Overexpression of kinesin superfamily members as prognostic biomarkers of breast cancer.

BACKGROUND: Kinesin superfamily (KIFs) has a long-reported significant influence on the initiation, development, and progress of breast cancer. However, the prognostic value of whole family members was poorly done. Our study intends to demonstrate the value of kinesin superfamily members as prognostic biomarkers as well as a therapeutic target of breast cancer. METHODS: Comprehensive bioinformatics analyses were done using data from TCGA, GEO, METABRIC, and GTEx. LASSO regression was done to select tumor-related members. Nomogram was constructed to predict the overall survival (OS) of breast cancer patients. Expression profiles were testified by quantitative RT-PCR and immunohistochemistry. Transcription factor, GO and KEGG enrichments were done to explore regulatory mechanism and functions. RESULTS: A total of 20 differentially expressed KIFs were identified between breast cancer and normal tissue with 4 (KIF17, KIF26A, KIF7, KIFC3) downregulated and 16 (KIF10, KIF11, KIF14, KIF15, KIF18A, KIF18B, KIF20A, KIF20B, KIF22, KIF23, KIF24, KIF26B, KIF2C, KIF3B, KIF4A, KIFC1) overexpressed. Among which, 11 overexpressed KIFs (KIF10, KIF11, KIF14, KIF15, KIF18A, KIF18B, KIF20A, KIF23, KIF2C, KIF4A, KIFC1) significantly correlated with worse OS, relapse-free survival (RFS) and distant metastasis-free survival (DMFS) of breast cancer. A 6-KIFs-based risk score (KIF10, KIF15, KIF18A, KIF18B, KIF20A, KIF4A) was generated by LASSO regression with a nomogram validated an accurate predictive efficacy. Both mRNA and protein expression of KIFs are experimentally demonstrated upregulated in breast cancer patients. Msh Homeobox 1 (MSX1) was identified as transcription factors of KIFs in breast cancer. GO and KEGG enrichments revealed functions and pathways affected in breast cancer. CONCLUSION: Overexpression of tumor-related KIFs correlate with worse outcomes of breast cancer patients and can work as potential prognostic biomarkers.

Occurrence of COVID-19 Symptoms During SARS-CoV-2 Infection Defines Waning of Humoral Immunity.

Approximately half of the SARS-CoV-2 infections occur without apparent symptoms, raising questions regarding long-term humoral immunity in asymptomatic individuals. Plasma levels of immunoglobulin G (IgG) and M (IgM) against the viral spike or nucleoprotein were determined for 25,091 individuals enrolled in a surveillance program in Wuhan, China. We compared 405 asymptomatic individuals who mounted a detectable antibody response with 459 symptomatic COVID-19 patients. The well-defined duration of the SARS-CoV-2 endemic in Wuhan allowed a side-by-side comparison of antibody responses following symptomatic and asymptomatic infections without subsequent antigen re-exposure. IgM responses rapidly declined in both groups. However, both the prevalence and durability of IgG responses and neutralizing capacities correlated positively with symptoms. Regardless of sex, age, and body weight, asymptomatic individuals lost their SARS-CoV-2-specific IgG antibodies more often and rapidly than symptomatic patients did. These findings have important implications for immunity and favour immunization programs including individuals after asymptomatic infections.

Improved hydrophobicity of carbon nanotube arrays with micropatterning.

The hydrophobicity of vertically aligned multiwalled carbon nanotubes (MWCNTs) was improved through the creation of a parallel array of microwalls via a laser pruning technique. Changes to the hydrophobic nature of the patterned MWCNTs due to artificially induced roughness through variations in both the widths of the walls and the distance between adjacent walls, channel width, were investigated. The sample became more hydrophobic whenever water droplets landed on one microwall 7 or 13 microm in width. However, when a droplet bridged two microwalls, the surface became less hydrophobic. The optimal superhydrophobic MWCNT surface corresponded to a parallel array of microwalls with a width of 13 microm and a channel width of approximately 50 microm. Such findings could possibly serve as value-add for further developments in the creation of water-repelling CNT surfaces via micropatterning.

Identifying cancer origin using circulating tumor cells.

Circulating tumor cells (CTCs) have become an established clinical evaluation biomarker. CTC count provides a good correlation with the prognosis of cancer patients, but has only been used with known cancer patients, and has been unable to predict the origin of the CTCs. This study demonstrates the analysis of CTCs for the identification of their primary cancer source. Twelve mL blood samples were equally dispensed on 6 CMx chips, microfluidic chips coated with an anti-EpCAM-conjugated supported lipid bilayer, for CTC capture and isolation. Captured CTCs were eluted to an immunofluorescence (IF) staining panel consisting of 6 groups of antibodies: anti-panCK, anti-CK18, anti-CK7, anti-TTF-1, anti-CK20/anti-CDX2, and anti-PSA/anti-PSMA. Cancer cell lines of lung (H1975), colorectal (DLD-1, HCT-116), and prostate (PC3, DU145, LNCaP) were selected to establish the sensitivity and specificity for distinguishing CTCs from lung, colorectal, and prostate cancer. Spiking experiments performed in 2mL of culture medium or whole blood proved the CMx platform can enumerate cancer cells of lung, colorectal, and prostate. The IF panel was tested on blood samples from lung cancer patients (n = 3), colorectal cancer patients (n = 5), prostate cancer patients (n = 5), and healthy individuals (n = 12). Peripheral blood samples found panCK(+) and CK18(+) CTCs in lung, colorectal, and prostate cancers. CTCs expressing CK7(+) or TTF-1(+), (CK20/ CDX2)(+), or (PSA/ PSMA)(+) corresponded to lung, colorectal, or prostate cancer, respectively. In conclusion, we have designed an immunofluorescence staining panel to identify CTCs in peripheral blood to correctly identify cancer cell origin.

