Regulation of cancer stem cells by CXCL1, a chemokine whose secretion is controlled by MCM2.

BACKGROUND: A high expression pattern of minichromosome maintenance 2 (MCM2) has been observed in various cancers. MCM2 is a protein involved in the cell cycle and plays a role in cancer growth and differentiation by binding to six members of the MCM subfamily. The MCM protein family includes MCM2 through MCM7. METHODS: MCM2 has shown high expression in both lung cancer stem cells (LCSCs) and glioma stem cells (GSCs). We investigated the characteristics of CSCs and the regulation of the epithelial-to-mesenchymal transition (EMT) phenomenon in LCSCs and GSCs by MCM2. Additionally, we explored secreted factors regulated by MCM2. RESULTS: There was a significant difference in survival rates between lung cancer patients and brain cancer patients based on MCM2 expression. MCM2 was found to regulate both markers and regulatory proteins in LCSCs. Moreover, MCM2 is thought to be involved in cancer metastasis by regulating cell migration and invasion, not limited to lung cancer but also identified in glioma. Among chemokines, chemokine (C-X-C motif) ligand 1 (CXCL1) was found to be regulated by MCM2. CONCLUSIONS: MCM2 not only participates in the cell cycle but also affects cancer cell growth by regulating the external microenvironment to create a favorable environment for cells. MCM2 is highly expressed in malignant carcinomas, including CSCs, and contributes to the malignancy of various cancers. Therefore, MCM2 may represent a crucial target for cancer therapeutics.

Impact of KIF4A on Cancer Stem Cells and EMT in Lung Cancer and Glioma.

Kinesin family member 4A (KIF4A) belongs to the kinesin 4 subfamily of kinesin-related proteins and is involved in the regulation of chromosome condensation and segregation during mitotic cell division. The expression of KIF4A in various types of cancer, including lung, breast, and colon cancer, has been found to be associated with poor prognosis in cancer patients. However, the exact mechanism by which it promotes tumorigenesis is not yet understood. In osteosarcoma, the expression of KIF4A has been shown to be associated with cancer stem cells (CSCs), whereas in breast cancer, it is not associated with the maintenance of CSCs but regulates the migratory ability of cells. In this light, we identified phenotypic phenomena affecting the malignancy of cancer in lung cancer and glioma, and investigated the mechanisms promoting tumorigenesis. As a result, we demonstrated that KIF4A affected lung cancer stem cells (LCSCs) and glioma stem cells (GSCs) and regulated CSC signaling mechanisms. In addition, the migratory ability of cells was regulated by KIF4A, and epithelial-to-mesenchymal transition (EMT) marker proteins were controlled. KIF4A regulated the expression of the secretory factor plasminogen activator inhibitor-1 (PAI-1), demonstrating that it sustains cancer malignancy through an autocrine loop. Taken together, these findings suggest that KIF4A regulates CSCs and EMT, which are involved in cancer recurrence and metastasis, indicating its potential value as a novel therapeutic target and prognostic marker in lung cancer and glioma.

A Novel Anticancer Peptide Derived from Bryopsis plumosa Regulates Proliferation and Invasion in Non-Small Cell Lung Cancer Cells.

The discovery of new highly effective anticancer drugs with few side effects is a challenge for drug development research. Natural or synthetic anticancer peptides (ACPs) represent a new generation of anticancer agents with high selectivity and specificity. The rapid emergence of chemoradiation-resistant lung cancer has necessitated the discovery of novel anticancer agents as alternatives to conventional therapeutics. In this study, we synthesized a peptide containing 22 amino acids and characterized it as a novel ACP (MP06) derived from green sea algae, Bryopsis plumosa. Using the ACP database, MP06 was predicted to possess an alpha-helical secondary structure and functionality. The anti-proliferative and apoptotic effects of the MP06, determined using the cytotoxicity assay and Annexin V/propidium iodide staining kit, were significantly higher in non-small-cell lung cancer (NSCLC) cells than in non-cancerous lung cells. We confirmed that MP06 suppressed cellular migration and invasion and inhibited the expression of N-cadherin and vimentin, the markers of epithelial-mesenchymal transition. Moreover, MP06 effectively reduced the metastasis of tumor xenografts in zebrafish embryos. In conclusion, we suggest considering MP06 as a novel candidate for the development of new anticancer drugs functioning via the ERK signaling pathway.

