Liquid Biopsies Making Impact in Lung Cancer, Other Tumor Types

Mark R. Bowling, MD

Mark R. Bowling, MD

The noninvasive testing that has long been the goal of solid tumor analysis is making its presence felt in lung cancer, and other malignancies likely will not be far behind, experts say. Although many potential uses for liquid biopsies are envisioned, much work remains to be done to establish the clinical utility of these tests.

At the Vidant Health System in North Carolina, patients with advanced lung cancer who are candidates for molecular analysis will receive plasma-based testing at the same time that they undergo a traditional biopsy. Reports from the “liquid biopsy” are available within 3 to 4 business days, and specialists are finding that the ability to obtain speedy results is making an impact on treatment decisions and possibly on outcomes.

“If it’s adenocarcinoma, knowing that there might be certain markers that can make a difference for that patient’s treatment upfront and quickly is very helpful,” says Mark R. Bowling, MD, the pulmonologist who performs many of the biopsies conducted at the center. “You can do the same thing on tissue, of course, but you usually have to wait 14 days.

“As much information as you can have on a patient to help you make decisions as soon as you can is very important,” he added. “To the patient, it’s an emergency. You’re not going to fix it just by biopsying it—you have to start therapy.”

The use of liquid biopsy testing at the center, which serves communities with a population of 1.2 million people, illustrates the rollout of a noninvasive approach that has long been the holy grail of solid tumor analysis. The era of the liquid biopsy has arrived in clinical practice in the United States—at least in lung cancer—with an expanding toolkit from commercial and academic providers and a growing body of clinical evidence.

The Cleveland Clinic has named the liquid biopsy, a term broadly used to describe blood-based testing for genetic mutations, as one of the top 10 medical innovations poised to make an impact on patient care in 2017.¹ As part of the Cancer Moonshot initiative, biopharmaceutical companies and academic institutions have agreed to collaborate on building a Blood Pro ling Atlas that will aggregate raw datasets from liquid biopsy studies along with relevant clinical data².

Many exciting uses for liquid biopsies that span the spectrum of cancer diagnosis and care that leverage existing and emerging platforms are under study, yet the clinical utility of such assays is an evolving question.

The liquid biopsy “is here today and it’s already established, but the uses of it are much more limited than people understand,” said Nathan A. Pennell, MD, PhD, director of the Lung Cancer Medical Oncology Program at the Cleveland Clinic’s Taussig Cancer Center. “The validated uses are much more narrow. That may expand dependent on our identifying clinically proven uses for it as opposed to doing it because we can. There aren’t that many cases where being able to detect something in the blood has been validated to impact patient outcomes and survival. That’s what’s missing from a lot of the liquid biopsies.”

Pennell said 2 areas where the utility of liquid biopsies has been established thus far are in monitoring treatment for emerging resistance and in identifying targetable genetic mutations or fusions to guide therapy.

As a noninvasive diagnostic, the ability to learn such information from a blood sample is particularly helpful for patients who cannot undergo a biopsy or whose tumor sample is exhausted, noted Pasi A. Jänne, MD, PhD, director of the Lowe Center for Thoracic Oncology at the Dana-Farber Cancer Institute. Jänne co-chaired a workshop on liquid biopsies that the American Association for Cancer Research (AACR) and the FDA sponsored in July 2016.

“Today we have technology that can look at both single genetic alterations from cell-free DNA from the plasma or can sequence cell-free DNA from the plasma and look at multiple genetic alterations,” said Jänne. “Both academic institutions and commercial vendors are providing that. The vast majority of what we need today are predictive markers. We need to be able to guide the care of patients who have advanced cancer.”

Jänne said there is a pressing need for studies that compare the utility of liquid biopsies with tumor biopsies. “That is the body of evidence that we need for more drugs,” he said. “There are some clinical trials that are allowing entry based on blood-based genotyping, not necessarily tumor-based genotyping. One of the questions is whether that is as good. That remains to be determined.”

In the immediate future, Jänne expects liquid biopsies to be most beneficial in solid tumor types where targeted therapies already have been identified, starting with non—small cell lung cancer (NSCLC) and including colon cancer, breast cancer, and melanoma.

Testing Choices Grow

“As we find more and more actionable mutations, this is going to become more and more useful,” said Pennell, noting the need to identify additional drugs that can target the growing body of mutations that researchers have characterized.In June 2016, the FDA approved the first blood-based assay for use in clinical decision making, the cobas EGFR Mutation Test v2, for patients with NSCLC.³ The test uses real-time polymerase chain reaction (PCR) technology on circulating-free tumor DNA derived from plasma.⁴

The assay is now approved as a companion diagnostic for the detection of EGFR exon 19 deletions and L858R substitutions as a means of selecting patients for frontline therapy with erlotinib (Tarceva) and for identifying the T790M resistance mutation as a screening tool for the use of osimertinib (Tagrisso) in individuals with progressive disease. The FDA said patients who test positive for the indicated mutations are eligible for the matching targeted therapy; however, those whose samples are negative on plasma-based testing should receive further testing with a tissue biopsy.

