Tumour cells circulating throughout the body in blood vessels have long been feared as harbingers of metastasizing cancer — even though most free-floating cancer cells will not go on to establish a new tumour.
But if these cast-offs could be accurately counted, they could provide an additional way to track treatment or screen for the disease.
New research by University of Wisconsin–Madison School of Pharmacy Professor Seungpyo Hong and his collaborators builds on several years of work in isolating these circulating tumour cells, or CTCs, by demonstrating improved methods for their capture on clinical samples for the first time. By forcing cancer cells to slow down and developing stronger molecular traps specific to CTCs, researchers were able to identify large numbers of the cells in cancer patients undergoing radiation therapy.
The number of CTCs dropped during therapy and subsequently rebounded in those patients that ended up requiring additional treatment — suggesting that this technology could supplement other techniques for tracking the progress of treatment.
Scientists have recognized CTCs as potentially useful metrics for tracking a patient’s disease for some time. But the cells are the proverbial needle-in-a-haystack, drowned out by billions of ordinary red blood cells and other cells found in the blood. Developing ways to specifically concentrate and trap CTCs has been technically challenging, with existing technologies only identifying a handful of cells from certain patients.
Hong’s team was inspired by the behaviour of CTCs in the blood, which attach themselves to blood vessel walls and begin tumbling along looking for suitable places to invade. This behaviour separates them from the oxygen-carrying cells floating by and is mimicked in the CapioCyte technology using an array of sticky proteins that force the CTCs to begin rolling, which slows them down.
The cells are then trapped using a series of three cancer-specific antibodies, proteins that tightly bind and hold onto the CTCs. To make the connection even stronger, the researchers developed a nanoscale structure shaped a little like a tree, with each branch tipped with an antibody. As a cancer cell passes nearby, many individual branches can latch on, increasing the strength of the attachment.
The cell rolling and multi-tipped branches helped the researchers capture an average of 200 CTCs from each millilitre of a patient’s blood, many times the number of cells captured with previous technology. They identified cancer cells in each of 24 patients undergoing treatment for head-and-neck, prostate, rectal or cervical cancer that enrolled in the study.
“The absolute numbers of CTCs don’t represent too much because there’s too much variation individually, but the more important thing we found was the trend — how the CTC numbers change over time upon treatment. So, for example, we’ve shown that the CTCs go down when the patients are responding really well to the radiotherapy,” says Hong.
Although the number of cells did not correlate with the stage, and thus severity, of the cancer, the reduction in cells was correlated with successful radiation therapy. In two of the three patients that had recurring or persistent disease, CTC numbers came back up.
“Our data suggest that we have a good chance of making CTCs a predictive biomarker or biomarker for surveillance for at least a few cancers, and that’s always exciting,” says Wang.
University of Wisconsin – Madisonnews.wisc.edu/improved-capture-of-cancer-cells-in-blood-could-help-track-disease/

High levels of cytoskeleton-associated protein 4 (CKAP4) have been identified in the blood of patients with lung cancer. In a novel study investigators found that CKAP4 levels were significantly higher in patients with lung cancer than in healthy individuals. They further determined that CKAP4 levels are already elevated in the blood of patients with stage I disease, making it a potential non-invasive diagnostic marker that could change current practices in the diagnosis and treatment of some types of lung cancer, including non-small-cell lung cancer and squamous cell carcinoma, and improve patient outcomes.
Lung cancer is the leading cause of cancer deaths in both men and women in the United States and worldwide. The disease is associated with a poor prognosis because most lung cancers are only diagnosed at an advanced stage.
"The identification of patients at an early stage of cancer when it can be treated surgically is extremely important to improve prognosis," explained Yuichi Sato, PhD, Department of Molecular Diagnostics, Kitasato University School of Allied Health Sciences, Sagamihara, Kanagawa, Japan, who led the study. "We need better biomarkers for early diagnosis."
Current biomarkers for lung cancer include carcinoma embryonic antigen (CEA), sialyl Lewis X antigen (SLX), squamous cell carcinoma (SCC) antigen, and cytokeratin fragment (CYFRA) 21-1, but these are not sensitive enough to detect tumors early, according to co-investigator Ryo Nagashio, PhD, from the Kitasato University School of Allied Health Sciences. "The results of our study provide evidence that the CKAP4 protein may be a novel early sero-diagnostic marker for lung cancer."
Researchers performed reverse-phase protein array analysis using a monoclonal antibody designated as KU-Lu-1 antibody on the blood of 271 lung cancer patients and 100 healthy individuals. KU-Lu-1 reacted only with tumor cells and tumor stromal fibroblasts in lung cancer tissues and not with normal lung tissues. Using immunoprecipitation and mass spectrometry, they confirmed that the KU-Lu-1 antibody recognized CKAP4 in lung cancer cells and tissues, and its secretion into the culture supernatant was also confirmed. In addition, a validation set consisting of samples from 100 patients with lung cancer and 38 healthy controls was also studied.
CKAP4 was recently identified as a receptor of Dickkopf1 (DKK1). Expressions of DKK1 and CKAP4 were frequently observed in tumor lesions of human pancreatic and lung cancers, and the simultaneous expression of both proteins in tumor tissues was inversely correlated with prognosis and relapse-free survival.
Across disease stages I-IV, the sensitivities of serum CEA, CYFRA, and SCCA are reported with 30 to 52, 17 to 82, and 24 to 39 percent, respectively. In this study, the sensitivity of serum CKAP4 was 81 percent in the training set and 69 percent in the validation set. These rates are higher than those of the current sero-diagnostic markers. Furthermore, the sensitivity of serum CKAP4 was also high even in stage I non-small-cell lung cancer and squamous cell carcinoma.
"The use of CKAP4 as a biomarker could change current practices regarding the treatment of lung cancer patients, and the diagnostic accuracies may be markedly improved by the combination of CKAP4 and conventional markers," concluded Dr. Sato.