BIRC5: A novel therapeutic target for lung cancer stem cells and glioma stem cells.

Baculoviral inhibitor of apoptosis repeat containing 5 (BIRC5) is also known as survivin. BIRC5, a member of the apoptosis inhibitor (IAP) family, negatively regulates apoptosis or programmed cell death by inhibiting caspase activation. Due to these properties, overexpression of BIRC5 enables specific survival and division associated with cancer malignancies. In addition, BIRC5 is highly expressed in stem cells, but not present at all in terminally differentiated cells. On this basis, there is speculation that BIRC5 may be involved in the regulation of cancer stem cells (CSCs), but few study results have been reported. In addition, the molecular mechanisms of BIRC5 regulation are not yet well understood. Through the present study, it was confirmed that BIRC5 is a key factor regulating CSCs and epithelial to mesenchymal transition (EMT). BIRC5 was simultaneously overexpressed in lung cancer stem cells (LCSCs) and glioma stem cells (GSCs), and when the expression was suppressed, the characteristics of CSCs disappeared. In addition, plasminogen activator inhibitor-1 (PAI-1), a secreted factor regulated by BIRC5, is involved in signaling mechanisms that regulate cancer stem cells and EMT, and PAI-1 forms an autocrine chain. Based on these results, BIRC5 is proposed as a novel therapeutic target protein for LCSCs and GSCs.

Regulation of Cancer Stem Cells and Epithelial-Mesenchymal Transition by CTNNAL1 in Lung Cancer and Glioblastoma.

CTNNAL1 is a protein known to be involved in cell-cell adhesion and cell adhesion. Alterations in the expression or function of CTNNAL1 have been reported to contribute to the development and progression of various types of cancer. In breast cancer, CTNNAL1 has been reported as a cancer suppressor gene, and in melanoma and lung cancer, it has been reported as a cancer driver gene. However, due to a lack of research, its function remains unclear. In this study, it is shown that CTNNAL1 regulates cancer stem cells (CSCs) in lung cancer and glioblastoma and modulates their migration and invasion abilities. CSCs are known to play an important role in the malignant transformation of cancer. They have the ability to resist chemotherapeutic drugs and irradiation, which is a known obstacle to cancer treatment. We found that CTNNAL1 regulates the ability to resist irradiation. In addition, we observed that CTNNAL1 regulates the ability of cells to migrate and invade, a key feature of the epithelial to mesenchymal transition phenomenon associated with cancer metastasis. CTNNAL1 was also involved in the secretion of C-C motif chemokine ligand 2 (CCL2), one of the chemokines. CCL2 plays a role in the recruitment of immune cells to the tumor microenvironment, but in cancer, it is known to influence malignancy and metastasis. CTNNAL1 may be a novel target for treating lung CSCs and glioma stem cells and may be used as a marker of malignancy.

RanBP1: A Potential Therapeutic Target for Cancer Stem Cells in Lung Cancer and Glioma.

Cancer stem cells (CSCs) are known to be one of the factors that make cancer treatment difficult. Many researchers are thus conducting research to efficiently destroy CSCs. Therefore, we sought to suggest a new target that can efficiently suppress CSCs. In this study, we observed a high expression of Ran-binding protein 1 (RanBP1) in lung cancer stem cells (LCSCs) and glioma stem cells (GSCs). Upregulated RanBP1 expression is strongly associated with the expression of CSC marker proteins and CSC regulators. In addition, an elevated RanBP1 expression is strongly associated with a poor patient prognosis. CSCs have the ability to resist radiation, and RanBP1 regulates this ability. RanBP1 also affects the metastasis-associated epithelial-mesenchymal transition (EMT) phenomenon. EMT marker proteins and regulatory proteins are affected by RanBP1 expression, and cell motility was regulated according to RanBP1 expression. The cancer microenvironment influences cancer growth, metastasis, and cancer treatment. RanBP1 can modulate the cancer microenvironment by regulating the cytokine IL-18. Secreted IL-18 acts on cancer cells and promotes cancer malignancy. Our results reveal, for the first time, that RanBP1 is an important regulator in LCSCs and GSCs, suggesting that it holds potential for use as a potential therapeutic target.

Effect of Copper Chelators via the TGF-ß Signaling Pathway on Glioblastoma Cell Invasion.

