Pathology Highlights From ASH 2020

The 2020 American Society of Hematology (ASH) annual meeting was held virtually from December 5-8, 2020, with pre- and post-meeting sessions beginning December 2 and ending December 10. The meeting drew over 30,000 registrants from all over the world and had multiple simultaneous broadcasts throughout the entire nine (9) days. Therefore, only a select few pathology-related abstracts or scientific sessions are highlighted below, grouped thematically. For those interested in more details, the abstract numbers are provided if applicable and include a summary of key points. Note, all abbreviations are listed at the bottom for non-hematology/hematopathology readers.

Overall, there were several pathology-related themes, not all of which are covered in this review, including COVID-19, CAR-T and immunotherapy, ctDNA, MRD, clonal hematopoiesis, single cell analyses, and AI.

Ari Melnick (Weill Cornell Medicine) and Courtney DiNardo (MD Anderson Cancer Center) tag-teamed the Beutler Lecture to provide a review of the bench-to bedside story of the discovery of IDH1/2 mutations in AML and their translation to FDA-approval of targeted inhibitors used as both as single agents and in combination, in a record 10 years.

Dr. Andrew Roberts (Walter and Aliza Hall Institute of Medical Research) highlighted a 20 year-long history of the discovery of BCL2, the development of small molecule BH3-mimetics, and the astounding success of Venetoclax in combination with other agents. Venetoclax is especially efficacious in t(11;14) myeloma, NPM1/IDH mutated AML, and primary resistance in complex karyotype AML and del(9p) in MCL. His talk also discussed the development of resistance mechanisms, including acquisition of TP53/del17p in CLL, kinase pathway mutations in AML, the effect of in vivo stroma, mutations of BCL2 (c.302G>T p.G101V) with 180x decreased binding to Venetoclax, overexpression of BCLXL, gains of the MCL1 locus, and alternate use of MCL1 and A1/BLF1 and other pro-survival members of the BCL2 family.

This discussion, led ty ASH president Dr. Stephanie Lee, marked the impact of SARS-CoV-2 on not only the past year, but on the research presented at the 2020 ASH annual meeting.

Pathogenic variants in some complement components (C3, ICD46, CFH, DGKE, ADAMTS13, etc.) are associated with severe disease in COVID-19 patients.

Use of convalescent plasma in COVID-19 therapy was demonstrated in Phase 1 to improve outcomes in COVID-19 (26% of events in the plasma group versus 46% in the control group).

The use of CAR-T therapy in DLBCL was found to be dependent upon functional CD58, and mutations in CD58 were associated with a failure to achieve a CR. KO of CD58 abrogates any curative potential of CAR-T therapy. This led to the exciting finding that providing CAR-T in trans with functional CD58 overcame any deleterious effects of a CD58 mutation.

Abstract #530 Key Points

ClonoSeq on CSF (both cellular fraction and cfDNA) was 100% sensitive in identifying the clonotype in patient with known CNS lymphoma. In new diagnosis patients with no known CNS involvement, no patients with negative baseline NGS-MRD assay in CSF had CNS relapse, whereas 2 of 10 with a positive assay had a CNS relapse (12-month incidence, 29%) despite both pts having negative CNS imaging, CSF cytology, flow cytometry, and IGH-PCR at diagnosis (100% PPV, 100% NPV).

Pre-SCT ctDNA was a better predictor of relapse post autoSCT than pre-SCT PET scan in DLBCL. 90% of patients with detectable ctDNA pre-SCT relapsed. In relapsed/refractory DLBCL, ctDNA positivity was an independent poor prognostic marker of response, PFS, OS in a study of BR+/- polatuzumab.

Dr. Davide Rossi (Instituto Oncologico della Svizzera Italiana) discussed the accepted use of MRD as a surrogate for progression free survival (PFS) in clinical trials (for instance in CLL), but that MRD is not yet accepted currently as a primary endpoint. In many disease types (CLL, DLBCL, cHL) there is clear evidence that MRD can be prognostically impactful. Dr. Ash Alizadeh (Stanford University) discussed his CAPP-Seq and Phase-Seq technologies and their clinical utility in monitoring ctDNA in lymphomas.

