Molecular Genetic Abnormalities Research Articles

Endocrine features of Prader-Willi syndrome: a narrative review focusing on genotype-phenotype correlation.

Prader-Willi syndrome (PWS) is a complex genetic disorder caused by three different types of molecular genetic abnormalities. The most common defect is a deletion on the paternal 15q11-q13 chromosome, which is seen in about 60% of individuals. The next most common abnormality is maternal disomy 15, found in around 35% of cases, and a defect in the imprinting center that controls the activity of certain genes on chromosome 15, seen in 1-3% of cases. Individuals with PWS typically experience issues with the hypothalamic-pituitary axis, leading to excessive hunger (hyperphagia), severe obesity, various endocrine disorders, and intellectual disability. Differences in physical and behavioral characteristics between patients with PWS due to deletion versus those with maternal disomy are discussed in literature. Patients with maternal disomy tend to have more frequent neurodevelopmental problems, such as autistic traits and behavioral issues, and generally have higher IQ levels compared to those with deletion of the critical PWS region. This has led us to review the pertinent literature to investigate the possibility of establishing connections between the genetic abnormalities and the endocrine disorders experienced by PWS patients, in order to develop more targeted diagnostic and treatment protocols. In this review, we will review the current state of clinical studies focusing on endocrine disorders in individuals with PWS patients, with a specific focus on the various genetic causes. We will look at topics such as neonatal anthropometry, thyroid issues, adrenal problems, hypogonadism, bone metabolism abnormalities, metabolic syndrome resulting from severe obesity caused by hyperphagia, deficiencies in the GH/IGF-1 axis, and the corresponding responses to treatment.

Abstract 1036: Development of a comprehensive RNA-based NGS panel for detecting tyrosine kinase fusion genes associated with primary hypereosinophilia

Abstract Hypereosinophilia (HE) is a medical condition diagnosed when the count of eosinophils in the blood is greater than 1.5 × 109 per liter. Primary HE is a clonal stem cell disorder manifesting as myeloid and/or lymphoid malignancies and is typically driven by aberrant fusion genes involving a tyrosine kinase (TK) gene (e.g., PDGFRA, PDGFRB, FGFR1, JAK2, FLT3, ABL1). The precise identification of the underlying molecular genetic abnormalities is crucial for timely diagnosing and managing primary HE. There has been no HE-specific assay that can concomitantly detect all known or novel TK fusion genes. We've developed an RNA-based panel for detecting TK fusion genes in HE. This panel, developed using 5' RACE technology, allows for the identification of fusion transcripts in total RNA extracted from whole blood sample. It covers 48 known TK fusion genes and also has the capability of detecting novel gene fusions. We balanced and optimized the panel using an in-house fusion RNA mix containing 11 fusion transcripts. We additionally validated the panel using samples from five patients with hematopoietic neoplasms and two fusion-positive cell lines to ensure its accuracy. The Limit of Detection (LoD) was established by analyzing the in-house RNA mix and normal whole blood RNA. It was found that the minimum RNA input required to detect all 11 fusion transcripts was 3540 copies in the presence of a background of 10 ng normal whole blood RNA. The panel successfully identified two fusion transcripts using 10 ng of total RNA obtained from cancer cell lines. The panel was then used to test five patient RNA samples extracted from whole blood. We successfully confirmed the presence of BCR-ABL1 fusion in 3 patient samples and ETV6-PDGFRB fusion in 1 patient sample, respectively. In summary, we developed HE-specific NGS panel for detecting all known and novel TK fusion genes and demonstrated its analytical accuracy and clinical effectiveness. The next phase of our study is to further validate the tests on a large patient cohort. Citation Format: Bongyong Lee, Andrew Chern, Aiguo Zhang, Chuanyi M. Lu, Michael Y. Sha. Development of a comprehensive RNA-based NGS panel for detecting tyrosine kinase fusion genes associated with primary hypereosinophilia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1036.

White blood cell count nadir to zero following intensive chemotherapy as a predictive factor for patients with acute myeloid leukemia

Predictors for response to intensive therapy in AML have focused on baseline factors: percent leukemic blasts in marrow, cytogenetic/molecular genetic abnormalities, and presence of secondary AML. Non-baseline dynamic factors, occurring after induction but before response, may be useful for decisions related to salvage chemotherapy. We hypothesized white blood cell (WBC) count nadir after induction may be a real time indicator of treatment efficacy. We also examined whether time to stem cell transplant (SCT) or baseline molecular genetic abnormalities are associated with a low nadir. Data showed WBC nadir = 0 was a negative predictor for response to intensive induction and was correlated with reduced overall survival and progression free survival. Patients with WBC nadir = 0 did not have a significantly longer time to SCT, and none of the mutations increased the likelihood of reaching WBC nadir = 0. WBC nadir may be a useful real-time monitor in AML patients receiving intensive induction chemotherapy.

The Prognostic Role of Genetic Aberrations in Mantle Cell Lymphoma: A Literature Review and Clinical Experience

Mantle cell lymphoma (MCL) is a type of peripheral B-cell non-Hodgkin’s lymphoma characterized by constitutive cyclin D1 overexpression leading to cell-cycle dysregulation and disruption of DNA damage repair. Apart from the typical translocation t(11;14)(q13;q32) and more rare variants, such as t(2;11)(p11;q13) and t(11;22)(q13;q11), a considerable number of patients quite often show secondary molecular and chromosomal aberrations underlying heterogeneity of the clinical course of MCL. Among a wide range of molecular genetic abnormalities, particular attention during the last years has been concentrated on studying the so-called double-hit MCL within a subgroup of patients with translocations involving CCND1 and MYC genes. Double-hit MCL is distinguished with rapid progression and tumor generalization at the time of diagnosis. Poor prognosis and low survival rates in most MCL patients call for the fastest possible diagnosis. Morphological and immunohistochemical as well as genetic methods (standard cytogenetic technique and fluorescence in situ hybridization) contribute to improving the quality of evidence-based diagnosis. The results of comprehensive diagnostic studies optimize prognosis assessment and treatment decision making in clinic.

