Emerging Role of Genetic Alterations Affecting Exosome Biology in Neurodegenerative Diseases.

The abnormal deposition of proteins in brain tissue is a common feature of neurodegenerative diseases (NDs) often accompanied by the spread of mutated proteins, causing neuronal toxicity. Exosomes play a fundamental role on their releasing in extracellular space after endosomal pathway activation, allowing to remove protein aggregates by lysosomal degradation or their inclusion into multivesicular bodies (MVBs), besides promoting cellular cross-talk. The emerging evidence of pathogenic mutations associated to ND susceptibility, leading to impairment of exosome production and secretion, opens a new perspective on the mechanisms involved in neurodegeneration. Recent findings suggest to investigate the genetic mechanisms regulating the different exosome functions in central nervous system (CNS), to understand their role in the pathogenesis of NDs, addressing the identification of diagnostic and pharmacological targets. This review aims to summarize the mechanisms underlying exosome biogenesis, their molecular composition and functions in CNS, with a specific focus on the recent findings invoking a defective exosome biogenesis as a common biological feature of the major NDs, caused by genetic alterations. Further definition of the consequences of specific genetic mutations on exosome biogenesis and release will improve diagnostic and pharmacological studies in NDs.

Genetic drivers of NF-κB deregulation in diffuse large B-cell lymphoma.

Diffuse large B cell lymphoma (DLBCL) is the most common form of B cell non-Hodgkin lymphoma worldwide and comprises a heterogeneous group of malignancies that originate from the malignant transformation of germinal center (GC) B cells. Over the past decade, significant improvement has been achieved in our understanding of the molecular pathogenesis underlying this disease, thanks in part to the implementation of powerful genomic technologies allowing genome-wide structural and functional analyses. These studies revealed the presence of multiple oncogenic alterations dysregulating signal transduction pathways that are normally required for the normal biology of the cells from which these tumors are derived. Among the pathways identified as recurrent targets of genetic lesions in DLBCL, NF-κB has emerged as a central player in the development and maintenance of this disease, particularly in the less curable, activated B cell (ABC)- like subtype. These lesions reveal vulnerabilities of the lymphoma cells that can be exploited for the design of more rationale therapeutic approaches. The purpose of this review is to summarize recent progresses in understanding the role of NF-κB deregulation in the pathogenesis of DLBCL, with emphasis on the genetic basis underlying its aberrant activation, in relationship to the normal biology of B lymphocytes, and the modelling of these lesions in the mouse.

Increased Th2 activity and diminished skin barrier function cooperate in allergic skin inflammation.

Atopic dermatitis (AD) is a chronic inflammatory skin disease induced by a complex interaction between susceptibility genes encoding skin barrier components and environmental allergen exposure that results in type 2 cytokine production. Although genetic lesions in either component can be risk factors for disease in patients, whether these pathways interact in the development of AD is not clear. To test this, we mated mice with T-cell specific expression of constitutively active Stat6 (Stat6VT) that spontaneously develop allergic skin inflammation with Flaky tail (Ft) mice that have mutations in Flg and Tmem79 genes that each affect skin barrier function. Our results demonstrate that over 90% of the Stat6VT transgenic mice carrying the Ft alleles (Stat6VTxFt-/- ) develop severe atopic dermatitis lesions by 3-5 months of age, compared with only 40% of Stat6VT mice that develop disease by 6-7 months of age. Further, histopathological analysis of skin tissues from Stat6VTxFt-/- mice revealed extensive thickening of the dermis with increased inflammatory infiltrates as compared with Stat6VT mice. Our study suggests that skin barrier defects and altered Th2 responses independently cooperate in the pathogenesis of allergic skin inflammation, similar to effects observed in patients with AD.

Feasibility to use whole-genome sequencing as a sole diagnostic method to detect genomic aberrations in pediatric B-cell acute lymphoblastic leukemia.

