BD SurePath Direct to Slide (DTS) cervical cytology: Migrating the benefits of liquid-based cytology to low-resource settings.

OBJECTIVES: The benefits of liquid-based cytology (LBC) in routine cervical cancer screening are often associated with the availability of instrumented platforms and economic considerations. A low-cost alternative to LBC in low-volume settings remains an unmet need. METHODS: A multisite evaluation of the BD SurePath (SurePath) LBC Direct to Slide (DTS) method was conducted. The DTS preparations were evaluated across 3 sites. Cytology features for DTS preparation included predetermined thresholds for total cellularity, cell distribution, cellular preservation, and stain quality. Rare event detection was evaluated using SiHa cells spiked into pools from negative cytology specimens. Concordance between Bethesda classification results was evaluated for SurePath LBC and DTS methods using routinely collected SurePath specimens in a split-sample study design. RESULTS: The DTS specimens met criteria for total cellularity, cell distribution, cellular preservation, and stain quality in more than 98% of all cases. Rare event detection was observed with an average detection of 5 SiHa cells per 2 mL of specimen. Concordant cervical cytology classifications were observed between SurePath LBC and DTS methods. CONCLUSIONS: The results demonstrate that the DTS process is suitable for routine cervical cytology evaluation. The procedure is reproducible and detected abnormal cervical cells in concordance with standard SurePath LBC preparation.

A comprehensive AI model development framework for consistent Gleason grading.

BACKGROUND: Artificial Intelligence(AI)-based solutions for Gleason grading hold promise for pathologists, while image quality inconsistency, continuous data integration needs, and limited generalizability hinder their adoption and scalability. METHODS: We present a comprehensive digital pathology workflow for AI-assisted Gleason grading. It incorporates A!MagQC (image quality control), A!HistoClouds (cloud-based annotation), Pathologist-AI Interaction (PAI) for continuous model improvement, Trained on Akoya-scanned images only, the model utilizes color augmentation and image appearance migration to address scanner variations. We evaluate it on Whole Slide Images (WSI) from another five scanners and conduct validations with pathologists to assess AI efficacy and PAI. RESULTS: Our model achieves an average F1 score of 0.80 on annotations and 0.71 Quadratic Weighted Kappa on WSIs for Akoya-scanned images. Applying our generalization solution increases the average F1 score for Gleason pattern detection from 0.73 to 0.88 on images from other scanners. The model accelerates Gleason scoring time by 43% while maintaining accuracy. Additionally, PAI improve annotation efficiency by 2.5 times and led to further improvements in model performance. CONCLUSIONS: This pipeline represents a notable advancement in AI-assisted Gleason grading for improved consistency, accuracy, and efficiency. Unlike previous methods limited by scanner specificity, our model achieves outstanding performance across diverse scanners. This improvement paves the way for its seamless integration into clinical workflows.

Gleason grading is a well-accepted diagnostic standard to assess the severity of prostate cancer in patients' tissue samples, based on how abnormal the cells in their prostate tumor look under a microscope. This process can be complex and time-consuming. We explore how artificial intelligence (AI) can help pathologists perform Gleason grading more efficiently and consistently. We build an AI-based system which automatically checks image quality, standardizes the appearance of images from different equipment, learns from pathologists' feedback, and constantly improves model performance. Testing shows that our approach achieves consistent results across different equipment and improves efficiency of the grading process. With further testing and implementation in the clinic, our approach could potentially improve prostate cancer diagnosis and management.

MicroRNA expression signature as a biomarker in the diagnosis of nodal T-cell lymphomas.