Using CRISPR in mice, the researchers targeted common LOF genes in CLL (ATM, MGA, SAMHD1, BIRC3, CHD2, and TP53) and found that TP53, MGA (regulator of MYC), and CHD2 mutations were most highly associated with Richter’s transformation.

The CLL-1100 project conducted comprehensive genomics in CLL cases and found mutations and copy number alterations associated with IGH somatically hypermutated CLL (M-CLL: CARD11, ITPKB, MD88, DIS3) and IGH unmutated CLL (U-CLL: KRAS, MGA, BRCC3, BCR, SAMHD1, SP140). Expression clusters could further subdivide prognosis within M-CLL and U-CLL. Finally, they studied the order of mutation emergence, with +12 early, ATM/SF3B1 intermediate, and FBXW7 late.

BIRC3 can be altered through 11q deletion of somatic mutations in 3-7% of CLL. Both biallelic and monoallelic hits in BIRC3 were associated with poor prognosis but sensitivity to venetoclax.

Somatic variants should be recognized as presumptive evidence of MDS. 36% of cytopenic patients had a pathogenic variant compared to 85% of MDS patients. DTA mutations were more common as a single genetic alteration in CCUS than in MDS. Mutations in “high risk CCUS” patterns or in SF3B1 (recommended to be presumptive dx of MDS already) allowed the “reclassification” of 20% of cytopenic patients (55% of CCUS) to presumptive MDS.

These two studies both stressed the lineage history of variants and found that some variants, specifically JAK2 and DNMT3A described, occur very early in life, even in utero, with slow expansion of the clones over the course of decades. In addition, clonal evolution with the acquisition of additional variants occurs over time, with additional acquisitions by decades. Prognostically, the rate of growth correlates inversely with the latency of diagnosis. Hence, it is less that, "aging promotes clonal hematopoiesis, but rather that it takes an age for the clones to sufficiently expand."

ETNK1, which catalyzes the conversion of ethanolamine to phophoethanolamine, is recurrently mutated in aCML, CMML, and SM with eosinophilia. ETNK1 mutations result in a mutator phenotype through increased reactive oxygen species that lead to increased 8-ox-guanine (and corresponding mutagenesis) as well as increased dsDNA breaks. This mutator phenotype can be reversed by treatment with phophoethanolamine.

The ASH Somatic Working group has created an online reference for variant tiering, initially posting 202 pathogenic/likely pathogenic (P/LP) variants in 42 genes for which there was high concordance of the tier assignment from all submitting laboratories. Ongoing efforts will continue to reconcile differences between participating laboratories and to expand to more P/LP variants as well as VUSes. Of note, this is a separate effort from the ClinVar Somatic Myeloid group (which presented at a scientific session) that has provided in-depth analysis of germline variants in RUNX1 and is currently doing the same for GATA2.

TET2 and IDH, but not SRSF2, mutations are enriched in patients with myelodysplastic syndrome/chronic myelomonocytic leukemia with systemic inflammatory and autoimmune disease (SIAD), a condition which is found in 20% of MDS and CMML cases. These mutations were associated with T cell imbalance in favor of effector memory T cells.

In cytopenic patients, 7 genes (in order of impact: TP53, SF3B1, U2AF1, ASXL1, TET2, STAG2, and SRSF2) were most informative in a discriminating diagnosis of MDS from Other.

Dr. Omar Abdel Wahab (Memorial Sloan Kettering Cancer Center) discussed the unique behavior of ZRSR2 of the recurrently mutated splicing components. Unlike the other components that act on the 99.7% of canonical splice-site containing introns, ZRSR2 works on the 0.3% that are U12 introns of the minor spliceosome and may have its transforming activity as a result of a retained minor intron of LZTR1 which results in increased RAS activity.

