Blasts-more than meets the eye: Evaluation of post-induction day 21 bone marrow in CBFB rearranged acute leukemia

Induction chemotherapy is often the first therapeutic intervention for acute myeloid leukemia (AML). Evaluation of post induction bone marrow provides critical information for clinical management; in general increased blast countsor increased marrow cellularity is an ominous sign, suggestive of ineffective therapy, and may warrant additional rounds of chemotherapy. However, increased blasts alone are not necessarily predictive of recurrent/persistent disease. Here we report a very unusual observation in a case of AML with a core binding factor beta (CBFB) rearrangement. In this case the day 21 post-induction marrow biopsy showed a high blast count (approximately 20%), however,subsequent fluorescence in-situ hybridization studies were negative for CBFB rearrangement. We compared this finding to post-induction marrows from a series of 6 AML cases with CBFB rearrangements, none of which showed an increased blast count. This case illustrates that increased blast counts, even those comprising 20% of cells, are not de facto evidence of induction failure, and that correlation with ancillary studies such as fluorescence in-situhybridization should be used to distinguish a persistent neoplastic clone, from a brisk marrow recovery

Sequencing-based measurable residual disease testing in acute myeloid leukemia

Next generation sequencing (NGS) methods have allowed for unprecedented genomic characterization of acute myeloid leukemia (AML) over the last several years. Further advances in NGS-based methods including error correction using unique molecular identifiers (UMIs) have more recently enabled the use of NGS-based measurable residual disease (MRD) detection. This review focuses on the use of NGS-based MRD detection in AML, including basic methodologies and clinical applications

Multidimensional biomarker predicts disease control in response to immunotherapy in recurrent or metastatic head and neck squamous-cell carcinoma

PURPOSE: Anti-PD-1 therapy provides clinical benefit in 40-50% of patients with relapsed and/or metastatic head and neck squamous cell carcinoma (RM-HNSCC). Selection of anti- PD-1 therapy is typically based on patient PD-L1 immunohistochemistry (IHC) which has low specificity for predicting disease control. Therefore, there is a critical need for a clinical biomarker that will predict clinical benefit to anti-PD-1 treatment with high specificity. METHODS: Clinical treatment and outcomes data for 103 RM-HNSCC patients were paired with RNA-sequencing data from formalin-fixed patient samples. Using logistic regression methods, we developed a novel biomarker classifier based on expression patterns in the tumor immune microenvironment to predict disease control with monotherapy PD-1 inhibitors (pembrolizumab and nivolumab). The performance of the biomarker was internally validated using out-of-bag methods. RESULTS: The biomarker significantly predicted disease control (65% in predicted non-progressors vs. 17% in predicted progressors, p \u3c 0.001) and was significantly correlated with overall survival (OS; p = 0.004). In addition, the biomarker outperformed PD-L1 IHC across numerous metrics including sensitivity (0.79 vs 0.64, respectively; p = 0.005) and specificity (0.70 vs 0.61, respectively; p = 0.009). CONCLUSION: This novel assay uses tumor immune microenvironment expression data to predict disease control and OS with high sensitivity and specificity in patients with RM-HNSCC treated with anti-PD-1 monotherapy

FGFR2 amplification in colorectal adenocarcinoma

FGFR2 is recurrently amplified in 5% of gastric cancers and 1%–4% of breast cancers; however, this molecular alteration has never been reported in a primary colorectal cancer specimen. Preclinical studies indicate that several FGFR tyrosine-kinase inhibitors (TKIs), such as AZD4547, have in vitro activity against the FGFR2-amplified colorectal cell line, NCI-H716. The efficacy of these inhibitors is currently under investigation in clinical trials for breast and gastric cancer. Thus, better characterizing colorectal tumors for FGFR2 amplification could identify a subset of patients who may benefit from FGFR TKI therapies. Here, we describe a novel FGFR2 amplification identified by clinical next-generation sequencing in a primary colorectal cancer. Further characterization of the tumor by immunohistochemistry showed neuroendocrine differentiation, similar to the reported properties of the NCI-H716 cell line. These findings demonstrate that the spectrum of potentially clinically actionable mutations detected by targeted clinical sequencing panels is not limited to only single-nucleotide polymorphisms and insertions/deletions but also to copy-number alterations.</jats:p

Acute EBV infection masquerading as "In-situ Follicular Lymphoma": a pitfall in the differential diagnosis of this entity