EurekAlert
www.eurekalert.org/pub_releases/2018-05/e-nbi050718.php

A group of investigators from Mayo Clinic and multiple academic research centres in Italy have identified a genetic model for predicting outcomes in patients with primary myelofibrosis who are 70 years or younger and candidates for stem cell transplant to treat their disease.
“Myelofibrosis is a rare type of chronic leukemia that disrupts the body’s normal production of blood cells,” says Dr. Tefferi. “Prior to this study, the most comprehensive predictive model for outcomes in myelofibrosis, utilized mostly clinical variables, such as age, hemoglobin level, symptoms, white blood cell count and the percentage of immature cells in the peripheral blood.”
Dr. Tefferi says he and his colleagues incorporated new genetic tests in the model for gene mutations including JAK2, CALR, and MPL, which are known to drive myelofibrosis. He says the new model also tests for the presence or absence of high-risk mutations such as ASXL1 and SRSF2. “Our model is also unique in that we developed it for patients who are age 70 years or younger who may still be candidates for a stem cell transplant to treat their disease,” Dr. Tefferi says.
Researchers studied 805 patients with primary myelofibrosis who were 70 years of age or younger. Patients were recruited from multiple centres in Italy and from Mayo Clinic in Minnesota. The Italian and Minnesota groups formed two independent learning and validation cohorts. “We were surprised by how similar the predictive models performed in two completely separate patient databases,” Dr. Tefferi says.
Dr. Tefferi says that genetic information is increasingly being used as a prognostic biomarker in patients with primary myelofibrosis and he anticipates the potential use of such an approach along with relevant clinical, cytogenetic and mutational data for other hematologic and non-hematologic cancers.
Mayo Clinic Cancer Centerhttps://tinyurl.com/y7zlxtqz