Eosinophil disorders: an update on diagnosis and management

Abstract Eosinophilia can be seen in almost all medical subspecialty patients. Delay in diagnostic workup and treatment is associated with significant morbidity and mortality. Clinical vigilance and timely referral for diagnostic evaluation are critical. Causes of hypereosinophilia (HE) are diverse and can be grouped under 3 categories: primary (neoplastic), secondary (reactive), and idiopathic. Advances in molecular genetic diagnostics have led to elucidation of the genetic basis for many neoplastic hypereosinophilic disorders. One common molecular feature is formation of a fusion gene, resulting in the expression of an aberrantly activated tyrosine kinase (TK). The World Health Organization endorsed a biologically oriented classification scheme and created a new major disease category, namely, myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions. Rearrangement of other TK genes and activating somatic mutation(s) in TK genes have also been reported in eosinophilic neoplasms. Diagnostic evaluation of HE involves a combination of clinical, histopathologic, and immunophenotypic analyses, as well as molecular genetic testing, including next-generation sequencing–based mutation panels. The management of primary HE is largely guided by the underlying molecular genetic abnormalities. Good knowledge of recent advances in HE is necessary to ensure timely and accurate diagnosis and to help optimize patient care.

B-Lymphoblastic Leukemia Presenting with Eosinophilia: A Case Report and Review

B-lymphoblastic leukemia (B-ALL) presenting with eosinophilia represents a rare entity. In particular, the unique clinical manifestations as well as the genetic and molecular pathogenesis of the associated eosinophilia have been topics of debate and investigation. Herein, we present a case of B-ALL presenting with concomitant eosinophilia and a review of the published cases, including 120 additional cases. The patient, at presentation, was a 20-year-old female who presented with cardiac, neurologic, and cutaneous signs of eosinophilia and with leukocytosis (WBC 51, 40% eosinophils, 0% blasts), anemia, and thrombocytopenia. Imaging revealed multi-focal brain infarcts, pulmonary infiltrates, and myocarditis. Empiric steroids, colchicine, and intravenous immunoglobulin were initiated. One week after presentation, bone marrow biopsy revealed a hypercellular (>95%) marrow with 50% blasts and marked eosinophilia, which on later review revealed an immunophenotype most consistent with B-ALL (positive for Pax5, CD10, CD19, CD20, cCD22, CD34, CD45, CD79a, HLA-DR, and TdT and negative for MPO and CD117). Conventional cytogenetic analysis of the bone marrow was unable to be performed. Fluorescence in situ hybridization (FISH) revealed an IGH rearrangement in 10.5% of nuclei analyzed. After three weeks of treatment with steroids and resolution of symptomatic organ involvement, a repeat bone marrow biopsy did not show any evidence of malignancy or genetic abnormalities. The patient was continued on a steroid taper, did not initiate further antileukemic therapy, and was lost to follow up. Ten weeks after initial presentation and after completing a steroid taper, the patient presented with diffuse bone pain. Repeat bone marrow biopsy showed B-ALL with 64% blasts and increased eosinophils, normal conventional cytogenetics and hematologic malignancy sequencing panel, and FISH with gain of Xp in 20.5% of nuclei analyzed; a lumbar puncture did not detect blasts in the cerebrospinal fluid. The patient began induction therapy with aBFM, rituximab, PEG-asparaginase, and intrathecal cytarabine. A bone marrow biopsy performed four months after her initial presentation revealed a mildly hypocellular marrow with 3% B-lymphoblasts and no eosinophilia, suggesting residual disease, with no evidence of cytogenetic or molecular genetic abnormalities. As a result, therapy with blinatumomab was initiated. A follow up bone marrow biopsy performed six months after her initial presentation revealed a hypocellular marrow with multilineage maturation and no excess blasts with normal cytogenetic and molecular genetic studies. A literature review identified 120 additional cases of ALL with eosinophilia. In summary, it tends to present in adolescent males, often with mild anemia and thrombocytopenia and no peripheral blasts, which can lead to a delay in diagnosis of ALL. The eosinophilia is often symptomatic, with cardiopulmonary involvement being the most common, and can lead to rapid mortality in a substantial number of cases. A significant proportion of cases of ALL with eosinophilia involve t(5;14)(q31;q32), leading to overexpression of IL-3, which was not definitively found in this patient; however, there are numerous other recurrent abnormalities that may be contributing to the pathogenesis, both concurrently with and in the absence of this translocation, including other rearrangements of IGH as well as IKZF1 and CDKN2A abnormalities. Prompt diagnosis, now commonly using sensitive molecular diagnostic assays, and initiation of glucocorticoids for symptom control and subsequent antileukemic therapy, are essential to avoiding complications of eosinophilia in such patients.

NUT carcinoma and thoracic SMARCA4-deficient undifferentiated tumour: facts and controversies.

NUT carcinoma and thoracic SMARCA4-deficient undifferentiated tumour are unique entities in the 5th edition of the World Health Organisation (WHO) Classification of Thoracic Tumours, whose definitions include molecular genetic abnormalities. These aggressive tumours require rapid work-ups on biopsies, but a broad list of differential diagnoses poses challenges for practising pathologists. This review provides an update on their key clinicopathological and molecular characteristics, as well as controversies regarding tumour classification and diagnostic strategy. Phenotypical assessment plays a substantial role in diagnosis because recurrent and predictable clinicopathological findings exist, including robust immunohistochemical phenotypes. Accurate diagnosis is crucial for appropriate management and a clearer understanding of the disease.

