Molecular Characterization Of Disease Research Articles

Therapeutic Targeting of TIM-4-L with Engineered T Cells for Acute Myeloid Leukemia.

Disruption of lipid bilayer asymmetry is a common feature observed in cancer cells and offers novel routes for therapeutic targeting. We used the natural immune receptor TIM-4 to interrogate for loss of plasma membrane phospholipid polarity in primary acute myelogenous leukemia (AML) samples and evaluated the anti-leukemic activity of TIM-4-L-directed T-cell therapy in preclinical AML models. We performed FACS analysis on 33 primary AML bone marrow specimens and correlated TIM-4-L expression frequency and intensity with molecular disease characteristics. Using Kasumi-1 and MV-4-11 AML cell lines, we further tested the anti-leukemic effects of TIM-4-L-directed engineered T cells in vitro and in vivo. We found that 86% of untreated AML blasts displayed upregulation of cell surface TIM-4-L. These observations were agnostic to AML genetic classification, as samples with mutations in TP53, ASXL1, and RUNX1 displayed TIM-4-L upregulation similar to that seen in favorable and intermediate subtypes. TIM-4-L dysregulation was also stably present in AML cell lines. To evaluate the potential of targeting upregulated TIM-4-L with adoptive T-cell therapy, we constructed TIM-4-L-directed engineered T cells, which demonstrated potent anti-leukemic effects, effectively eliminating AML cell lines with a range of endogenous TIM-4-L expression levels both in vitro and in vivo. These results highlight TIM-4-L as a highly prevalent target on AML across a range of genetic classifications and novel target for T-cell-based therapy in AML. Further investigations into the role of TIM-4-L in AML pathogenesis and its potential as an anti-leukemic target for clinical development are warranted.

DOP54 Guselkumab and golimumab combination induction therapy in Ulcerative Colitis results in early local tissue healing that is sustained through guselkumab maintenance therapy

Abstract Background Combination induction therapy with guselkumab (GUS), an interleukin (IL)-23p19 subunit antagonist, and golimumab (GOL), a tumor necrosis factor (TNFα) antagonist, induced higher rates of clinical remission, endoscopic, and histologic outcomes than either monotherapy at Week (WK)12 in patients (pts) with ulcerative colitis (VEGA NCT03662542). Data through WK38 suggested the continued benefit of combination induction even after a transition to GUS monotherapy at WK12. We explored early molecular changes in colon tissue in a subset of pts at WK4 to define mechanistic contributions of each monotherapy and combination; these parameters were evaluated again at WK38 to assess potential carry-over of efficacy. Methods Colon biopsies were obtained at baseline (n=195), WK4 (n=42 substudy), and WK38 (n=172). Transcriptional profiles were generated with RNA sequencing (RNAseq). Gene correlation network analysis was applied in conjunction with publicly available single cell RNAseq data to define biologically relevant bulk and cell type-specific transcriptional modules associated with molecular features of disease. Gene set variation analysis (GSVA) was used to quantitatively assess changes in biologic modules with treatment. Results At WK4, combination induction (n=10) showed significant (p<0.05) decreases in molecular features compared to GUS (n=19) and GOL (n=13), including transcriptomic modules representing the IL-23 pathway, interferon response, and inflammatory epithelial and myeloid transcriptional states associated with endoscopic improvement at WK4 (Mayo endoscopy subscore of 0 or 1) (combination 5/11, GOL 1/13; GUS 2/19). Reduction (p<0.05) in inflammatory modules persisted through WK38 with combination vs monotherapy induction. Module changes between treatments at WK4 indicated a stronger correlation between GUS and combination (R=0.8; p=2.2e-16) than GOL and combination (R=0.52; p=4.1e-12), with processes associated with epithelial and stromal biology, and mucosal inflammation. In contrast, the combination effect on specific neutrophil/myeloid biology at WK4 was more similar to GOL than GUS, supporting the early role of GOL in targeting innate inflammation. Conclusion Combination induction with GUS and GOL showed significant reductions in major inflammatory features of disease as early as WK4 which persisted through WK38 with GUS maintenance. Correlative analysis supports the role of GUS as the primary driver of tissue healing, with GOL further contributing to innate inflammatory activity, which demonstrate differential and complementary mechanisms of action of TNFα and IL-23p19 subunit blockade.

Bladder cancer.

