cHL subtypes - Clinical and prognostic significance

23 Feb 2023

In the 64th American Society of Hematology (ASH) Annual Meeting and Exposition 2022, Dr. Stefan K. Alig, from Stanford University, the United States, shared his group’s study on the distinct molecular subtypes of classic Hodgkin lymphoma (cHL) identified by comprehensive noninvasive profiling.1

Pathologically, cHL is challenging to genotype due to its uniqueness, with only 0.1%-10% of the tumor bulk being Hodgkin Reed-Sternberg (HRS) cells, while the remaining are immune infiltrate, i.e.,  non-malignant cells.1 This means that genetic studies are limited technically, despite that liquid biopsies may play a role in overcoming such challenges.1

It was first found that mutation caused by plasma is highly tumor-specific and plasma fraction exceeds those in the tumor in three-quarters of patients, showing that noninvasive genotype is more sensitive than bulk tissue genotyping.1 Overall, the circulating tumor deoxyribonucleic acid (ctDNA) levels in both cHL and large B-cell lymphoma (LBCL) cells are similar, but the amount of ctDNA shed by HRS cells was approximately 2.5-fold greater than that of LBCL.1 The ctDNA of cHL is also shorter than that of LBCL, a left shift in a density plot, which is not evident in wild-type molecules, thus suggesting a tumor-specific result.1 The expression of deoxyribonuclease 1 like 3 (DNASE1L3) is also much greater in cHL than in LBCL, leading to a shorter fragment size and greater DNASE1L3-specific end motifs.1 This means that DNASE1L3 is likely the cause of greater ctDNA shedding in HRS cells.1

This non-invasive study cohort focused on pre-treatment plasma cells from 366 patients, and 99% of them with an initial diagnosis and a median age of 32.1 All samples (n=366) underwent targeted sequencing using Cancer Personalized Profiling by deep Sequencing (CAPP-Seq), with some selected (n=119) additional plasma whole-exon sequencing (WES).1 Samples (n=293) were then integrated into mutation cause, simple nucleotide variant (SNV), and indel somatic copy number aberration (SCNA) to identify genetic subtypes with Latent Dirichlet Allocation (LDA).1 Two clear subtypes were identified - cluster H1 consisting of mutations and genes involved in nuclear factor kappa light chain enhancer of activated B cells/signal transducer and activator of transcription/phosphatidylinositol-3 kinase (NFkB/STAT/PI3K) pathways; and cluster H2 of genomic instability mediated through Epstein-Barr virus (EBV), transformation-related protein 53 (TP53) and epigenetic modifier inactivation (e.g., KMT2D).1 There was a greater mutational burden in H1 (p<0.001), higher age distribution in younger ages, whereas there was a higher fraction of genome affected by SCNA (p<0.0001) and bimodal age distribution in H2.1 Compared with H1, H2 had a greater amount of EBV-positive status (40% vs. 18%; p<0.0001), mixed cellularity (MC) subtype (23% vs. 9%; p<0.01), ctDNA level (p<0.001), and an inferior progression of survival independent of ctDNA levels (p<0.01).1

In the study cohort, interleukin-4 receptor (IL4R), which was recognized to be repeatedly and significantly mutated in approximately 10% of cHL cases, mediates STAT activation and is involved in IL4 and IL13 signaling through dimerization of either common gamma chain or interleukin-13 receptor (IL13R) alpha chain.1 Mutations in this study on cHL were distinctive from primary mediastinal B-cell lymphoma (PMBL) and clustered at the C-terminals of the protein, with most mutations resulting in a stop codon in proximity to the immunoreceptor tyrosine-based inhibitory motif (ITIM) domain.1 This led to the belief that these mutations will cause a gain-of-function through ITIM domain disruption.1 A separate study found that IL4R mutations were gain-of-functions and upregulate IL13, but not IL4 signaling in activated STAT6 (pSTAT6) positive cells. More importantly, an increased STAT signaling was also identified across the patients.1


Of note, ribonucleic acid (RNA) sequencing between cHL and LBCL indicated that IL13 is one of the greatest expressed genes in cHL and LBCL, and cHL with IL4R mutations had a greater copy number of chromosome 5q31.1, which consists of IL13 locus, meaning that IL13 is a hallmark of cHL and mediates IL4R mutation selection.1 Contrary to non-Hodgkin lymphomas (NHL), cHL IL4R mutants are susceptible to IL4R-targeting drugs presently used in atopic disease treatment and thus, are readily available.1

Similar to the findings in LBCL, the ctDNA level in cHL correlates with the clinical risk factors and disease burden, including advanced stage, bulky disease, B-symptoms, total metabolic tumor volume (TMTV), and EBV positivity.1 Baseline ctDNA levels can also independently determine the prognosis of progression-free survival (PFS) in cHL as evident in both univariable and multivariable multi-analysis.1 ctDNA monitoring with phased variant enrichment and detection sequencing (PhasED-Seq) showed that ctDNA kinetics were also prognostic of PFS.1 More importantly, patients (n=50) overall underwent a rapid decrease in ctDNA after half a cycle of treatment, including one-third of patients being undetectable in cycle 1, day 15.1 In cycle 3, day 1, most patients tested negative for ctDNA.1 In addition, ctDNA was prognostic in patients independent of positron emission tomography scan after cycle 2 (PET2) status.1

In summary, most cHL cases have a higher mutation load in plasma than in bulk tumors, with greater DNASE1L3 likely causing greater ctDNA shedding.1 Clinical and prognostic consequences are significantly different in the 2 cHL genetic subtypes.1 Since truncating IL4R gain-of-function mutations are IL13 cytokine-dependent, they are therapeutically targetable.1 Baseline ctDNA is both correlated with tumor burden measurements and independently prognostic, along with phasED-Seq minimal residual disease (MRD) with an effective prognostic utility independent of interim positron emission tomography-computed tomography (PET-CT).1 Prospectively, studies may focus more on the predictive value of ctDNA for the cHL therapy options.1

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