Outcomes of Patients with FLT3 Positive Acute Myeloid Leukaemia; an Experience from a Tertiary Care Hospital in Karachi, Pakistan.

Molecular genetic abnormalities in acute myeloid leukaemia (AML) are essential for disease diagnosis and determining prognosis and clinical course. Mutations in FLT3 and nucleophosmin (NPM) genes are the most frequent genetic abnormalities, which are also known to impact disease outcomes. FLT3 mutations have been identified in approximately 30% of de novo AML patients and are associated with poor prognoses. This study aimed to determine the response to induction chemotherapy, overall survival (OS) and relapse rate (RR) in patients with FLT3-positive AML. In this study, a retrospective analysis was performed of 75 newly diagnosed patients with AML registered between January 2015 and July 2022. Patient demographics and clinical-haematological parameters were noted and molecular analysis for FLT3 ITD/TKD and NPM mutations was performed. All the patients received standard induction chemotherapy and their response to treatment, OS and RR were assessed. A total of 75 cases of AML were analysed. The mean age of the sample was 34.9 years, of which 65.3% were males and 34.7% were females. The patients were stratified into two groups: Those who were positive for FLT3 while negative for NPM (FLT3+/NPM-), representing 17.3% and those who were negative for both FLT3 and NPM (FLT3-/NPM-), representing 82.7% of cases. On day 28 post-induction, the complete remission rate was 69.2% in the FLT3 positive group and 77.4% in the FLT3 negative group. In the FLT3+/NPM- group, 55.6% of cases who were in remission at day 28 subsequently relapsed, compared to 50.0% of FLT3-/NPM- cases. The overall median survival time for the cohort and FLT3+ group was 1467 days, while that of the FLT3-group could not be estimated due to the very high survival rate. No significant differences in outcomes were observed in patients who were FLT3 positive compared to those who were FLT3 negative.

Recent Research Progress of Extramedullary Plasmacytoma --Review

Extramedullary plasma cell tumor (EMP) is a kind of plasma cell tumor, and its pathogenesis is not completely clear. According to whether it is independent of myeloma disease, it can be divided into primary and secondary EMP, which have different biological and clinical characteristics. Primary EMP has low invasion, fewer cytogenetic and molecular genetic abnormalities and good prognosis, and surgery and / or radiotherapy are the mainly treatments. Secondary EMP, as the extramedullary invasive progression of multiple myeloma (MM), is often accompanied by high-risk cellular and molecular genetic abnormalities and poor prognosis, chemotherapy, immunotherapy and hematopoietic stem cell transplantation are the mainly treatment. This paper reviews the latest research progress of EMP in the pathogenesis, cytogenetics molecular genetics and treatment, so as to provide reference for clinical work.

Correlation between Expression Patterns of CD117/CD200 in Plasma Cells and Molecular Genetic Prognostic Parameters in Multiple Myeloma

To investigate the correlation between the expression of CD117 and CD200 in plasma cells and molecular genetic abnormalities in patients with multiple myeloma (MM). 100 newly diagnosed MM patients were selected, and fresh bone marrow fluid was collected from the patients. The immunophenotypes and chromosomal structural variations of plasma cells were detected by flow cytometry (FCM) and fluorescence in situ hybridization (FISH). The positive expression frequencies of CD117 and CD200 in abnormal plasma cells of all MM patients were 44.0% and 44.0%, respectively. At least one molecular genetic abnormality was detected in 53 of the 75 patients who underwent FISH testing, and the overall detection rate was 70.7% (53/75). The detection rates of 1q21 (CKS1B ) duplication, 1p32 (CDKN2C ) deletion, p53 deletion and IgH rearrangement were 48.6% (36/74), 10.6% (7/66), 11.1% (8/72) and 32.9% (24/73), respectively. The incidence of IgH rearrangement in CD117+ patients was significantly lower than that in CD117- patients (P<0.05), and the proportion of 1p32 (CDKN2C ) deletion in CD200- patients was significantly lower than that in CD200+ patients (P<0.05). According to the expressions of CD117 and CD200, the patients were divided into 4 groups: CD117+CD200+, CD117+CD200-, CD117-CD200+ and CD117-CD200-. Further analysis showed that the incidence of IgH rearrangement in the CD117+CD200- group was significantly lower than that in the CD117-CD200+ group (P<0.05), and the deletion rate of 1p32 (CDKN2C ) gene in CD117+CD200- group was significantly lower than that in CD117+CD200+ group and CD117-CD200+ group (P<0.05). The difference in the expression patterns of CD117 combined with CD200 shows important value in judging the prognosis of MM patients, and the MM patients with CD117-CD200+ expression patterns in abnormal plasma cells have a worse prognosis.

RUNX1 Mutated AML and MDS: Similarities, Differences and Molecular Factors Leading to Disease Progression