Bladder cancer is a global health issue with sex differences in incidence and prognosis. Bladder cancer has distinct molecular subtypes with multiple pathogenic pathways depending on whether the disease is non-muscle invasive or muscle invasive. The mutational burden is higher in muscle-invasive than in non-muscle-invasive disease. Commonly mutated genes include TERT, FGFR3, TP53, PIK3CA, STAG2 and genes involved in chromatin modification. Subtyping of both forms of bladder cancer is likely to change considerably with the advent of single-cell analysis methods. Early detection signifies a better disease prognosis; thus, minimally invasive diagnostic options are needed to improve patient outcomes. Urine-based tests are available for disease diagnosis and surveillance, and analysis of blood-based cell-free DNA is a promising tool for the detection of minimal residual disease and metastatic relapse. Transurethral resection is the cornerstone treatment for non-muscle-invasive bladder cancer and intravesical therapy can further improve oncological outcomes. For muscle-invasive bladder cancer, radical cystectomy with neoadjuvant chemotherapy is the standard of care with evidence supporting trimodality therapy. Immune-checkpoint inhibitors have demonstrated benefit in non-muscle-invasive, muscle-invasive and metastatic bladder cancer. Effective management requires a multidisciplinary approach that considers patient characteristics and molecular disease characteristics.

Phase I Study of Ruxolitinib in Combination with Abemaciclib for Patients with Primary or Post-Polycythemia Vera/Essential Thrombocythemia Myelofibrosis

Introduction and preclinical rationale: Patients with primary and secondary myelofibrosis (MF) have a reduced life expectancy compared to age-matched controls. The prognosis of individual patients is defined by clinical, cytogenetic, and molecular disease characteristics. Several oral Janus Kinase (JAK)-1/2 inhibitors such as ruxolitinib are approved by the US FDA for the treatment of patients with intermediate or high-risk MF based on improvements in symptom burden and spleen volume reduction compared to physician choice of best available therapy. However, these inhibitors often have a limited duration of response, and survival in patients after ruxolitinib discontinuation is poor (median survival of 13 months). Novel mechanism-based therapies for patients with suboptimal response or progression on ruxolitinib are thus needed. Preclinical studies have demonstrated that JAK2 V617F mutations promote transition from G1 to S-phase of the cell cycle via increased expression of CDC25A. Notably, CDC25A expression has been shown to be upregulated in primary MPN patient samples. Additional cell cycle regulators such as CDK6 and Cyclin D have also been demonstrated to play a role in MPN pathogenesis and to drive myeloproliferation. Preclinical studies testing the combination of the CDK4/6 inhibitor ribociclib and ruxolitinib demonstrated reduction in spleen and liver size when compared to ruxolitinib alone in murine models, suggesting synergism (Rampal et al., Clinical Cancer Research 2021). Methods: Based on these promising preclinical data, we hypothesize that combining ruxolitinib with a CDK4/6 inhibitor could have synergistic effects in patients with primary or secondary MF who have an inadequate response to ruxolitinib monotherapy. In this multicenter, phase I dose-escalation trial we evaluate the safety of combination therapy with ruxolitinib and the CDK4/6 inhibitor abemaciclib. Patients with primary or secondary MF with intermediate-1/2 or high-risk disease by DIPSS who require treatment and had an inadequate response to ruxolitinib are eligible for enrollment. An inadequate response will be defined by (I) palpable splenomegaly ≥5 cm below the left costal margin at study entry AND/OR (II) active MPN symptoms [presence of one symptom score ≥5 or two symptom scores ≥3 using the screening MPN-SAF TSS (Emanuel at al. JCO 2012). (Table 1). This study follows a conventional "3+3” dose-escalation design, in which patients on a stable dose of ruxolitinib (10mg or 15mg BID) with sufficient baseline platelet and absolute neutrophil count will be treated with increasing doses of abemaciclib. The pre-study dose of ruxolitinib will be maintained to determine the maximum tolerated dose of the combination of ruxolitinib and abemaciclib (Figure 1). The primary endpoint of this study is to determine the safety and tolerability, and to identify the recommended phase II dose of abemaciclib in MF patients on a stable, standard dose of ruxolitinib. Secondary endpoints include overall response rate (defined as a composite of complete remission [CR], partial remission [PR], and clinical improvement [CI; defined as anemia, spleen or symptom response]), rates of CR, PR, and CI as well as progression-free survival (PFS), overall survival (OS), and duration of response (DOR) as defined in accordance with the modified 2013 consensus definitions proposed by the International Working Group - Myeloproliferative Neoplasms Research and Treatment (IWG-MRT) and the European Leukemia Net (ELN). As part of this clinical trial several correlative studies are planned to identify biomarkers predictive of response to ruxolitinib and abemaciclib as well as mechanisms of resistance (e.g., MAP kinase pathway activation). These include evaluation of changes in variant allele fraction of driver mutations, gene expression profile, cell cycle, clonal architecture, cytokine profile, and bone marrow fibrosis, aiming to inform patient selection as well as other alternative combination therapies for future clinical trials. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Circulating miRNA Expression Profiling in Breast Cancer Molecular Subtypes: Applying Machine Learning Analysis in Bioinformatics.

