PAFC in vitro. We demonstrate PAFC’s capability to detect rare leukemia, squamous carcinoma, melanoma, and bulk and stem breast CTCs and its clusters in preclinical animal models in blood, lymph, bone, and cerebrospinal fluid, as well as the release of CTCs from primary tumors triggered by palpation, biopsy or surgery, increasing the risk of metastasis. CTC lifetime as a balance between intravasation and extravasation rates was in the range of 0.5–4 h depending on a CTC metastatic potential. We introduced theranostics of CTCs as an integration of nanobubble-enhanced PA diagnosis, photothermal therapy, and feedback through CTC counting. In vivo data were verified with in vitro PAFC demonstrating a higher sensitivity (1 CTC/40 mL) and throughput (up to 10 mL/min) than conventional assays. Further developments include detection of circulating cancer-associated microparticles, and super-resolution PAFC beyond the diffraction and spectral limits.

Circulating tumor cell technologies

“…To date, no clinical studies applying CytoTrack's technology have been published. Another unique, label-free method to detect CTCs is photoacoustic flow cytometry (PAFC) (Galanzha and Zharov, 2013). PAFC uses laser-based technology for real-time detection of CTCs in veins by interrogating the blood flow through the skin.…”

“…Absorption spectra Non-invasive label-free interrogation of large blood volumes (Galanzha and Zharov, 2013) Functional Assays Viable CTC enrichment based on bioactivity of cells, such as protein secretion or cell adhesion. epithelial-to-mesencyhmal transition (EMT) and stem cell markers in CTCs has given researchers pause as they struggle to determine the significance of different CTC subpopulations.…”

High frame rate photoacoustic imaging at 7000 frames per second using clinical ultrasound system

“…Imaging CTC is most commonly performed ex vivo only. However, recently in vivo photoacoustic flow cytometry has been demonstrated with the injection of external contrast agent (gold nanoparticles) with good PA contrast [29,30]. Since, melanoma has strong light absorption in the NIR wavelengths; it may not require any external contrast for the detection.…”

Label-Free Photoacoustic Cell-Tracking In Real-Time

“…The ability to follow the distribution and migration of cells in living organisms is crucial for both the development of metastatic circulating tumor cells (CTCs) detection and cell-based therapies [1,2]. Several technologies exist that are able to noninvasively track cells in vivo, each with its own advantages and disadvantages in terms of cell tracking performance.…”