First-hit SETBP1 mutations cause a myeloproliferative disorder with bone marrow fibrosis.

ABSTRACT: SETBP1 mutations are found in various clonal myeloid disorders. However, it is unclear whether they can initiate leukemia, because SETBP1 mutations typically appear as later events during oncogenesis. To answer this question, we generated a mouse model expressing mutated SETBP1 in hematopoietic tissue: this model showed profound alterations in the differentiation program of hematopoietic progenitors and developed a myeloid neoplasm with megakaryocytic dysplasia, splenomegaly, and bone marrow fibrosis, prompting us to investigate SETBP1 mutations in a cohort of 36 triple-negative primary myelofibrosis (TN-PMF) cases. We identified 2 distinct subgroups, one carrying SETBP1 mutations and the other completely devoid of somatic variants. Clinically, a striking difference in disease aggressiveness was noted, with patients with SETBP1 mutation showing a much worse clinical course. In contrast to myelodysplastic/myeloproliferative neoplasms, in which SETBP1 mutations are mostly found as a late clonal event, single-cell clonal hierarchy reconstruction in 3 patients with TN-PMF from our cohort revealed SETBP1 to be a very early event, suggesting that the phenotype of the different SETBP1+ disorders may be shaped by the opposite hierarchy of the same clonal SETBP1 variants.

De novo UBE2A mutations are recurrently acquired during chronic myeloid leukemia progression and interfere with myeloid differentiation pathways.

Despite the advent of tyrosine kinase inhibitors, a proportion of chronic myeloid leukemia patients in chronic phase fail to respond to imatinib or to second-generation inhibitors and progress to blast crisis. Until now, improvements in the understanding of the molecular mechanisms responsible for chronic myeloid leukemia transformation from chronic phase to the aggressive blast crisis remain limited. Here we present a large parallel sequencing analysis of 10 blast crisis samples and of the corresponding autologous chronic phase controls that reveals, for the first time, recurrent mutations affecting the ubiquitin-conjugating enzyme E2A gene (UBE2A, formerly RAD6A). Additional analyses on a cohort of 24 blast crisis, 41 chronic phase as well as 40 acute myeloid leukemia and 38 atypical chronic myeloid leukemia patients at onset confirmed that UBE2A mutations are specifically acquired during chronic myeloid leukemia progression, with a frequency of 16.7% in advanced phases. In vitro studies show that the mutations here described cause a decrease in UBE2A activity, leading to an impairment of myeloid differentiation in chronic myeloid leukemia cells.

Evolutionary signatures of human cancers revealed via genomic analysis of over 35,000 patients.

Recurring sequences of genomic alterations occurring across patients can highlight repeated evolutionary processes with significant implications for predicting cancer progression. Leveraging the ever-increasing availability of cancer omics data, here we unveil cancer's evolutionary signatures tied to distinct disease outcomes, representing "favored trajectories" of acquisition of driver mutations detected in patients with similar prognosis. We present a framework named ASCETIC (Agony-baSed Cancer EvoluTion InferenCe) to extract such signatures from sequencing experiments generated by different technologies such as bulk and single-cell sequencing data. We apply ASCETIC to (i) single-cell data from 146 myeloid malignancy patients and bulk sequencing from 366 acute myeloid leukemia patients, (ii) multi-region sequencing from 100 early-stage lung cancer patients, (iii) exome/genome data from 10,000+ Pan-Cancer Atlas samples, and (iv) targeted sequencing from 25,000+ MSK-MET metastatic patients, revealing subtype-specific single-nucleotide variant signatures associated with distinct prognostic clusters. Validations on several datasets underscore the robustness and generalizability of the extracted signatures.

Characterization of SARS-CoV-2 Mutational Signatures from 1.5+ Million Raw Sequencing Samples.

We present a large-scale analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) substitutions, considering 1,585,456 high-quality raw sequencing samples, aimed at investigating the existence and quantifying the effect of mutational processes causing mutations in SARS-CoV-2 genomes when interacting with the human host. As a result, we confirmed the presence of three well-differentiated mutational processes likely ruled by reactive oxygen species (ROS), apolipoprotein B editing complex (APOBEC), and adenosine deaminase acting on RNA (ADAR). We then evaluated the activity of these mutational processes in different continental groups, showing that some samples from Africa present a significantly higher number of substitutions, most likely due to higher APOBEC activity. We finally analyzed the activity of mutational processes across different SARS-CoV-2 variants, and we found a significantly lower number of mutations attributable to APOBEC activity in samples assigned to the Omicron variant.

Validation of a new NGS-based myeloid panel in acute myeloid leukemia: A single-center experience.

Acute myeloid leukemia (AML) identifies a heterogeneous group of clonal disorders, both clinically and genetically. A large number of mutations have been described in AML, although only a few are currently employed in clinical practice. Next generation sequencing (NGS) allows for better understanding of the complex genetic background in AML and may direct individualized therapies. In this study, we aim to identify molecular aberrations that are not routinely investigated in AML using an NGS-based panel encompassing 101 genes and to evaluate how their oncogenic potential correlates with survival. Forty consecutive patients with newly diagnosed AML were enrolled between January 2018 and April 2020. We performed targeted NGS and detected 96 mutations in 36 patients (90%), while 14 fusion genes were detected in 13 patients (32%). Each mutation was weighed using OncoScore, a text-mining tool ranking genes according to their oncogenic potential. An OncoScore ≥ 100 was associated with shorter PFS among our patients (p = 0.05). In 11 patients with no available MRD markers at diagnosis, we were able to perform NGS-based MRD monitoring using targeted deep sequencing. Overall, our study shows that NGS is a powerful tool in AML and should be employed both in routine diagnostic workup and follow up.