Breast cancer is a leading worldwide cause of female cancer-related morbidity and mortality. Since molecular characteristics increasingly guide disease management, demystifying breast tumor miRNA signature emerges as an essential step toward personalized care. This study aimed to investigate the variations in circulating miRNA expression profiles between breast cancer subtypes and healthy controls and to identify relevant target genes and molecular functions. MiRNA expression was tested by miScript™ miRNA PCR Array Human Cancer Pathway Finder kit, and subsequently, a machine learning approach was applied for miRNA profiling of the various breast cancer molecular subtypes. Serum samples from patients with primary breast cancer (n=66) and healthy controls (n=16) were analyzed. MiR-21 was the single common molecule among all breast cancer subtypes. Furthermore, several miRNAs were found to be differentially expressed explicitly in the different subtypes; luminal A (miR-23b, miR-142, miR-29a, miR-181d, miR-16, miR-29b, miR-155, miR-181c), luminal B (miR-148a, let-7d, miR-92a, miR-34c, let-7b, miR-15a), HER2+ (miR-125b, miR-134, miR-98, miR-143, miR-138, miR-135b) and triple negative breast cancer (miR-17, miR-150, miR-210, miR-372, let-7f, miR-191, miR-133b, miR-146b, miR-7). Finally, miRNA-associated target genes and molecular functions were identified. Applying a machine learning approach to delineate miRNA signatures of various breast cancer molecular subtypes allows further understanding of molecular disease characteristics that can prove clinically relevant.

MDS-519 Outcome Predictors and Timing Considerations for Allogeneic Stem Cell Transplantation in Chronic Myelomonocytic Leukemia

Chronic myelomonocytic leukemia (CMML) outcomes are poor in general with a median overall survival (mOS) of 3 years. Allogeneic transplant (HSCT) offers the only curative option. Considerations as to when to pursue HSCT, identification of relapse predictors and assessment of whether transplant-related, clinical and molecular disease features have an impact on outcomes remain crucial. To characterize best timing to HSCT, best therapy to achieve CR prior to transplant, and analyze associations between molecular and transplant-related variables and relapse risk Design: Retrospective collection of clinical and molecular data of transplanted CMML patients treated at Moffitt Cancer Center between 1995-2022. Median duration of follow-up was 93 months. A total of 84 patients went to HSCT: 30 (36%) in BP-CMML (sAML), 36(43%) in CMML-0/ CMML-1(EarlyStage) and 15(18%) in CMML-2. Median age at HSCT was 63years (18-76). mOS for the transplant cohort was 62 months (95%CI 35-89 months). Patients who went to transplant earlier had better outcomes: mOS for ES-CMML: 202 months (95%CI NR), vs 62 months for BP-CMML (95%CI 39-84 moths) (p=0.019). There was no association between relapse rate and number of treatment lines prior to HSCT (p=.112), type of donor (p=.491), stem cell source (p=.768) or type of conditioning regimen (MA vs RIC/NMA) (p=.565). There was a trend toward better outcomes for those receiving intensive chemotherapy prior to transplant (p=.08) and with lower blast burden (mOS:80 vs 64 months, (p=.92)). Regarding specific therapies both standard 7+3/AML-like induction and CLAG regimens were equally effective in inducing CR and superior to single-agent HMA (p<0.05). 28 patients (34%) experienced relapse. Relapse rate for those with lower blast percentage at time of HSCT was 30% (n=18) compared to 40% (p=0.458). Patients who developed GvHD were less likely to relapse (85.7% vs 14.3%, p=0.000). There was an association between DNMT3A mutations and risk of relapse (p=0.04) with a trend also favoring NRAS (p=0.08). Patients with CMML who went to transplant earlier in the disease course had better outcomes. There was a trend toward better outcomes for those receiving intensive chemotherapy. CR was more likely with CLAG and 7+3 induction. Relapse was less likely for those who developed GvHD and more likely to be associated with DNMT3A mutations.

Pyoderma gangrenosum: A systematic review of the molecular characteristics of disease.

Pyoderma gangrenosum is a painful recurrent ulcerative neutrophilic dermatosis in which the pathogenesis is incompletely defined. Current evidence suggests that PG is associated with dysregulation of components of both the innate and adaptive immune system with dysregulation of neutrophil function and contribution of the Th17 immune axis. PG can be present in numerous heterogeneous clinical presentations and be associated with multiple inflammatory conditions including rheumatoid arthritis, inflammatory bowel disease and hidradenitis suppurativa. However, no critical evaluation of the observed molecular characteristics in PG studies in association with their clinical findings has been assessed. Additionally, emerging evidence suggests a potential role for other cell types and immune pathways including B cells, macrophages, autoantibodies and the complement system in PG, although these have not yet been integrated into the pathogenesis of disease. This systematic review aims to critically evaluate the current molecular observations regarding the pathogenesis of PG and discuss associations with clinical characteristics as well as the evidence supporting novel cell types and immune pathways in PG.

Pediatric versus Adult Medulloblastoma: Towards a Definition That Goes beyond Age.