Glioblastoma multiforme (GBM) is a fast-growing and aggressive type of brain cancer. Unlike normal brain cells, GBM cells exhibit epithelial-mesenchymal transition (EMT), which is a crucial biological process in embryonic development and cell metastasis, and are highly invasive. Copper reportedly plays a critical role in the progression of a variety of cancers, including brain, breast, and lung cancers. However, excessive copper is toxic to cells. D-penicillamine (DPA) and triethylenetetramine (TETA) are well-known copper chelators and are the mainstay of treatment for copper-associated diseases. Following treatment with copper sulfate and DPA, GBM cells showed inhibition of proliferation and suppression of EMT properties, including reduced expression levels of N-cadherin, E-cadherin, and Zeb, which are cell markers associated with EMT. In contrast, treatment with copper sulfate and TETA yielded the opposite effects in GBM. Genes, including TGF-ß, are associated with an increase in copper levels, implying their role in EMT. To analyze the invasion and spread of GBM, we used zebrafish embryos xenografted with the GBM cell line U87. The invasion of GBM cells into zebrafish embryos was markedly inhibited by copper treatment with DPA. Our findings suggest that treatment with copper and DPA inhibits proliferation and EMT through a mechanism involving TGF-ß/Smad signaling in GBM. Therefore, DPA, but not TETA, could be used as adjuvant therapy for GBM with high copper concentrations.

miR-519d-3p suppresses tumorigenicity and metastasis by inhibiting Bcl-w and HIF-1α in NSCLC.

Bcl-w, a member of the Bcl-2 family, is highly expressed in various solid tumor, including lung cancer, suggesting that it is involved in cancer cell survival and carcinogenesis. Solid cancer-induced hypoxia has been reported to increase angiogenesis, growth factor, gene instability, invasion, and metastasis. Despite many studies on the treatment of non-small cell lung cancer (NSCLC) with a high incidence rate, the survival rate of patients has not improved because the cancer cells acquired resistance to treatment. This study investigated the correlation between Bcl-w expression and hypoxia in tumor malignancy of NSCLC. Meanwhile, microRNAs (miRNAs) are involved in a variety of key signaling mechanisms associated with hypoxia. Therefore, we discovered miR-519d-3p, which inhibits the expression of Bcl-w and hypoxia-inducing factor (HIF)-1α, and found that it reduces hypoxia-induced tumorigenesis. Spearman's correlation analysis showed that the expression levels of miR-519d-3p and Bcl-w/HIF-1α were negatively correlated, respectively. This showed that miR-519d-3p can be used as a diagnostic biomarker and target therapy for NSCLC.

Epithelial membrane protein 3 regulates lung cancer stem cells via the TGF­ß signaling pathway.

Epithelial membrane protein 3 (EMP3) is a transmembrane glycoprotein that contains a peripheral myelin protein 22 domain. EMP3 first received attention as a tumor suppressor, but accumulating evidence has since suggested that it may exhibit a tumor­promoting function. Nonetheless, the biological function of EMP3 remains largely unclear with regards to its role in cancer. Herein, it was shown that EMP3 expression is upregulated in non­small cell lung cancer (NSCLC) cells overexpressing aldehyde dehydrogenase 1 (ALDH1). EMP3 was shown to be involved in cell proliferation, the formation of cancer stem cells (CSCs) and in epithelial­mesenchymal transition (EMT). The ability to resist irradiation, one of the characteristics of CSCs, decreased when the EMP3 mRNA expression was knocked down using small interfering RNA. In addition, when EMP3 knockdown reduced the migratory ability of cells, a characteristic of EMT. Additionally, it was shown that the TGF­ß/Smad signaling axis was a target of EMP3. EMP3 was found to interact with TGF­ß receptor type 2 (TGFBR2) upon TGF­ß stimulation in lung CSCs (LCSC). As a result, binding of EMP3­TGFBR2 regulates TGF­ß/Smad signaling activation and consequently affects CSCs and EMT. Kaplan­Meier analysis results confirmed that patients with high expression of EMP3 had poor survival rates. Taken together, these findings showed that EMP3 may be a potential target for management of LCSCs with high expression of ALDH1, and that EMP3 is involved in TGF­ß/Smad signaling activation where it promotes acquisition of cancerous properties in tumors.

Targeting therapy-resistant lung cancer stem cells via disruption of the AKT/TSPYL5/PTEN positive-feedback loop.