Introduction: The suitability of whole-genome sequencing (WGS) as the sole method to detect clinically relevant genomic aberrations in B-cell acute lymphoblastic leukemia (ALL) was investigated with the aim of replacing current diagnostic methods. Methods: For this purpose, we assessed the analytical performance of 150 bp paired-end WGS (90x leukemia/30x germline). A set of 88 retrospective B-cell ALL samples were selected to represent established ALL subgroups as well as ALL lacking stratifying markers by standard-of-care (SoC), so-called B-other ALL. Results: Both the analysis of paired leukemia/germline (L/N)(n=64) as well as leukemia-only (L-only)(n=88) detected all types of aberrations mandatory in the current ALLTogether trial protocol, i.e., aneuploidies, structural variants, and focal copy-number aberrations. Moreover, comparison to SoC revealed 100% concordance and that all patients had been assigned to the correct genetic subgroup using both approaches. Notably, WGS could allocate 35 out of 39 B-other ALL samples to one of the emerging genetic subgroups considered in the most recent classifications of ALL. We further investigated the impact of high (90x; n=58) vs low (30x; n=30) coverage on the diagnostic yield and observed an equally perfect concordance with SoC; low coverage detected all relevant lesions. Discussion: The filtration of the WGS findings with a short list of genes recurrently rearranged in ALL was instrumental to extract the clinically relevant information efficiently. Nonetheless, the detection of DUX4 rearrangements required an additional customized analysis, due to multiple copies of this gene embedded in the highly repetitive D4Z4 region. We conclude that the diagnostic performance of WGS as the standalone method was remarkable and allowed detection of all clinically relevant genomic events in the diagnostic setting of B-cell ALL.

Richter Syndrome: From Molecular Pathogenesis to Druggable Targets.

Richter syndrome (RS) represents the occurrence of an aggressive lymphoma, most commonly diffuse large B-cell lymphoma (DLBCL), in patients with chronic lymphocytic leukemia (CLL). Most cases of RS originate from the direct transformation of CLL, whereas 20% are de novo DLBCL arising as secondary malignancies. Multiple molecular mechanisms contribute to RS pathogenesis. B-cell receptor (BCR) overreactivity to multiple autoantigens is due to frequent stereotyped BCR configuration. Genetic lesions of TP53, CDKN2A, NOTCH1 and c-MYC deregulate DNA damage response, tumor suppression, apoptosis, cell cycle and proliferation. Hyperactivation of Akt and NOTCH1 signaling also plays a role. Altered expression of PD-1/PD-L1 and of other immune checkpoints leads to RS resistance to cytotoxicity exerted by T-cells. The molecular features of RS provide vulnerabilities for therapy. Targeting BCR signaling with noncovalent BTK inhibitors shows encouraging results, as does the combination of BCL2 inhibitors with chemoimmunotherapy. The association of immune checkpoint inhibitors with BCL2 inhibitors and anti-CD20 monoclonal antibodies is explored in early phase clinical trials with promising results. The development of patient-derived xenograft mice models reveals new molecular targets for RS, exemplified by ROR1. Although RS still represents an unmet medical need, understanding its biology is opening new avenues for precision medicine therapy.

PAX5 alterations in B-cell acute lymphoblastic leukemia.

PAX5, a master regulator of B cell development and maintenance, is one of the most common targets of genetic alterations in B-cell acute lymphoblastic leukemia (B-ALL). PAX5 alterations consist of copy number variations (whole gene, partial, or intragenic), translocations, and point mutations, with distinct distribution across B-ALL subtypes. The multifaceted functional impacts such as haploinsufficiency and gain-of-function of PAX5 depending on specific variants have been described, thereby the connection between the blockage of B cell development and the malignant transformation of normal B cells has been established. In this review, we provide the recent advances in understanding the function of PAX5 in orchestrating the development of both normal and malignant B cells over the past decade, with a focus on the PAX5 alterations shown as the initiating or driver events in B-ALL. Recent large-scale genomic analyses of B-ALL have identified multiple novel subtypes driven by PAX5 genetic lesions, such as the one defined by a distinct gene expression profile and PAX5 P80R mutation, which is an exemplar leukemia entity driven by a missense mutation. Although altered PAX5 is shared as a driver in B-ALL, disparate disease phenotypes and clinical outcomes among the patients indicate further heterogeneity of the underlying mechanisms and disturbed gene regulation networks along the disease development. In-depth mechanistic studies in human B-ALL and animal models have demonstrated high penetrance of PAX5 variants alone or concomitant with other genetic lesions in driving B-cell malignancy, indicating the altered PAX5 and deregulated genes may serve as potential therapeutic targets in certain B-ALL cases.