Simple SummaryMedulloblastoma is a rare brain tumor that affects children and adults. Treatment with surgery, radiation, and chemotherapy currently cures most patients; however, ~30% of all patients have poor clinical outcomes despite treatment. Prospective clinical trials have historically excluded older patients, while recent advances in molecular diagnostics have enhanced our understanding of tumorigenesis. The aim of this literature review is to discuss the history of clinical trials in medulloblastoma and to argue in favor of prioritizing molecular drivers of disease as trial inclusion features rather than an arbitrary age cutoff.Medulloblastoma is a rare malignant brain tumor that predominantly affects children but also occurs in adults. The incidence declines significantly after age 15, and distinct tumor molecular features are seen across the age spectrum. Standard of care treatment consists of maximal safe surgical resection followed by adjuvant radiation and/or chemotherapy. Adjuvant treatment decisions are based on individual patient risk factors and have been informed by decades of prospective clinical trials. These trials have historically relied on arbitrary age cutoffs for inclusion (age 16, 18, or 21, for example), while trials that include adult patients or stratify patients by molecular features of disease have been rare. The aim of this literature review is to review the history of clinical trials in medulloblastoma, with an emphasis on selection criteria, and argue in favor of rational and inclusive trials based on molecular features of disease as opposed to chronological age. We performed a scoping literature review for medulloblastoma and clinical trials and include a summary of those results. We also discuss some of the significant advances made in understanding the molecular biology of medulloblastoma within the past decade, most notably the identification of four distinct subgroups based on gene expression profiling. We will also cite the recent experiences of childhood leukemia and the emergence of tissue-agnostic therapies as examples of successes of rationally designed, inclusive trials translating to improved clinical outcomes for patients across the age spectrum. Despite the prior trial history and recent molecular advances outcomes remain poor for ~30% of medulloblastoma patients. We believe that defining patients by the specific molecular alterations their tumors harbor is the best way to ensure they can access potentially efficacious therapies on clinical trials.

Prevalence and Management Practices of Lumpy Skin Disease (LSD) in Cattle at Natore District of Bangladesh

Lumpy skin disease (LSD) is a potentially contagious viral disease in cattle, caused by the Lumpy skin disease virus (LSDV), which belongs to the family Poxviridae. The virus is suspected to spread via biological vectors such as mosquitoes, flies, ticks, and direct contact. It is a significant economic disease caused by a virus that causes significant losses in milk production, fertility, abortions, import restrictions, and, in some cases, death in the livestock animals. This study aimed to assess the current state of LSD and its management on several farms in the north-western region of Bangladesh. Data were obtained from two Upazilas in the Natore district (Natore Sadar and Baraigram) using a structured questionnaire. During the study period of 1 June to 20 December 2020, a total of 34 small farms and 87 animals were monitored. The OIE guidelines directed the diagnosis of LSD based on current clinical indicators. MS Excel and SPSS statistical software were used to evaluate all of the data. Morbidity, mortality, and case-fatality rates were found to be 64.70%, 2.94 %, and 4.53 % in Natore Sadar and 83.02 %, 3.77 %, and 4.55 % in Baraigram Upazilas, respectively. Female animals that were newly matured (2-4 years) were a larger prone to LSD infection (43.1%) than animals of other ages. A total of 34.43% illness was found in young bull cattle aged (2-4). Bull and heifer calves are also vulnerable populations, with reports of LSD infection leading to mortality. Limb swelling is a common clinical symptom, and LSD-positive young mature (2-4 year) cattle had the most significant rate (18.39%) of limb swelling. The farm hygiene practice is a critical determinant in the spread of LSDV, and a large proportion of cattle (48.27 %) infected with LSD were on farms with poor hygienic management, compared to good (01.14 %) and medium (01.14 %) hygienic management practices (26.43 %). Despite the fact that mosquito nets help prevent mosquitoes, most owners (91.17 %) did not use one in their cattle barn at night. More study is needed in Bangladesh to improve the clinical management of LSD, identify risk factors, and understand the molecular characteristics of diseases.

Integrated multi-omic analyses reveals clinical relevance of endometrial cancer cell line models