Background: Mutations in the myeloid transcription factor RUNX1 are frequently detected in AML and MDS. Differences have been described regarding additional cytogenetic and molecular genetic abnormalities, however the molecular factors responsible for progression of RUNX1 mutated MDS to AML are yet not fully elucidated. Aim: Analysis of genetic (1) similarities and (2) differences of RUNX1 mutated AML and MDS cases, (3) identification of factors potentially causing disease progression from RUNX1 mutated MDS to AML. Methods: Whole-genome sequencing (WGS) was performed for 773 AML and 747 MDS patients (median coverage 100x). 151bp paired-end reads were generated on NovaSeq 6000 and HiSeqX machines (Illumina, San Diego, CA). As no sample specific normal tissue was available, a so-called Tumor/Unmatched normal (TUN) workflow was used to reduce technical artefacts and germline calls. All reported p-values are two-sided and were considered significant at p<0.05. Results: Mutations in RUNX1 (RUNX1mut) were detected at comparable frequencies in AML and MDS (AML: 97/773 cases, 13%; MDS: 71/747, 10%). In MDS, an association of the RUNX1mut frequency with higher blast counts was observed (0-5% blasts: 15/419 with RUNX1mut, 4%; ≥5,<10%: 16/175, 9%; ≥10,<20%: 40/153, 26%). However, blast count did not influence overall survival (OS) in RUNX1mut subgroups (0-5% blasts: 18 months (m); ≥5,<10%: 25 m; ≥10,<20%: 18 m; AML: 21 m). The variant allele frequency (VAF) of RUNX1mut differed significantly dependent on the blast count (median VAF in cases <5% blasts: 0.22; ≥5%: 0.42; p<0.001). Regarding cytogenetic abnormalities, RUNX1mut MDS showed a higher frequency of normal karyotype (NK) compared to AML (66% vs. 38%, p<0.001) but similar frequencies of trisomies (18% vs. 22%) and complex karyotype (3% vs. 5%), although differences regarding the trisomy type were detected (MDS: only trisomy 8; AML: trisomies 8, 11 and 13). In 10% of AML cases with RUNX1mut a rearrangement involving MECOM was detected (5% GATA::MECOM, 5% other MECOM rearrangements), which was absent in MDS with RUNX1mut. Similarly, KMT2A-PTD and FLT3-ITD alterations were detected in AML but not in MDS with RUNX1mut (KMT2A-PTD: 7% in AML vs. 0% in MDS, p=0.021; FLT3-ITD: 13% vs. 0%, p=0.001). By contrast, TP53mut were found at comparably low frequencies (MDS: 4/71, 6%; AML: 3/97, 3%). For both AML and MDS, a strong association of RUNX1mut with mutations in spliceosome genes (SF3B1, SRSF2, U2AF1, ZRSR2) was detected (AML: 58/97 = 60% in RUNX1mut vs. 139/676 = 21% in RUNX1wild type (wt); p<0.001; MDS: 52/71 = 73% vs. 346/676 = 51%, p<0.001). However, while SRSF2mut in MDS were associated with increasing blast counts similar to RUNX1mut (0-5% blasts: 25/419 with SRSF2mut, 6%; ≥5,<10%: 130/175, 17%; ≥10,<20%: 50/153, 32%), SF3B1mut were negatively correlated to blast counts and thus to RUNX1mut (0-5% blasts: 179/419, 43%; ≥5,<10%: 28/175, 16%; ≥10,<20%: 11/153, 7%). In the total AML cohort, in 24/773 of cases a CN-LOH involving RUNX1 was detected, 21 of these also showed a RUNX1mut (22% (21/97) of RUNX1mut cases). By contrast, in the total MDS cohort in only 2/747 cases a CN-LOH covering RUNX1 was found, none of them harboring a RUNX1mut. The VAF of cases with RUNX1mut + CN-LOH (RUNX1mutLOH) was >0.5 for all cases and was significantly higher than the VAF of RUNX1mut cases without CN-LOH (0.89 (range 0.54-0.97) vs. 0.42 (range 0.05-0.85), p<0.001). Moreover, cases with RUNX1mutLOH were found to be significantly associated with FLT3-ITD (RUNX1mutLOH, 6/21 = 29%; RUNX1mut without CN-LOH, 7/76 = 9%; p=0.032). Conclusions: (1) RUNX1mut occur at similar frequencies in AML and MDS and genetically show an MDS signature, reflected e.g. by association with mutations in spliceosome genes. OS is similar in all subgroups independent of blast counts. (2) In MDS, RUNX1mut are associated with higher blast counts, thus potentially functioning as driver towards progression to AML. (3) Additional cytogenetic/molecular genetic alterations that might function as progression factors from RUNX1mut MDS to AML include: (i) karyotype, especially MECOM rearrangements, trisomy 11 and trisomy 13 (ii) KMT2A-PTD, (iv) FLT3-ITD, (v) and CN-LOH covering RUNX1 (Figure). (4) By contrast, TP53mut do not seem to play a role in progression of RUNX1 mutated MDS to AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

R-DA-Edoch/R-DHAP Alteration Could Overcome High Risk Factors in Younger Mantle Cell Lymphoma

