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Oncologists’ Guide to Genomics

Stay current on the latest trends in genomics and molecular diagnostics for oncology.

Wednesday, May 5, 2021

By Susan Jenks​

Five years after the first liquid biopsy test entered the marketplace, several others that track the molecular fingerprints of cancer in the blood have won approval from the FDA. Although the most recent FDA approvals expand the reach of these blood-based tests, identifying more genetic mutations in several new cancers, they fall short of detecting early-stage disease in otherwise healthy individuals—still a distant goal, according to researchers.

“Liquid biopsy is definitely a game-changing technology,' said Phillip Castle, PhD, MPH, Director of the National Cancer Institute's Division of Cancer Prevention in Bethesda, MD. “But, it's much easier—though far from easy—to work from the starting point, where a person already has cancer."

In fact, all but one of the five tests now available measures DNA snippets and other genetic materials that tumors shed in the blood for clues to cancers in individuals battling advanced disease. Also, each of these tests, except for an early screening product for colon cancer, pairs liquid biopsy to traditional tissue biopsies as companion diagnostics, not as stand-alone tests.

Investigators' interest in moving liquid biopsy closer to a simple blood draw, however, has intensified in recent years with advances in next-generation sequencing, which allows for large-scale DNA analyses. Eventually, the hope is that such a test could identify not just a single cancer, but a wide spectrum of cancers, long before symptoms appear.

“That's the Holy Grail of research," Castle noted, given that improvements in prevention go hand-in-hand with treatment advances. “Imagine if we could find cancers earlier, how much better treatments would work," or even that some cancers might be cured, he said.

Researchers are actively pursuing early detection blood tests. While Galleri is a methylation-based multi-cancer screening test, CancerSEEK scours the blood for incipient disease, using mutated DNA and cancer-related proteins.

“These are the two tests out front in early detection," Castle said. Neither test has gained FDA approval, nor is it yet clear the benefit to patients, he noted. “And, until we're sure of that benefit, we're not recommending them."

Ultimately, Castle and others stressed that, to a reach liquid biopsy's full-screening potential, scientists need a much better understanding of tumor biology.

“Genetic abnormalities are only one piece of the puzzle," said William Cance, MD, a surgical oncologist and Chief Medical and Scientific Officer of the American Cancer Society in Atlanta. “We need to look comprehensively at tumors for the best therapy," from their metabolic changes and protein signatures in the blood to the epigenetic modifications that may occur, as cancers take hold, he said. “It's not just shed DNA in the blood."

Many issues remain unsettled, such as which testing approach works best. One approach nmeasures intact circulating tumor cells, while another —furtherest along, given its apparent improved sensitivity—looks at tumor-related DNA. A third searches for exosomes, an assortment of genetic debris, including DNA and RNA molecules and metabolites in the blood.

Investigators also don't know whether to use these tests in urine, cerebral spinal fluid, or even saliva in the future to monitor changes signaling worsening disease or a new cancer. “There are a variety of biomarkers out there and there's no need to limit yourself," Castle said. “The most important thing is—is it a good test?"

Meanwhile, the more immediate benefits of liquid biopsy continue to evolve. Although tissue biopsy remains the gold standard for guiding patient care, liquid biopsy is considered

cheaper and far easier to do, especially if patients are too ill to undergo tissue sampling or have tumors that are inaccessible to treatment.

“The ability to do these tests quickly (and often) is a big advantage in the sickest patients," Cance stated. Not only does liquid biopsy eliminate the need for further tissue biopsies, he said, “it may find mutations where there's a possible new drug available, opening up options that might help or make a difference in a patient's care."

Liquid biopsy also provides a way to monitor tumors as they progress. Because cancer cells change so quickly and reprogram quickly to escape immune defenses, Cance said, tracking their molecular trail in the blood through repeated blood draws offers a way to keep pace, shifting treatment strategies as needed.

“I think liquid biopsies are here to stay," though still in the early days of development, he said. “They're all part of precision medicine, tailored to the individual."

But, as with all cancer-detection efforts, he said researchers need to know more about which tumors shed the most DNA into the blood, whether some cancers shed none, and most importantly, “whether the DNA we're measuring is from the tumor we're targeting and trying to treat."

That uncertainty lies at the heart of ongoing research. Even normal aging can create DNA mutations mistaken for cancer, according to the FDA, blurring results and raising the risk and potential harm of overtreatment.

Susan Jenks is a contributing writer.