<h3>Objectives:</h3> Endometrial cancer (EC) cell lines models established from diverse ancestral backgrounds and reflective of <i>in situ</i> disease characteristics are necessary to support preclinical investigations with broad generalizability. We performed multi-omic analyses of EC cell line models established from women representing diverse ancestries and compared these with clinical and molecular profiling data from the TCGA UCEC patient cohort to assess the relevance of models to <i>in situ</i> clinical and molecular disease characteristics. <h3>Methods:</h3> Whole genome sequencing (WGS), transcriptome (RNAseq) and proteome (LC-MS/MS and RPPA) analyses were performed for primary cell line models established from endometrioid EC patients (NCI-EC1, ACI-80, ACI-181, ACI-52, ACI-61 and ACI-68) as well as commercial EC models (MFE-296, SNG-M, HEC1A, RL95-2, Ishikawa, AN3-CA, KLE). Microsatellite instability (MSI) was determined from WGS data using MSISensor2 and standard ancestry estimates by comparison with reference populations from the 1000 Genomes Project. Hierarchical cluster analyses was performed using Pvclust and differential analyses using LIMMA. TCGA UCEC clinical and transcriptome data (n=371 patients) were downloaded from the TCGA UCEC manuscript supplement or cbioportal.org and odds ratio significance was assessed using Fisher's exact test (p<0.05). <h3>Results:</h3> EC cell line models are MSI-high except ACI-61, ACI-68 and KLE cells which are microsatellite stable. Standard ancestry estimates showed most cell lines are from women of European ancestry except Ishikawa, HEC1A and SNG-M which are of East Asian ancestry and NCI-EC1, ACI-181 and ACI-80 which are of African ancestry. Unsupervised hierarchical analysis of proteome data revealed distinct clusters comprising MFE-296, AN3-CA, ACI-68, ACI-61, NCI-EC1 cells (cluster 1) and ACI-80, ACI-52, HEC1A, SNG-M, RL95-2, KLE, ACI-181 and Ishikawa cells (cluster 2). Differential analyses showed elevated levels of mesenchymal markers (eg, vimentin) and lower levels of epithelial markers (eg, E-cadherin) in cluster 1 versus 2 cells. Correlation of transcript-level data with TCGA UCEC patients showed all cell lines are significantly correlated with endometrioid versus mixed/ uterine serous carcinoma tumors except NCI-EC1, ACI-52, HEC1A and KLE cells. Most cell lines significantly correlated with MSI-H patient tumors and largely trended as being correlated with tumors harboring high mutational loads, except ACI-68 and KLE cells. Cluster 2 cell lines are largely correlated with tumors exhibiting low somatic copy-number alterations except KLE, Ishikawa and HEC1A cells and are significantly correlated with tumors classified as hormonal versus mitotic or immunoreactive molecular subtypes except KLE cells. <h3>Conclusions:</h3> We describe EC cell line models established from women of diverse patient ancestries including those from women of African descent (i.e. NCI-EC1, ACI-181 and ACI-80 cells) that represent novel preclinical resources to investigate ancestry-linked molecular alterations underlying racial disparities between European and African-American women diagnosed with EC. We further define EC cell line models correlating with endometrioid tumors that exhibit more epithelial-like (elevation of E-Cadherin) as well as hormonal characteristics that will benefit preclinical investigations of endometrioid EC disease biology.

Shape decomposition algorithms for laser capture microdissection

BackgroundIn the context of biomarker discovery and molecular characterization of diseases, laser capture microdissection is a highly effective approach to extract disease-specific regions from complex, heterogeneous tissue samples. For the extraction to be successful, these regions have to satisfy certain constraints in size and shape and thus have to be decomposed into feasible fragments.ResultsWe model this problem of constrained shape decomposition as the computation of optimal feasible decompositions of simple polygons. We use a skeleton-based approach and present an algorithmic framework that allows the implementation of various feasibility criteria as well as optimization goals. Motivated by our application, we consider different constraints and examine the resulting fragmentations. We evaluate our algorithm on lung tissue samples in comparison to a heuristic decomposition approach. Our method achieved a success rate of over 95% in the microdissection and tissue yield was increased by 10–30%.ConclusionWe present a novel approach for constrained shape decomposition by demonstrating its advantages for the application in the microdissection of tissue samples. In comparison to the previous decomposition approach, the proposed method considerably increases the amount of successfully dissected tissue.

Rationale and design of the Kidney Precision Medicine Project

Chronic kidney disease (CKD) and acute kidney injury (AKI) are common, heterogeneous, and morbid diseases. Mechanistic characterization of CKD and AKI in patients may facilitate a precision-medicine approach to prevention, diagnosis, and treatment. The Kidney Precision Medicine Project aims to ethically and safely obtain kidney biopsies from participants with CKD or AKI, create a reference kidney atlas, and characterize disease subgroups to stratify patients based on molecular features of disease, clinical characteristics, and associated outcomes. An additional aim is to identify critical cells, pathways, and targets for novel therapies and preventive strategies. This project is a multicenter prospective cohort study of adults with CKD or AKI who undergo a protocol kidney biopsy for research purposes. This investigation focuses on kidney diseases that are most prevalent and therefore substantially burden the public health, including CKD attributed to diabetes or hypertension and AKI attributed to ischemic and toxic injuries. Reference kidney tissues (for example, living-donor kidney biopsies) will also be evaluated. Traditional and digital pathology will be combined with transcriptomic, proteomic, and metabolomic analysis of the kidney tissue as well as deep clinical phenotyping for supervised and unsupervised subgroup analysis and systems biology analysis. Participants will be followed prospectively for 10 years to ascertain clinical outcomes. Cell types, locations, and functions will be characterized in health and disease in an open, searchable, online kidney tissue atlas. All data from the Kidney Precision Medicine Project will be made readily available for broad use by scientists, clinicians, and patients.

