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Serum alpha-fetoprotein and radiologic imaging are the most commonly used tests for early diagnosis and dynamic monitoring of treatment response in hepatocellular carcinoma (HCC). However, the accuracy of these tests is limited, and they may not reflect the underlying biology of the tumor. Thus, developing highly accurate novel HCC biomarkers reflecting tumor biology is a clinically unmet need. Circulating tumor cells (CTCs) have long been proposed as a noninvasive biomarker in clinical oncology. Most CTC assays utilize immunoaffinity-based, size-based, and/or enrichment-free mechanisms followed by immunocytochemical staining to characterize CTCs. The prognostic value of HCC CTC enumeration has been extensively validated. Subsets of CTCs expressing mesenchymal markers are also reported to have clinical significance. In addition, researchers have been devoting their efforts to molecular characterizations of CTCs (e.g. genetics and transcriptomics) as molecular profiling can offer a more accurate readout and provide biological insights. As new molecular profiling techniques, such as digital polymerase chain reaction, are developed to detect minimal amounts of DNA/RNA, several research groups have established HCC CTC digital scoring systems to quantify clinically relevant gene panels. Given the versatility of CTCs to provide intact molecular and functional data that reflects the underlying tumor, CTCs have great potential as a noninvasive biomarker in HCC. Large-scale, prospective studies for HCC CTCs with a standardized protocol are necessary for successful clinical translation.

Circulating tumor cell (CTC) clusters are present in cancer patients with severe metastasis, resulting in poor clinical outcomes. However, CTC clusters have not been studied as extensively as single CTCs, and the clinical utility of CTC clusters remains largely unknown. In this study, we aim sought to explore the feasibility of NanoVelcro Chips to simultaneously detect both single CTCs and CTC clusters with negligible perturbation to their intrinsic properties in neuroendocrine tumors (NETs). We discovered frequent CTC clusters in patients with advanced NETs and examined their potential roles, together with single NET CTCs, as novel biomarkers of patient response following peptide receptor radionuclide therapy (PRRT). We observed dynamic changes in both total NET CTCs and NET CTC cluster counts in NET patients undergoing PRRT which correlated with clinical outcome. These preliminary findings suggest that CTC clusters, along with single CTCs, offer a potential non-invasive option to monitor the treatment response in NET patients undergoing PRRT.

Placenta accreta spectrum (PAS) is a high-risk obstetrical condition associated with significant morbidity and mortality. Current clinical screening modalities for PAS are not always conclusive. Here, we report a nanostructure-embedded microchip that efficiently enriches both single and clustered circulating trophoblasts (cTBs) from maternal blood for detecting PAS. We discover a uniquely high prevalence of cTB-clusters in PAS and subsequently optimize the device to preserve the intactness of these clusters. Our feasibility study on the enumeration of cTBs and cTB-clusters from 168 pregnant women demonstrates excellent diagnostic performance for distinguishing PAS from non-PAS. A logistic regression model is constructed using a training cohort and then cross-validated and tested using an independent cohort. The combined cTB assay achieves an Area Under ROC Curve of 0.942 (throughout gestation) and 0.924 (early gestation) for distinguishing PAS from non-PAS. Our assay holds the potential to improve current diagnostic modalities for the early detection of PAS.

Transcriptomic profiling of tumor tissues introduces a large database, which has led to improvements in the ability of cancer diagnosis, treatment, and prevention. However, performing tumor transcriptomic profiling in the clinical setting is very challenging since the procurement of tumor tissues is inherently limited by invasive sampling procedures. Here, we demonstrated the feasibility of purifying hepatocellular carcinoma (HCC) circulating tumor cells (CTCs) from clinical patient samples with improved molecular integrity using Click Chips in conjunction with a multimarker antibody cocktail. The purified CTCs were then subjected to mRNA profiling by NanoString nCounter platform, targeting 64 HCC-specific genes, which were generated from an integrated data analysis framework with 8 tissue-based prognostic gene signatures from 7 publicly available HCC transcriptomic studies. After bioinformatics analysis and comparison, the HCC CTC-derived gene signatures showed high concordance with HCC tissue-derived gene signatures from TCGA database, suggesting that HCC CTCs purified by Click Chips could enable the translation of HCC tissue molecular profiling into a noninvasive setting.