BACKGROUND: The diagnosis of T-cell lymphomas is typically established through a multiparameter approach that combines clinical, morphologic, immunophenotypic, and genetic features, utilizing a variety of histopathologic and molecular techniques. However, accurate diagnosis of such lymphomas and distinguishing them from reactive lymph nodes remains challenging due to their low prevalence and heterogeneous features, hence limiting the confidence of pathologists. We investigated the use of microRNA (miRNA) expression signatures as an adjunctive tool in the diagnosis and classification of T-cell lymphomas that involve lymph nodes. This study seeks to distinguish reactive lymph nodes (RLN) from two types of frequently occurring nodal T-cell lymphomas: nodal T-follicular helper (TFH) cell lymphomas (nTFHL) and peripheral T-cell lymphomas, not otherwise specified (nPTCL). METHODS: From the formalin-fixed paraffin-embedded (FFPE) samples from a cohort of 88 subjects, 246 miRNAs were quantified and analyzed by differential expression. Two-class logistic regression and random forest plot models were built to distinguish RLN from the nodal T-cell lymphomas. Gene set enrichment analysis was performed on the target genes of the miRNA to identify pathways and transcription factors that may be regulated by the differentially expressed miRNAs in each subtype. RESULTS: Using logistic regression analysis, we identified miRNA signatures that can distinguish RLN from nodal T-cell lymphomas (AUC of 0.92 ± 0.05), from nTFHL (AUC of 0.94 ± 0.05) and from nPTCL (AUC of 0.94 ± 0.08). Random forest plot modelling was also capable of distinguishing between RLN and nodal T-cell lymphomas, but performed worse than logistic regression. However, the miRNA signatures are not able to discriminate between nTFHL and nPTCL, owing to large similarity in miRNA expression patterns. Bioinformatic analysis of the gene targets of unique miRNA expression revealed the enrichment of both known and potentially understudied signalling pathways and genes in such lymphomas. CONCLUSION: This study suggests that miRNA biomarkers may serve as a promising, cost-effective tool to aid the diagnosis of nodal T-cell lymphomas, which can be challenging. Bioinformatic analysis of differentially expressed miRNAs revealed both relevant or understudied signalling pathways that may contribute to the progression and development of each T-cell lymphoma entity. This may help us gain further insight into the biology of T-cell lymphomagenesis.

Identification of PRDX5 as A Target for The Treatment of Castration-Resistant Prostate Cancer.

Treatment of castration-resistant prostate cancer (CRPC) is a long-standing clinical challenge. Traditionally, CRPC drugs work by either reducing dihydrotestosterone biosynthesis or blocking androgen receptor (AR) signaling. Here it is demonstrated that AR inhibitor treatment gives rise to a drug-tolerant persister (DTP) state. The thioredoxin/peroxiredoxin pathway is up-regulated in DTP cells. Peroxiredoxin 5 (PRDX5) promotes AR inhibitor resistance and CRPC development. Inhibition of PRDX5 suppresses DTP cell proliferation in culture, dampens CRPC development in animal models, and stabilizes PSA progression and metastatic lesions in patients. Therefore, the study provides a novel mechanism and potential target for the management of castration-resistant prostate cancer.

Dynamic altruistic cooperation within breast tumors.

BACKGROUND: Social behaviors such as altruism, where one self-sacrifices for collective benefits, critically influence an organism's survival and responses to the environment. Such behaviors are widely exemplified in nature but have been underexplored in cancer cells which are conventionally seen as selfish competitive players. This multidisciplinary study explores altruism and its mechanism in breast cancer cells and its contribution to chemoresistance. METHODS: MicroRNA profiling was performed on circulating tumor cells collected from the blood of treated breast cancer patients. Cancer cell lines ectopically expressing candidate miRNA were used in co-culture experiments and treated with docetaxel. Ecological parameters like relative survival and relative fitness were assessed using flow cytometry. Functional studies and characterization performed in vitro and in vivo include proliferation, iTRAQ-mass spectrometry, RNA sequencing, inhibition by small molecules and antibodies, siRNA knockdown, CRISPR/dCas9 inhibition and fluorescence imaging of promoter reporter-expressing cells. Mathematical modeling based on evolutionary game theory was performed to simulate spatial organization of cancer cells. RESULTS: Opposing cancer processes underlie altruism: an oncogenic process involving secretion of IGFBP2 and CCL28 by the altruists to induce survival benefits in neighboring cells under taxane exposure, and a self-sacrificial tumor suppressive process impeding proliferation of altruists via cell cycle arrest. Both processes are regulated concurrently in the altruists by miR-125b, via differential NF-κB signaling specifically through IKKß. Altruistic cells persist in the tumor despite their self-sacrifice, as they can regenerate epigenetically from non-altruists via a KLF2/PCAF-mediated mechanism. The altruists maintain a sparse spatial organization by inhibiting surrounding cells from adopting the altruistic fate via a lateral inhibition mechanism involving a GAB1-PI3K-AKT-miR-125b signaling circuit. CONCLUSIONS: Our data reveal molecular mechanisms underlying manifestation, persistence and spatial spread of cancer cell altruism. A minor population behave altruistically at a cost to itself producing a collective benefit for the tumor, suggesting tumors to be dynamic social systems governed by the same rules of cooperation in social organisms. Understanding cancer cell altruism may lead to more holistic models of tumor evolution and drug response, as well as therapeutic paradigms that account for social interactions. Cancer cells constitute tractable experimental models for fields beyond oncology, like evolutionary ecology and game theory.

