Novel Germline TET2 Mutations in Two Unrelated Patients with Autoimmune Lymphoproliferative Syndrome-Like Phenotype and Hematologic Malignancy.

Somatic mutations in the ten-eleven translocation methylcytosine dioxygenase 2 gene (TET2) have been associated to hematologic malignancies. More recently, biallelic, and monoallelic germline mutations conferring susceptibility to lymphoid and myeloid cancer have been described. We report two unrelated autoimmune lymphoproliferative syndrome-like patients who presented with T-cell lymphoma associated with novel germline biallelic or monoallelic mutations in the TET2 gene. Both patients presented a history of chronic lymphoproliferation with lymphadenopathies and splenomegaly, cytopenias, and immune dysregulation. We identified the first compound heterozygous patient for TET2 mutations (P1) and the first ALPS-like patient with a monoallelic TET2 mutation (P2). P1 had the most severe form of autosomal recessive disease due to TET2 loss of function resulting in absent TET2 expression and profound increase in DNA methylation. Additionally, the immunophenotype showed some alterations in innate and adaptive immune system as inverted myeloid/plasmacytoid dendritic cells ratio, elevated terminally differentiated effector memory CD8 + T-cells re-expressing CD45RA, regulatory T-cells, and Th2 circulating follicular T-cells. Double-negative T-cells, vitamin B12, and IL-10 were elevated according to the ALPS-like suspicion. Interestingly, the healthy P1's brother carried a TET2 mutation and presented some markers of immune dysregulation. P2 showed elevated vitamin B12, hypergammaglobulinemia, and decreased HDL levels. Therefore, novel molecular defects in TET2 confirm and expand both clinical and immunological phenotype, contributing to a better knowledge of the bridge between cancer and immunity.

Leukaemic Presentation of Small-Cell Alk-Positive Anaplastic Large Cell Lymphoma in a Young Woman-Report of a Case with 9-Year Survival.

Anaplastic large cell lymphoma (ALCL) with leukaemic presentation (either ab initio or along the course of the disease) has been rarely reported. Irrespective of ALK expression in the neoplastic cells, it features a dismal prognosis. We report a rare case of leukaemic, small cell variant ALK-positive ALCL with 9-year survival in a young woman who was treated upfront with corticosteroids and standard chemotherapy, and review thoroughly the previously published cases. Such an unexpected, good outcome hints at the existence of different clinical subgroups in the leukaemic variant of ALK-positive ALCL.

Double CD4/CD8-Positive, Nonpoikilodermic Mycosis Fungoides Expressing CD56 in a Young Man.

ABSTRACT: We report a case of mycosis fungoides (MF) in an 18-year-old man whose neoplastic T cells expressed CD4, CD8, and CD56, with no evidence of TCR-delta or Epstein-Barr virus (EBER) expression. Clinically, neither hypopigmentation nor hyperpigmentation nor poikilodermatous skin lesions were present, and the lesions subsided with oral corticoids and retinoids and environmental solar ultraviolet exposure. Our case represents the oldest patient reported so far with nonpoikilodermatous, CD8/CD56 MF and adds to the phenotypic diversity of MF in the pediatric population. This distinct phenotype does not seem to be linked to a more aggressive course than the classic CD-4 positive one.

Epstein-Barr virus-associated large B-cell lymphoma transformation in marginal zone B-cell lymphoma: a series of four cases.

AIMS: To present four examples of clonally related Epstein-Barr virus (EBV)-associated large-cell transformation of marginal zone lymphoma (MZL) (of nodal, extranodal and splenic types), occurring 120, 11 and 5 months after the initial diagnosis in three instances, and concurrently in one case; and to discuss several interesting features of EBV infection. METHODS AND RESULTS: Somatic mutations were detected by use of a customised panel for next-generation sequencing and polymerase chain reaction studies of IgH in both low-grade and high-grade components of each case. In case 1, the initial biopsy of nodal MZL showed scattered EBV-positive cells, which might constitute an indication of EBV-induced progression. Case 2 showed heterogeneous EBV expression, a phenomenon attributable to loss of the EBV episomes during cell division, or to a secondary superinfection or reactivation of the virus. In case 3, p53 overexpression related to gene mutation and EBV-encoded small RNAs were identified in the same neoplastic component. In case 4, the mucosa-associated lymphoid tissue-type MZL and the high-grade component were identified concurrently in a patient previously treated with methotrexate for an autoimmune disorder. CONCLUSION: These data suggest that the presence of EBV should be added to the list of potential markers to be analysed for MZL prognosis.