Thursday, March 25, 2021

Urinalysis has long been a staple of physical exams to detect and manage a number of diseases and disorders, but not cancer. What if it were that easy, though, and cancer was detected in its very earliest stages when the disease responds more favorably to treatment and improved outcomes are more likely?

That was the question posed by scientists at the Translational Genomics Research Institute (TGen), an affiliate of City of Hope, who have found a way of zeroing in on early-stage cancer by analyzing short strands of cell-free DNA in urine. Their study's findings were published today in the scientific journal Science Translational Medicine (2021: doi: 10.1126/scitranslmed.aaz3088).

Previous thought once held that DNA fragments in urine were degraded at random and were too short to provide any meaningful information about a disease as complex as cancer. TGen and City of Hope researchers and their colleagues from Baylor University and Phoenix Children's Hospital found that these DNA fragments are not random at all, and can clearly indicate a difference between healthy individuals and those with cancer.

"There are many steps between where we are now and where we want to go—detecting cancer from a urine sample—but without doubt this is an encouraging first step," said Muhammed Murtaza, MBBS, PhD, an Associate Professor and Co-Director of TGen's Center for Noninvasive Diagnostics, and the study's senior author.

Murtaza previously led a team of TGen scientists who pioneered the use of circulating tumor DNA in blood, using genetic fragments to detect cancer with a simple blood draw. This "liquid biopsy" method sidesteps the need for many surgical biopsies of suspected tumors, and means that physicians can monitor cancer in their patients more frequently given the less invasive nature of the procedure.

Collecting a urine sample reduces the physical invasiveness to zero, Murtaza explained, and may eliminate a lab visit, given that the sample could be collected at home and mailed in for analysis.

By studying tissue samples from children with various cancers, whose malignancies often move extraordinarily fast, and adults with pancreatic cancer, whose early detection is critical to their disease outcomes, researchers mapped the DNA fragmentation profiles in their urine.

"We found that certain regions of the genome are protected from fragmentation in urine from healthy individuals, but the same regions are more fragmented in patients with cancer," Murtaza said.

The fragmentation profiles were remarkably similar across multiple individuals; the length of the DNA fragments were similar, the regions of the genome where the fragmentation occurred were consistent, and informed researchers what type of cells contributed the fragments.

Ajay Goel, PhD, Chair of the Department of Molecular Diagnostics and Experimental Therapeutics and Associate Director for Basic Science at City of Hope, a world-renowned independent research and treatment center for cancer and diabetes, is one of the study's authors. He is a leading expert in developing early-detection blood tests for colon, pancreatic and ovarian cancers.

"If the study results come to fruition, our urinalysis technology would be a remarkable breakthrough in the detection of many cancers, especially in pancreatic cancer," Goel said. "If cancer is detected early, it could substantially lower the mortality rate for what is currently the third leading cause of cancer death in the U.S."

While early results are promising, the researchers indicate the need to test their findings in much larger populations of cancer patients and identify differences between men and women, young and old, and those with co-morbidities, such as diabetes and other chronic diseases.

"This is a fundamental new finding and provides a potentially dynamic path forward for the early diagnosis of cancer, given that urine is one of the easiest samples to collect," said Daniel D. Von Hoff, MD, TGen Distinguished Professor and one of the paper's authors. "If follow-on studies yield positive results, I could one day see this test becoming an integral part of one's annual physical."​

Friday, February 19, 2021

By Mark L. Fuerst

Assessments of circulating tumor DNA (ctDNA) can detect minimal residual disease (MRD) and help stratify colorectal cancer (CRC) patients into high and low risk of recurrence after resection, and it may also guide adjuvant chemotherapy.

Timely detection of recurrence and identification of CRC patients at high risk of recurrence after surgery and after completion of adjuvant therapy remain major challenges. From 20 percent to 30 percent of these patients relapse.

“If we could identify MRD, we could enable a better recurrent risk assessment. Earlier detection of recurrence could increase the proportion of patients that we treat with curative intent after the recurrence, thereby hopefully improving the survival of this entire patient group. We believe that ctDNA is a promising marker for MRD detection," said Tenna V. Henriksen, a PhD candidate at Aarhus University in Denmark, at the 2021 ASCO Gastrointestinal Cancers Symposium (Abstract 11).

Identifying High Risk

In a prospective, multicenter study, researchers set out to determine whether serial postsurgical ctDNA analysis could identify the patients at high risk of recurrence, provide an assessment of adjuvant therapy efficacy, and detect relapse earlier than standard-of-care radiological imaging. The study included 265 stage I-III CRC patients, the largest cohort assessed for ctDNA to date, Henriksen noted. All patients had the tumor resected and a subset of 166 patients was additionally treated with adjuvant chemotherapy.