Therapeutic Vulnerabilities of Transcription Factors in AML.

Acute myeloid leukemia (AML) is characterized by impaired myeloid lineage differentiation, uncontrolled proliferation, and inhibition of proapoptotic pathways. In spite of a relatively homogeneous clinical disease presentation, risk of long-term survival in AML varies from 20% to 80% depending on molecular disease characteristics. In recognition of the molecular heterogeneity of AML, the European Leukemia Net (ELN) and WHO classification systems now incorporate cytogenetics and increasing numbers of gene mutations into AML prognostication. Several of the genomic AML subsets are characterized by unique transcription factor alterations that are highlighted in this review. There are many mechanisms of transcriptional deregulation in leukemia. We broadly classify transcription factors based on mechanisms of transcriptional deregulation including direct involvement of transcription factors in recurrent translocations, loss-of-function mutations, and intracellular relocalization. Transcription factors, due to their pleiotropic effects, have been attractive but elusive targets. Indirect targeting approaches include inhibition of upstream kinases such as TAK1 for suppression of NFκB signaling and downstream effectors such as FGF signaling in HOXA-upregulated leukemia. Other strategies include targeting scaffolding proteins like BrD4 in the case of MYC or coactivators such as menin to suppress HOX expression; disrupting critical protein interactions in the case of β-catenin:TCF/LEF, and preventing transcription factor binding to DNA as in the case of PU.1 or FOXM1. We comprehensively describe the mechanism of deregulation of transcription factors in genomic subsets of AML, consequent pathway addictions, and potential therapeutic strategies.

Role of Racial, Demographic and Socioeconomic Disparities in Treatment Patterns and Outcomes in AML

Role of Racial, Demographic and Socioeconomic Disparities in Treatment Patterns and Outcomes in AML

A Randomized Trial of Bortezomib in Late Antibody-Mediated Kidney Transplant Rejection.

Late antibody-mediated rejection (ABMR) is a leading cause of kidney allograft failure. Uncontrolled studies have suggested efficacy of the proteasome inhibitor bortezomib, but no systematic trial has been undertaken to support its use in ABMR. In this randomized, placebo-controlled trial (the Bortezomib in Late Antibody-Mediated Kidney Transplant Rejection [BORTEJECT] Trial), we investigated whether two cycles of bortezomib (each cycle: 1.3 mg/m2 intravenously on days 1, 4, 8, and 11) prevent GFR decline by halting the progression of late donor-specific antibody (DSA)-positive ABMR. Forty-four DSA-positive kidney transplant recipients with characteristic ABMR morphology (median time after transplant, 5.0 years; pretransplant DSA documented in 19 recipients), who were identified on cross-sectional screening of 741 patients, were randomly assigned to receive bortezomib (n=21) or placebo (n=23). The 0.5-ml/min per 1.73 m2 per year (95% confidence interval, -4.8 to 5.8) difference detected between bortezomib and placebo in eGFR slope (primary end point) was not significant (P=0.86). We detected no significant differences between bortezomib- and placebo-treated groups in median measured GFR at 24 months (33 versus 42 ml/min per 1.73 m2; P=0.31), 2-year graft survival (81% versus 96%; P=0.12), urinary protein concentration, DSA levels, or morphologic or molecular rejection phenotypes in 24-month follow-up biopsy specimens. Bortezomib, however, associated with gastrointestinal and hematologic toxicity. In conclusion, our trial failed to show that bortezomib prevents GFR loss, improves histologic or molecular disease features, or reduces DSA, despite significant toxicity. Our results reinforce the need for systematic trials to dissect the efficiency and safety of new treatments for late ABMR.

Predictive Simulation Modeling in JAK2-Mutant MPNs Identified Ruxolitinib and Sorafenib As a Synergistic Drug Combination for Disease Modifying Therapy