WNTinib is a multi-kinase inhibitor with specificity against ß-catenin mutant hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. ß-Catenin (CTNNB1)-mutated HCC represents 30% of cases of the disease with no precision therapeutics available. Using chemical libraries derived from clinical multi-kinase inhibitor (KI) scaffolds, we screened HCC organoids to identify WNTinib, a KI with exquisite selectivity in CTNNB1-mutated human and murine models, including patient samples. Multiomic and target engagement analyses, combined with rescue experiments and in vitro and in vivo efficacy studies, revealed that WNTinib is superior to clinical KIs and inhibits KIT/mitogen-activated protein kinase (MAPK) signaling at multiple nodes. Moreover, we demonstrate that reduced engagement on BRAF and p38α kinases by WNTinib relative to several multi-KIs is necessary to avoid compensatory feedback signaling-providing a durable and selective transcriptional repression of mutant ß-catenin/Wnt targets through nuclear translocation of the EZH2 transcriptional repressor. Our studies uncover a previously unknown mechanism to harness the KIT/MAPK/EZH2 pathway to potently and selectively antagonize CTNNB1-mutant HCC with an unprecedented wide therapeutic index.

Patterns of Oncogene Coexpression at Single-Cell Resolution Influence Survival in Lymphoma.

Cancers often overexpress multiple clinically relevant oncogenes, but it is not known if combinations of oncogenes in cellular subpopulations within a cancer influence clinical outcomes. Using quantitative multispectral imaging of the prognostically relevant oncogenes MYC, BCL2, and BCL6 in diffuse large B-cell lymphoma (DLBCL), we show that the percentage of cells with a unique combination MYC+BCL2+BCL6- (M+2+6-) consistently predicts survival across four independent cohorts (n = 449), an effect not observed with other combinations including M+2+6+. We show that the M+2+6- percentage can be mathematically derived from quantitative measurements of the individual oncogenes and correlates with survival in IHC (n = 316) and gene expression (n = 2,521) datasets. Comparative bulk/single-cell transcriptomic analyses of DLBCL samples and MYC/BCL2/BCL6-transformed primary B cells identify molecular features, including cyclin D2 and PI3K/AKT as candidate regulators of M+2+6- unfavorable biology. Similar analyses evaluating oncogenic combinations at single-cell resolution in other cancers may facilitate an understanding of cancer evolution and therapy resistance. SIGNIFICANCE: Using single-cell-resolved multiplexed imaging, we show that selected subpopulations of cells expressing specific combinations of oncogenes influence clinical outcomes in lymphoma. We describe a probabilistic metric for the estimation of cellular oncogenic coexpression from IHC or bulk transcriptomes, with possible implications for prognostication and therapeutic target discovery in cancer. This article is highlighted in the In This Issue feature, p. 1027.

Targeting RNA Exonuclease XRN1 Potentiates Efficacy of Cancer Immunotherapy.

