Phenotypic Plasticity in Circulating Tumor Cells Is Associated with Poor Response to Therapy in Metastatic Breast Cancer Patients.

Circulating tumor cells (CTCs) are indicators of metastatic spread and progression. In a longitudinal, single-center trial of patients with metastatic breast cancer starting a new line of treatment, a microcavity array was used to enrich CTCs from 184 patients at up to 9 timepoints at 3-month intervals. CTCs were analyzed in parallel samples from the same blood draw by imaging and by gene expression profiling to capture CTC phenotypic plasticity.

Gene expression profiling of circulating tumor cells captured by MicroCavity Array is superior to enumeration in demonstrating therapy response in patients with newly diagnosed advanced and locally advanced non-small cell lung cancer.

Background: Circulating tumor cells (CTCs) are a promising non-invasive tool for monitoring therapy response. The only Food and Drug Administration (FDA)-approved test is limited to enumeration of epithelial CTC without further characterization and is not approved for the management of non-small cell lung cancer (NSCLC). Here we use a MicroCavity Array (MCA) system to capture CTC agnostic of epithelial markers for further molecular testing in NSCLC.

A Multi-Center Clinical Study to Harvest and Characterize Circulating Tumor Cells from Patients with Metastatic Breast Cancer Using the Parsortix PC1 System.

Circulating tumor cells (CTCs) captured from the blood of cancer patients may serve as a surrogate source of tumor material that can be obtained via a venipuncture (also known as a liquid biopsy) and used to better understand tumor characteristics. However, the only FDA-cleared CTC assay has been limited to the enumeration of surface marker-defined cells and not further characterization of the CTCs. In this study, we tested the ability of a semi-automated device capable of capturing and harvesting CTCs from peripheral blood based on cell size and deformability, agnostic of cell-surface markers (the Parsortix PC1 System), to yield CTCs for evaluation by downstream techniques commonly available in clinical laboratories.

Ensemble of nucleic acid absolute quantitation modules for copy number variation detection and RNA profiling.

Current gold standard for absolute quantitation of a specific DNA sequence is droplet digital PCR (ddPCR), which has been applied to copy number variation (CNV) detection. However, the number of quantitation modules in ddPCR is limited by fluorescence channels, which thus limits the CNV sensitivity due to sampling error following Poisson distribution. Here we develop a PCR-based molecular barcoding NGS approach, quantitative amplicon sequencing (QASeq), for accurate absolute quantitation scalable to over 200 quantitation modules.

Enumeration and molecular characterization of circulating tumor cells enriched by microcavity array from stage III non-small cell lung cancer patients.

Background: Various methods of liquid biopsy through the sampling of blood in cancer patients allow access to minuscule amounts of tumor that can easily be sampled repeatedly throughout therapy. Circulating tumor cells (CTCs) represent shed tumor cells that can be characterized by imaging or molecular techniques using an amenable enrichment platform. Here we validate the Hitachi Chemical Micro Cavity Array (MCA) for the enrichment of CTCs from the blood of patients diagnosed with stage III non-small cell lung cancer (NSCLC).

Antigen-agnostic microfluidics-based circulating tumor cell enrichment and downstream molecular characterization.

Circulating tumor cells (CTC) isolated from the peripheral blood of cancer patients by a minimally invasive procedure provide surrogate markers of the tumor that can be repeatedly sampled. However, the selection and enumeration of CTCs by traditional methods based on surface proteins like EPCAM may not detect CTCs with a mesenchymal phenotype. Here, we employed an antibody-agnostic platform, the Parsortix® PR1 system, which enriches CTCs based on cell size and membrane deformability.

Exogenous Restoration of TUSC2 Expression Induces Responsiveness to Erlotinib in Wildtype Epidermal Growth Factor Receptor (EGFR) Lung Cancer Cells through Context Specific Pathways Resulting in Enhanced Therapeutic Efficacy.

Expression of the tumor suppressor gene TUSC2 is reduced or absent in most lung cancers and is associated with worse overall survival. In this study, we restored TUSC2 gene expression in several wild type EGFR non-small cell lung cancer (NSCLC) cell lines resistant to the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor erlotinib and analyzed their sensitivity to erlotinib in vitro and in vivo. A significant inhibition of cell growth and colony formation was observed with TUSC2 transient and stable expression.

Phase I clinical trial of systemically administered TUSC2(FUS1)-nanoparticles mediating functional gene transfer in humans.

Background: Tumor suppressor gene TUSC2/FUS1 (TUSC2) is frequently inactivated early in lung cancer development. TUSC2 mediates apoptosis in cancer cells but not normal cells by upregulation of the intrinsic apoptotic pathway. No drug strategies currently exist targeting loss-of-function genetic abnormalities.

Analysis of protein-protein interaction using proteinchip array-based SELDI-TOF mass spectrometry.