Abstract Background: Despite the identification of a number of genes that underlie the genetic complexity of the myelofibrosis (MF) phenotype and even with the subsequent FDA approval of a targeted therapy for MF that is ruxolitinib, the median overall survival time of MF patients is still only 5-6 years. This unchanged survival may be explained, in part, via the complex mutation patterns that contribute to MF. Consequently, treating MF patients remains a significant challenge in the hematology clinic. Aim: To identify new drugs for MF patients, we used predictive simulation modeling of JAK2-V617F expressing MPN cells that could readily be re-purposed for MF. Methods: : To identify combinations of other drugs that can enhance the effect of Ruxolitinib, we (1) employed predictive simulation to model a JAK2-V617F bearing cell line (2) identified the key molecular disease characteristics of the JAK2-driven disease network, and (3) experimentally validated the predictions. The predictive simulation approach from Cellworks provides a representation of disease physiology incorporating signaling and metabolic networks with an integrated phenotype view. We modelled the JAK2-V617F expressing SET2 cell line based on its genomic aberration data. We then screened a library of drugs on the computational model in combination with Ruxolotinib, by assessing the impact on disease specific biomarkers and phenotypes of proliferation, viability, and apoptosis. Results: The modeling and digital drug screening identified a short list of drugs that were predicted to inhibit Jak2 signaling. Validation studies were conducted with the representative JAK2 driven SET2 cell line which harbors the JAK2-V617F mutation and was predicted to lead to subsequent activation of STAT3, STAT5, ERK via SHC1, AKT via IRS1-PIK3CA and STAT1 (Fig 1). JAK2 is known to phosphorylate STAT3 on Y705. Predictive modeling also discovered activation of parallel signaling via mutations of ALK, IRS2 and RPTOR. ALK is also known to activate STAT3 via Y705 phosphorylation and ERK via SHC1. IRS2 leads to activation of PIK3CA-AKT-MTOR. SET2 cells also harbor a RPTOR mutation which also activates MTOR pathway further. With higher ERK and MTOR signaling, there was S727 phosphorylation of STAT3. Using digital drug screening on the SET2 simulation model, sorafenib mitigated the effect of JAK2 mutation by inhibiting pathways downstream of JAK2, rather than decreasing the JAK2 mutation burden itself. Sorafenib is a small molecule kinase inhibitor that targets multiple proteins including VEGFR, PDGFR, and several Raf family kinases. It is approved for use in renal, liver, and thyroid cancers and is under clinical investigation for a number of other neoplasias. A potential role for sorafenib in MF has not been fully investigated. Sorafenib is known to inhibit ERK via inhibition of RAF. Sorafenib was also predicted to inhibit STAT3, STAT5, STAT1 and AKT1 via activation of PTPN6 in the SET2 model. With down-regulation of ERK and AKT1-MTOR pathway, Sorafenib was predicted to inhibit STAT3 S727 phosphorylation. Hence, with sorafenib and ruxolitinib, not only was JAK2 burden reduced, but also parallel pathway signaling via ALK, IRS2 and RPTOR. Using in vitro validation, we found that sorafenib was highly efficacious in reducing Jak2-V617F cell viability in a dose-dependent manner (GI50 = 6 uM) . Moreover, when Jak2-V617F cells were treated with both ruxolitinib and sorafenib, there was synergy, suggesting that the compounds were acting on multiple, cooperating signaling pathways. A prior report also showed synergistic MPN regression with ruxolitinib and sorafenib in BaF3/Jak2-V617F and HEL cell lines and primary specimens (PMID: 23560534), thereby providing independent evidence for the rationale use of these compounds in MF patients harboring mutation of PDGFRA/B, NRAS, KRAS in addition to JAK2 mutations. Conclusion: Thus, in addition to identifying a novel drug regimen for MF, we demonstrate that successful treatment of MF likely requires targeting multiple, non-redundant, constitutively active, and cooperative signaling pathways, rather than a one-biomarker/one-drug approach as is the current clinical standard. Further, we demonstrate how predictive simulation modeling can be used for precision medicine purposes in MF to inform clinical treatment decision-making of FDA approved drugs, with the intent of providing significant health and cost benefits. Download : Download high-res image (220KB) Download : Download full-size image Figure 1 . Disclosures Singh: Cellworks Research India Pvt. Ltd: Employment. Pampana: CellWorks: Employment. Usmani: Cellworks: Employment. Sauban: Cellworks: Employment. Kushwaha: CellWorks: Employment. Ramakrishnan: CellWorks: Employment. Abbasi: Cellworks Group Inc.: Employment. Vali: Cellworks Group Inc.: Employment. Cogle: Celgene: Other: Membership on Steering Committee for Connect MDS/AML Registry.

Clinical features of essential thrombocythemia and primary myelofibrosis, depending on the molecular characteristics of disease

The aim of the present paper was to evaluate the clinical features and risk of thrombotic events (TE) in patients with essential thrombocythemia (ET) and primary myelofibrosis (PMF), depending on the molecular characteristics of disease. Clinical data and laboratory parameters were analyzed in 50 ET patients and 50 PMF ones who had been followed up at the Department for Standardization of Treatments, National Research Center for Hematology, Ministry of Health of the Russian Federation, from February 2015 to September 2016. The patients with ET and those with PMF were found to have a high risk of TE. The risk for TE in the patients with ET is higher (24% in the entire group) than in those with PMF (14% in the study group). In ET, there is a high thrombosis risk in the detection of JAK2 and CALR gene mutations as compared with triple-negative cases. The PMF patients with JAK2 V617F mutations are at high risk for TE compared to those who are CALR mutation carriers and in triple-negative cases. There was no significant association of TE with high thrombocytosis. A factor, such as age, was found to be of no negative prognostic value in the patients with PMF.

