Researchers have demonstrated that an optical technique known as Raman spectroscopy can be used to differentiate between benign and cancerous thyroid cells. The new study shows Raman spectroscopy’s potential as a tool to improve the diagnosis of thyroid cancer, which is the ninth most common cancer with more than 50,000 new cases diagnosed in the United States each year.
“Our encouraging results show that Raman spectroscopy could be developed into a new optical modality that can help avoid invasive procedures used to diagnose thyroid cancer by providing biochemical information that isn’t currently accessible,” said James W. Chan from the University of California, Davis, U.S.A. “This could have a major impact in the field of pathology and could lead to new ways to diagnose other diseases.”
A lump — or nodule — in the neck is a common symptom of thyroid cancer. However, most thyroid nodules aren’t cancerous. Ultrasound-guided fine needle aspiration biopsies are typically used to check for cancer by inserting a thin needle into the nodule to obtain cells that are prepared on a microscope slide, stained and analysed by a pathologist. For about 15 to 30 percent of cases, the pathologist cannot determine whether cells acquired from the biopsy are benign or malignant. For these cases, a surgical procedure known as a thyroidectomy is required to remove tissue, which provides more information for a more accurate diagnosis. The researchers turned to Raman spectroscopy as a possible solution because it is a non-invasive technique that requires no sample preparation or staining to determine subtle differences in the molecular composition of complex samples such as cells.
“We would like to use Raman spectroscopy to improve the pathologist’s analysis of the cells obtained with fine needle aspiration to reduce the number of thyroidectomies necessary,” said Chan. “This would both minimize surgical complications and reduce healthcare costs.”
For the new study, the researchers used a line-scan Raman microscope that allowed them to rapidly acquire Raman signals from an entire cell volume. This allowed them to more accurately capture the chemical composition of entire cells compared to other approaches that acquire a Raman spectrum from only part of a cell’s volume. Multivariate statistical methods and classification methods were then used to analyse the Raman data and classify the cells in an objective, unbiased manner.
The researchers applied this Raman spectroscopy approach to individual cells isolated from 10 patient thyroid nodules diagnosed as benign or cancerous. The data analysis identified unique spectral differences that could distinguish cancerous cells from benign with 97 percent diagnostic accuracy. They also showed that other subtypes could be identified by their spectral differences.
“These preliminary results are exciting because they involve single cells from human clinical samples, but more work will need to be done to take this from a research project to final clinical use,” said Chan.
The Optical Society (OSA) https://tinyurl.com/y6hw35z8