Despite the remarkable clinical responses achieved with immune checkpoint blockade therapy, the response rate is relatively low and only a subset of patients can benefit from the treatment. Aberrant RNA accumulation can mediate IFN signaling and stimulate an immune response, suggesting that targeting RNA decay machinery might sensitize tumor cells to immunotherapy. With this in mind, we identified an RNA exoribonuclease, XRN1, as a potential therapeutic target to suppress RNA decay and stimulate antitumor immunity. Silencing of XRN1 suppressed tumor growth in syngeneic immunocompetent mice and potentiated immunotherapy efficacy, while silencing of XRN1 alone did not affect tumor growth in immunodeficient mice. Mechanistically, XRN1 depletion activated IFN signaling and the viral defense pathway; both pathways play determinant roles in regulating immune evasion. Aberrant RNA-sensing signaling proteins (RIG-I/MAVS) mediated the expression of IFN genes, as depletion of each of them blunted the elevation of antiviral/IFN signaling in XRN1-silenced cells. Analysis of pan-cancer CRISPR-screening data indicated that IFN signaling triggered by XRN1 silencing is a common phenomenon, suggesting that the effect of XRN1 silencing may be extended to multiple types of cancers. Overall, XRN1 depletion triggers aberrant RNA-mediated IFN signaling, highlighting the importance of the aberrant RNA-sensing pathway in regulating immune responses. These findings provide the molecular rationale for developing XRN1 inhibitors and exploring their potential clinical application in combination with cancer immunotherapy. SIGNIFICANCE: Targeting XRN1 activates an intracellular innate immune response mediated by RNA-sensing signaling and potentiates cancer immunotherapy efficacy, suggesting inhibition of RNA decay machinery as a novel strategy for cancer treatment.

Tissue-Specific microRNA Expression Profiling to Derive Novel Biomarkers for the Diagnosis and Subtyping of Small B-Cell Lymphomas.

Accurate diagnosis of the most common histological subtypes of small B-cell lymphomas is challenging due to overlapping morphological features and limitations of ancillary testing, which involves a large number of immunostains and molecular investigations. In addition, a common diagnostic challenge is to distinguish reactive lymphoid hyperplasia that do not require additional stains from such lymphomas that need ancillary investigations. We investigated if tissue-specific microRNA (miRNA) expression may provide potential biomarkers to improve the pathology diagnostic workflow. This study seeks to distinguish reactive lymphoid proliferation (RL) from small B-cell lymphomas, and to further distinguish the four main subtypes of small B-cell lymphomas. Two datasets were included: a discovery cohort (n = 100) to screen for differentially expressed miRNAs and a validation cohort (n = 282) to develop classification models. The models were evaluated for accuracy in subtype prediction. MiRNA gene set enrichment was also performed to identify differentially regulated pathways. 306 miRNAs were detected and quantified, resulting in 90-miRNA classification models from which smaller panels of miRNAs biomarkers with good accuracy were derived. Bioinformatic analysis revealed the upregulation of known and other potentially relevant signaling pathways in such lymphomas. In conclusion, this study suggests that miRNA expression profiling may serve as a promising tool to aid the diagnosis of common lymphoid lesions.

Downregulation of hsa-miR-548d-3p and overexpression of HOXA9 in diffuse large B-cell lymphoma patients and the risk of R-CHOP chemotherapy resistance and disease progression.