Background: Mantle-cell lymphoma (MCL) is a rare subtype of non-Hodgkin's lymphoma, with a median overall survival of only 3~5 years in the era of immunochemotherapy. Although first-line intensive chemotherapy with rituximab (R) and high dose cytarabine combined with autologous transplantation (ASCT) has significantly prolonged the survival of patients with MCL, some patients still die within few years. Our published data show that Chinese patients with MCL are younger and have a higher proportion of TP53 deletion and MYC abnormality [Oncotarget, 2015 Dec 8;6(39):42362-71]. Considering the efficacy and tolerability of the dose-enhanced immunochemotherapy R-DA-EDOCH regimen in patients with multiple high-risk aggressive B-cell lymphomas and the potential higher proliferative activity of tumor cells in Chinese patients, we conducted a prospective clinical study of alternate R-DA-EDOCH/R-DHAP induction therapy for young patients with newly-diagnosed MCL (NCT02858804). Here, we analyzed the efficacy and safety of this regimen. Methods: The main inclusion criteria for patients with newly-diagnosed MCL included age ≤ 65 years, Ann Arbor stage II~IV and ECOG ≤ 1. During induction, patients received alternate R-DA-EDOCH/R-DHAP for two cycles. Patients who achieved less than partial remission would quit the trial, while the other patients would continue another one cycle of treatment. After finishing the 3 cycles of inductive treatment, patients would decide whether to continue ASCT consolidation therapy or another cycle of consolidation. R maintenance therapy was recommended to all patients. The primary endpoint of this study was the progression-free survival (PFS). Key secondary endpoints were complete response rate (CRR) and objective response rate (ORR) at the end of induction (EOI), overall survival (OS), minimal residual disease (MRD) status and the incidence of adverse events (AEs). Results: In this study, we recruited 55 patients with a median age of 53 years (range, 37~65 years) and a male/female ratio of 2.2:1. Totally, 61.8% of patients were of at least one risk factor (HR) according to risk stratification, pathological classification and molecular genetic abnormalities. All patients received at least two cycles of alternate R-DA-EDOCH/R-DHAP immunochemotherapy, and one patient dropped out due to the poor response. The ORR of the alternate R-DA-EDOCH/R-DHAP inductive treatment was 98% (95% CI 90-100), and the CRR was 89% (95%CI 78-96). There was no significant difference in CRR and ORR among patients with different prognostic characteristics. Moreover, patients rapidly achieved MRD-negative and the rate of MRD-negative after 2 cycles of treatment, the end of the inductive therapy and the end of the overall treatment was 79.2%, 88.7% and 90.6%, respectively. The median follow-up of the cohort was 31 months (range 4-86). The 3-year PFS and OS rates were 66.3% and 85.4%, respectively. Sequential ASCT consolidation and/or R maintenance slightly increased CRR but significantly prolonged PFS of patients (3-year PFS rate: ASCT+R 90.4% versus R 64.6% versus without ASCT/R 33.3%, P<0.001). There was no significance difference in PFS and OS between patients with (n=34) and without (n=9) HR (PFS P=0.537, OS P=0.408). For patients receiving ASCT and R maintenance, OS was even longer in patients with HR (mOS not reach versus 47m, P=0.007). Univariate analysis showed that age≥60 years, blastoid/pleomorphic pathologic morphology and TP53 double-allele events were still the risk factors for PFS in this therapeutic pattern. However, high tumor proliferative activity (such as Ki-67≥30% and MYC abnormality) and specific genetic abnormalities (such as 17p deletion, 9p deletion, etc) showed no significant adverse prognostic influence. In particular, patients who achieved MRD-negative by treatment had longer PFS (P<0.001). The alternate R-DA-EDOCH/R-DHAP regimen was well tolerated, and no patients were discontinued due to intolerance of treatment-related AEs. AEs were mainly hematologic and at least one hematologic AEs occurred in 85.5% of the patients. Grade 3-4 leukopenia and neutropenia were found in 49.1% and 58.2% of the patients, respectively. Conclusion: The alternate R-DA-EDOCH/R-DHAP induction therapy showed great efficacy and tolerability in young patients with newly diagnosed MCL and could overcome the adverse prognostic impact of some high-risk factors. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

High Tumor Mutation Burden in Epigenetic Regulatory Genes Predicts Decreased Overall Survival in Nodal Peripheral T-Cell Lymphomas

Introduction: Nodal peripheral T-cell lymphomas (nPTCL) constitute a heterogeneous group of rare malignancies with aggressive biological behavior and poor prognosis. Epigenetic phenomena involving genes that control DNA-methylation and histone deacetylation, such as IDH2, DNMT3A, TET2 and RhoA, play a central role in their pathogenesis. Tumor mutation burden (TMB) has emerged as a prognostic biomarker in numerous solid neoplasms. Furthermore, TMB reflects the process of clonal evolution, with progressive acquisition of additional molecular-genetic abnormalities. Recent studies were capable to establish a relationship between TMB and response to anti-cancer treatments. In 2021 Fachi et cols. established a correlation between high TMB in epigenetic regulatory genes and higher response rates to 5-azacytidine and romidepsin in nPTCL. However, the potential prognostic impact of epigenetic TMB remains unknown in nPTCL. Based on this premise, here we aimed, in a pioneering way, to search for a correlation between epigenetic high TMB and adverse prognosis in nPTCL. Methods: This is a retrospective, observational and single-center study, involving 59 patients with nPTCL diagnosed and treated at the Department of Hematology, University of São Paulo, Brazil, from January 2000 to December 2019. Formalin-fixed paraffin-embedded (FFPE) diagnostic tumor samples were submitted to Sanger sequencing with a target-panel, involving IDH2, DNMT3A, TET2 and RhoA genes to search for recurrent mutations. Only non-synonymous mutations were considered for prognostic association. Survival curves were constructed using the Kaplan-Meier method and the Log-Rank test was used to establish a relationship between TMB and outcomes. Results were presented as hazard ratio (HR) and 95% confidence interval (95% CI), and a p-value < 0.10 was considered statistically significant. Results: The median age was 50 years (IqR 38-61), and 57.6% (34/59) were male. Thirty-nine percent (23/59) had bulky disease ≥ 7 cm, 83% (49/59) had B-symptoms, 94.9% (56/59) had advanced stage III/IV, 27.1% (16/59) had ECOG ≥ 2, 27.1% (16/59) had ≥ 2 extranodal sites involved by lymphoma, and 55.9% (33/59) had intermediate-high/high-risk IPI. Seventy-six percent (45/59) received up-front therapy with CHOP-like regimens (CHOP or CHOEP), 23.7% (14/59) experienced radiotherapy, and 33.3% (20/59) were consolidated with ASCT. With a median follow-up of 3.7 years (95% CI: 0.9-12.4), the estimated 2-year OS and PFS were 57.1% (95% CI: 45.5-70.4) and 49.2% (95% CI: 34.0-59.2), respectively. Missense, non-sense or frameshift mutations in the IDH2 gene were observed in 3.3% (2/59) of cases, DNMT3A in 3.3% (2/59), RhoA in 23.7% (14/59) and TET2 in 37.2% (22/59). Among the 59 patients included, 23/59 (38.9%) had no mutations in the target-genes, 25/59 (42.4%) had 1 mutation, and 11/59 (18.6%) had two or more mutations. Therefore, the cases were categorized into low TMB (< 2 mutations) [n=48/59 - 81.3%] or high TMB (≥ 2 mutations) [n=11/59 - 18.7%]. The median overall survival was 51.4 months (95% CI: 0-124.1) for low TMB and only 3.1 months (95% CI: 3.1-15.2) for high TMB, p=0.08. The estimated 2-year OS was 64.3% (95% CI: 50.6-78.0) for low TMB and 36.4% (95% CI: 8.0-65.0) for high TMB [Figure 1]. Similarly, median progression-free survival was 13.3 months (95% CI: 0-29.4) for low TMB and 6.3 months (95% CI: 0.9-11.6) for high TMB, p=0.319. The estimated 2-year PFS was 46.1% (95% CI: 20.0-72.1) for low TMB and 25.0% (95% CI: 17.5-67.5) for high TMB. Conclusion: In a pioneering way, we demonstrated that the high epigenectic tumor mutation burden (TMB), defined by the presence of 2 or more mutations involving epigenetic regulatory genes, was associated with decreased overall survival in Brazilian patients with nodal PTCL. Although preliminary, these data support the potential impact of epigenetic TMB as a prognostic biomarker in nPTCL. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Molecular genetic abnormalities in patients with T-cell acute lymphoblastic leukemia: a literature review