The patients were recruited from surgical centers in Denmark and Spain. Blood samples were collected before and directly after surgery, and in some patients every 3 months after surgery for up to 3 years. These patients were also monitored by CT scans at 12 and 36 months after surgery. “The CT scans form the clinical basis for which we compared our ctDNA data from blood samples," said Henriksen.

Examining blood samples that were drawn within 2 months after surgery and before the start of neoadjuvant chemotherapy, the researchers were able to stratify patients into high and low risk of recurrence.

“When we look at the current rates, we can also see that there is a very low recurrence rate amongst the ctDNA-negative patients compared to the higher recurrence rate in the ctDNA-positive patients," explained Henriksen.

Not all ctDNA-positive patients experienced disease recurrence. “In fact, the four patients who did not experience disease recurrence but were ctDNA-positive all received adjuvant chemotherapy," she said. “This indicates that the outcome here was modified by the adjuvant chemotherapy, and that perhaps these patients were cured for any type of residual disease that was left after the surgery by adjuvant chemotherapy. Hopefully this could indicate that there is a benefit for ctDNA-positive patients in some cases to receive adjuvant chemotherapy."

The researchers also looked into stratifying patients into high and low risk of recurrence at the end of treatment. “Again, we can see a very good stratification into high and low risk of recurrence, and low recurrence rates in the ctDNA-negative patients versus the ctDNA-positive patients with even higher recurrence rates that we saw before," stated Henriksen.

“If we include longitudinal samples drawn every 3 months after surgery, we can shift a portion of the ctDNA-negative patients to being ctDNA-positive at a later time point, simply by including more sampling. We saw the recurrence rates in the ctDNA-negative patients drop to about 3 percent, which is very low. This indicates that serial sampling has a benefit over just single time-point sampling."

They also found that ctDNA had a median lead time of about 8 months compared to clinical CT scans. In addition, a multivariable analysis showed that ctDNA was a stronger biomarker compared to an already established biomarker, carcinoembryonic antigen (CEA), at least in terms of relapse-free survival.

Detecting Recurrences Earlier

In conclusion, Henriksen said: “We saw the patients with ctDNA detected immediately after surgery had a very high risk of recurrence. Also, we could see that longitudinal monitoring increased the predictive power of ctDNA and that we had a lead time of 8 months before radiological detection of recurrence using ctDNA. And finally, we could see that longitudinal testing with ctDNA outperformed CEA in recurrence-free survival prediction."

Before ctDNA is implemented in clinics, it needs to be tested in randomized trials. She noted that two Danish trials of ctDNA-guided clinical management have begun, IMPROVE-IT and IMPROVE-IT 2.  Henrisksen suggested that patients could be stratified according to ctDNA—ctDNA-positive patients would receive adjuvant treatment, but ctDNA-negative patients would not.

“Currently, all stage III patients receive adjuvant chemotherapy, but it's estimated that 60 percent of them were cured by surgery alone. These patients were in a sense overtreated with adjuvant chemotherapy. If you stratified them according to their real risk with ctDNA in mind, maybe you could spare some patients the toxic side effects of adjuvant chemotherapy," she noted.

She also suggested that using ctDNA could improve radiological surveillance. “Currently, we scan patients at 12 and 36 months—according to the Danish guidelines, at least—and only 20 percent of detected recurrences are offered curative intent treatment. If you switch to a ctDNA-based system, sampling ctDNA maybe every 3 months, and if they become ctDNA-positive you switch them over to an intensified radiological surveillance, [then] we could detect recurrences sooner because we have the lead time of ctDNA of around 8 months and thereby shift a larger proportion of patients into receiving curative intent treatment," Henriksen concluded.

Mark L. Fuerst is a contributing writer.​

Thursday, February 4, 2021

By Susan Jenks

In a small, proof-of-concept study, researchers have identified molecular changes in tumors that may partially explain why some cancer patients respond to treatments far outside the expected norm.

Using modern genomic tools, scientists at the National Cancer Institute collaborated with investigators from around the country to analyze the tumor samples of 111 patients with various types of advanced cancer—all identified as “exceptional responders." In roughly a fourth of them, the researchers found that distinct molecular features seemed to provide a rare survival edge after cancer treatment, with a few patients surviving many years past diagnosis.

