Scientists in Spain and Germany have identified a genetic signature of 27 microbiome organisms in stool samples that can identify people at high risk of pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer. The researchers, headed by Núria Malats, PhD, at the Spanish National Cancer Research Centre (CNIO), and Peer Bork, PhD, at the European Molecular Biology Laboratory (EMBL) in Heidelberg, say that while the high predictive value of the gene signature could serve as a biomarker to define people at high risk of the disease, if findings from the newly reported study can be validated in clinical trials, the panel could also represent a non-invasive method of diagnosing PDAC, even at its early stages.

“Sophisticated biostatistical and bioinformatics analyses have allowed us to construct a signature of 27 stool-derived microbes, mostly bacteria, that discriminates very well between cases with pancreatic cancer and controls, both in their most advanced and earliest stages, Malats and Bork noted.

The researchers reported on their work in Gut, in a paper titled, “A fecal microbiota signature with high specificity for pancreatic cancer.” A patent application has been made to develop a rapid, cost-effective, and non-invasive pancreatic cancer diagnostic kit.

Pancreatic cancer is the 12th most common cancer worldwide, but is set to become even more prevalent over the next couple of decades. The outlook for patients with PDAC is particularly poor, with fewer than 1 in 20 of those affected surviving five or more years.

This is largely because diagnosis is often made when the disease is advanced, and in the absence of effective treatment options. Scans, tissue specimens, and urine and blood samples are used to diagnose PDAC. But less invasive ways of diagnosis, and particularly at an early stage, are urgently needed, said the researchers. “In many cases, once pancreatic cancer is detected, it is too late,” they noted. “We need to diagnose the disease at a much earlier stage, before symptoms appear. To do this, we need to identify and define the population at risk and have good screening tests to detect the cancer when it is still curable.”

In their paper, they further stated, “Small-scale studies have proposed PDAC markers based on pancreatic tissue, urine, and blood serum with limited applicability. Yet there are currently no screening tools for PDAC in the clinic—in particular, for early disease stages … Sensitive and affordable tests for an early detection of PDAC could therefore improve outcome.”

Recent evidence suggests that changes in the microbiome—the trillions of bacteria, fungi, and other microbes that inhabit the digestive tract—may have a role in both the development and progression of pancreatic cancer. However, the authors noted, translating research findings into PDAC-specific microbiome signatures for use in clinical settings has remained “largely unexplored.”

To study the possible relationship between the microbiome and PDAC in greater detail, the researchers conducted a case-control study involving data from 136 individuals, including 57 newly diagnosed patients, 50 controls, and 27 patients with chronic pancreatitis, which is a known risk factor for pancreatic cancer. The participants were all deeply characterized at epidemiological and clinical levels and samples of saliva, feces, and pancreatic tissue were taken to analyze their microbiome. The subjects were recruited from two Spanish hospitals in Madrid (Ramon y Cajal Hospital) and Barcelona (Vall Hebron Hospital).

Contrary to what might have been expected, the oral microbiome was not found to be associated with pancreatic cancer, but fecal microbes were. The investigators identified a distinct microbial profile in the stool samples of people with PDAC when compared with the individuals with chronic pancreatitis and those without either disease. Pancreatic cancer is a disease with a very complex etiology and multiple risk factors such as age, obesity, diabetes, chronic pancreatitis, smoking, high alcohol consumption, blood type, and family history of cancer. To avoid biases and to ensure that the microbes identified are associated with pancreatic cancer and not with obesity, diabetes, or other risk factors, the authors controlled for these clinical and demographic variables in the analysis. “This level of analysis is unprecedented in pancreatic cancer metagenome studies,” the investigators said.

In red, Bacteroides, one of the microorganisms identified in this work, inside the nucleus of a pancreatic cell, in blue. [CNIO]

Machine learning techniques identified a characteristic enrichment of certain species and a relative scarcity of others. Methanobrevibacter smithii, Fusobacterium nucleatum, Alloscardovia omnicolens, Veillonella atypica, and Bacteroides finegoldii were abundant in the stool samples of the cancer patients while Faecalibacterium prausnitzii, Bacteroides coprocola, Bifidobacterium bifidum, or Romboutsia timonensis were depleted.