T-cell acute lymphoblastic leukemia/lymphoma (T-ALL) is an aggressive hematological disease. Modern polychemotherapy protocols allow achieving a 5-year overall survival of 60–90 % in different age groups, however, relapses and refractory forms of T-ALL remain incurable. Over the past decades, the pathogenesis of this variant of leukemia has been studied in many trials, and it has been found that various signaling pathways are involved in the multi-step process of leukemogenesis. This opens the way for targeted therapy.In this review, we provide an update on the pathogenesis of T-ALL, opportunities for introducing targeted therapies, and issues that remain to be addressed.

Integrated clinical genotype-phenotype characteristics of early T-cell precursor acute lymphoblastic leukemia.

Early T-cell precursor acute lymphoblastic leukemia (ETP-ALL) is a distinct subtype of T-ALL with a unique immunophenotype and high treatment failure rate. The molecular genetic abnormalities and their prognostic impact in ETP-ALL patients are poorly understood. The authors performed systematic analyses of the clinicopathologic features with an emphasis on molecular genetic aspects of 32 patients with ETP-ALL. The median age was 43years (range, 16-71). The blasts were positive for cytoplasmic CD3 and CD7 and negative for CD1a and CD8. Other markers expressed included CD34 (88%), CD33 (72%), CD117 (68%), CD13 (58%), CD5 (partial, 56%), CD2 (38%), CD10 (25%), CD56 (partial, 19%), and CD4 (6%). Cytogenetic analyses revealed a diploid karyotype in 10 patients, simple (1-2) abnormalities in 10 patients, and complex karyotype in 10 patients. Next-generation sequencing for 21 patients demonstrated that all had gene mutations (median, four mutations per patient). The most frequently mutated genes were WT1 (38%), NOTCH1 (29%), NRAS (29%), PHF6 (25%), TP53 (24%), ASXL1 (19%), FLT3 (19%), and IKZF1 (19%). All patients except one received multi-agent chemotherapy, and 22 patients underwent allogeneic stem cell transplantation. Thrombocytopenia, an abnormal karyotype, and TP53 mutation were associated with markedly shortened overall survival. Stem cell transplantation significantly improved overall survival. Patients with ETP-ALL often have high mutation burden with increased genomic instability. TP53 mutation was the only molecular prognostic marker and was associated with complex karyotype and greater than or equal to five mutations. These patients may benefit from stem cell transplantation, and recurrent gene mutations may be novel therapeutic markers.

HIGH TUMOR MUTATION BURDEN IN EPIGENETIC REGULATORY GENES PREDICTS DECREASED OVERALL SURVIVAL IN NODAL PERIPHERAL T-CELL LYMPHOMAS