INTRODUCTION: Routine categorization of DLBCL patients into GCB and non-GCB groups by Hans' criteria could not accurately predict chemotherapy resistance and disease progression in patients treated with standard R-CHOP therapy. There is a need to identify better biomarker predictors to enhance assisted selection of chemotherapy regimens for DLBCL patients. AIM OF THE STUDY: To identify dysregulated miRNAs and mRNAs that are predictive of resistance to R-CHOP chemotherapy or disease progression in patients with DLBCL. METHODS: miRNA and mRNA profiling were performed on archival FFPE samples of the DLBCL patients. miRabel and miRNet bioinformatic tools were applied to determine experimental validated miRNA-mRNA target interaction. The significance of the genomic predictive values was assessed using adjusted odds ratios (AOR) and 95% confidence intervals (CI). RESULTS: 19/36 were R-CHOP therapy-resistant whilst 17/36 were R-CHOP therapy-sensitive. Ten dysregulated miRNAs and 12 dysregulated mRNAs were identified in therapy-resistant DLBCL patients. These dysregulated miRNAs and mRNA cause therapy resistance and disease progression in DLBCL patients, most likely via upregulation of the anti-apoptotic protein bcl2, activation of the JAK/STAT signalling pathway and dysregulation of p53 pathway. Downregulation of hsa-miR-548d-3p and overexpression of HOXA9 mRNA were significantly associated with therapy resistance and disease progression in DLBCL patients [hsa-miR-548d-3p AOR: 0.258, 95%CI: 0.097-0.684, p = 0.006]. CONCLUSION: DLBCL patients with downregulation of hsa-miR-548d-3p and overexpression of HOXA9 mRNA are more likely to experience R-CHOP therapy resistance and disease progression.

Cellular Origins and Pathogenesis of Gastrointestinal NK- and T-Cell Lymphoproliferative Disorders.

The intestinal immune system, which must ensure appropriate immune responses to both pathogens and commensal microflora, comprises innate lymphoid cells and various T-cell subsets, including intra-epithelial lymphocytes (IELs). An example of innate lymphoid cells is natural killer cells, which may be classified into tissue-resident, CD56bright NK-cells that serve a regulatory function and more mature, circulating CD56dim NK-cells with effector cytolytic properties. CD56bright NK-cells in the gastrointestinal tract give rise to indolent NK-cell enteropathy and lymphomatoid gastropathy, as well as the aggressive extranodal NK/T cell lymphoma, the latter following activation by EBV infection and neoplastic transformation. Conventional CD4+ TCRαß+ and CD8αß+ TCRαß+ T-cells are located in the lamina propria and the intraepithelial compartment of intestinal mucosa as type 'a' IELs. They are the putative cells of origin for CD4+ and CD8+ indolent T-cell lymphoproliferative disorders of the gastrointestinal tract and intestinal T-cell lymphoma, NOS. In addition to such conventional T-cells, there are non-conventional T-cells in the intra-epithelial compartment that express CD8αα and innate lymphoid cells that lack TCRs. The central feature of type 'b' IELs is the expression of CD8αα homodimers, seen in monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL), which primarily arises from both CD8αα+ TCRαß+ and CD8αα+ TCRγδ+ IELs. EATL is the other epitheliotropic T-cell lymphoma in the GI tract, a subset of which arises from the expansion and reprograming of intracytoplasmic CD3+ innate lymphoid cells, driven by IL15 and mutations of the JAK-STAT pathway.

IRF4 drives clonal evolution and lineage choice in a zebrafish model of T-cell lymphoma.

IRF4 is a master regulator of immunity and is also frequently overexpressed in mature lymphoid neoplasms. Here, we demonstrate the oncogenicity of IRF4 in vivo, its potential effects on T-cell development and clonal evolution using a zebrafish model. IRF4-transgenic zebrafish develop aggressive tumors with massive infiltration of abnormal lymphocytes that spread to distal organs. Many late-stage tumors are mono- or oligoclonal, and tumor cells can expand in recipient animals after transplantation, demonstrating their malignancy. Mutation of p53 accelerates tumor onset, increases penetrance, and results in tumor heterogeneity. Surprisingly, single-cell RNA-sequencing reveals that the majority of tumor cells are double-negative T-cells, many of which express tcr-γ that became dominant as the tumors progress, whereas double-positive T-cells are largely diminished. Gene expression and epigenetic profiling demonstrates that gata3, mycb, lrrn1, patl1 and psip1 are specifically activated in tumors, while genes responsible for T-cell differentiation including id3 are repressed. IRF4-driven tumors are sensitive to the BRD inhibitor.

Immune pathway upregulation and lower genomic instability distinguish EBV-positive nodal T/NK-cell lymphoma from ENKTL and PTCL-NOS.