Scientists have known for decades that the Hox family of transcription factors are key regulators in the formation of blood cells and the development of leukemia. Exactly how this large family of genes, which are distributed in four separate chromosomal clusters named A through D, is regulated has been less clear. Now, new research from the Stowers Institute for Medical Research reveals that a DNA regulatory element within the Hoxb cluster globally mediates signals to the majority of Hoxb genes to control their expression in blood-forming stem cells.
“It’s like we found a general control that simultaneously turns the lights on and off in many rooms, rather than having a single switch that controls each individual room,” says Stowers Investigator Linheng Li, PhD, who co-led the study along with Stowers Scientific Director and Investigator Robb Krumlauf, PhD. These findings also help explain why a particular form of leukemia resists treatment and points to potential new therapeutic avenues.
In mammals, the blood system contains a number of mature cell types — white blood cells, red blood cells, platelets — that arise from blood-forming, or hematopoietic, stem cells (HSCs). HSCs renew themselves and differentiate into other cells to replenish the body’s blood supply in a process called hematopoiesis. Hox genes, which are well known for their roles in establishing the body plan of developing organisms, are also important for HSCs to maintain their critical balancing act in the adult blood system, and have been implicated in the development of leukemia.
In an article Li, Krumlauf, and co-authors including first author Pengxu Qian, PhD, second author Bony De Kumar, PhD, and other collaborators provide new details as to how Hox genes are regulated in HSCs. They report that a single cis-regulatory element, DERARE, works over a long range to control the majority of Hoxb genes in HSCs in a coordinated manner. The researchers found that the loss of the DERARE decreased Hoxb expression and altered the types of blood cells arising from HSCs, whereas “turning on” DERARE allowed Hoxb cluster gene expression in progenitor cells and increased the progression of leukemia.
Genes can be regulated by non-coding DNA sequences termed cis-regulatory sequences. These sequences get input from multiple types of molecules, such as transcription factors, histone modifiers, or various morphogens. The DERARE, or distal element RARE (retinoic acid response element), is a cis-regulatory element that responds to signals from the vitamin A derivative retinoic acid and determines the fate of HSCs.
Using human leukemia cell lines and mouse models, the Stowers researchers and collaborators have identified a mechanism for how the retinoid-sensitive DERARE maintains normal hematopoiesis and prevents acute myeloid leukemia (AML) by regulating Hoxb cluster genes in a methylation-dependent manner.
Methylation is the process of adding methyl groups to the DNA molecule, which can change the activity of the DNA segment. The researchers demonstrated that DNA methyltransferases mediate DNA methylation on DERARE, leading to reduced Hoxb cluster expression. AML patients with mutations in the DNA methyltransferase DNMT3A exhibit reduced DERARE methylation, elevated Hoxb expression, and adverse outcomes.
“In two human AML cell lines carrying a DNMT3A mutation, we used an adaptation of genome editing technology called dCas9-DNMT3A to specifically increase the DNA methylation on DERARE. This targeted methylation technique was able to reduce Hoxb cluster expression and alleviate the progression of leukemia,” says Qian. “It is known that Hoxb cluster genes show a dramatic increase in expression in patients with DNMT3A-mutated AML. Our work provides mechanistic insights into the use of DNA methylation on the DERARE as a potential screening tool for therapeutic drugs that target DNMT3A-mutated AML, thus leading to the development of new drugs for treating AML, in which DNA methylation is abnormal.”

Stowers Institutehttps://tinyurl.com/yarf3xfd

Hong Kong Baptist University (HKBU) Chemistry scholars have invented a new class of multifunctional cyanine compounds that can be used for detection, imaging and thus treatment of Alzheimer’s disease. The discovery has been granted four US patents and a patent by the Chinese government.
The research team was jointly led by Professor Ricky Wong Man-shing and Associate Professor Dr Li Hung-wing with members from the Department of Chemistry of HKBU. By making use of the proprietary compounds, the HKBU team, on one hand, has proved that the cyanine compounds applied onto a “nano”-detection platform can quantify trace amounts of Alzheimer’s disease related protein biomarkers present in human fluids such as cerebrospinal fluid, serum, saliva, and urine. It is a rapid, low-cost and ultrasensitive detection assay. On the other hand, the compounds also serve as an imaging agent for in vivo detection and monitoring of disease progression and understanding the disease pathogenesis as well as a drug candidate for treatment of the disease.
Alzheimer’s disease is the most common neurodegenerative disorder, it is incurable and the underlying cause is still not well understood. Alzheimer’s disease is characterized by the formation of amyloid plaque in human brains. Clinical evaluation, cognitive tests and neuroimaging (monitoring the brain’s structural changes) are commonly used to diagnose Alzheimer’s disease, but are only effective after symptoms appear. Moreover, neuroimaging, such as magnetic resonance imaging (MRI), requires injecting contrast agents into a person that may bring health risks.
The proteins of interest, namely beta amyloid peptide, tau, and p-tau, in human’s cerebrospinal fluid are linked to Alzheimer’s disease. The versatile detection assay using the compounds developed by the team requires only a minute amount of the sample fluids (a few microliters) to reliably quantify the target proteins. The detection assay developed by the team is fast, cheaper and more sensitive than traditional commercially available biological methods.
Detection is based on the specific immuno-interactions between the target antigen and detection antibody that is immobilised on the surface of magnetic nanoparticles. The sandwiched immuno-assembly is then labelled with a newly developed turn-on cyanine compound that enhances the fluorescence signal, which is quantified by an imaging system.
Dr Li said, “This newly developed assay will be particularly useful as a low-cost yet accurate diagnostic and prognostic tool for Alzheimer’s disease. It can also serve as a novel alternative non-invasive tool for population-wide screening for the disease. This scientific detection assay has a high potential to serve as a practical diagnosis tool.”
Dr Li said that the new approach is universal and general enough to be readily modified and elaborated further, such as replacing the antibodies with other disease-associated antibodies, nucleic acids, for a broad range of biomedical research and disease diagnostics.   
Hong Kong Baptist Universityhkbuenews.hkbu.edu.hk/?t=press_release_details/2196