LIQUID BIOPSIES FOR LYMPHOMAS

The sequencing of circulating tumor DNA (ctDNA) in blood samples – so-called liquid biopsies ― has emerged as a promising and noninvasive approach to detect the clonal evolution of lymphoma. Previously, the only way to follow the disease as it evolved was by tissue biopsy.

The new findings come from a study conducted at Stanford University, where researchers carried out cancer personalized profiling by deep sequencing (CAPP-Seq) on lymphoma tissue samples and ctDNA from blood samples. They showed that molecular profiling seen in blood samples matched that seen in lymphoma tissue.

In addition, they were able to identify specific genetic variations associated with adverse outcomes, likelihood of response to current therapy, and emergence of resistance, and they could predict the likelihood of relapse before it occurred.

"We can now measure several unique features of B-cell lymphomas in the blood that were once only possible to do with tissue samples," Ash A. Alizadeh, MD, PhD, assistant professor of medicine at Stanford University, told Medscape Medical News.

"The blood biopsy could also measure risk-associated features of the disease, differentiate between subtypes, and monitor changes from low-grade to high-grade lymphoma," he added.

"Watching how the lymphoma changes over time and seeing this much earlier in the blood provides us a window of opportunity to intervene sooner before disease burden sets in," Maximilian Diehn, MD, PhD, assistant professor of radiation oncology at Stanford University, told Medscape Medical News.

Dr Alizadeh and Dr Diehn share senior authorship of the article, which was published online November 9 in Science Translational Medicine.

"This is a landmark paper and represents a significant technological advance in managing patients with lymphoma," said lymphoma expert Mark Roschewski, MD, from the Center for Cancer Research at the National Cancer Institute. He was not involved in the study and was approached for comment by Medscape Medical News.

Diffuse large B-cell Lymphoma (DLBCL) evolves differently in different patients. Dr Roschewski explained that the heterogeneity of DLBCL is at the molecular level and cannot be seen under the microscope. Once it begins, treatment adds selection pressure, and "DLBCL has the ability to further evolve and change its molecular biology," he added.

"Now for the first time in lymphoma, we have seen a broad panel noninvasively interrogate genes and other molecular aberrations that are specific to DLBCL and other lymphoma subtypes. The ability to pick up individual mutations that confer drug resistance could prove to be very valuable in the clinical management of these patients," Dr Roschewski commented.

Unmasking the Landscape of DLBCL

For their study, which included 92 patients, the Stanford team profiled DLBCL at "various disease milestones" to evaluate whether CAPP-Seq could identify DLBCL genotypes.

By profiling 76 diagnostic DLBCL tumor biopsy specimens and 144 longitudinal blood samples (45 obtained before treatment), the researchers were able to show somatic changes in 100% of the tumors (134 variants, which included the V(D)J rearrangement) and 89% of translocations typically identified through fluorescence in situ hybridization (FISH). Pretreatment blood samples could detect ctDNA in 100% of patients with 99.8% specificity.

Several specific driver mutations and FISH-confirmed translocations previously identified from tissue samples could then be identified from ctDNA extracted from blood samples.

The researchers also showed that these mutations seen in tumor biopsy specimens and pretreatment blood samples could also be captured over time in serial blood samples of patients.

"These data suggest that, in most DLBCL patients, ctDNA is a robust surrogate for direct assessment of primary tumor genotypes," the researchers note.

CAPP-Seq genotyping of blood samples applied to patients whose disease progressed on treatment with ibrutinib (Imbruvica, Janssen Biotech) — a Bruton kinase inhibitor under investigation in the treatment of DLBCL — showed that the technique could capture mutations in BTK previously seen only in ibrutinib-refractory chronic lymphocytic leukemia and mantle cell lymphoma. The clonal dynamics of the tumors was different in different patients and suggested that the technique used on blood samples could follow clonal evolution of the tumors.

CAPP-Seq was also able to detect clinical risk equivalents. ctDNA concentrations in pretreatment blood samples were higher in patients with higher blood lactate dehydrogenase levels or higher metabolic tumor volume and correlated with Ann Arbor stage — all risk equivalents in DLBCL.

"Pretreatment ctDNA in DLBCL can complement traditional clinical indices and serve as an independent prognostic biomarker," the authors write.

The researchers showed that CAPP-Seq could detect occult disease long before it could be seen in the clinic using current radiographic techniques. They molecularly profiled blood samples taken from patients who had shown a complete radiographic response, from those whose disease recurred, and from those who ultimately experienced disease progression. Although ctDNA was not able to detect mutations in patients who had a complete response, the molecular profile at relapse compared well with the currently used assay to detect V(D)J rearrangements.

Indeed, patients with detectable ctDNA showed inferior progression-free survival. What was most significant was the observation that in those patients who eventually relapsed, CAPP-Seq was able to capture minimal residual disease with a mean lead time of longer than 2 months. The mean time between the first positive ctDNA time point and clinical relapse was 188 days (>6 months).

"CAPP-Seq can identify patients who will relapse, often months before recurrence can be clinically identified." Dr Diehn told Medscape Medical News