by Dr Allison B. Chambliss
The diagnosis of acute pancreatitis has long relied on elevations in serum amylase or lipase. Recent test utilization efforts have called f or the discontinuation of amylase in acute pancreatitis, favouring the higher specificity and longer elevation of lipase. However, neither biomarker correlates with disease severity, and early recognition of severe cases remains a diagnostic challenge.
Introduction to acute pancreatitis
Acute pancreatitis (AP) represents one of the most common gastrointestinal-related causes for hospital admissions. AP refers to an inflammatory condition of the pancreas commonly associated with a severe, rapid onset of abdominal pain. Patients may also experience other non-specific symptoms, including fever, tachycardia, nausea and vomiting. AP may be classified as mild, moderate or severe based on the degree of organ failure and systemic complications, a system referred to as the revised Atlanta classification (Table 1) [1].
The most frequent cause of AP is gallstones, which are hardened deposits of bile. Gallstones may account for 40–70% or more of AP cases, depending on the geographic region [2]. Gallstone pancreatitis typically resolves upon spontaneous or endoscopic removal of the stone. Once recovered, gallstone pancreatitis patients typically undergo cholecystectomy, the surgical removal of the gallbladder, to prevent recurrent AP episodes. Alcohol abuse is typically ranked as the second most frequent cause of AP (25–35% of cases), followed by a variety of other rarer causes such as metabolic abnormalities, drugs and toxins, and trauma.
Treatment for most patients involves supportive care, including fluid resuscitation, pain control and monitoring. Although patients with mild disease may recover within a few days without complications, the most severe cases may involve systemic inflammatory response syndrome with the failure of multiple organs, including acute respiratory failure, shock, and/or renal failure. Rapid diagnosis of AP and assessment of risk for disease severity, both of which rely on laboratory testing, are critical to guide patient management. Recurrent episodes of AP may progress to chronic pancreatitis.
Increases in disease prevalence
The annual incidence of AP is estimated at 20–40 per 100¦000 worldwide [3]. Interestingly, the incidence has increased over the past few decades, particularly in Western countries [4]. One study found an increase of 13.2% in AP-related hospital admissions in 2009–2012 compared to 2002–2005 across the USA [5]. Although these epidemiological trends are not entirely understood, several reasons for the overall increasing incidence of AP have been proposed. One hypothesis is the global epidemic of obesity, which may promote gallstone formation. Increases in alcohol consumption could also play a role in some countries. Other experts suggest that the wider availability and increased frequency of laboratory testing may be major factors. This latter concept is in alignment with the fact that although cases in AP have risen, the mortality rate of the disease has, in fact, declined [5]. Nevertheless, mortality remains high in the severe case category.
Biomarkers for AP
Serum amylase and lipase are well-established as the primary biomarkers for the diagnosis of AP. Both amylase and lipase are digestive enzymes; amylase hydrolyses complex carbohydrates to simple sugars, and lipase catalyses the hydrolysis of triglycerides. Although lipase is synthesized predominantly by the pancreas, amylase is produced both by the pancreas (P-type) and the salivary glands (S-type) and is found in several other organs and tissues. Both enzymes are released into the circulation at the onset of AP, and elevations of both are typically observed within 3-6|h [6, 7]. Multiple clinical societies and guidelines recommend a serum amylase or lipase test result greater than three times the upper reference limit as a diagnostic criterion for AP, in addition to characteristic symptoms and imaging findings [2, 8]. Both biomarkers are widely measured by automated enzymatic methods and are thus commonly found in routine hospital laboratories, permitting rapid diagnoses. Notably, most routine assays do not distinguish between P-type and S-type amylase. This distinction requires the analysis of amylase isoenzymes, which is typically limited to reference laboratories.
Questioning the value of amylase
Serum amylase and lipase are well-established as the primary biomarkers for the diagnosis of AP. Both amylase and lipase are digestive enzymes; amylase hydrolyses complex carbohydrates to simple sugars, and lipase catalyses the hydrolysis of triglycerides. Although lipase is synthesized predominantly by the pancreas, amylase is produced both by the pancreas (P-type) and the salivary glands (S-type) and is found in several other organs and tissues. Both enzymes are released into the circulation at the onset of AP, and elevations of both are typically observed within 3-6|h [6, 7]. Multiple clinical societies and guidelines recommend a serum amylase or lipase test result greater than three times the upper reference limit as a diagnostic criterion for AP, in addition to characteristic symptoms and imaging findings [2, 8]. Both biomarkers are widely measured by automated enzymatic methods and are thus commonly found in routine hospital laboratories, permitting rapid diagnoses. Notably, most routine assays do not distinguish between P-type and S-type amylase. This distinction requires the analysis of amylase isoenzymes, which is typically limited to reference laboratories.
Questioning the value of amylase
In contrast to amylase, lipase is reabsorbed by the tubules of the kidney and is not excreted into the urine. Thus, lipase tends to remain elevated for longer than amylase, which may allow for a longer diagnostic window for AP. This advantage, in addition to lipase’s higher specificity for the pancreas, has led some organizations to recommend lipase over amylase for the diagnosis of AP. The American Board of Internal Medicine Foundation’s Choosing Wisely® campaign, in collaboration with the American Society for Clinical Pathology, has recommended: “Do not test for amylase in cases of suspected acute pancreatitis. Instead, test for lipase” [9].
Despite these recommendations, many hospital laboratories still maintain assays for amylase. We performed a retrospective audit at our institution to determine the ordering patterns of amylase relative to lipase in cases of AP. We found that in a cohort of 438 consecutive patients admitted with AP, lipase was ordered for all patients, while amylase was only ordered for 12% of patients [10]. We observed that most of the amylase orders stemmed from patients with gallstone pancreatitis who were referred for laparoscopic cholecystectomy procedures and who were under the care of the surgical team. We speculated that amylase may have been co-ordered with lipase in this subgroup of patients to check for biomarker normalization. Laparoscopic cholecystectomy is ideally to be performed as early as possible when gallstone AP resolves, and normalization of amylase or lipase may be used to document that resolution. Because amylase is believed to fall more rapidly than lipase after AP, trending amylase over time could possibly allow for a quicker documentation of biomarker normalization. However, our study also showed that there was no significant difference in amylase versus lipase in the time for the biomarker to fall below three times the upper reference limit. These observations led us to further question the added value of amylase relative to lipase alone in the diagnosis and management of AP.
Lipase does have limitations that may preclude it from being the AP biomarker of choice in some cases. Lipase may be elevated in non-pancreatic conditions such as renal insufficiency and cholecystitis (Table 2). Both amylase and lipase may rarely be non-specifically elevated due to complexes with immunoglobulins, termed macroamylasemia and macrolipasemia. Further, amylase may be useful in the workup of other pancreatic diseases and, unlike lipase, can be measured in the urine. Quantitation of amylase in body fluids, such as pancreatic fluid and peritoneal fluid, can aid in the evaluation of pancreatic cysts and pancreatic ascites [11]. For these reasons, many laboratories choose to maintain amylase assays.
An unmet need for biomarkers for AP severity
Although AP may be easily diagnosed with elevations in amylase or lipase, there is an unmet need for biomarkers or algorithms that can specifically identify severe forms of AP early in the disease course. Twenty to thirty percent of AP patients may develop a moderate or severe form of the disease involving single or multiple organ dysfunction or failure and requiring intensive care. Identifying the severe cases early such that treatment may be tailored to minimize complications remains one of the major challenges of AP. Risk factors such as old age and obesity often correlate with disease severity. However, neither amylase nor lipase levels correlate with disease severity, and no other laboratory tests are consistently accurate to predict severity in patients with AP.
In 2019, the World Society of Emergency Surgery (WSES) published guidelines for the management of severe AP [12]. These guidelines indicate that C-reactive protein (CRP), an acute phase reactant synthesized by the liver and a non-specific indicator of inflammation, may have a role as a prognostic factor for severe AP. However, CRP may not reach peak levels for 48 to 72|h, limiting it as an early severity indicator. Specifically, WSES recommended that a CRP result greater than or equal to 150|mg/L on the third day after AP onset could be used as a prognostic factor for severe disease. Elevated or rising blood urea nitrogen, hematocrit, lactate dehydrogenase, and procalcitonin have also demonstrated predictive value for pancreatic necrosis infections.
Other biomarkers have been investigated to distinguish mild from non-mild forms of AP. Interleukin-6 has shown good discriminatory capability in combination with CRP [13]. Resistin is a more recently discovered peptide hormone that was first described as a contributor to insulin resistance (hence the name). Resistin is secreted by adipocytes and may play a role in obesity, hypertriglyceridemia, and inflammatory cytokine reactions. A prospective observational study found that resistin levels were better than CRP for predicting severe AP on the third day and for predicting the development of necrosis [14]. However, more studies are needed before resistin can be recommended as a prognostic indicator, and clinical resistin testing is not widely available. Thus, there still remains a need for prognostic severity biomarkers that rise early (prior to 48|h) in the course of AP.
The authors
Allison B. Chambliss PhD, DABCC
Department of Pathology, Keck School of Medicine of the University of Southern California, Los Angeles, CA 90033, USA