We present the case of a 30 year-old man who was referred for evaluation of diffuse lymphadenopathy. Six weeks prior, he noticed darkening of his urine associated with pale stools, nausea and an eventual 30 lb weight loss within a month. The initial laboratory findings showed elevation of the liver enzymes. A CT scan showed mesenteric and periaortic lymphadenopathy with the largest lymph node measuring 2.8 cm. Other laboratory results were otherwise unremarkable (including a normal LDH) with the exception of positive serum antibodies against Epstein-Barr virus (EBV) associated antigens (IgM+ and IgG+). An excisional biopsy of 4 of the small neck lymph nodes showed a normal architecture with prominent follicles and an intact capsule. But, by immunohistochemistry two of the follicles showed aberrant coexpression of BCL-2, in addition to CD10 and BCL-6. In-situ hybridization for early Epstein-Barr virus mRNA (EBER) and immunohistochemistry for latent membrane protein-1 (LMP-1) stained both scattered positive cells, as well as BCL-2 positive B-cells. Although an original diagnosis of in-situ follicular lymphoma was favored at an outside facility, additional interphase fluorescence in situ hybridization (FISH) studies for t(14;18);(IGH-BCL2) rearrangement (performed on the BCL-2 + follicles microdissected from the tissue block; Abott probe dual colour fusion) and molecular studies (IGH gene rearrangement by PCR, also performed on the microdissected follicles) were negative. Serologic studies (positive EBV antibodies) and immunostains in conjunction with the molecular studies confirmed the reactive nature of the changes. Our case also shows direct immunopathogenic evidence of BCL-2 expression among the EBV-infected cells, which has to our knowledge not been previously documented in vivo. A diagnosis of EBV infection should, therefore, be considered when confronted with BCL-2 expression in germinal centers, particularly in younger individuals, as the diagnosis of FLIS may lead to extensive and invasive haematologic work-ups. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/132365631894006

Performance of Common Analysis Methods for Detecting Low-Frequency Single Nucleotide Variants in Targeted Next-Generation Sequence Data

Next-generation sequencing (NGS) is becoming a common approach for clinical testing of oncology specimens for mutations in cancer genes. Unlike inherited variants, cancer mutations may occur at low frequencies because of contamination from normal cells or tumor heterogeneity and can therefore be challenging to detect using common NGS analysis tools, which are often designed for constitutional genomic studies. We generated high-coverage (>1000×) NGS data from synthetic DNA mixtures with variant allele fractions (VAFs) of 25% to 2.5% to assess the performance of four variant callers, SAMtools, Genome Analysis Toolkit, VarScan2, and SPLINTER, in detecting low-frequency variants. SAMtools had the lowest sensitivity and detected only 49% of variants with VAFs of approximately 25%; whereas the Genome Analysis Toolkit, VarScan2, and SPLINTER detected at least 94% of variants with VAFs of approximately 10%. VarScan2 and SPLINTER achieved sensitivities of 97% and 89%, respectively, for variants with observed VAFs of 1% to 8%, with >98% sensitivity and >99% positive predictive value in coding regions. Coverage analysis demonstrated that >500× coverage was required for optimal performance. The specificity of SPLINTER improved with higher coverage, whereas VarScan2 yielded more false positive results at high coverage levels, although this effect was abrogated by removing low-quality reads before variant identification. Finally, we demonstrate the utility of high-sensitivity variant callers with data from 15 clinical lung cancers

A common founding clone with TP53 and PTEN mutations gives rise to a concurrent germ cell tumor and acute megakaryoblastic leukemia

We report the findings from a patient who presented with a concurrent mediastinal germ cell tumor (GCT) and acute myeloid leukemia (AML). Bone marrow pathology was consistent with a diagnosis of acute megakaryoblastic leukemia (AML M7), and biopsy of an anterior mediastinal mass was consistent with a nonseminomatous GCT. Prior studies have described associations between hematological malignancies, including AML M7 and nonseminomatous GCTs, and it was recently suggested that a common founding clone initiated both cancers. We performed enhanced exome sequencing on the GCT and the AML M7 from our patient to define the clonal relationship between the two cancers. We found that both samples contained somatic mutations in PTEN (C136R missense) and TP53 (R213 frameshift). The mutations in PTEN and TP53 were present at ∼100% variant allele frequency (VAF) in both tumors. In addition, we detected and validated five other shared somatic mutations. The copy-number analysis of the AML exome data revealed an amplification of Chromosome 12p. We also identified a heterozygous germline variant in FANCA (S858R), which is known to be associated with Fanconi anemia but is of uncertain significance here. In summary, our data not only support a common founding clone for these cancers but also suggest that a specific set of distinct genomic alterations (in PTEN and TP53) underlies the rare association between GCT and AML. This association is likely linked to the treatment resistance and extremely poor outcome of these patients. We cannot resolve the clonal evolution of these tumors given limitations of our data

A case of acute myeloid leukemia with promyelocytic features characterized by expression of a novel RARG-CPSF6 fusion

Key Points Novel RARG-CPSF6 fusion in an AML case with promyelocytic features and no evidence of PML-RARA or X-RARA fusion. Gene fusions involving RARG can initiate AML with promyelocytic morphological features.</jats:p

Melanoma in a patient with DNMT3A overgrowth syndrome

Alterations in epigenetic regulators are increasingly recognized as early events in tumorigenesis; thus, patients with acquired or inherited variants in epigenetic regulators may be at increased risk for developing multiple types of cancer. DNMT3A overgrowth syndrome (DOS), caused by germline pathogenic variants in the DNA methyltransferase gen