Hepatocellular carcinoma (HCC) is among the leading causes of worldwide cancer-related morbidity and mortality. Poor prognosis of HCC is attributed primarily to tumor presentation at an advanced stage when there is no effective treatment to achieve the long term survival of patients. Currently available tests such as alpha-fetoprotein have limited accuracy as a diagnostic or prognostic biomarker for HCC. Liver biopsy provides tissue that can reveal tumor biology but it is not used routinely due to its invasiveness and risk of tumor seeding, especially in early-stage patients. Liver biopsy is also limited in revealing comprehensive tumor biology due to intratumoral heterogeneity. There is a clear need for new biomarkers to improve HCC detection, prognostication, prediction of treatment response, and disease monitoring with treatment. Liquid biopsy could be an effective method of early detection and management of HCC. Circulating tumor cells (CTCs) are cancer cells in circulation derived from the original tumor or metastatic foci, and their measurement by liquid biopsy represents a great potential in facilitating the implementation of precision medicine in patients with HCC. CTCs can be detected by a simple peripheral blood draw and potentially show global features of tumor characteristics. Various CTC detection platforms using immunoaffinity and biophysical properties have been developed to identify and capture CTCs with high efficiency. Quantitative abundance of CTCs, as well as biological characteristics and genomic heterogeneity among the CTCs, can predict disease prognosis and response to therapy in patients with HCC. This review article will discuss the currently available technologies for CTC detection and isolation, their utility in the clinical management of HCC patients, their limitations, and future directions of research.

A major metastasis suppressing mechanism is the rapid apoptotic death of cancer cells upon detachment from extracellular matrix, a process called anoikis. Focal adhesion kinase (PTK2/FAK) is a key enzyme involved in evasion of anoikis. We show that loss of the Cub-domain containing protein-1 (CDCP1), paradoxically stimulates FAK activation in the detached state of prostate cancer cells. In CDCP1low DU145 and PC3 prostate cancer cells, detachment-activation of FAK occurs through local production of PI(4,5)P2. PI(4,5)P2 is generated by the PIP5K1c-201 splicing isoform of PIP5K1c, which contains a unique SRC phosphorylation site. In the detached state, reduced expression of CDCP1 and an alternative CDCP1-independent SRC activation mechanism triggers PIP5K1c-pY644 phosphorylation by SRC. This causes a switch of Talin binding from β1-integrin to PIP5K1c-pY644 and leads to activation of PIP5K1c-FAK. Reduced CDCP1 expression also inactivates CDK5, a negative regulator of PIP5K1c. Furthermore, immersion of prostate cancer cells in 10% human plasma or fetal bovine serum is required for activation of PIP5K1c-FAK. The PIP5K1c induced detachment-activation of FAK in preclinical models sensitizes CDCP1low prostate cancer cells to FAK inhibitors. In patients, CDCP1High versus CDCP1low circulating tumor cells differ in expression of AR-v7, ONECUT2 and HOXB13 oncogenes and TMPRSS2 and display intra-patient heterogeneity of FAK-pY397 expression. Taken together, CDCP1low and CDCP1high detached prostate cancer cells activate distinct cytoplasmic kinase complexes and targetable transcription factors, which has important therapeutic implications.