Nodal peripheral T-cell lymphomas (nPTCL) constitute a heterogeneous group of rare malignancies with aggressive biological behavior and poor prognosis. Epigenetic phenomena involving genes that control DNA-methylation and histone deacetylation, such as IDH2, DNMT3A, TET2 and RhoA , play a central role in their pathogenesis. Tumor mutation burden (TMB) has emerged as a prognostic biomarker in numerous solid neoplasms. Furthermore, TMB reflects the process of clonal evolution, with progressive acquisition of additional molecular-genetic abnormalities. Recent studies were capable to establish a relationship between TMB and response to anti-cancer treatments. In 2021 Fachi et cols. established a correlation between high TMB in epigenetic regulatory genes and higher response rates to 5-azacytidine and romidepsin in nPTCL. However, the potential prognostic impact of epigenetic TMB remains unknown in nPTCL. Based on this premise, here we aimed, in a pioneering way, to search for a correlation between epigenetic high TMB and adverse prognosis in nPTCL. This is a retrospective, observational and single-center study, involving 59 patients with nPTCL diagnosed and treated at the Department of Hematology, University of São Paulo, Brazil, from January 2000 to December 2019. FFPE diagnostic tumor samples were submitted to Sanger sequencing with a target-panel, involving IDH2, DNMT3A, TET2 and RhoA genes to search for recurrent mutations. The median age was 50 years (IqR 38-61), and 57.6% (34/59) were male. Thirty-nine percent (23/59) had bulky disease ≥ 7 cm, 83% (49/59) had B-symptoms, 94.9% (56/59) had advanced stage III/IV, 27.1% (16/59) had ECOG ≥ 2, 27.1% (16/59) had ≥ 2 extranodal sites involved by lymphoma, and 55.9% (33/59) had intermediate-high/high-risk IPI. Seventy-six percent (45/59) received up-front therapy with CHOP-like regimens (CHOP or CHOEP), 23.7% (14/59) experienced radiotherapy, and 33.3% (20/59) were consolidated with ASCT. With a median follow-up of 3.7 years (95% CI: 0.9-12.4), the estimated 2-year OS and PFS were 57.1% (95% CI: 45.5-70.4) and 49.2% (95% CI: 34.0-59.2), respectively. Missense, non-sense or frameshift mutations in the IDH2 gene were observed in 3.3% (2/59) of cases, DNMT3A in 3.3% (2/59), RhoA in 23.7% (14/59) and TET2 in 37.2% (22/59). Among the 59 patients included, 23/59 (38.9%) had no mutations in the target-genes, 25/59 (42.4%) had 1 mutation, and 11/59 (18.6%) had two or more mutations. Therefore, the cases were categorized into low TMB (< 2 mutations) [n = 48/59 – 81.3%] or high TMB (≥ 2 mutations) [n = 11/59 – 18.7%]. The median overall survival was 51.4 months (95% CI: 0-124.1) for low TMB and only 3.1 months (95% CI: 3.1-15.2) for high TMB, p = 0.08. The estimated 2-year OS was 64.3% (95% CI: 50.6-78.0) for low TMB and 36.4% (95% CI: 8.0-65.0) for high TMB. Similarly, median progression-free survival was 13.3 months (95% CI: 0-29.4) for low TMB and 6.3 months (95% CI: 0.9-11.6) for high TMB, p = 0.319. The estimated 2-year PFS was 46.1% (95% CI: 20.0-72.1) for low TMB and 25.0% (95% CI: 17.5-67.5) for high TMB. In a pioneering way, we demonstrated that the high epigenectic TMB, defined by the presence of 2 or more mutations involving epigenetic regulatory genes, was associated with decreased overall survival in Brazilian patients with nodal PTCL. Although preliminary, these data support the potential impact of epigenetic TMB as a prognostic biomarker in nPTCL.

ReDisX, a machine learning approach, rationalizes rheumatoid arthritis and coronary artery disease patients uniquely upon identifying subpopulation differentiation markers from their genomic data.

Diseases originate at the molecular-genetic layer, manifest through altered biochemical homeostasis, and develop symptoms later. Hence, symptomatic diagnosis is inadequate to explain the underlying molecular-genetic abnormality and individual genomic disparities. The current trends include molecular-genetic information relying on algorithms to recognize the disease subtypes through gene expressions. Despite their disposition toward disease-specific heterogeneity and cross-disease homogeneity, a gap still exists in describing the extent of homogeneity within the heterogeneous subpopulation of different diseases. They are limited to obtaining the holistic sense of the whole genome-based diagnosis resulting in inaccurate diagnosis and subsequent management. Addressing those ambiguities, our proposed framework, ReDisX, introduces a unique classification system for the patients based on their genomic signatures. In this study, it is a scalable machine learning algorithm deployed to re-categorize the patients with rheumatoid arthritis and coronary artery disease. It reveals heterogeneous subpopulations within a disease and homogenous subpopulations across different diseases. Besides, it identifies granzyme B (GZMB) as a subpopulation-differentiation marker that plausibly serves as a prominent indicator for GZMB-targeted drug repurposing. The ReDisX framework offers a novel strategy to redefine disease diagnosis through characterizing personalized genomic signatures. It may rejuvenate the landscape of precision and personalized diagnosis and a clue to drug repurposing.

The presence of Auer rods in neutrophils and monocytes besides blasts in a case of acute myeloid leukemia with CSF3R mutation