MYD88 L265P mutation in cutaneous involvement by Waldenström macroglobulinemia.

Cutaneous manifestations of Waldenström macroglobulinemia (WM) may occur because of several mechanisms, the least common being direct skin infiltration by neoplastic cells. We report a case of patient that after 4-year history of indolent WM developed skin infiltration by lymphoplasmacytoid cells in the form of a small, mildly indurated plaque on the anterior chest. MYD88 L265P mutation was detected both in the previous bone marrow biopsy and in the cutaneous lesion. We review the impact of this new genetic tool in the diagnosis and treatment of lymphoplasmacytic proliferations.

Primary cutaneous follicular helper T-cell lymphoma.

Follicular helper T-cells (TFH) represent a specific subset of CD4-positive helper T-cells that help B-cells to differentiate into long-lived antibody-secreting plasma cells or memory B-cells. The expression of TFH markers in neoplastic T-cells, traditionally related to the angioimmunoblastic (AITL) subgroup of peripheral T-cell lymphomas, is nowadays well-known to be more widespread than previously thought. We report hereby a case of cutaneous T-cell lymphoma in a 75-year-old woman, whose morphological and immunophenotypical features raises the differential diagnosis between cutaneous involvement by AITL and the recently described primary cutaneous T-cell lymphoma with follicular helper-phenotype.

Hyperphosphorylation of PP2A in colorectal cancer and the potential therapeutic value showed by its forskolin-induced dephosphorylation and activation.

BACKGROUND: The tumor suppressor protein phosphatase 2A (PP2A) is frequently inactivated in human cancer and phosphorylation of its catalytic subunit (p-PP2A-C) at tyrosine-307 (Y307) has been described to inhibit this phosphatase. However, its molecular and clinical relevance in colorectal cancer (CRC) remains unclear. METHODS: p-PP2A-C Y307 was determined by immunoblotting in 7 CRC cell lines and 35 CRC patients. CRC cells were treated with the PP2A activator forskolin alone or combined with the PP2A inhibitor okadaic acid, 5-fluorouracil and oxaliplatin. We examined cell growth, colonosphere formation, caspase activity and AKT and ERK activation. RESULTS: PP2A-C was found hyperphosphorylated in CRC cell lines. Forskolin dephosphorylated and activated PP2A, impairing proliferation and colonosphere formation, and inducing activation of caspase 3/7 and changes in AKT and ERK phosphorylation. Moreover, forskolin showed additive effects with 5-fluorouracil and oxaliplatin treatments. Analysis of p-PP2A-C Y307 in primary tumors confirmed the presence of this alteration in a subgroup of CRC patients. CONCLUSIONS: Our data show that PP2A-C hyperphosphorylation is a frequent event that contributes to PP2A inhibition in CRC. Antitumoral effects of forskolin-mediated PP2A activation suggest that the analysis of p-PP2A-C Y307 status could be used to identify a subgroup of patients who would benefit from treatments based on PP2A activators.

PP2A inhibition as a novel therapeutic target in castration-resistant prostate cancer.

Protein phosphatase 2A (PP2A) is a well-known tumor suppressor frequently inhibited in human cancer. Alterations affecting PP2A subunits together with the deregulation of endogenous PP2A inhibitors such as CIP2A and SET have been described as contributing mechanisms to inactivate PP2A in prostate cancer. Moreover, recent findings highlight that functional inactivation of PP2A could represent a key event in the acquisition of castration-resistant phenotype and a novel molecular target with high impact at both clinical and therapeutic levels in prostate cancer.

Activation of the Tumor Suppressor PP2A Emerges as a Potential Therapeutic Strategy for Treating Prostate Cancer.