This microbial profile consistently identified patients with the disease, irrespective of how far it had progressed, suggesting that characteristic microbiome signatures emerge early on and that the stool microbiome might pick up early-stage disease, the researchers suggested.

Predictive ability was assessed using the area under the receiver operating characteristic (AUROC) curve, a graphic representation of how well a test identifies and excludes a given condition. An area beneath the ROC curve of 0.5 corresponds to random chance, whereas 1 equals perfect accuracy. In this case, the AUROC curve reached 0.84.

Accuracy increased still further, to 0.94, when the microbial profile was combined with blood levels of carbohydrate antigen 19-9, a biomarker that is indicative of pancreatic cancer, but also various other conditions, and which is the only current non-invasive test approved by the FDA for monitoring pancreatic cancer progression.

The predictive ability of the microbial profile was then validated in a separate group of 76 German people, 44 of whom had pancreatic ductal cancer and 32 of whom didn’t. The team separately validated the gene signature against publicly available data from 25 studies involving 5,792 samples covering nine different health conditions, including other cancers and type 2 diabetes, a risk factor for pancreatic cancer. “We moreover validated our metagenomic classifiers against the independently sampled, yet consistently processed, DE [German] population and against external populations of various health states from 25 different studies (n=5792).”

Nuria Malats, PhD [Laura M. Lombardía. CNIO]

The microbial profile of mouth, stool, and pancreatic tissue samples of patients with ductal pancreatic cancer were similar, suggesting that they may be closely linked. “Our data are strictly observational and cross-sectional,” the researchers acknowledged. “Nevertheless, there are strong indications that the identified fecal microbiome shifts are not merely a consequence of impaired pancreatic function or systemic effects thereof, although indirect effects cannot be ruled out.”

And while currently, screening programs are targeted to families with pancreatic cancer aggregation, which represent only 10% of the burden of the disease, including into these screening programs a stool analysis to identify the identified microbial signature could help to detect the rest of the population at risk, they pointed out.  “In summary, the described fecal microbiome signatures enabled robust metagenomic classifiers for PDAC detection at high disease specificity, complementary to existing markers, and with potential towards cost-effective PDAC screening and monitoring.”

The team also noted that, given previous research on a possible link between pancreatic cancer and the gut microbiome, “we believe that the presented panel of PDAC-associated bacterial species may be relevant beyond their use for diagnosis, providing promising future entry points for disease prevention and therapeutic intervention.”

In a linked commentary, Rachel Newsome, PhD, and Christian Jobin, PhD, of the University of Florida, cautioned: “Although promising, these findings have limited clinical value due to the cross-sectional nature of the study, and thus the predictive markers will need to be tested using a prospective cohort before reaching a conclusion on their clinical impact.” Further research would be needed to find out if the microbial profile is specific for pancreatic cancer and not shared with other types of cancer, they added.

Notwithstanding, the research represents “an important contribution toward the generation of predictive markers for [ductal pancreatic cancer] and highlights the key role of microbiota in cancer surveillance … and represents significant progress for non-invasive cancer detection,” Newsome and Jobin wrote. They suggested the work “brings clarity to the connection between microbiome signature and PDAC and represents significant progress for non-invasive cancer detection.”

Helen Rippon, PhD, chief executive at Worldwide Cancer Research, which partly funded the study, further commented, “This new breakthrough builds on the growing evidence that the microbiome—the collection of microorganisms that live side by side with the cells inside our body—is linked to the development of cancer. What’s amazing about this discovery is that the microbiome of stool samples from patients could be used to help diagnose pancreatic cancer early. Early detection and diagnosis are just as important an approach to starting new cancer cures as developing treatments.

“Pancreatic cancer is one of those cancers that we have seen very little improvement in survival over the last few decades. It’s often diagnosed late when it has already spread to other parts of the body and treatment options for people are extremely limited. This research provides hope that an effective, non-invasive way to diagnose pancreatic cancer early is on the horizon.”