Synergistic Drug Combinations Prevent Resistance in ALK+ Anaplastic Large Cell Lymphoma.

Anaplastic lymphoma kinase-positive (ALK+) anaplastic large-cell lymphoma (ALCL) is a subtype of non-Hodgkin lymphoma characterized by expression of the oncogenic NPM/ALK fusion protein. When resistant or relapsed to front-line chemotherapy, ALK+ ALCL prognosis is very poor. In these patients, the ALK inhibitor crizotinib achieves high response rates, however 30-40% of them develop further resistance to crizotinib monotherapy, indicating that new therapeutic approaches are needed in this population. We here investigated the efficacy of upfront rational drug combinations to prevent the rise of resistant ALCL, in vitro and in vivo. Different combinations of crizotinib with CHOP chemotherapy, decitabine and trametinib, or with second-generation ALK inhibitors, were investigated. We found that in most cases combined treatments completely suppressed the emergence of resistant cells and were more effective than single drugs in the long-term control of lymphoma cells expansion, by inducing deeper inhibition of oncogenic signaling and higher rates of apoptosis. Combinations showed strong synergism in different ALK-dependent cell lines and better tumor growth inhibition in mice. We propose that drug combinations that include an ALK inhibitor should be considered for first-line treatments in ALK+ ALCL.

An Imatinib-non-responsive patient with an ABL Leu387Trp mutation achieves cytogenetic and molecular response under bosutinib: Case report and biological characterization.

Leu387Trp mutation, aroused in an imatinib-non-responsive CML patient, was selected by imatinib treatment along with other unknown factors responsible for resistance, and then it was overcome by bosutinib. These results will be useful for treating patients with this rare mutation and will advise against automatically considering a new mutation as the cause of TKI resistance.

ETNK1 mutations induce a mutator phenotype that can be reverted with phosphoethanolamine.

Recurrent somatic mutations in ETNK1 (Ethanolamine-Kinase-1) were identified in several myeloid malignancies and are responsible for a reduced enzymatic activity. Here, we demonstrate in primary leukemic cells and in cell lines that mutated ETNK1 causes a significant increase in mitochondrial activity, ROS production, and Histone H2AX phosphorylation, ultimately driving the increased accumulation of new mutations. We also show that phosphoethanolamine, the metabolic product of ETNK1, negatively controls mitochondrial activity through a direct competition with succinate at mitochondrial complex II. Hence, reduced intracellular phosphoethanolamine causes mitochondria hyperactivation, ROS production, and DNA damage. Treatment with phosphoethanolamine is able to counteract complex II hyperactivation and to restore a normal phenotype.

Long-term Results After Hyaluronan-based MACT for the Treatment of Cartilage Lesions of the Patellofemoral Joint.

BACKGROUND: Cartilage lesions of the patellofemoral joint are a challenging condition. Hyaluronan-based matrix-assisted autologous chondrocyte transplantation (MACT) has been shown to offer a significant improvement in the short term but has a tendency to worsen at midterm follow-up. HYPOTHESIS: Patients treated with MACT for lesions of the articular surface of the patellofemoral joint will present further clinical worsening at long-term follow-up. STUDY DESIGN: Case series; Level of evidence, 4. METHODS: Thirty-two patients with full-thickness chondral lesions in the patellofemoral joint were treated with hyaluronan-based MACT and were prospectively evaluated preoperatively and at 2-, 5-, and 10-year follow-up. The mean defect size was 4.45 cm(2). There were 20 lesions located on the patella and 8 on the trochlea, and 4 patients had multiple lesions: 3 with patellar and trochlear lesions and 1 with patellar and lateral femoral condyle lesions. Results were evaluated using International Knee Documentation Committee (IKDC) subjective scores, EuroQol visual analog scale (EQ VAS) scores, and Tegner scores. Surgical and clinical failures were documented. RESULTS: All scores showed a statistically significant improvement at 2-, 5-, and 10-year follow-up with respect to the preoperative level. No worsening was observed at the last follow-up, and results were stable up to 10 years. The improvement in mean (±SD) outcome scores from preoperatively to 2-, 5-, and 10-year follow-up was as follows: IKDC, from 46.0 ± 19.8 to 77.1 ± 17.4, 72.0 ± 20.4, and 78.6 ± 16.4, respectively; Tegner, from 2.5 ± 1.4 to 4.7 ± 1.8, 4.7 ± 1.6, and 4.4 ± 1.5, respectively; and EQ VAS, from 56.9 ± 18.4 to 81.7 ± 13.2, 79.2 ± 17.9, and 78.9 ± 1.7, respectively. Four patients did not achieve significant clinical improvement, and 1 of these patients required further surgical treatment. All failures were female patients with patellar defects, and 3 of them had degenerative lesions and underwent a previous or combined realignment procedure. CONCLUSION: The clinical results of hyaluronan-based MACT treatment of chondral lesions of the patellofemoral joint do not worsen over time but remain stable and show a low rate of failure at long-term follow-up.