Although that test has gained the agency’s stamp of approval as a companion diagnostic, the menu of commercially available assays is growing outside of that specific category. The assay that doctors at the Vidant center have chosen for liquid biopsies is GeneStrat, which uses droplet digital PCR technology to analyze circulating tumor DNA and RNA in plasma from a whole-blood sample.⁵

The test can detect EGFR sensitizing and resistance mutations, EML4-ALK fusions, and KRAS mutations, with sensitivities ranging from approximately 85% for the gene fusions to nearly 96% for EGFR alterations, according to Biodesix, the Colorado-based company that developed the test.⁶

For an analysis of turnaround times using the assay, Bowling and colleagues reviewed 179 samples from 5 oncology centers.⁷ They found that the average time from receipt of test kit to biomarker results was about 33 hours compared with guideline recommendations of a maximum 10 business days for test reports.

Bowling said the information gleaned from GeneStrat not only helps direct the choice of therapy targeted to a specific alteration but also may affect whole brain radiotherapy recommendations, for example, for patients with KRAS mutations with brain lesions. The liquid biopsy is part of an aggressive treatment approach that may pay dividends throughout a patient’s cancer journey, Bowling noted.

“What we’re finding preliminarily is that by treating these patients very aggressively upfront, they’re getting out of the hospital much faster,” said Bowling, who also is an assistant professor of medicine and director of Interventional Pulmonology and Pulmonary Diagnostic Services at East Carolina University School of Medicine. “We believe what it’s going to show is that they’re not getting secondary pneumonia staying in the hospital, which means their performance status is better and they can tolerate their chemotherapy much better as outpatients.

“It’s relatively new and there needs to be a lot more data gathered but I think the impact of liquid biopsies is going to be huge,” added Bowling. “We’re one center with a very organized multi- disciplinary program that utilizes this technology and we think in a very effective way. That’s going to be the future—getting as much information as you can quickly to direct you to therapies.”

Assay Types and Challenges

In the current landscape, blood-based liquid biopsy tests fall into 1 of 2 categories: PCR-based tests that identify known mutations and next-generation sequencing (NGS) assays that detect a broader range of alterations including gene fusions. PCR testing is faster and less expensive but yields more limited information, Pennell noted.

He said the choice of test type depends upon the clinical situation. For example, if an oncologist has a patient with an EGFR mutation who is progressing, using a PCR-based assay to look for the T790M resistance mutation would be a valid initial step. “On the other hand, if you have a nonsmoker who you’re highly suspicious may have some kind of targetable mutation and you’ve never been able to test them and you cannot get a biopsy safely or they don’t have enough tissue to do it, then doing a more broad NGS-based plasma test in order to assay a wide range of defects including fusions would be a better choice than a PCR-based test,” he said.

Regardless of the technology utilized, there are challenges with liquid biopsy testing. Geoffrey Oxnard, MD, a thoracic oncologist at the Dana-Farber Cancer Institute, said during the FDA-AACR workshop that assays are better able to detect mutations in patients with more advanced disease.⁸

Oxnard discussed the advantages and disadvantages of using plasma-based genotyping in 3 case study examples in patients with NSCLC: newly diagnosed NSCLC of an unknown genotype, acquired resistance to EGFR inhibitor therapy with an unknown mechanism of resistance, and suspected recurrence of NSCLC with surveillance.

Although testing was valuable for the newly diagnosed patient, the sensitivity of the assay was approximately 80%, creating the potential for false negatives. For the patient with acquired resistance, there is a “clear clinical need for plasma genotyping” but there is an unclear “reference standard” comparing these tests with tumor biopsies.

For the patient with prior treatment of NSCLC and suspicious signs of recurrence with a CT scan and bone scan, there are no “clear data” yet on the clinical utility of plasma genomics for making a diagnosis, Oxnard said. However, he said tumor NGS is proving increasingly useful in amplifying the standard pathological evaluation.

Overall, sensitivity and reference standards have emerged as 2 challenges for the plasma-based testing. “As you have more disease, your sensitivity goes up,” said Oxnard. “If you have stage IV lung-only disease, your sensitivity was in the range of 50% or 60% because a lot of these patients just aren’t shedding DNA into their blood for you to detect. If you start getting brain, bone, liver metastasis, if you start getting sick with a high-volume disease, your chance of shed DNA goes up and the chance of a liquid biopsy being successful goes up.”

In the area of reference standards for resistance testing, Oxnard said “a single biopsy does not represent resistance. We need a better sense of what a reference standard should be and maybe that is, in fact, treatment outcome. Many of the available assays have not been optimized because of these complexities.”

Oxnard also said mutation test results can be difficult to interpret.⁸ “It’s hard to know what a positive and negative is with these assays,” he said. “There are low-level results you are seeing; you are not sure if it’s in the noise range or not. Rigorously defining a positive for your assay is really important, and it’s a place where a lot of these assays are struggling.”

