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Background: To date, fewer than 20 patients have been identified as having germline biallelic mutations in the coronin-1A gene (CORO1A) and its protein with clinical features of combined immunodeficiency characterized by T-cell lymphopenia ranging from the severe phenotype to the mild phenotype, recurrent infections, and lymphoproliferative disorders. However, the effects of CORO1A protein disruption on actin-dependent functions in primary cells have not been fully delineated. Objective: We sought to characterize the underlying defects of actin-dependent cellular functions in a female patient with combined immunodeficiency caused by a novel missense variant in the CORO1A gene in combination with a de novo heterozygous microdeletion of chromosome 16p11.2 and also to provide evidence of the pathogenicity of this gene mutation. Methods: To identify the genetic defect, next-generation sequencing followed by Sanger confirmation and array comparative genomic hybridization were performed. Western blot and quantitative PCR tests were used to assess the effects on the protein. Flow cytometry and live microscopy were performed to investigate cellular motility and immune cell counts and function. Results: We demonstrated that the CORO1A hemizygous variant c.19C>T, p. A7C induces significant decreases in cellular levels of the CORO1A protein while leaving mRNA concentrations unaffected. The observed mutation resulted in impaired natural killer cell cytotoxicity and platelet calcium signaling. In addition, primary granulocytes and mesenchymal cells showed significant defects in motility. Conclusion: Collectively, we added new data about the CORO1A gene as a key player in actin cytoskeleton dynamics and cell signaling. Our findings expand the clinical spectrum regarding CORO1A protein deficiency and confirm the importance of a personalized therapeutic approach for each patient.

Abstract Background Accurate variant detection in the coding regions of the human genome is a key requirement for molecular diagnostics of Mendelian disorders. Efficiency of variant discovery from next-generation sequencing (NGS) data depends on multiple factors, including reproducible coverage biases of NGS methods and the performance of read alignment and variant calling software. Although variant caller benchmarks are published constantly, no previous publications have leveraged the full extent of available gold standard whole-genome (WGS) and whole-exome (WES) sequencing datasets. Results In this work, we systematically evaluated the performance of 4 popular short read aligners (Bowtie2, BWA, Isaac, and Novoalign) and 9 novel and well-established variant calling and filtering methods (Clair3, DeepVariant, Octopus, GATK, FreeBayes, and Strelka2) using a set of 14 “gold standard” WES and WGS datasets available from Genome In A Bottle (GIAB) consortium. Additionally, we have indirectly evaluated each pipeline’s performance using a set of 6 non-GIAB samples of African and Russian ethnicity. In our benchmark, Bowtie2 performed significantly worse than other aligners, suggesting it should not be used for medical variant calling. When other aligners were considered, the accuracy of variant discovery mostly depended on the variant caller and not the read aligner. Among the tested variant callers, DeepVariant consistently showed the best performance and the highest robustness. Other actively developed tools, such as Clair3, Octopus, and Strelka2, also performed well, although their efficiency had greater dependence on the quality and type of the input data. We have also compared the consistency of variant calls in GIAB and non-GIAB samples. With few important caveats, best-performing tools have shown little evidence of overfitting. Conclusions The results show surprisingly large differences in the performance of cutting-edge tools even in high confidence regions of the coding genome. This highlights the importance of regular benchmarking of quickly evolving tools and pipelines. We also discuss the need for a more diverse set of gold standard genomes that would include samples of African, Hispanic, or mixed ancestry. Additionally, there is also a need for better variant caller assessment in the repetitive regions of the coding genome.

Simultaneous multilineage hematologic malignancies are uncommon and associated with poorer prognosis than single-lineage leukemia or lymphoma. Here, we describe a concomitant malignant neoplasm in a 4-year-old boy. The child presented with massive lymphoproliferative syndrome, nasal breathing difficulties, and snoring. Morphological, immunocytochemical, and flow cytometry diagnostics showed coexistence of acute myeloid leukemia (AML) and peripheral T-cell lymphoma (PTCL). Molecular examination revealed a rare t(9;9)(q34;q34)/SET::NUP214 translocation as well as common TCR clonal rearrangements in both the bone marrow and lymph nodes. The disease showed primary refractoriness to both lymphoid and myeloid high-dose chemotherapy as well as combined targeted therapy (trametinib + ruxolitinib). Hence, HSCT was performed, and the patient has since been in complete remission for over a year. This observation highlights the importance of molecular techniques for determining the united nature of complex SET::NUP214-positive malignant neoplasms arising from precursor cells with high lineage plasticity.

IntroductionIn this report we firstly describe undifferentiated embryonal sarcoma of the liver (UESL) in a patient with neurofibromatosis type 1 (NF1), fatally complicated by synchronous malignant peripheral nerve sheath tumor (MPNST) with a highly aggressive metastatic course. The case also represents our first experience of chemoperfusion involving the transcatheter arterial chemoembolization (TACE) in a pediatric patient, applied as a treatment for UESL.Case presentationA 13-year-old girl was diagnosed with NF1 and presented with a liver tumor identified as UESL by histological assessment. The tumor was refractive to the conventional first-line chemotherapy. The patient received hepatic chemoperfusion with TACE, which was efficacious; however, the overall curative outcome was unsatisfactory due to synchronous unresectable retroperitoneal MPNST with mesenteric metastases and ultimate progression of the UESL.ConclusionThis is the first reported case of UESL in a patient with NF1. The results demonstrate the efficacy of hepatic chemoperfusion with TACE in pediatric UESL.

We report incidence and deep molecular characteristics of lineage switch in 182 pediatric patients affected by B-cell precursor acute lymphoblastic leukemia (BCP-ALL), who were treated with blinatumomab. We documented six cases of lineage switch that occurred after or during blinatumomab exposure. Therefore, lineage conversion was found in 17.4% of all resistance cases (4/27) and 3.2% of relapses (2/63). Half of patients switched completely from BCP-ALL to CD19-negative acute myeloid leukemia, others retained CD19-positive B-blasts and acquired an additional CD19-negative blast population: myeloid or unclassifiable. Five patients had KMT2A gene rearrangements; one had TCF3::ZNF384 translocation. The presented cases showed consistency of gene rearrangements and fusion transcripts across initially diagnosed leukemia and lineage switch. In two of six patients, the clonal architecture assessed by IG/TR gene rearrangements was stable, while in others, loss of clones or gain of new clones was noted. KMT2A-r patients demonstrated very few additional mutations, while in the TCF3::ZNF384 case, lineage switch was accompanied by a large set of additional mutations. The immunophenotype of an existing leukemia sometimes changes via different mechanisms and with different additional molecular changes. Careful investigation of all BM compartments together with all molecular –minimal residual disease studies can lead to reliable identification of lineage switch.