Protein phosphatase 2A (PP2A) is a tumor suppressor complex that has recently been reported as a novel and highly relevant molecular target in prostate cancer (PCa). However, its potential therapeutic value remains to be fully clarified. We treated PC-3 and LNCaP cell lines with the PP2A activators forskolin and FTY720 alone or combined with the PP2A inhibitor okadaic acid. We examined PP2A activity, cell growth, prostasphere formation, levels of PP2A phosphorylation, CIP2A and SET expression, and AKT and ERK activation. Interestingly, both forskolin and FTY720 dephosphorylated and activated PP2A, impairing proliferation and prostasphere formation and inducing changes in AKT and ERK phosphorylation. Moreover, FTY720 led to reduced CIP2A levels. Treatment with okadaic acid impaired PP2A activation thus demonstrating the antitumoral PP2A-dependent mechanism of action of both forskolin and FTY720. Levels of PP2A phosphorylation together with SET and CIP2A protein expression were studied in 24 PCa patients and both were associated with high Gleason scores and presence of metastatic disease. Altogether, our results suggest that PP2A inhibition could be involved in PCa progression, and the use of PP2A-activating drugs might represent a novel alternative therapeutic strategy for treating PCa patients.

Actionable mutational profiling in solid tumors using hybrid-capture-based next-generation sequencing in a real-world setting in Spain.

OBJECTIVE: This study aimed to describe the performance of a next-generation sequencing (NGS) panel for the detection of precise genomic alterations in cancer in Spanish clinical practice. The impact of tumor characteristics was evaluated on informative NGS and actionable mutation rates. MATERIALS AND METHODS: A cross-sectional study was conducted at the Fundación Jiménez Díaz University Hospital (May 2021-March 2022) where molecular diagnostic of 537 Formalin-Fixed Paraffin-Embedded (FFPE) tissue samples of diverse solid tumors (lung, colorectal, melanoma, gastrointestinal stromal, among others) was performed using AVENIO Tumor Tissue Targeted Kit. A descriptive analysis of the features of all samples was carried out. Multivariable logistic analysis was conducted to assess the impact of sample characteristics on NGS performance defined by informative results rate (for all tumors and for lung tumors), and on actionable mutations rate (for lung tumors only). RESULTS: AVENIO performance rate was 75.2% in all tumor samples and 75.3% in lung cancer samples, and the multivariable analysis showed that surgical specimens are most likely to provide informative results than diagnostic biopsies. Regarding the mutational findings, 727 pathogenic, likely pathogenic, or variant of unknown significance mutations were found in all tumor samples. Single nucleotide variant was the most common genomic alteration, both for all tumor samples (85.3% and 81.9% for all solid tumors and lung samples, respectively). In lung tumors, multivariable analysis showed that it is more likely to find actionable mutations from non-smokers and patients with adenocarcinoma, large cell, or undifferentiated histologies. CONCLUSION: This is the largest cohort-level study in Spain to profile the analyses of biopsy samples of different tumors using NGS in routine clinical practice. Our findings showed that the use of NGS routinely provides good rates of informative results and can improve tumor characterization and identify a greater number of actionable mutations.

Anaplastic large cell lymphomas with the 6p25.3 rearrangement are a heterogeneous group of tumours with a diverse molecular background.

ALK-negative anaplastic large cell lymphoma (ALCL) cases with 6p25.3 rearrangement are characterized by peculiar morphological and immunohistochemical features compare to 6p25.3-negative ALK-negative ALCL cases. A subgroup of 6p25.3-positive ALK-negative ALCL cases show the t(6,7) (p25.3;q32.3) rearrangement. Aims: To analyse the differences between 6p25.3-rearranged cases with and without t(6,7) (p25.3;q32.3). Using RNA-sequencing we studied a series of 17 samples showing 6p25.3-rearrangement, identified by FISH, consisting of seven systemic and eight primary cutaneous cases including two examples of secondary skin involvement by systemic ALCL. RNA-sequencing exclusively detected a translocation involving a gene in the 6p25.3 region (either IRF4 or DUSP22) in 7/14 cases (50%). In six of these seven cases the partner proved to be the LINC-PINT region in chromosome 7, while an EXOC2::DUSP22 rearrangement was found in one case. All cases but one were primary cutaneous ALCLs. They all were CD3 positive and BCL2 negative, while most of them expressed p-STAT3. On the contrary, cases without the t(6,7) (p25.3;q32.3) were mainly systemic (71%, 5/7) against just two pcALCL. In general, they lose CD3 (50% positive) and p-STAT3 (25% positive) expression, being all of them BCL2 positive. Moreover, in 60% of them other gene fusions were found. At the transcriptional level, they were characterized by the overexpression of TCF3 (TCF7L1/E2A), DLL3, CD58 and BCL2 genes 75%(6/8) of pcALCL with 6p25.3 rearrangement featured the so-called "biphasic morphologic pattern, which was not found in cutaneous involvement from systemic ALCL. 83% (5/6) of the pcALCL cases with the "biphasic morphologic pattern" showed the t(6,7) (p25.3;q32.3) rearrangement. ALK-negative ALCL cases with 6p25.3 rearrangement are a subgroup of tumours that are heterogeneous with respect to the presence or absence of the t(6,7) (p25.3;q32.3) translocation.