Primary Epstein-Barr virus (EBV)-positive nodal T/NK-cell lymphoma (PTCL-EBV) is a poorly understood disease which shows features resembling extranodal NK/T-cell lymphoma (ENKTL) and is currently not recognized as a distinct entity but categorized as a variant of primary T-cell lymphoma not otherwise specified (PTCL-NOS). Herein, we analyzed copynumber aberrations (n=77) with a focus on global measures of genomic instability and homologous recombination deficiency and performed gene expression (n=84) and EBV miRNA expression (n=24) profiling as well as targeted mutational analysis (n=16) to further characterize PTCL-EBV in relation to ENKTL and PTCL-NOS. Multivariate analysis revealed that patients with PTCL-EBV had a significantly worse outcome compared to patients with PTCL-NOS (P=0.002) but not to those with ENKTL. Remarkably, PTCL-EBV exhibited significantly lower genomic instability and homologous recombination deficiency scores compared to ENKTL and PTCL-NOS. Gene set enrichment analysis revealed that many immune-related pathways, interferon α/γ response, and IL6_JAK_STAT3 signaling were significantly upregulated in PTCLEBV and correlated with lower genomic instability scores. We also identified that NFκB-associated genes, BIRC3, NFKB1 (P50) and CD27, and their proteins are upregulated in PTCL-EBV. Most PTCL-EBV demonstrated a type 2 EBV latency pattern and, strikingly, exhibited downregulated expression of most EBV miRNA compared to ENKTL and their target genes were also enriched in immune-related pathways. PTCL-EBV also showed frequent mutations of TET2, PIK3CD and STAT3, and are characterized by microsatellite stability. Overall, poor outcome, low genomic instability, upregulation of immune pathways and downregulation of EBV miRNA are distinctive features of PTCL-EBV. Our data support the concept that PTCL-EBV could be considered as a distinct entity, provide novel insights into the pathogenesis of the disease and offer potential new therapeutic targets for this tumor.

Effective Killing of Acute Myeloid Leukemia by TIM-3 Targeted Chimeric Antigen Receptor T Cells.

Acute myeloid leukemia (AML) is an aggressive disease with poor outcomes, overwhelmingly due to relapse. Minimal residual disease (MRD), defined as the persistence of leukemic cells after chemotherapy treatment, is thought to be the major cause of relapse. The origins of relapse in AML have been traced to rare therapy-resistant leukemic stem cells (LSCs) that are already present at diagnosis. Effective treatment strategies for long-term remission are lacking, as it has been difficult to eliminate LSCs with conventional therapy. Here, we proposed a new approach based on the chimeric antigen receptor (CAR)-directed T lymphocytes, targeting T-cell immunoglobulin, and mucin domain 3 (TIM-3) to treat MRD in patients with AML. TIM-3 is selected as the target because it is highly expressed on AML blasts and LSCs in most subtypes regardless of the patient's genetic characteristics and treatment course. Moreover, it is absent in the normal hematopoietic stem cells, granulocytes, naïve lymphocytes, and most normal nonhematopoietic tissues. Using a naïve human Fab phage display library, we isolated an anti-human TIM-3 antibody and designed a second-generation anti-TIM-3. Our anti-TIM-3 CAR T cells exhibit potent antileukemic activity against AML cell lines and primary AML blasts, and in the mouse models. More importantly, we demonstrate efficient killing of the primary LSCs directly isolated from the patients. Hence, eradication of the LSCs present in the MRD by anti-TIM-3 CAR T-cell therapy following the first-line treatment may improve the clinical outcomes of patients with AML.

Diagnostic approach to T- and NK-cell lymphoproliferative disorders in the gastrointestinal tract.