Cancer stem cells (CSCs) are regarded as essential targets to overcome tumor progression and therapeutic resistance; however, practical targeting approaches are limited. Here, we identify testis-specific Y-like protein 5 (TSPYL5) as an upstream regulator of CSC-associated genes in non-small cell lung cancer cells, and suggest as a therapeutic target for CSC elimination. TSPYL5 elevation is driven by AKT-dependent TSPYL5 phosphorylation at threonine-120 and stabilization via inhibiting its ubiquitination. TSPYL5-pT120 also induces nuclear translocation and functions as a transcriptional activator of CSC-associated genes, ALDH1 and CD44. Also, nuclear TSPYL5 suppresses the transcription of PTEN, a negative regulator of PI3K signaling. TSPYL5-pT120 maintains persistent CSC-like characteristics via transcriptional activation of CSC-associated genes and a positive feedback loop consisting of AKT/TSPYL5/PTEN signaling pathway. Accordingly, elimination of TSPYL5 by inhibiting TSPYL5-pT120 can block aberrant AKT/TSPYL5/PTEN cyclic signaling and TSPYL5-mediated cancer stemness regulation. Our study suggests TSPYL5 be an effective target for therapy-resistant cancer.

HSPA1L Enhances Cancer Stem Cell-Like Properties by Activating IGF1Rß and Regulating ß-Catenin Transcription.

Studies have shown that cancer stem cells (CSCs) are involved in resistance and metastasis of cancer; thus, therapies targeting CSCs have been proposed. Here, we report that heat shock 70-kDa protein 1-like (HSPA1L) is partly involved in enhancing epithelial-mesenchymal transition (EMT) and CSC-like properties in non-small cell lung cancer (NSCLC) cells. Aldehyde dehydrogenase 1 (ALDH1) is considered a CSC marker in some lung cancers. Here, we analyzed transcriptional changes in genes between ALDH1high and ALDH1low cells sorted from A549 NSCLC cells and found that HSPA1L was highly expressed in ALDH1high cells. HSPA1L played two important roles in enhancing CSC-like properties. First, HSPA1L interacts directly with IGF1Rß and integrin αV to form a triple complex that is involved in IGF1Rß activation. HSPA1L/integrin αV complex-associated IGF1Rß activation intensified the EMT-associated cancer stemness and γ-radiation resistance through its downstream AKT/NF-κB or AKT/GSK3ß/ß-catenin activation pathway. Secondly, HSPA1L was also present in the nucleus and could bind directly to the promoter region of ß-catenin to function as a transcription activator of ß-catenin, an important signaling protein characterizing CSCs by regulating ALDH1 expression. HSPA1L may be a novel potential target for cancer treatment because it both enhances IGF1Rß activation and regulates γß-catenin transcription, accumulating CSC-like properties.

Crosstalk between GBM cells and mesenchymal stemlike cells promotes the invasiveness of GBM through the C5a/p38/ZEB1 axis.

BACKGROUND: Mesenchymal stemlike cells (MSLCs) have been detected in many types of cancer including brain tumors and have received attention as stromal cells in the tumor microenvironment. However, the cellular mechanisms underlying their participation in cancer progression remain largely unexplored. The aim of this study was to determine whether MSLCs have a tumorigenic role in brain tumors. METHODS: To figure out molecular and cellular mechanisms in glioma invasion, we have cultured glioma with MSLCs in a co-culture system. RESULTS: Here, we show that MSLCs in human glioblastoma (GBM) secrete complement component C5a, which is known for its role as a complement factor. MSLC-secreted C5a increases expression of zinc finger E-box-binding homeobox 1 (ZEB1) via activation of p38 mitogen-activated protein kinase (MAPK) in GBM cells, thereby enhancing the invasion of GBM cells into parenchymal brain tissue. CONCLUSION: Our results reveal a mechanism by which MSLCs undergo crosstalk with GBM cells through the C5a/p38 MAPK/ZEB1 signaling loop and act as a booster in GBM progression. KEY POINTS: 1. MSLCs activate p38 MAPK-ZEB1 signaling in GBM cells through C5a in a paracrine manner, thereby boosting the invasiveness of GBM cells in the tumor microenvironment.2. Neutralizing of C5a could be a potential therapeutic target for GBM by inhibition of mesenchymal phenotype.

Tescalcin/c-Src/IGF1Rß-mediated STAT3 activation enhances cancer stemness and radioresistant properties through ALDH1.