“We all know a metastatic stage IV patient who's alive 7 years later and that this shouldn't happen," said Louis Staudt, MD, PhD, Director of the NCI's Center for Cancer Genomics, who co-led the study, along with Percy Ivy, MD, of NCI's Division of Cancer Treatment and Diagnosis.

To understand why such outliers occur, researchers developed stringent study criteria, classifying molecular mechanisms into four different categories: 1) the ability to repair DNA damage, 2) intracellular signaling pathway activity, 3) immune responses, and 4) prognostic genetics, in which a “nasty" tumor seen under the microscope proved less aggressive upon genomic analysis, with a known favorable outcome.

In all, the final analysis focused on 26 patients whose results appeared in a recent issue of Cancer Cell (2020; https://doi.org/10.1016/j.ccell.2020.10.015). Although that number might seem small, Staudt conceded, “it's actually large with respect to the difficulty in identifying these types of patients," usually studied one at a time. And, eventually, as further data accumulates about them, he said routine molecular analysis “will inform the care of a great number of cancer patients we would not know about," without benefit of a clinical trial.

However, because cancer is so heterogeneous a disease, he said “it's literally impossible to do all the clinical trials we would need to do," especially when a genetic mutation may be a “one-off"—useful, perhaps, to only a few patients.

“We would never do a clinical trial of the 1 percent of patients in whom PARP inhibitors work," for example, he said, so the NCI analysis provides a different type of evidence: better guidance in clinical decision-making and more precise personalized medicine in the future.

The investigators defined exceptional responders in the study as patients who had partial or complete responses to treatment—primarily combination chemotherapy—that ordinarily would be effective in less than 10 percent of those on similar drugs. They defined an exceptional response as a treatment response lasting at least 3 times longer than the median response to the same cancer drugs, before a cancer's return.

No gender, racial, or age distinctions have yet emerged in the NCI initiative. Most exceptional responders in the study skewed older—“not vastly different from the general demographics of cancer patients," Ivy said, ruling out the notion that youth alone might explain an exceptional response.

And using the NCI criteria, established through a review of medical literature and input from experts familiar with patient responses “way outside the norm," she noted, investigators in many other countries now have begun their own studies of exceptional responders. Unlike the NCI focus, however, scientists in the United Kingdom, France, and Italy, among others, are looking at exceptional responders in specific cancers, such as breast cancers in France and ovarian cancer in Australia.

Only six patients in the NCI analysis underwent treatment with immunotherapy before being classified as exceptional responders, primarily because the study predated the immunotherapy era, according to the investigators.

But one case, Staudt suggested, illustrates how immunotherapy already has changed the treatment landscape in cancer and future studies will include them. A bladder cancer patient, for example, who was placed on a checkpoint inhibitor after failing other treatments had a longer-than-expected response time, he said, with his cancer cells recruiting T-cell cytokines to attack his tumors. While such an unusual response, seen in only about 3 percent of bladder cancer patients, likely is one of those “one offs" he described, Staud said it underscores the importance of the immune system in cancer surveillance.

Other findings, highlighted in the study, included rare combinations of genomic errors that resulted in the death of tumor cells during treatment, known as synthetic lethality. In one instance, the researchers identified genetic mutations in the DNA repair genes of two cancer patients—one with bile duct cancer and the other with rectal cancer. Mutations in their BRCA1 or BRCA2 genes—rarely seen in these cancers—may have thwarted the tumors' ability to repair damaged DNA, the researchers theorized, allowing platinum-based chemotherapy drugs to kill cancer cells more efficiently.

The new NCI research, part of the institute's Exceptional Responders Initiative, was first launched in 2014. The hypotheses developed since then about the molecular underpinnings of exceptional responders will need to be validated by larger studies going forward, Staudt said, with a number of groups expected to publish in the next few years.

Susan Jenks is a contributing writer.

Tuesday, December 22, 2020

By Warren Froelich

The genetic makeup of prostate tumors in African-American men exhibits a higher frequency of damaging mutations, potentially contributing to the significant disparity in prostate cancer cases and death when compared to White males.

In a retrospective study published in the online issue of Molecular Cancer Research, a journal of the American Association for Cancer Research (AACR), a research team from NorthShore University HealthSystem in Illinois found more than 35 percent of prostate tumors in African-American men harbored potentially damaging mutations in several genes, a higher frequency than White men (2020; doi: 10.1158/1541-7786.MCR-20-0648).

What's more, prostate tumors from African-American men had a higher frequency of altered genetic regions than White men, including deletions and gains that may contribute to more aggressive prostate cancer.

