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Hodgkin Lymphoma Subtypes, Treatment Response Untangled with Liquid Biopsy Assays

NEW YORK – New research suggests that cell-free DNA-based liquid biopsy assays are well suited to characterize classic Hodgkin lymphoma (cHL) tumors, which typically contain low levels of malignant cells that are eclipsed by infiltrating tumor cells.

"Published cohorts in cHL are relatively small, and the profiling methods employed have been limited in both genomic breadth and depth," senior and corresponding author Ash Alizadeh, a researcher at Stanford University, and his colleagues wrote in Nature on Monday. "A major reason for these limitations is the relative paucity of malignant cells within cHL tumors, which typically account for [around 1 percent] of the bulk tumor tissue cellularity."

Using CAPP-seq on matched tumor and blood plasma samples from 24 cHL patients, together with single-cell RNA sequencing on more than 87,500 cells from nine biopsy samples, the team showed that cHL alterations are enriched in ctDNA compared to the tumor itself. The reason, they noted, is DNA cleavage activity by the enzyme encoded by the DNASE1L3 gene.

From there, the investigators turned to exome sequencing or other targeted sequencing strategies to assess ctDNA samples from 366 pretreatment pediatric or adult cHL patients.

In the process, they identified previously unappreciated subtypes of cHL, dubbed H1 and H2, while showing that cHL ctDNA serves as a sensitive method for tracking relapse-related features, particularly the presence of minimal residual disease. They also highlighted potential treatment targets, including gain-of-function mutations affecting the interleukin-4 receptor-coding gene IL4R.

"Collectively, these results support the utility of noninvasive strategies for genotyping and dynamic monitoring of cHL as well as capturing molecularly distinct subtypes with diagnostic, prognostic, and therapeutic potential," the authors reported, noting that the "rich genomic landscape of somatic variants allowed us to apply clustering methods to identify two distinct cHL genetic subtypes (H1 and H2), characterized by a high mutational burden and genomic instability, respectively."

When it came to exploring ctDNA clues to residual disease and relapse, meanwhile, the team used a method called PhasED-seq for ultrasensitive MRD detection in 319 blood plasma samples collected over time from 109 adult cHL patients, combining the approach with a variant phasing algorithm called RePhyNER to track treatment responses and disease remission.

In cHL patients who went on to experience lasting remission, for example, the researchers saw a relatively steep treatment-related dip in ctDNA levels, marked by particularly low ctDNA measurements starting early in treatment. In contrast, relapse risk increased and progression-free survival times diminished in patients with ctDNA that persisted during and after treatment, even when levels of the tumor DNA in their blood plasma dropped significantly.

"Compared with what was previously known about the role of ctDNA MRD in this disease, this study is distinguished by our profiling of a significantly larger and unselected cohort of patients of diverse age and risk profiles from the US and Europe, and by our application of PhasED-seq as an ultrasensitive ctDNA monitoring method," Alizadeh explained in an email.

Together, the results point to the possibility of using baseline ctDNA levels, H1/H2 subtype classification, IL4R genotyping, and ctDNA-based MRD measurements to assess cHL patients' most promising treatment options, treatment responses, and management strategies, he added.

"For example, ctDNA MRD levels could help guide future risk-adapted trials to reduce treatment duration of systemic therapy, or trials to integrate novel therapies in the frontline, including PD-L1 antibodies such as pembrolizumab, nivolumab, tislelizumab, or other therapies," Alizadeh suggested. "Separately, ctDNA MRD levels could help inform trials testing selective omission of high dose chemotherapy and autologous stem cell transplantation after successful remissions are induced for relapsed disease."