Tescalcin (TESC; also known as calcineurin B homologous protein 3, CHP3) has recently reported as a regulator of cancer progression. Here, we showed that the elevation of TESC in non-small cell lung cancer (NSCLC) intensifies epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) properties, consequently enhancing the cellular resistance to γ-radiation. TESC expression and the phosphorylation (consequent activation) of signal transducer and activator of transcription 3 (STAT3) were upregulated in CSC-like ALDH1high cells than in ALDH1low cells sorted from A549 NSCLC cells. Knockdown of TESC suppressed CSC-like properties as well as STAT3 activation through inhibition of insulin-like growth factor 1 receptor (IGF1R), a major signaling pathway of lung cancer stem cells. TESC activated IGF1R by the direct recruitment of proto-oncogene tyrosine kinase c-Src (c-Src) to IGF1Rß complex. Treatment of IGF1R inhibitor, AG1024, also suppressed c-Src activation, implicating that TESC mediates the mutual activation of c-Src and IGF1R. STAT3 activation by TESC/c-Src/IGF1R signaling pathway subsequently upregulated ALDH1 expression, which enhanced EMT-associated CSC-like properties. Chromatin immunoprecipitation and luciferase assay demonstrated that STAT3 is a potential transcription activator of ALDH1 isozymes. Ultimately, targeting TESC can be a potential strategy to overcome therapeutic resistance in NSCLC caused by augmented EMT and self-renewal capacity.

FBXL14 abolishes breast cancer progression by targeting CDCP1 for proteasomal degradation.

Understanding the molecular mechanisms that underlie the aggressive behavior and relapse of breast cancer may help in the development of novel therapeutic interventions. CUB-domain-containing protein 1 (CDCP1), a transmembrane adaptor protein, is highly maintained and required in the context of cellular metastatic potential in triple-negative breast cancer (TNBC). For this reason, gene expression levels of CDCP1 have been considered as a prognostic marker in TNBC. However, not rarely, transcript levels of genes do not reflect always the levels of proteins, due to the post-transcriptional regulation. Here we show that miR-17/20a control the FBXL14 E3 ligase, establishing FBXL14 as an upstream regulator of the CDCP1 pathway. FBXL14 acts as an novel interaction partner of CDCP1, and facilitates its ubiquitination and proteasomal degradation with an enhanced capacity to suppress CDCP1 protein stability that eventually prevents CDCP1 target genes involved in breast cancer metastasis. Our findings first time uncovers the regulatory mechanism of CDCP-1 protein stabilization, more predictable criteria than gene expression levels for prognosis of breast cancer patients.

Urokinase-derived peptide UP-7 suppresses tumor angiogenesis and metastasis through inhibition of FAK activation.

The recombinant kringle domain of urokinase (UK1) has been shown to inhibit angiogenesis and brain tumor growth in vivo. To avoid limitations in application due to mass production of recombinant protein, we attempted to develop a novel peptide inhibitor from UK1 sequence consisting of 83 amino acids that contains α-helices, loops and ß-sheets. We dissected UK1 sequence to seven peptides based on structure and amino acid characteristics, and examined the anti-angiogenic activities for the constructed peptides. Among the tested peptides, UP-7 most potently inhibited the proliferation and migration of endothelial cells (ECs) in vitro, and also potently inhibited in vivo angiogenesis in the mouse matrigel plug assay. Such anti-angiogenic activities were not exerted by the scrambled peptide. At molecular level, UP-7 inhibited growth factor-induced phosphorylation of FAK and ERK1/2. It also suppressed formation of stress fibers and focal adhesions and also inhibited the attachment and spreading of ECs onto immobilized fibronectin. In a lung cancer animal model xenografted with non-UP-7-sensitive NCI-H460 cells, systemic treatment of UP-7 effectively suppressed tumor growth through inhibition of angiogenesis. Interestingly, breast cancer cells such as LM-MDA-MB-231 cells were moderately sensitive to UP-7 in proliferation differently from other cancer cells. UP-7 also inhibited migration, invasion and FAK phosphorylation of LM-MDA-MB-231 cells. Accordingly, UP-7 potently inhibited lung metastatic growth of LM-MDA-MB-231 cells in an experimental metastasis model. Taken together, these results suggest that novel peptide UP-7 can be effectively used for treatment of breast cancer metastatic growth through inhibition of angiogenesis and invasion.

