Introduction: Truncating mutations in Additional Sex Combs-Like 1 (ASXL1) are associated with a h... more Introduction: Truncating mutations in Additional Sex Combs-Like 1 (ASXL1) are associated with a high-risk disease phenotype in myeloid malignancies. In chronic myelomonocytic leukemia (CMML), truncating ASXL1 mutations are known to increase transcriptional activity of leukemic driver genes and have been associated with gene body hypermethylation. We interrogated the transcriptome and methylome of patients with ASXL1-mutant (MT) and -wildtype (WT) CMML using a multi-omics approach to test the hypothesis that gene expression is mediated through gene body methylation. Methods: Bone marrow mononuclear cells from patients with ASXL1 WT (n=8) and MT (n=8) CMML were subjected to targeted NGS of DNA, whole transcriptome shotgun sequencing (RNA-seq), and immunoprecipitation of DNA methyl residues (DIP-seq). After quality control all samples were sequenced on an Illumina HiSeq 4000 before further processing and data analysis. Differential gene expression analysis was performed to identify gen...
Introduction: Additional Sex Combs-Like 1 (ASXL1) is a chromatin modifier frequently affected by ... more Introduction: Additional Sex Combs-Like 1 (ASXL1) is a chromatin modifier frequently affected by truncating mutations in myeloid malignancies. In chronic myelomonocytic leukemia (CMML), truncating ASXL1 mutations are associated with the overexpression of leukemogenic driver genes, however the molecular mechanisms underlying this transcriptional activation remain controversial. We performed single-cell chromatin accessibility studies of ASXL1-mutant (MT) and -wildtype (WT) CMML to identify cis-regulatory elements (CREs) such as distal enhancer elements that may explain the observed transcriptional activity in MT CMML. Methods: Bone marrow mononuclear cells from a patient with WT and a patient with MT CMML were subjected to single-cell DNA transposase accessibility studies (scATAC-seq). The cryopreserved cells were thawed and resuspended and approximately 100000 viable mononuclear cells per sample were subjected to transposase assays before proceeding to single-cell partitioning into ...
Anal squamous cell carcinoma (ASCC) is a rare malignancy, with most cases associated with human p... more Anal squamous cell carcinoma (ASCC) is a rare malignancy, with most cases associated with human papilloma virus and an increased incidence in immunocompromised patients. Progress in management of ASCC has been limited not only due to its rarity, but also the associated lack of research funding and social stigma. Historically, standard of care for invasive ASCC has been highly morbid surgical resection, requiring a permanent colostomy. Surgery was associated with disease recurrence in approximately half of the patients. However, the use of chemotherapy (5-fluorouracil and mitomycin C) concomitantly with radiation in the 1970s resulted in disease regression, curing a subset of patients and sparing them from morbid surgery. Validation of the use of systemic therapy in prospective trials was not achieved until approximately 20 years later. In this review, advancements and shortcomings in the use of systemic therapy in the management of ASCC will be discussed. Not only will standard-of-c...
Key Points Systemic immune microenvironment signatures in CMML indicate an altered T- and natural... more Key Points Systemic immune microenvironment signatures in CMML indicate an altered T- and natural killer cell balance. CMML bone marrow dendritic cell aggregates associate with disease progression and systemic regulatory T-cell phenotypic switch.
The Hedgehog-regulated transcription factors GLI1 and GLI2 play overlapping roles in development ... more The Hedgehog-regulated transcription factors GLI1 and GLI2 play overlapping roles in development and disease; however, the mechanisms underlying their interplay remain elusive. We report for the first time that GLI1 and GLI2 physically and functionally interact in cancer cells. GLI1 and GLI2 were shown to co-immunoprecipitate in PANC1 pancreatic cancer cells and RMS13 rhabdomyosarcoma cells. Mapping analysis demonstrated that the zinc finger domains of both proteins are required for their heteromerization. RNAi knockdown of either GLI1 or GLI2 inhibited expression of many well-characterized GLI target genes (BCL2, MYCN, PTCH2, IL7 and CCND1) in PANC1 cells, whereas PTCH1 expression was only inhibited by GLI1 depletion. qPCR screening of a large set of putative canonical and non-canonical Hedgehog/GLI targets identified further genes (e.g. E2F1, BMP1, CDK2) strongly down-regulated by GLI1 and/or GLI2 depletion in PANC1 cells, and demonstrated that ANO1, AQP1 and SOCS1 are up-regulate...
Proliferative chronic myelomonocytic leukemia (pCMML), an aggressive CMML subtype, is associated ... more Proliferative chronic myelomonocytic leukemia (pCMML), an aggressive CMML subtype, is associated with dismal outcomes. RAS pathway mutations, mainly NRASG12D, define the pCMML phenotype as demonstrated by our exome sequencing, progenitor colony assays and a Vav-Cre-NrasG12D mouse model. Further, these mutations promote CMML transformation to acute myeloid leukemia. Using a multiomics platform and biochemical and molecular studies we show that in pCMML RAS pathway mutations are associated with a unique gene expression profile enriched in mitotic kinases such as polo-like kinase 1 (PLK1). PLK1 transcript levels are shown to be regulated by an unmutated lysine methyl-transferase (KMT2A) resulting in increased promoter monomethylation of lysine 4 of histone 3. Pharmacologic inhibition of PLK1 in RAS mutant patient-derived xenografts, demonstrates the utility of personalized biomarker-driven therapeutics in pCMML.