Plasmacytoid Dendritic Cell Dermatosis Associated to Myeloproliferative/Myelodysplastic Neoplasms.

Cutaneous lesions in the setting of myeloproliferative neoplasms and myelodysplastic syndromes are poorly understood. We report 6 patients with pruritic papular eruptions composed of mature T-lymphocytes with large clusters of CD123-positive cells. Double immunohistochemical studies demonstrated a lack of myeloid cell nuclear differentiation antigen in the CD123-positive cells, which expressed SPIB, confirming that they were mature plasmacytoid dendritic cells. Four patients were diagnosed with chronic myelomonocytic leukemia and 2 with myelodysplastic syndromes (AREB-I and myelodysplastic syndromes with 5q deletion, respectively). All patients had a long history of hematological alterations, mainly thrombocytopenia, preceding the cutaneous disorder. Nevertheless, the skin lesions developed in all cases coincidentally with either progression or full-establishment of their hematological disease. Most cutaneous lesions disappeared spontaneously or after corticosteroid treatment. Molecular studies performed in both bone marrow and cutaneous lesions in 2 patients demonstrated the same mutational profile, confirming the specific, neoplastic nature of these mature plasmacytoid dendritic cells-composed cutaneous lesions.

Redefining the high-grade B cell lymphoma with double/triple rearrangements of MYC and BCL2/BCL6 genes. Learning from a case report.

We report a patient initially diagnosed with a triple hit high-grade B cell lymphoma (HGBL-TH), in which further morphologic, immunohistochemical, and next-generation sequencing studies of subsequent specimens disclosed it to be a germinal center diffuse large B cell lymphoma (GC-DLBCL) with BCL2/BCL6 gene translocations, PVT1-deletion, and gain of MYC genes evolving from a previous follicular lymphoma. However, fluorescence in situ hybridization (FISH) studies with the break-apart probe for MYC gene showed a fusion and two separated signals (red and green, respectively) leading to the interpretation of MYC gene translocation and a false diagnosis of a TH-lymphoma, according to the recent WHO classification. Nevertheless, PVT1 deletion plus MYC gain/amplification has been described as a cause of the double-hi transcription profile. These data highlight the need for new criteria to identify these highly aggressive lymphomas.

Comprehensive Approach to Genomic and Immune Profiling: Insights of a Real-World Experience in Gynecological Tumors.

Gynecological cancer accounts for an elevated incidence worldwide requiring responsiveness regarding its care. The comprehensive genomic approach agrees with the classification of certain tumor types. We evaluated 49 patients with gynecological tumors undergoing high-throughput sequencing to explore whether identifying alterations in cancer-associated genes could characterize concrete histological subtypes. We performed immune examination and analyzed subsequent clinical impact. We found 220 genomic aberrations mostly distributed as single nucleotide variants (SNV, 77%). Only 3% were classified as variants of strong clinical significance in BRCA1 and BRCA2 of ovarian high-grade serous (HGSC) and uterine endometrioid carcinoma. TP53 and BRCA1 occurred in 72% and 28% of HGSC. Cervical squamous cell carcinoma was entirely HPV-associated and mutations occurred in PIK3CA (60%), as well as in uterine serous carcinoma (80%). Alterations were seen in PTEN (71%) and PIK3CA (60%) of uterine endometrioid carcinoma. Elevated programmed death-ligand 1 (PD-L1) was associated with high TILs. Either PD-L1 augmented in deficient mis-matched repair (MMR) proteins or POLE mutated cases when compared to a proficient MMR state. An 18% received genotype-guided therapy and a 4% immunotherapy. The description of tumor subtypes is plausible through high-throughput sequencing by recognizing clinically relevant alterations. Additional concomitant assessment of immune biomarkers identifies candidates for immunotherapy.