Jänne agrees that additional steps must be taken to improve testing. “One of the things that deserves further work is making sure that there is a common set of standard principles by which these tests have been validated and that that’s a transparent process,” he said. “That’s the only way you’re going to be able to understand the performance characteristics of test A versus test B versus test C. It’s both the analytical validation and ultimately you need some clinical validation. I don’t think that [standard] exists at the moment.”

Beyond Lung Cancer

Oxnard believes the standards set in lung cancer for reference testing will set the pace for other tumor types. “What we are establishing in lung cancer will be emulated across oncology,” he said.Although liquid biopsies are most advanced in lung cancer, researchers are actively delving into their potential in other solid tumor types. At the 2016 San Antonio Breast Cancer Symposium, Massimo Cristofanilli, MD, of the Robert H. Lurie Comprehensive Cancer Center, presented a retrospective study of 91 patients with locally and advanced and metastatic breast cancer.⁹

Using Guardant360, an NGS panel that tests circulating tumor DNA (ctDNA) for more than 50 genes, Cristofanilli and colleagues found a statistically significant difference by log rank test in progression-free survival and overall survival associated with the percentage of ctDNA at baseline (<0.5 vs ≥0.5; P = .003 and P = .012, respectively) and number of mutations at baseline (<2 vs ≥2; P = .059 borderline and P = .0015). Targeted therapy was initiated for 16 patients (19%) based on the mutation analysis.

Investigators concluded that the biopsy provides real-time information useful for treatment planning, disease monitoring, and prognostic evaluation.

In prostate cancer, Howard I. Scher, MD, has been leading investigations into the use of liquid biopsies to translate growing knowledge about the biology of the tumor type to better direct therapies. In an interview with OncLive^®, Scher noted the difficulty of conducting tissue biopsies in the malignancy, which most frequently metastasizes to the bone.

“The ability to do consistent molecular profiling is actually quite low. Our experience using directed biopsies, where we know exactly where the lesion is, is only about 50%. It’s an invasive procedure and is costly,” Scher said. “If a patient has 10 individual lesions, they are not all biologically the same. Inadvertently, we may biopsy a lesion, identify a specific gene or pathway, and that pathway is not the key driver of the resistant cell population.”

Scher said there are several tumor biomarkers that can be extracted from a blood draw, including circulating tumor cells (CTCs), DNA, RNA, and exosomes. His team has been focusing on using technology developed by Epic Sciences to study an androgen receptor (AR) splice variant, AR-V7, that drives prostate cancer progression and its association with response to therapies.¹⁰

"We have seen that the frequency of this splice variant is relatively low in the first-line setting," Scher said. "It gets higher with each course. Each time we identify it, the patients treated with AR-signaling directed therapy do not respond. In contrast, there is no relationship between the presence of AR-V7 and response to chemotherapy—in this case, docetaxel or cabazitaxel (Jevtana). We have shown that the survival of patients is improved in those with AR-V7 present who receive a taxane, and it is inferior for those who receive AR-directed therapy.”

Studies such as those that Scher is conducting are part of the future face of liquid biopsies, said Jänne. “This is a rapidly evolving area in terms of clinical use and the technology. It’s giving us insights into cancer that we’ve never previously had and I think that’s the exciting part from the cancer basic biology side and from the therapeutic side. This is an alternative that may help guide clinical care.”

References

1. Top 10 for 2017. Cleveland Clinic. http://innovations.clevelandclinic.org/Summit/Top-10-Medical-Innovations/Top-10-for-2017.aspx. Accessed January 22, 2017.
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3. FDA grants first liquid biopsy approval to the Roche cobas® EGFR Mutation Test v2 [news release]. Pleasanton, CA: Roche; June 1, 2016. https://molecular.roche.com/news/fda-grants-first-liquid-biopsy-approval-to-the-roche-cobas-egfr-mutation-test-v2/. Accessed January 22, 2017.
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7. Bowling M, Mattingley J, Bhadra K, et al. Shortening time from diagnosis to treatment in NSCLC: are blood-based biopsies the answer? J Thorac Oncol. 2016;11(11 suppl):S278-S279. Abstract PS01.16. doi:http://dx.doi.org/10.1016/j.jtho.2016.09.051.
8. FDA-AACR Liquid Biopsies in Oncology Drug and Device Development Workshop; July 19, 2016; Washington, DC. http://www.aacr.org/AdvocacyPolicy/GovernmentA airs/Pages/FDA-AACR-liquid-biopsies-in-oncology-drug-and-device-development.aspx#. WIYkrrYrJPU. Accessed January 22, 2017.
9. Rossi G, Austin LK, Nagy RJ, et al. Circulating tumor DNA (ctDNA): a real-time application of precision medicine to the management of metastatic breast cancer (MBC). Poster presented at: 2016 San Antonio Breast Cancer Symposium; December 6-10, 2016; San Antonio, TX. Abstract PD1-02.
10. Scher HI, Lu D, Schreiber NA, et al. Association of AR-V7 on circulating tumor cells as a treatment-specific biomarker with outcomes and survival in castration-resistant prostate cancer. JAMA Oncol. 2016;2(11):1441-1449. doi:10.1001/jamaoncol.2016.1828.