Dr. Vijay Sankaran (Boston Children’s Hospital) demonstrated the power of mtDNA sequencing since mutations rates are 10-100x higher than in the genome, with only 16.6 kb (37 genes) to cover, allowing for accurate clonal tree reconstruction across all cell lineages at only 194x coverage per cell. Dr. Margaret Goodall (Baylor College of Medicine) examine the biological ramification of different mutations in DNMT3A. Loss of DNMT3A function results in increased stem cell self-renewal and decreased differentiation, but it is a gradation of DNMT3A fitness with many stable mutations demonstrating minor deficits in DNMT3A activity, unstable mutations resulting in intermediate deficits due to heterozygous loss of function, and p.R882 mutations resulting in a dominant negative function and only 20% residual DNMT3A activity. These results explain the prevalence of p.R882 mutations in 50-60% of DNMT3A mutations of myeloid neoplasms but only 15-20% of those in clonal hematopoiesis. Dr. Konstanze Dohner (University Hospital of Ulm) discussed the use of NGS MRD as an independent prognostic indicator in AML with a clinical applicability of >90% of cases (similar to flow cytometry) but with superior analytic sensitivity (10-3 to 10-6 versus 10-3-10-4). Specific discussion of the DTA genes, NPM1, and FLT3-ITD were discussed.

Use of CTX-001 (CRISPR silencing of BCL11A), resulted in dramatic sustained elevations of HgF in thalassemic and Sickle Cell disease patients with resultant astounding results in transfusion independence and lack of venous occlusive disease crises, respectively.

New first in class drug asciminib that specifically targets ABL1 myristoyl pocket (STAMP, not the ATP binding pocket) showed improved efficacy and better safety profile than bosutinib in CML-CP.

In patients with no established cause of the bleeding, this study found that blood group O is associated with increased bleeding and decreased vWF.

WES in teenage females identified somatic variants in a number of genes (such as CD36, vWF, GP6, ANK1, ADAMTS13, CPO, …) associated with low vWF and heavy menstrual bleeding as well as three polymorphisms demonstrating linkage disequilibrium (such as KIND2). This study suggested that these may not only be pathogenic in bleeding but directly in anemia.

A novel subgroup of acute leukemias of ambiguous lineage was identified by RNAseq, associated with monoallelic expression of BCL11B due to structural variants with a high association with FLT3-ITDs. One-third of the translocations involved ARID1B, with other translocation partners including ETV6, RUNX1, ZEB2, SATB1, etc. The breakpoints in BCL11B occur at the BCL11B Enhancer Tandem Amplification (BETA) that results in ectopic high levels of BCL11B expression.

aCML: atypical chronic myeloid leukemia; AML: acute myeloid leukemia; CCUS: clonal cytopenias of uncertain significance; cfDNA: cell free DNA; CH: clonal hematopoiesis; cHL: classic Hodgkin lymphoma; CLL: chronic lymphocytic leukemia; CML-CP: chronic myeloid leukemia- chronic phase; CMML: chronic myelomonocytic leukemia; CNS: central nervous system; CSF: cerebrospinal fluid; ctDNA: circulating tumor DNA; DLBCL: diffuse large B cell lymphoma; DTA: used to denote three genes (DNMT3A, TET2, ASXL1); MCL: mantle cell lymphoma; MDS: myelodysplastic syndromes; mtDNA: mitochondrial DNA; NGS: next generation sequencing; SM: systemic mastocytosis; vWF: von Willebrand’s factor

Annette S. Kim, MD, PhD, FCAP is the Henry Clay Bryant Professor and Division Head of Diagnostic Genetics and Genomics at the University of Michigan. Dr. Kim’s research program has focused on the study of hematolymphoid malignancies, including miRNAs in myelodysplastic syndromes, myeloid and lymphoid mutational patterns, and test utilization management. She has served as a member of Molecular Oncology Committee and is currently Vice-Chair of the Personized Healthcare Committee for the College of American Pathologists. She is the Vice-Chair of the ASH Precision Medicine committee. In 2019, Dr. Kim was awarded the CAP Public Service Award.