Programming Thermoresponsiveness of NanoVelcro Substrates Enables Effective Purification of Circulating Tumor Cells in Lung Cancer Patients

Abstract

Unlike tumor biopsies that can be constrained by problems such as sampling bias, circulating tumor cells (CTCs) are regarded as the “liquid biopsy” of the tumor, providing convenient access to all disease sites, including primary tumor and fatal metastases. Although enumerating CTCs is of prognostic significance in solid tumors, it is conceivable that performing molecular and functional analyses on CTCs will reveal much significant insight into tumor biology to guide proper therapeutic intervention. We developed the Thermoresponsive NanoVelcro CTC purification system that can be digitally programmed to achieve an optimal performance for purifying CTCs from non-small cell lung cancer (NSCLC) patients. The performance of this unique CTC purification system was optimized by systematically modulating surface chemistry, flow rates, and heating/cooling cycles. By applying a physiologically endurable stimulation (i.e., temperature between 4 and 37 °C), the mild operational parameters allow minimum disruption to CTCs’ viability and molecular integrity. Subsequently, we were able to successfully demonstrate culture expansion and mutational analysis of the CTCs purified by this CTC purification system. Most excitingly, we adopted the combined use of the Thermoresponsive NanoVelcro system with downstream mutational analysis to monitor the disease evolution of an index NSCLC patient, highlighting its translational value in managing NSCLC.

Beyond the Capture of Circulating Tumor Cells: Next‐Generation Devices and Materials

“…Thef irst methods developed for CTC analysis were designed to facilitate the identification of these cells via immunostaining,b ut the use of destructive characterization was quickly recognized as ac onstraint that would limit downstream analysis of the genetics and proteomics of these cells.Releasing viable cells allows for further analysis such as quantitative PCR, whole genome sequencing,a nd xenograft studies, [54,63,64] which are essential for fully understanding cancer metastases.T his has prompted as earch for gentle conditions that could be used to release fragile CTCs from capture devices.O ver the last several years,av ariety of systems have permitted the efficient recovery of cancer cells after capture by using chemical, [65] enzymatic, [66,67] selfassembly, [68] mechanosensitive, [69] and thermal release [70,71] mechanisms (Figure 4). High levels of cellular viability have been achieved for cancer cells isolated with low levels of contaminating cells.This is anew capability that will enhance our understanding of the biological properties of CTCs and their medical relevance.…”

Microfluidics in Malignant Glioma Research and Precision Medicine

“…[41] The isolation of these cells from the blood stream serves as a minimally invasive, multiple time-point liquid biopsy that can inform patient status without invasive measures. These cells are present in the blood in very low numbers (less than 100 cells mL −1 of whole blood), are heterogeneous, and are very difficult to isolate as pure populations, inspiring the design and fabrication of novel microfluidic platforms to improve the efficacy of CTC capture through both affinity-based and affinity-free technologies [42–45] (Figure 3). Affinity-based cell separation relies on the presence of unique biomarkers on the cancer cell surface that can be recognized by a magnetic or a polymer bead coupled to a high-affinity ligand to selectively separate bound cells of interest.…”

“…CTC separation devices using tumor-specific labeling technologies have been used successfully to separate CTCs from blood samples from pancreatic, prostate, breast, colon, melanoma, and lung cancer patients to provide prognostic information. [42,51] However, the innate heterogeneity of biomarker expression and the uncertainty introduced by epithelial-to-mesenchymal transitions of CTCs could limit the efficacy of affinity-based methods. Alternatively, affinity-free methods including those based on filtration, [52] acoustophoresis, [53] dielectrophoresis, [54] dean flow, [55] and vortex technology [56] exploit the intrinsic physical differences among cell types to deplete non-CTCs from blood and enrich cancer cells.…”

Profilierung zirkulierender Tumorzellen mit Apparaturen und Materialien der nächsten Generation

“…Während der letzten Jahre wurde eine Reihe von Systemen entwickelt, die die effiziente Gewinnung von Tu morzellen nach der Einfangreaktion erlauben. Die Ablçsung erfolgt über chemische [65] und enzymatische Mechanismen, [66,67] über Selbstorganisation, [68] mechanosensitiv [69] oder thermisch [70,71] (Abbildung 4). Dabei wurde ein hoher Grad an Lebensfä-higkeit der Tu morzellen bei gleichzeitig geringer Kontamination durch andere Zellen erreicht.…”

“…[69] Der zweite Ansatz kann auch verwendet werden, um einzelne CTCs mechanisch freizusetzen. Eine weitere auf thermische Einflüsse reagierende Technik wird durch immobilisierte Polymerbürsten mçglich, die den anhängenden Antikçrper bei niedrigen Temperaturen internalisieren, [70,71] ein Effekt, mit dem man CTCs ablçsen kann, indem man ein entsprechend modifiziertes Substrat abkühlt. So lassen sich CTCs aus Patientenproben isolieren und die tumorrelevanten Mutationen durch Sequenzierung ermitteln.…”