Major Molecular Signaling Pathways in Oral Cancer Associated With Therapeutic Resistance.

Oral cancer is a sub-category of head and neck cancers that primarily initiates in the oral cavity. The primary treatment option for oral cancer remains surgery but it is associated with massive disfigurement, inability to carry out normal oral functions, psycho-social stress and exhaustive rehabilitation. Other treatment options such as chemotherapy and radiotherapy have their own limitations in terms of toxicity, intolerance and therapeutic resistance. Immunological treatments to enhance the body's ability to recognize cancer tissue as a foreign entity are also being used but they are new and underdeveloped. Although substantial progress has been made in the treatment of oral cancer, its complex heterogeneous nature still needs to be explored, to elucidate the molecular basis for developing resistance to therapeutic agents and how to overcome it, with the aim of improving the chances of patients' survival and their quality of life. This review provides an overview of up-to-date information on the complex role of the major molecules and associated signaling, epigenetic changes, DNA damage repair systems, cancer stem cells and micro RNAs in the development of therapeutic resistance and treatment failure in oral cancer. We have also summarized the current strategies being developed to overcome these therapeutic challenges. This review will help not only researchers but also oral oncologists in the management of the disease and in developing new therapeutic modalities.

Targeting Kinase-activating Genetic Lesions to Improve Therapy of Pediatric Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) is the most common hematologic malignancy in children, characterized by an abnormal proliferation of immature lymphoid cells. Thanks to risk-adapted combination chemotherapy treatments currently used, survival at 5 years has reached 90%. ALL is a heterogeneous disease from a genetic point of view: patients' lymphoblasts may harbor in fact several chromosomal alterations, some of which have prognostic and therapeutic value. Of particular importance is the translocation t(9;22)(q34;q11.2) that leads to the formation of the BCR-ABL1 fusion gene, encoding a constitutively active chimeric tyrosine kinase (TK): BCR-ABL1 that is present in ~3% of pediatric ALL patients with B-immunophenotype and is associated with a poor outcome. This type of ALL is potentially treatable with specific TK inhibitors, such as imatinib. Recent studies have demonstrated the existence of a subset of BCR-ABL1 like leukemias (~10-15% of Bimmunophenotype ALL), whose blast cells have a gene expression profile similar to that of BCR-ABL1 despite the absence of t(9;22)(q34;q11.2). The precise pathogenesis of BCR-ABL1 like ALL is still to be defined, but they are mainly characterized by the activation of constitutive signal transduction pathways due to chimeric TKs different from BCR-ABL1. BCR-ABL1 like ALL patients represent a group with unfavorable outcome and are not identified by current risk criteria. In this review, we will discuss the design of targeted therapy for patients with BCR-ABL1 like ALL, which could consider TK inhibitors, and discuss innovative approaches suitable to identify the presence of patient's specific chimeric TK fusion genes, such as targeted locus amplification or proteomic biosensors.

Co-occurring KRAS mutation/LKB1 loss in non-small cell lung cancer cells results in enhanced metabolic activity susceptible to caloric restriction: an in vitro integrated multilevel approach.