An 81-year-old male with a history of diabetes, hypertension, hypercholesterolemia, previous DVT not on anticoagulation, and arthritis presented to the emergency department with a chief complaint of generalized weakness. Complete blood counts revealed white blood cells (WBC) 165.3 × 109/L, hemoglobin (Hb) 4.4 g/dl, and platelet (PLT) < 10 × 109/L, with 62% blasts. Review of peripheral blood showed blasts of variable sizes with a moderate amount of basophilic cytoplasm (Figure 1A–D; peripheral blood; Leishman stain; 100× objective), and pale-salmon-pink-colored cytoplasmic granules in a subset (Figure 1A, see arrows). Auer rods are easily identified in blasts (Figure 1A,B, see asterisk), occasionally seen in monocytes (Figure 1C, see asterisk) and neutrophils (Figure 1d, see asterisk). Bone marrow was examined to establish diagnosis, showing marked hypercellularity with 72% blasts on aspirate smears with morphologic features similar to those in peripheral blood (Figure 1E [bone marrow biopsy; hematoxylin and eosin, 40× objective] and F [bone marrow aspirate; Leishman stain; 50× objective]). Granulocytic dysplasia with pelgeroid morphology and abnormal nuclear chromatin clumping was seen in both peripheral blood and bone marrow (> 10% and < 50%). Concurrent flow cytometry showed increased myeloblasts with expression of CD34, CD13, CD 117, CD33, HLA-DR, CD19 (dim; minor subset), CD11b (dim; subset), CD14 (dim; minor subset), CD15 (dim; minor subset), myeloperoxidase (dim; subset), and terminal deoxynucleotidyl transferase (dim; minor subset). Conventional cytogenetics showed normal male karyotype. Concurrent FISH studies using acute myeloid leukemia (AML) and myelodysplastic syndrome panels of probes are negative for all loci studied, including 8q22 and 21q22 probes for RUNX1-RUNX1T1 rearrangement. Next generation sequencing showed mutation on the CSF3R gene (p.T640N, variant allele frequency 48.4%). An integrated diagnosis of AML with maturation was established (French–American–British Classification Subtype M2). Auer rods are a morphologic feature in leukemic blasts defining myeloid differentiation. The presence of Auer rods in neutrophils and monocytes are rarely reported, most commonly in association with acute promyelocytic leukemia (APL) with PML-RARA and AML with RUNX1-RUNX1T1, occasionally seen in other subtypes of AML and myelodysplastic syndrome (MDS) in the literature as well [1]. We presented a case of AML with Auer rods in non-blast cells as well as morphologic and immunophenotypic features resembling AML with RUNX1-RUNX1T1: blasts with basophilic cytoplasm, salmon-pink (pale) cytoplasmic granules and perinuclear Hof besides Auer rods as well as granulocytic dysplasia in the background. However, no recurrent cytogenetic and molecular genetic abnormalities as defined by World Health Organization (WHO) classification are detected, while a CSF3R mutation seen. Although mutations in the colony stimulating factor 3 receptor (CSF3R) occurs rarely in AML, comprising only about 1% of adults with AML, it is suggested that CSF3R can be a potential therapeutic target in AML [2]. In summary, we presented for the first time a case of AML with a few rare morphologic features including the presence of Auer rods in non-blast cells coinciding with a potential therapeutic target of CSF3R mutation. Whether the association between the morphologic features and the CSF3R mutation can be established warrants additional studies. In addition, a potential diagnostic pitfall to be taken into consideration in this case is that no reverse transcription-polymerase chain reaction (RT-PCR) was done to further exclude RUNX1-RUNX1T1 rearrangement, although the likelihood of having undetected rearrangement by karyotyping and FISH studies is extremely low. What is the new aspect of your work? Here, we for the first time presented a case of AML with morphologic features resembling AML with RUNX1-RUNX1T1, including Auer rods in non-blast cells, coinciding with CSF3R mutation, which is a rare mutation in AML while representing a potential novel therapeutic target. What is the central finding of your work? A rare case of CSF3R-mutated AML with no recurrent cytogenetic abnormality, showing unusual morphologic features resembling AML with RUNX1-RUNX1T1, including Auer rods in non-blast cells. What is (or could be) the specific clinical relevance of your work? The rare morphological features present in this AML could potentially serve as a trigger to identify cases of AML with CSF3R mutation for targeted therapy. None. The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported. The authors received no specific funding for this work. Not applicable. Not applicable. Not applicable. The data that support the findings of this study are available from the corresponding author upon reasonable request.

Evaluation of the Efficacy and Safety of Chemotherapy in Elderly Patients with Multiple Myeloma under Different Frailty Scores

To compare the efficacy and safety of different chemotherapy regimens in elderly multiple myeloma (MM) patients with different Frailty scores. The clinical data of elderly patients with MM were retrospectively analyzed, including age, treatment regimen, efficacy, adverse reactions, and the Frailty score included in the activity of daily living score, the instrumental activity of daily living scale and the Charlson comorbidity index. The patients were divided into fit group, mediate fit group and frail group according to the scoring standard. The treatment efficiency and adverse reaction rates of elderly MM with different physical conditions treated by different chemotherapy regimens were analyzed. Among the 70 patients, the effective rates of the patients in fit group, the mediate fit group, and the frail group were 79.5%, 81%, and 40%, and the effective rates of the fit patients in double and triple groups were 54.5% and 89.3%, 70% and 90.9% for mediate fit patients, 42.9% and 33.3% for frail patients, the triple regimen in fit patients showed obvious advantages, and the difference showed statistically significant (P<0.05), while the efficacy for mediate patients and frail patients showed no significant difference. During the induction of bortezomib, the incidence of adverse reactions for the patients in the triple group (78.6%) was higher than 67.9% in the double group, and the difference showed no statistically significant (P>0.05).There was no significant difference in the 1-year overall survival rate of the patients and with molecular genetic abnormalities among each groups. The therapeutic effect is related to the patient's physical condition. For patients with healthy physique, the triple regimen should be used first. For patients with weak physical constitution, the chemotherapy regimen with low drug toxicity should be selected for safety.

Prenatal exome sequencing and chromosomal microarray analysis in fetal structural anomalies in a highly consanguineous population reveals a propensity of ciliopathy genes causing multisystem phenotypes.

Fetal abnormalities are detected in 3% of all pregnancies and are responsible for approximately 20% of all perinatal deaths. Chromosomal microarray analysis (CMA) and exome sequencing (ES) are widely used in prenatal settings for molecular genetic diagnostics with variable diagnostic yields. In this study, we aimed to determine the diagnostic yield of trio-ES in detecting the cause of fetal abnormalities within a highly consanguineous population. In families with a history of congenital anomalies, a total of 119 fetuses with structural anomalies were recruited and DNA from invasive samples were used together with parental DNA samples for trio-ES and CMA. Data were analysed to determine possible underlying genetic disorders associated with observed fetal phenotypes. The cohort had a known consanguinity of 81%. Trio-ES led to diagnostic molecular genetic findings in 59 fetuses (with pathogenic/likely pathogenic variants) most with multisystem or renal abnormalities. CMA detected chromosomal abnormalities compatible with the fetal phenotype in another 7 cases. Monogenic ciliopathy disorders with an autosomal recessive inheritance were the predominant cause of multisystem fetal anomalies (24/59 cases, 40.7%) with loss of function variants representing the vast majority of molecular genetic abnormalities. Heterozygous de novo pathogenic variants were found in four fetuses. A total of 23 novel variants predicted to be associated with the phenotype were detected. Prenatal trio-ES and CMA detected likely causative molecular genetic defects in a total of 55% of families with fetal anomalies confirming the diagnostic utility of trio-ES and CMA as first-line genetic test in the prenatal diagnosis of multisystem fetal anomalies including ciliopathy syndromes.