Protein-protein interactions are key elements in the assembly of cellular regulatory and signaling protein complexes that integrate and transmit signals and information in controlling and regulating various cellular processes and functions. Many conventional methods of studying protein-protein interaction, such as the immuno-precipitation and immuno-blotting assay and the affinity-column pull-down and chromatographic analysis, are very time-consuming and labor intensive and lack accuracy and sensitivity. We have developed a simple, rapid, and sensitive assay using a ProteinChip array and SELDI-TOF mass spectrometry to analyze protein-protein interactions and map the crucial elements that are directly involved in these interactions.

High throughput profiling of serum phosphoproteins/peptides using the SELDI-TOF-MS platform.

Protein phosphorylation is a dynamic post-translational modification that plays a critical role in the regulation of a wide spectrum of biological events and cellular functions including signal transduction, gene expression, cell proliferation, and apoptosis. Determination of the sites and magnitudes of protein phosphorylation has been an essential step in the analysis of the control of many biological systems. A high throughput analysis of phosphorylation of proteins would provide a simple, logical, and useful tool for a functional dissection and prediction of biological functions and signaling pathways in association with these important molecular events.

Regulation of tumor suppressor gene FUS1 expression by the untranslated regions of mRNA in human lung cancer cells.

FUS1, also known as tumor suppressor candidate 2 (TUSC2), is a tumor suppressor gene located in the human chromosome 3p21.3 region. FUS1 mRNA transcripts could be detected on Northern blots in both normal lung and some lung cancer cell lines, but no endogenous FUS1 protein could be detected in a majority of lung cancer cell lines and small cell and non-small cell lung tumor tissues.

NPRL2 sensitizes human non-small cell lung cancer (NSCLC) cells to cisplatin treatment by regulating key components in the DNA repair pathway.

NPRL2, one of the tumor suppressor genes residing in a 120-kb homozygous deletion region of human chromosome band 3p21.3, has a high degree of amino acid sequence homology with the nitrogen permease regulator 2 (NPR2) yeast gene, and mutations of NPRL2 in yeast cells are associated with resistance to cisplatin-mediated cell killing. Previously, we showed that restoration of NPRL2 in NPRL2-negative and cisplatin-resistant cells resensitize lung cancer cells to cisplatin treatment in vitro and in vivo.

Fragile histidine triad-mediated tumor suppression of lung cancer by targeting multiple components of the Ras/Rho GTPase molecular switch.

The fragile histidine triad (FHIT) gene has been shown to function as a tumor suppressor gene in vitro and in vivo. However, the mechanism of its action is still largely unknown. To elucidate the molecular mechanism and biological pathway in FHIT-mediated tumor suppression, we used a complementary gene and protein expression profiling with DNA microarray and ProteinChip technologies to quantitatively monitor cellular changes in gene and protein expression and discover the molecular targets of FHIT in non-small cell lung carcinoma (NSCLC) cells.

Tumor-specific activation of human telomerase reverses transcriptase promoter activity by activating enhancer-binding protein-2beta in human lung cancer cells.

The up-regulated expression and telomerase activity of human telomerase reverse transcriptase (hTERT) are hallmarks of tumorigenesis. The hTERT promoter has been shown to promote hTERT gene expression selectively in tumor cells but not in normal cells. However, little is known about how tumor cells differentially activate hTERT transcription and induce telomerase activity.

Tumor suppressor 101F6 and ascorbate synergistically and selectively inhibit non-small cell lung cancer growth by caspase-independent apoptosis and autophagy.

101F6 is a candidate tumor suppressor gene harbored on chromosome 3p21.3, a region with frequent and early allele loss and genetic alterations in many human cancers. We previously showed that enforced expression of wild-type 101F6 by adenoviral vector-mediated gene transfer significantly inhibited tumor cell growth in 3p21.

Synergistic tumor suppression by coexpression of FUS1 and p53 is associated with down-regulation of murine double minute-2 and activation of the apoptotic protease-activating factor 1-dependent apoptotic pathway in human non-small cell lung cancer cells.

FUS1 is a novel tumor suppressor gene identified in human chromosome 3p21.3 region. Loss of expression and deficiency of posttranslational modification of FUS1 protein have been found in a majority of human lung cancers.

Decreased survival of mosquito cells after stable transfection with a Drosophila ecdysteroid response element: possible involvement of a 40 kDa DNA binding protein.

Homologous transfection systems provide a useful tool for characterizing promoters and other regulatory elements from cloned genes. We have used cultured Aedes albopictus C7-10 mosquito cells to evaluate expression of 20-hydroxyecdysone-inducible genes. Although this cell line has previously been shown to synthesize components of the ecdysteroid receptor and ecdysone-inducible proteins, the well-characterized ecdysteroid response element (EcRE) from the Drosophilahsp27 promoter failed to confer a substantial 20-hydroxyecdysone mediated induction in transfected mosquito cells.

Ribosomal protein P0 from Aedes albopictus mosquito cells: cDNA cloning and analysis of expression.

Ribosomal protein P0 is one of the highly conserved phosphorylated proteins in the large subunit of eukaryotic ribosomes. P0 has been shown in Drosophila to be a multifunctional protein that associates with elongation factor eEF2 to facilitate translation and also plays a role in DNA repair. In this paper we describe the cloning and characterization of the full-length cDNA encoding P0 from the mosquito, Aedes albopictus.