Unsupervised hierarchical clustering of surface antigen expression to identify normal karyotype AML patients with distinct disease characteristics and poor outcome.

7042 Background: Surface antigen expression evaluation is part of the standard work-up at acute myeloid leukemia (AML) diagnosis. The biological &amp; prognostic implications of surface antigen expression patterns in normal karyotype (NK) AML patients (pts) remain unknown. Methods: The diagnostic antigen expression patterns of mononuclear cells in bone marrow (BM) of 111 NK-AML pts were assessed using a standard flow cytometric panel. At diagnosis common AML gene mutations (mut) &amp; expression levels were analyzed. Pts received stem cell transplantation (SCT, 98% allogeneic, 2% autologous; median age 63 years [y, range 26-74y]) after induction therapy at our institution. Median follow up was 3.3y. With R’s gplot package unsupervised hierarchical clustering of surface antigens was performed &amp; revealed 4 distinct clusters. Results: Pts in cluster 1 (n = 36) had higher expression of immature, in cluster 2 (n = 31) of thrombocytic/T-cell/erythroid, in cluster 3 (n = 24) of monocytic &amp; in cluster 4 (n = 20) of myeloid surface antigens. All 4 clusters associated with distinct clinical &amp; molecular features. At diagnosis, compared to all others, pts in cluster 1 had a higher CD34+/CD38- cell burden ( P&lt; .001), higher blood blasts ( P&lt; .03) &amp; BM blasts ( P&lt; .06) by trend. They had less NPM1 mut ( P&lt; .001) &amp; DNMT3A mut ( P= .02), were more likely to be EVI1 positive ( P= .03) &amp; had higher EZH2 ( P= .02), RUNX1 ( P= .009), BAALC ( P&lt; .001), ERG ( P= .02) &amp; MN1 ( P&lt; .001) expression. Compared to all others, pts in cluster 1 had a higher cumulative incidence of relapse (CIR, P= .002, at 1y 41% vs 15%) &amp; shorter event-free survival (EFS, P= .02, at 1y 50% vs 69%). In multivariate analysis, cluster 1 pts had a significantly higher CIR (Hazard Ratio [HR] 5.4, P= .01) after adjustment for FLT3-ITD &amp; shorter EFS (HR 2.1, P= .02) after adjustment for FLT3-ITD, age &amp; disease status at SCT. Conclusions: Pts in cluster 1 had high expression of immature surface antigens (eg CD34, CD117, CD13), genes involved in stem cell renewal &amp; worse outcome. Our data indicate a relationship between easily accessible surface antigen expression patterns at diagnosis, molecular disease features &amp; aggressiveness of the NK-AML phenotype.

Personalizing Chinese medicine by integrating molecular features of diseases and herb ingredient information: application to acute myeloid leukemia.

Traditional Chinese Medicine (TCM) has been widely used as a complementary medicine in Acute Myeloid Leukemia (AML) treatment. In this study, we proposed a new classification of Chinese Medicines (CMs) by integrating the latest discoveries in disease molecular mechanisms and traditional medicine theory. We screened out a set of chemical compounds on basis of AML differential expression genes and chemical-protein interactions and then mapped them to Traditional Chinese Medicine Integrated Database. 415 CMs contain those compounds and they were categorized into 8 groups according to the Traditional Chinese Pharmacology. Pathway analysis and synthetic lethality gene pairs were applied to analyze the dissimilarity, generality and intergroup relations of different groups. We defined hub CM pairs and alternative CM groups based on the analysis result and finally proposed a formula to form an effective anti-AML prescription which combined the hub CM pairs with alternative CMs according to patients’ molecular features. Our method of formulating CMs based on patients’ stratification provides novel insights into the new usage of conventional CMs and will promote TCM modernization.

FocusHeuristics \u2013 expression-data-driven network optimization and disease gene prediction

To identify genes contributing to disease phenotypes remains a challenge for bioinformatics. Static knowledge on biological networks is often combined with the dynamics observed in gene expression levels over disease development, to find markers for diagnostics and therapy, and also putative disease-modulatory drug targets and drugs. The basis of current methods ranges from a focus on expression-levels (Limma) to concentrating on network characteristics (PageRank, HITS/Authority Score), and both (DeMAND, Local Radiality). We present an integrative approach (the FocusHeuristics) that is thoroughly evaluated based on public expression data and molecular disease characteristics provided by DisGeNet. The FocusHeuristics combines three scores, i.e. the log fold change and another two, based on the sum and difference of log fold changes of genes/proteins linked in a network. A gene is kept when one of the scores to which it contributes is above a threshold. Our FocusHeuristics is both, a predictor for gene-disease-association and a bioinformatics method to reduce biological networks to their disease-relevant parts, by highlighting the dynamics observed in expression data. The FocusHeuristics is slightly, but significantly better than other methods by its more successful identification of disease-associated genes measured by AUC, and it delivers mechanistic explanations for its choice of genes.