Tumor-derived extracellular vesicles (EVs) play essential roles in intercellular communication during tumor growth and metastatic evolution. Currently, little is known about the possible roles of tumor-derived EVs in sarcoma because the lack of specific surface markers makes it technically challenging to purify sarcoma-derived EVs. In this study, a specific purification system is developed for Ewing sarcoma (ES)-derived EVs by coupling covalent chemistry-mediated EV capture/ release within a nanostructure-embedded microchip. The purification platform-ES-EV Click Chip-takes advantage of specific anti-LINGO-1 recognition and sensitive click chemistry-mediated EV capture, followed by disulfide cleavage-driven EV release. Since the device is capable of specific and efficient purification of intact ES EVs with high purity, ES-EV Click Chip is ideal for conducting downstream functional studies of ES EVs. Absolute quantification of the molecular hallmark of ES (i.e., EWS rearrangements) using reverse transcription Droplet Digital PCR enables specific quantification of ES EVs. The purified ES EVs can be internalized by recipient cells and transfer their mRNA cargoes, exhibiting their biological intactness and potential role as biological shuttles in intercellular communication.

Hepatocellular carcinoma (HCC) is a leading cause of mortality. Checkpoint inhibitors of programmed cell death protein-1 (PD-1) and programmed death-ligand 1 (PD-L1) have shown great efficacy, but lack biomarkers that predict response. Circulating tumor cells (CTCs) have promise as a liquid-biopsy biomarker; however, data on HCC CTCs expressing PD-L1 have not been reported. We sought to detect PD-L1-expressing HCC-CTCs and investigated their role as a prognostic and predictive biomarker. Using an antibody-based platform, CTCs were enumerated/phenotyped from a prospective cohort of 87 patients with HCC (49 early-stage, 22 locally advanced, and 16 metastatic), 7 patients with cirrhosis, and 8 healthy controls. Immunocytochemistry identified total HCC CTCs (4',6-diamidino-2-phenylindole-positive [DAPI+]/cytokeratin-positive [CK+]/clusters of differentiation 45-negative [CD45-]) and a subpopulation expressing PD-L1 (DAPI+/CK+/PD-L1+/CD45-). PD-L1+ CTCs were identified in 4 of 49 (8.2%) early-stage patients, but 12 of 22 (54.5%) locally advanced and 15 of 16 (93.8%) metastatic patients, accurately discriminating early from locally advanced/metastatic HCC (sensitivity = 71.1%, specificity = 91.8%, area under the receiver operating characteristic curve = 0.807; P

We report a covalent chemistry-based hepatocellular carcinoma (HCC)-specific extracellular vesicle (EV) purification system for early detection of HCC by performing digital scoring on the purified EVs. Earlier detection of HCC creates more opportunities for curative therapeutic interventions. EVs are present in circulation at relatively early stages of disease, providing potential opportunities for HCC early detection. We develop an HCC EV purification system (i.e., EV Click Chips) by synergistically integrating covalent chemistry-mediated EV capture/release, multimarker antibody cocktails, nanostructured substrates, and microfluidic chaotic mixers. We then explore the translational potential of EV Click Chips using 158 plasma samples of HCC patients and control cohorts. The purified HCC EVs are subjected to reverse-transcription droplet digital PCR for quantification of 10 HCC-specific mRNA markers and computation of digital scoring. The HCC EV-derived molecular signatures exhibit great potential for noninvasive early detection of HCC from at-risk cirrhotic patients with an area under receiver operator characteristic curve of 0.93 (95% CI, 0.86 to 1.00; sensitivity = 94.4%, specificity = 88.5%).