PRMT8 Controls the Pluripotency and Mesodermal Fate of Human Embryonic Stem Cells By Enhancing the PI3K/AKT/SOX2 Axis.

Basic fibroblast growth factor (bFGF) supplementation is critical to maintain the pluripotency of human pluripotent stem cells (hPSCs) through activation of PI3K/AKT, rather than MEK/ERK pathway. Thus, elaborate molecular mechanisms that preserve PI3K/AKT signaling upon bFGF stimulation may exist in hPSCs. Protein arginine methyltransferase 8 (PRMT8) was expressed and then its level gradually decreased during spontaneous differentiation of human embryonic stem cells (hESCs). PRMT8 loss- or gain-of-function studies demonstrated that PRMT8 contributed to longer maintenance of hESC pluripotency, even under bFGF-deprived conditions. Direct interaction of membrane-localized PRMT8 with p85, a regulatory subunit of PI3K, was associated with accumulation of phosphoinositol 3-phosphate and consequently high AKT activity. Furthermore, the SOX2 induction, which was controlled by the PRMT8/PI3K/AKT axis, was linked to mesodermal lineage differentiation. Thus, we propose that PRMT8 in hESCs plays an important role not only in maintaining pluripotency but also in controlling mesodermal differentiation through bFGF signaling toward the PI3K/AKT/SOX2 axis. Stem Cells 2017;35:2037-2049.

Low-dose radiation decreases tumor progression via the inhibition of the JAK1/STAT3 signaling axis in breast cancer cell lines.

Breast cancer is a widely distributed type of cancer in women worldwide, and tumor relapse is the major cause of breast cancer death. In breast cancers, the acquisition of metastatic ability, which is responsible for tumor relapse and poor clinical outcomes, has been linked to the acquisition of the epithelial-mesenchymal transition (EMT) program and self-renewal traits (CSCs) via various signaling pathways. These phenomena confer resistance during current therapies, thus creating a major hurdle in radiotherapy/chemotherapy. The role of very low doses of radiation (LDR) in the context of EMT has not yet to be thoroughly explored. Here, we report that a 0.1 Gy radiation dose reduces cancer progression by deactivating the JAK1/STAT3 pathway. Furthermore, LDR exposure also reduces sphere formation and inhibits the self-renewal ability of breast cancer cells, resulting in an attenuated CD44+/CD24- population. Additionally, in vivo findings support our data, providing evidence that LDR is a promising option for future treatment strategies to prevent cancer metastasis in breast cancer cases.

Radiation driven epithelial-mesenchymal transition is mediated by Notch signaling in breast cancer.

Epithelial to mesenchymal transition (EMT) is developmental process associated with cancer metastasis. Here, we found that breast carcinoma cells adopt epithelial-to-mesenchymal transition (EMT) in response to fractionated-radiation. Importantly, we show that Notch signaling is highly activated in fractionally-irradiated tumors as compared to non-irradiated tumors that are accompanied by an EMT. Moreover, we uncovered the mechanism of Notch-driven EMT, in which Notch enhanced EMT through IL-6/JAK/STAT3 signaling axis in mammary tumor cells. Collectively, we present converging evidence from our studies that Notch2 is a critical mediator of radiation-induced EMT and responsible for induced malignant tumor growth.

Clogging-free microfluidics for continuous size-based separation of microparticles.

In microfluidic filtration systems, one of the leading obstacles to efficient, continuous operation is clogging of the filters. Here, we introduce a lateral flow microfluidic sieving (µ-sieving) technique to overcome clogging and to allow continuous operation of filter based microfluidic separation. A low frequency mechanical oscillation was added to the fluid flow, which made possible the release of aggregated unwanted polystyrene (PS) particles trapped between the larger target PS particles in the filter demonstrating continuous µ-sieving operation. We achieved collection of the target PS particles with 100% separation efficiency. Also, on average, more than 98% of the filtered target particles were retrieved after the filtration showing high retrieval rates. Since the oscillation was applied to the fluid but not to the microfluidic filter system, mechanical stresses to the system was minimized and no additional fabrication procedures were necessary. We also applied the µ-sieving technique to the separation of cancer cells (MDA-MB-231) from whole blood and showed that the fluidic oscillations prevented the filters from being blocked by the filtered cancer cells allowing continuous microfluidic separation with high efficiency.