BACKGROUND: Non-small-cell lung cancer (NSCLC) is a heterogeneous disease, with multiple different oncogenic mutations. Approximately 25-30% of NSCLC patients present KRAS mutations, which confer poor prognosis and high risk of tumor recurrence. About half of NSCLCs with activating KRAS lesions also have deletions or inactivating mutations in the serine/threonine kinase 11 (LKB1) gene. Loss of LKB1 on a KRAS-mutant background may represent a significant source of heterogeneity contributing to poor response to therapy. METHODS: Here, we employed an integrated multilevel proteomics, metabolomics and functional in-vitro approach in NSCLC H1299 isogenic cells to define their metabolic state associated with the presence of different genetic background. Protein levels were obtained by label free and single reaction monitoring (SRM)-based proteomics. The metabolic state was studied coupling targeted and untargeted mass spectrometry (MS) strategy. In vitro metabolic dependencies were evaluated using 2-deoxy glucose (2-DG) treatment or glucose/glutamine nutrient limitation. RESULTS: Here we demonstrate that co-occurring KRAS mutation/LKB1 loss in NSCLC cells allowed efficient exploitation of glycolysis and oxidative phosphorylation, when compared to cells with each single oncologic genotype. The enhanced metabolic activity rendered the viability of cells with both genetic lesions susceptible towards nutrient limitation. CONCLUSIONS: Co-occurrence of KRAS mutation and LKB1 loss in NSCLC cells induced an enhanced metabolic activity mirrored by a growth rate vulnerability under limited nutrient conditions relative to cells with the single oncogenetic lesions. Our results hint at the possibility that energy stress induced by calorie restriction regimens may sensitize NSCLCs with these co-occurring lesions to cytotoxic chemotherapy.

Gene mutations and actionable genetic lesions in mantle cell lymphoma.

Mutations and epigenetic alterations are key events in transforming normal cells to cancer cells. Mantle cell lymphoma (MCL), a non-Hodgkin's lymphoma of the B-cell, is an aggressive malignancy with poor prognosis especially for those patients who are resistant to the frontline drugs. There is a great need to describe the molecular basis and mechanism of drug resistance in MCL to develop new strategies for treatment. We reviewed frequent somatic mutations and mutations involving the B-cell pathways in MCL and discussed clinical trials that attempted to disrupt these gene pathways and/or epigenetic events. Recurrent gene mutations were discussed in the light of prognostic and therapeutic opportunity and also the challenges of targeting these lesions. Mutations in the ATM, CCND1, TP53, MLL2, TRAF2 and NOTCH1 were most frequently encountered in mantle cell lymphoma. Translational models should be built that would assess mutations longitudinally to identify important compensatory, pro-survival and anti-apoptic pathways and actionable genetic targets.

Modern treatment in chronic lymphocytic leukemia: impact on survival and efficacy in high-risk subgroups.

Treatment of chronic lymphocytic leukemia (CLL) has dramatically changed over the last years, with significant improvement in overall survival (OS) and increased efficacy in genetically defined "high-risk" disease. Besides prospective clinical trials usually enrolling young and fit patients, retrospective studies were performed comparing the outcome of patients belonging to different age groups and showing longer survival in patients diagnosed in the most recent periods. In patients younger than 70 years the 10-year relative survival was 43-53% in the 1980s as compared with 59-63% in the 2000s. Likewise, the 10-year relative survival in patients >70 years was 22-42% in the 1980s and 46-55% in the 2000s. Improved outcome derived in part by the introduction of effective regimens in genetically defined "high-risk" disease (i.e., 17p-, 11q-, TP53, NOTCH1, SF3B1 mutations), especially in the younger and/or fit patients. The unfavorable prognostic significance of 11q- was overcome by chemoimmunotherapy. High-dose steroids with anti-CD52 appeared to improve the response rate in 17p-/TP53 mutated cases and allogeneic transplantation achieved prolonged disease control irrespective of high-risk disease. Further improvement is being generated by the new anti-CD20 obinutuzumab in the elderly and by mechanism-based treatment using kinase-targeting agents or anti-BCL2 molecules yielding high-response rate and impressive progression-free survival in the chemorefractory setting as well as in previously untreated patients.