E-mail: abchambl@usc.edu

In symptom-free individuals, the detection of misfolded amyloid-β protein in the blood indicated a considerably higher risk of Alzheimer’s disease – up to 14 years before a clinical diagnosis was made. Amyloid-β folding proved to be superior to other risk markers evaluated, as shown by scientists from the German Cancer Research Center (DKFZ), Ruhr University Bochum (RUB), the Saarland Cancer Registry, and the Network Aging Research at Heidelberg University.
There is currently still no effective treatment for Alzheimer’s disease. For many experts, this is largely due to the fact that the disease cannot be clinically diagnosed until long after the biological onset of disease when characteristic symptoms such as forgetfulness appear. However, the underlying brain damage may already be advanced and irreversible by this stage.
"Everyone is now pinning their hopes on using new treatment approaches during this symptom-free early stage of disease to take preventive steps. In order to conduct studies to test these approaches, we need to identify people who have a particularly high risk of developing Alzheimer’s disease," explained Hermann Brenner from DKFZ. In patients with Alzheimer’s disease, misfolding of the amyloid-β protein may occur 15–20 years before the first clinical symptoms are observed. The misfolded proteins accumulate and form amyloid plaques in the brain. A technique devised by Klaus Gerwert from RUB can determine whether amyloid proteins are misfolded in blood plasma.

In a previous study, Gerwert and Brenner showed that the amyloid-β changes in the blood can be demonstrated many years before the clinical onset of disease. They also showed that demonstration of misfolded amyloid-β in the blood correlates with plaque formation in the brain. The researchers now wanted to investigate whether analysis of amyloid-β can be used to predict the risk of developing Alzheimer’s disease and how the risk marker performs in comparison to other known and suspected risk factors. To do so, they re-examined blood samples collected as part of ESTHER, a cohort study led by Hermann Brenner and conducted in collaboration with the Saarland Cancer Registry. The cohort study was initiated back in the year 2000.