“These distinct genetic differences may help us understand the racial disparity in prostate cancer cases and provide the future basis for genetic testing to better predict the prognosis of tumors at time of diagnosis and inform targeted treatment options," said Jianfeng Xu, DrPH, Vice President of Translational Research at NorthShore and senior author of the study.

According to the American Cancer Society, one in seven or 14.8 percent of African-American men will develop prostate cancer in his lifetime, with one in 25 (4%) dying from their disease. Overall, African-American men are 1.8 times more likely to be diagnosed—and 2.2 times more likely to die—from prostate cancer than White men.

“Prostate cancer incidence and mortality are highest in African-American men, but the exact reasons for the disparity are not fully understood," said Xu. “The disparity is likely due to multiple factors, including socioeconomic differences and biology. We suspect the differences in genetic changes that occur within tumors may play a critical role."

Genetic Research

Xu and first author Weenuan Liu, PhD, along with their colleagues, sought to sequence and compare DNA extracted from prostate cancer tumor tissues from African-American and White men to explore the genetic differences.

This can be challenging since the segment of DNA of interest containing the mutation only makes up a small proportion of the DNA present in the samples. The vast majority of DNA may come from normal cells, other clones of the primary tumor present in the sample, or the non-mutated copy of the gene of interest.

These challenges, along with an insufficient number of specimens for study, previously have hindered the identification of genetic factors responsible for more aggressive prostate cancer and higher mortality rates in African-American males.

To overcome this hurdle, Xu and colleagues turned to “deeper next-generation sequencing" to better separate mutated genes that drive prostate tumor growth and proliferation from all the other genetic noise.

“In deeper next-generation sequencing used in our study, we copy and read all the DNA segments greater than 2,000 times so that we can better detect the mutated segments we are interested in from all the other segments of DNA," Xu said in an interview.

Study Details

In their study, the research team focused on two types of major genetic changes: point mutations, a change in a single DNA position; and changes (deletions and gains) in a stretch of DNA affecting an entire gene or several genes, also known as “copy number alterations."

“These changes can be useful when predicting how the cancer will progress, whether it will metastasize, and how it will respond to certain drugs and treatments," Xu stated.

In the first part of the study, the NorthShore team extracted and sequenced 39 driver genes of interest in tumors and matched normal DNA from 77 African-American men with prostate cancer. After filtering out tumors containing less than 10 percent mutated genes, the team identified the top 22 mutated genes for analysis.

More than 35 percent of these patients' tumors had point mutations in several genes known to affect cancer growth and progression, including DNA repair genes ATM, BRCA2, and ZMYM3, among others.

ZMTM3, which regulates chromatin and DNA repair, was found to be among the most frequently mutated genes in patients, with nine of the 77 African-American patients (11.7%) harboring mutations in ZMYM3, compared to 2.7 percent of tumors from 410 white patients whose data was included in the Genomic Data Commons database.

“ZMYM3 promotes BRCA1 to the location of the damaged chromatin to facilitate DNA repair," said Xu. “Knockout of ZMYM3 in mice and in cells, respectively, causes defects in spindle assembly and impairs homologous recombination DNA repair leading to genomic instability."

In the second part of their study, the researchers compared the copy number alterations—where genetic material is gained or lost—in the prostate tumors of African-American men with White men. The study included a pool of 171 African-American patients and 860 White patients from several public databases. They found distinct copy number alterations between African-American and White patients in the more aggressive, high-grade prostate tumors (Gleeson score 7 or higher), but not in low-grade tumors.

High-grade tumors from African-American patients were more likely to have additional copies of the MYC oncogene and deletions of the LRP1B, MAP3K7, BNIP3L, and RB1 genes than tumors from White patients. Gain in MYC and loss of MAP3K7 or RB1 were also associated with more advanced tumor stage.

“Taken together, these results show that there are unique differences in prostate tumors of African-American and European American (White) men that could affect how we manage and treat prostate cancer between these two groups," Xu noted.

In the future, the research team is interested in expanding its study to include more African-American men from multiple institutions. In addition, they want to collect additional clinical data such as recurrence/relapse, metastasis, treatment, and prostate cancer-specific death, and then analyze their associations with distinct genetic alterations in African-American men. And then, the team plans to develop tests to detect these genetic changes.

“We have already developed and tested the feasibility of such type of assays for Caucasian patients," Xu stated. (Prostate 2020; https://doi.org/10.1002/pros.24057).

Warren Froelich is a contributing writer.