Most gastrointestinal NK and T cell lymphomas are aggressive in behavior, although in recent years a subset of indolent lymphoproliferative disorders have been described, which must be distinguished from their more malignant mimics. Intestinal T-cell lymphomas may arise from intra-epithelial lymphocytes and display epitheliotropism, such as enteropathy-associated T-cell lymphoma and monomorphic epitheliotropic intestinal T-cell lymphoma. They are both aggressive in behavior but differ in their clinic-pathological features. On the other hand, intra-epithelial lymphocytes are not prominent in intestinal T-cell lymphoma, NOS, which is a diagnosis of exclusion and probably represents a heterogeneous group of entities. Indolent lymphoproliferative disorders of NK- and T-cells of both CD8 and CD4 subsets share a chronic, recurring clinical course but display differences from each other. CD8+ T-cell lymphoproliferative disorder of GI tract has a low proliferative fraction and does not progress nor undergo large cell transformation. Whilst NK-cell enteropathy runs an indolent clinical course, it may display a high proliferation fraction. On the other hand, CD4+ indolent T-cell lymphoproliferative disorder displays variable proliferation rates and may progress or transform after a number of years. In Asia and South America, it is not uncommon to see involvement of the gastrointestinal tract by EBV-associated extranodal NK/T cell lymphoma, nasal type, which must be distinguished from NK cell enteropathy and EBV-associated mucocutaneous ulcers.

T-Cell Lymphoma Clonality by Copy Number Variation Analysis of T-Cell Receptor Genes.

T-cell lymphomas arise from a single neoplastic clone and exhibit identical patterns of deletions in T-cell receptor (TCR) genes. Whole genome sequencing (WGS) data represent a treasure trove of information for the development of novel clinical applications. However, the use of WGS to identify clonal T-cell proliferations has not been systematically studied. In this study, based on WGS data, we identified monoclonal rearrangements (MRs) of T-cell receptors (TCR) genes using a novel segmentation algorithm and copy number computation. We evaluated the feasibility of this technique as a marker of T-cell clonality using T-cell lymphomas (TCL, n = 44) and extranodal NK/T-cell lymphomas (ENKTLs, n = 20), and identified 98% of TCLs with one or more TCR gene MRs, against 91% detected using PCR. TCR MRs were absent in all ENKTLs and NK cell lines. Sensitivity-wise, this platform is sufficiently competent, with MRs detected in the majority of samples with tumor content under 25% and it can also distinguish monoallelic from biallelic MRs. Understanding the copy number landscape of TCR using WGS data may engender new diagnostic applications in hematolymphoid pathology, which can be readily adapted to the analysis of B-cell receptor loci for B-cell clonality determination.

Pharmacological significance of the non-canonical NF-κB pathway in tumorigenesis.

The understanding of the impact of the non-canonical NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) pathway in several human diseases including autoimmune, inflammatory and cancers has been on the rise. This pathway induces the expression of several important genes involved in diverse biological processes. Though progress has been made in understanding the activation, regulation and biological functions of the non-canonical NF-κB signaling mechanism, no specific drug has been approved to target NF-κB inducing kinase (NIK), the key signaling molecule in this pathway. The inhibition of NIK can serve as a potential therapeutic strategy for various ailments, especially for the treatment of different types of human cancers. There are other targetable downstream molecules in this pathway as well. This review highlights the possible role of the non-canonical NF-κB pathway in normal physiology as well as in different cancers and discusses about various pharmacological strategies to modulate the activation of this pathway.

Targeting N-myristoylation for therapy of B-cell lymphomas.

Myristoylation, the N-terminal modification of proteins with the fatty acid myristate, is critical for membrane targeting and cell signaling. Because cancer cells often have increased N-myristoyltransferase (NMT) expression, NMTs were proposed as anti-cancer targets. To systematically investigate this, we performed robotic cancer cell line screens and discovered a marked sensitivity of hematological cancer cell lines, including B-cell lymphomas, to the potent pan-NMT inhibitor PCLX-001. PCLX-001 treatment impacts the global myristoylation of lymphoma cell proteins and inhibits early B-cell receptor (BCR) signaling events critical for survival. In addition to abrogating myristoylation of Src family kinases, PCLX-001 also promotes their degradation and, unexpectedly, that of numerous non-myristoylated BCR effectors including c-Myc, NFκB and P-ERK, leading to cancer cell death in vitro and in xenograft models. Because some treated lymphoma patients experience relapse and die, targeting B-cell lymphomas with a NMT inhibitor potentially provides an additional much needed treatment option for lymphoma.