Sensitive and Specific Biomimetic Lipid Coated Microfluidics to Isolate Viable Circulating Tumor Cells and Microemboli for Cancer Detection.

Here we presented a simple and effective membrane mimetic microfluidic device with antibody conjugated supported lipid bilayer (SLB) "smart coating" to capture viable circulating tumor cells (CTCs) and circulating tumor microemboli (CTM) directly from whole blood of all stage clinical cancer patients. The non-covalently bound SLB was able to promote dynamic clustering of lipid-tethered antibodies to CTC antigens and minimized non-specific blood cells retention through its non-fouling nature. A gentle flow further flushed away loosely-bound blood cells to achieve high purity of CTCs, and a stream of air foam injected disintegrate the SLB assemblies to release intact and viable CTCs from the chip. Human blood spiked cancer cell line test showed the ~95% overall efficiency to recover both CTCs and CTMs. Live/dead assay showed that at least 86% of recovered cells maintain viability. By using 2 mL of peripheral blood, the CTCs and CTMs counts of 63 healthy and colorectal cancer donors were positively correlated with the cancer progression. In summary, a simple and effective strategy utilizing biomimetic principle was developed to retrieve viable CTCs for enumeration, molecular analysis, as well as ex vivo culture over weeks. Due to the high sensitivity and specificity, it is the first time to show the high detection rates and quantity of CTCs in non-metastatic cancer patients. This work offers the values in both early cancer detection and prognosis of CTC and provides an accurate non-invasive strategy for routine clinical investigation on CTCs.

Circulating Tumor Cell Count Correlates with Colorectal Neoplasm Progression and Is a Prognostic Marker for Distant Metastasis in Non-Metastatic Patients.

Enumeration of circulating tumor cells (CTCs) has been proven as a prognostic marker for metastatic colorectal cancer (m-CRC) patients. However, the currently available techniques for capturing and enumerating CTCs lack of required sensitivity to be applicable as a prognostic marker for non-metastatic patients as CTCs are even more rare. We have developed a microfluidic device utilizing antibody-conjugated non-fouling coating to eliminate nonspecific binding and to promote the multivalent binding of target cells. We then established the correlation of CTC counts and neoplasm progression through applying this platform to capture and enumerate CTCs in 2 mL of peripheral blood from healthy (n = 27), benign (n = 21), non-metastatic (n = 95), and m-CRC (n = 15) patients. The results showed that the CTC counts progressed from 0, 1, 5, to 36. Importantly, after 2-year follow-up on the non-metastatic CRC patients, we found that those who had ≥5 CTCs were 8 times more likely to develop distant metastasis within one year after curable surgery than those who had <5. In conclusion, by employing a sensitive device, CTC counts show good correlation with colorectal neoplasm, thus CTC may be as a simple, independent prognostic marker for the non-metastatic CRC patients who are at high risk of early recurrence.

Efficient elusion of viable adhesive cells from a microfluidic system by air foam.

We developed a new method for releasing viable cells from affinity-based microfluidic devices. The lumen of a microchannel with a U-shape and user-designed microstructures was coated with supported lipid bilayers functionalized by epithelial cell adhesion molecule antibodies to capture circulating epithelial cells of influx solution. After the capturing process, air foam was introduced into channels for releasing target cells and then carrying them to a small area of membrane. The results show that when the air foam is driven at linear velocity of 4.2 mm/s for more than 20 min or at linear velocity of 8.4 mm/s for more than 10 min, the cell releasing efficiency approaches 100%. This flow-induced shear stress is much less than the physiological level (15 dyn/cm(2)), which is necessary to maintain the intactness of released cells. Combining the design of microstructures of the microfluidic system, the cell recovery on the membrane exceeds 90%. Importantly, we demonstrate that the cells released by air foam are viable and could be cultured in vitro. This novel method for releasing cells could power the microfluidic platform for isolating and identifying circulating tumor cells.

Antibody conjugated supported lipid bilayer for capturing and purification of viable tumor cells in blood for subsequent cell culture.

Interest in the identification and isolation of circulating tumor cells (CTCs) has been growing since the introduction of CTCs as an alternative to the tumor tissue biopsy, which can potentially be important indices for prognosis and cancer treatment. However, the contamination of non-specific binding of normal hematologic cells makes high purity CTCs detection problematic. Furthermore, preserving the viability of CTCs remains a challenge. In this study, we proposed to construct an anti-EpCAM functionalized supported lipid bilayer (SLB), a biomimetic and non-fouling membrane coating, for CTCs capturing, purification and maintaining the viability. Healthy human blood spiked with pre-stained colorectal cancer cell lines, HCT116 and colo205, were used to investigate interaction of cells with the anti-EpCAM functionalized SLB surfaces. Over 97% of HCT116, and 72% of colo205 were captured and adhered by the surface anti-EpCAM; conversely, the majority of blood cells were easily removed by gentle buffer exchange, with the overall purity of cancer cells exceeding 95%. The bound cancer cells were subsequently detached for cell culture. Both HCT116 and colo205 continued to proliferate over 2-week observation period, indicating that the anti-EpCAM functionalized SLB platform providing a simple strategy for capturing, purifying, and releasing viable targeted rare cells.