Rationale: Our objective was to develop a circulating tumor cell (CTC)-RNA assay for characterizing clinically relevant RNA signatures for the assessment of androgen receptor signaling inhibitor (ARSI) sensitivity in metastatic castration-resistant prostate cancer (mCRPC) patients. Methods: We developed the NanoVelcro CTC-RNA assay by combining the Thermoresponsive (TR)-NanoVelcro CTC purification system with the NanoString nCounter platform for cellular purification and RNA analysis. Based on the well-validated, tissue-based Prostate Cancer Classification System (PCS), we focus on the most aggressive and ARSI-resistant PCS subtype, i.e., PCS1, for CTC analysis. We applied a rigorous bioinformatic process to develop the CTC-PCS1 panel that consists of prostate cancer (PCa) CTC-specific RNA signature with minimal expression in background white blood cells (WBCs). We validated the NanoVelcro CTC-RNA assay and the CTC-PCS1 panel with well-characterized PCa cell lines to demonstrate the sensitivity and dynamic range of the assay, as well as the specificity of the PCS1 Z score (the likelihood estimate of the PCS1 subtype) for identifying PCS1 subtype and ARSI resistance. We then selected 31 blood samples from 23 PCa patients receiving ARSIs to test in our assay. The PCS1 Z scores of each sample were computed and compared with ARSI treatment sensitivity. Results: The validation studies using PCa cell line samples showed that the NanoVelcro CTC-RNA assay can detect the RNA transcripts in the CTC-PCS1 panel with high sensitivity and linearity in the dynamic range of 5-100 cells. We also showed that the genes in CTC-PCS1 panel are highly expressed in PCa cell lines and lowly expressed in background WBCs. Using the artificial CTC samples simulating the blood sample conditions, we further demonstrated that the CTC-PCS1 panel is highly specific in identifying PCS1-like samples, and the high PCS1 Z score is associated with ARSI resistance samples. In patient bloods, ARSI-resistant samples (ARSI-R, n=14) had significantly higher PCS1 Z scores as compared with ARSI-sensitive samples (ARSI-S, n=17) (Rank-sum test, P=0.003). In the analysis of 8 patients who were initially sensitive to ARSI (ARSI-S) and later developed resistance (ARSI-R), we found that the PCS1 Z score increased from the time of ARSI-S to the time of ARSI-R (Pairwise T-test, P=0.016). Conclusions: Using our new methodology, we developed a first-in-class CTC-RNA assay and demonstrated the feasibility of transforming clinically-relevant tissue-based RNA profiling such as PCS into CTC tests. This approach allows for detecting RNA expression relevant to clinical drug resistance in a non-invasive fashion, which can facilitate patient-specific treatment selection and early detection of drug resistance, a goal in precision oncology.

The fomite transmission of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) has drawn attention because of its highly contagious nature. Therefore, surfaces that can prevent coronavirus contamination are an urgent and unmet need during the coronavirus disease 2019 (COVID‐19) pandemic. Conventional surfaces are usually based on superhydrophobic or antiviral coatings. However, these coatings may be dysfunctional because of biofouling, which is the undesired adhesion of biomolecules. A superhydrophobic surface independent of the material content and coating agents may serve the purpose of antibiofouling and preventing viral transmission. Doubly reentrant topology (DRT) is a unique structure that can meet the need. This study demonstrates that the DRT surfaces possess a striking antibiofouling effect that can prevent viral contamination. This effect still exists even if the DRT surface is made of a hydrophilic material such as silicon oxide and copper. To the best of our knowledge, this work first demonstrates that fomite transmission of viruses may be prevented by minimizing the contact area between pathogens and surfaces even made of hydrophilic materials. Furthermore, the DRT geometry per se features excellent antibiofouling ability, which may shed light on the applications of pathogen elimination in alleviating the COVID‐19 pandemic. [ABSTRACT FROM AUTHOR]

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease in the world. NAFLD is a hepatic manifestation of insulin resistance, the core pathophysiology of diabetes. Multiple clinical studies show that diabetes increases the risk of liver disease progression and cirrhosis development in patients with NAFLD. Diabetes has causal associations with many different cancers, including hepatocellular carcinoma (HCC). More recent studies demonstrate that diabetes increases the risk of HCC in patients with underlying NAFLD cirrhosis, confirming the direct hepatocarcinogenic effect of diabetes among cirrhosis patients. Diabetes promotes hepatocarcinogenesis via the activation of inflammatory cascades producing reactive oxygen species and proinflammatory cytokines, leading to genomic instability, cellular proliferation, and inhibition of apoptosis. Given the global increase in the burden of NAFLD and HCC, high-risk patients such as older diabetic individuals should be carefully monitored for HCC development. Future larger studies should explore whether the effect of diabetes on HCC risk in NAFLD cirrhosis is modifiable by the type of antidiabetic medication and the effectiveness of diabetes control.