In the current study, the researchers looked at the initial blood samples of 150 ESTHER participants in whom dementia was subsequently diagnosed during the 14-year follow-up period. These samples were compared with those of 620 randomly selected control participants not known to have been diagnosed with dementia who correlated with the dementia participants in terms of age, sex, and level of education.
Participants with Aβ misfolding had a 23-fold increased odds of Alzheimer’s disease diagnosis within 14 years. In patients with other types of dementia, such as those caused by reduced blood supply to the brain, the study did not demonstrate an increased risk, supporting Alzheimer’s disease specificity.

The researchers also included a number of other possible risk predictors in their analysis, including a particular variant of the gene for apolipoprotein E (APOE Ɛ4) and pre-existing diseases (diabetes, high blood pressure, de-pression) or lifestyle factors (bodyweight, level of education). With the exception of the APOE4 status, which showed a 2.4 times higher risk in those people who later went on to develop Alzheimer’s disease, none of the factors studied correlated with the risk of disease.
In predicting the risk of disease, it was largely irrelevant whether 0–8 or 8–14 years had passed between the time the blood sample was obtained and the clinical onset of dementia.
"This work was not about the use of amyloid-β folding as a diagnostic marker. Instead, we wanted to examine whether this marker could be used for risk stratification in the Alzheimer’s disease therapeutic development setting. Amyloid-β misfolding proved to be a far superior risk marker compared to the other potential risk factors," explained lead author Hannah Stocker from DKFZ and the University of Heidelberg’s Network Aging Research.
The German Cancer Research Center (DKFZ) www.dkfz.de/en/presse/pressemitteilungen/2019/dkfz-pm-19-46-Protein-misfolding-as-a-risk-marker-for-Alzheimers-disease-up-to-14-years-before-the-diagnosis.php

A team led by Johns Hopkins Kimmel Cancer Center investigators reports that a new laboratory test they developed to identify chemical changes to a group of cancer-related genes can accurately detect which breast tumours are cancerous or benign, and do it in far less time than gold-standard tests on biopsied breast tissue.
Although the findings are preliminary and need further validation in larger groups of people, the investigators say the test has the potential to dramatically reduce the time (minimum by one month, maximum by 15 months) generally needed to make a definitive breast cancer diagnosis in poorer countries.  A quick diagnosis has already been definitively proven to boost survival for all cancers by reducing wait times to surgical and other treatments. A report on the test, which exploits the tendency of some cancer-related genes to undergo the attachment of a chemical group, by a process known as methylation, has been published.
“Diagnosis is a huge bottleneck to starting treatment, especially in developing countries that have a small number of pathologists available to review breast cancer biopsies who serve a huge population,” says study leader Saraswati Sukumar, Ph.D., professor of oncology and pathology at the Johns Hopkins Kimmel Cancer Center.  “That means a test like ours could be especially useful in places with fewer resources and where mortality rates from breast cancer are much higher compared to the developed world.”
Breast cancer cases are rising around the world, Sukumar notes. Globally, breast cancer incidence is steadily increasing. In 1980, GLOBOCAN reported 641,000 new cases of breast cancer worldwide. In 2018, the estimated incidence of breast cancer worldwide rose to 2.1 million cases (a 3.2% annual rate of increase) with 626,000 deaths due to this cancer.
The reasons for higher death rates in the developing world include social stigmas that prevents many women from seeking timely treatment and a lack of healthcare resources. However, a major factor is time between biopsies and delivery of a diagnosis, which can be as long as 15 months in places with fewer resources compared to a few days or weeks in the United States.  
Seeking to shrink the time from biopsy to diagnosis, Sukumar and her colleagues in the Johns Hopkins Kimmel Cancer Center, Johns Hopkins University School of Medicine’s departments of pathology, surgery, and radiology, and the Johns Hopkins Bloomberg School of Public Health and collaborators from Cepheid developed a novel technology platform.  Here, a patient’s biopsy sample is loaded into cartridges and inserted in a machine that tests levels of gene methylation—a chemical addition to genes that results in changes in gene activity. This platform returns methylation marker results within five hours.
These results suggest that the test holds promise as a “first pass” to distinguish between malignant and benign breast tumours, Sukumar says. With the 5-hour-long return on results, low skill required to run the test, and relatively low expense, it could offer hope of speeding diagnosis for thousands of women worldwide.
Sukumar cautions that the team’s molecular test cannot replace expert analysis by a pathologist, whose skill will be necessary to review core biopsies of the breast lesion for a definitive diagnosis and optimal therapy recommendations.
John Hopkins University https://tinyurl.com/yxkg5sjy