Understanding the molecular events controlling melanoma progression is of paramount importance fo... more Understanding the molecular events controlling melanoma progression is of paramount importance for the development of alternative treatment options for this devastating disease. Here we report a mechanism regulated by the oncogenic SOX2-GLI1 transcriptional complex driving melanoma invasion through the induction of the sialyltransferase ST3GAL1. Using in vitro and in vivo studies, we demonstrate that ST3GAL1 drives melanoma metastasis. Silencing of this enzyme suppresses melanoma invasion and significantly reduces the ability of aggressive melanoma cells to enter the blood stream, colonize distal organs, seed and survive in the metastatic environment. Analysis of glycosylated proteins reveals that the receptor tyrosine kinase AXL is a major effector of ST3GAL1 pro-invasive function. ST3GAL1 induces AXL dimerization and activation that, in turn, promotes melanoma invasion. Our data support a key role of the ST3GAL1-AXL axis as driver of melanoma metastasis, and highlight the therapeu...
Chromosome region maintenance protein1 (CRM1) mediates protein export from the nucleus and is a n... more Chromosome region maintenance protein1 (CRM1) mediates protein export from the nucleus and is a new target for anti-cancer therapeutics. Broader application of KPT-330 (selinexor), a first in class CRM1 inhibitor recently approved for relapsed multiple myeloma and diffuse large B-cell lymphoma, have been limited by substantial toxicity. We discovered that salicylates markedly enhance the anti-tumor activity of CRM1 inhibitors by extending the mechanisms of action beyond CRM1 inhibition. Using salicylates in combination enables targeting of a range of blood cancers with a much lower dose of selinexor, thereby potentially mitigating prohibitive clinical adverse effects. Choline salicylate (CS) with low-dose KPT-330 (K+CS) had potent, broad activity across high-risk hematological malignancies and solid organ cancers ex vivo and in vivo. The K+CS combination was not toxic to non-malignant cells as compared to malignant cells and was safe without inducing toxicity to normal organs in mic...
In this issue, Hou and colleagues present their exciting work demonstrating that, through remodel... more In this issue, Hou and colleagues present their exciting work demonstrating that, through remodeling of the local tumor microenvironment (TME), pancreatic ductal adenocarcinoma forms a tumor-supportive niche capable of liberating cancer cells from dependence on oncogenic KRAS signaling. Through extensive experimentation both in vitro and in vivo, the authors reveal that the HDAC5-CCL2 axis drives the recruitment of tumor-associated macrophages to the TME to provide trophic signaling.See related article by Hou et al., p. 1058.
Malignant melanoma, the most aggressive form of skin cancer, is characterized by high prevalence ... more Malignant melanoma, the most aggressive form of skin cancer, is characterized by high prevalence of BRAF/NRAS mutations and hyperactivation of extracellular signal-regulated kinase 1 and 2 (ERK1/2), mitogen-activated protein kinases (MAPK), leading to uncontrolled melanoma growth. Efficacy of current targeted therapies against mutant BRAF or MEK1/2 have been hindered by existence of innate or development of acquired resistance. Therefore, a better understanding of the mechanisms controlled by MAPK pathway driving melanogenesis will help develop new treatment approaches targeting this oncogenic cascade. Here, we identify E3 ubiquitin ligase PARK2 as a direct target of ELK1, a known transcriptional effector of MAPK signaling in melanoma cells. We show that pharmacological inhibition of BRAF-V600E or ERK1/2 in melanoma cells increases PARK2 expression. PARK2 overexpression reduces melanoma cell growth in vitro and in vivo and induces apoptosis. Conversely, its genetic silencing increas...
RAS pathway mutations are driver mutations in several cancers including myeloid neoplasms. In par... more RAS pathway mutations are driver mutations in several cancers including myeloid neoplasms. In particular, in myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN) overlap syndromes, RAS pathway mutations are associated with a proliferative phenotype, increased rates of leukemic transformation (LT) and adverse outcomes [1, 2]. In fact, juvenile myelomonocytic leukemia (JMML), a pediatric MDS/MPN overlap syndrome, is a bona fide RASopathy, with germline and somatic mutations in RAS pathway genes giving rise to disease [3]. Whole genome and exome sequencing (WES) studies in MDS/ MPN overlap syndromes, including chronic myelomonocytic leukemia (CMML), have shown recurrent somatic mutations involving NRAS, CBL, KRAS, PTPN11, and NF1 [4]. In CMML, RAS pathway mutations are seen in 30% of patients and are commonly associated with MPN-CMML, increased rate of LT and shortened survival [5]. We carried out this study in patients with MDS/MPN overlap neoplasms to (i) define the prevalence and phenotypic correlates of RAS pathway mutations and (ii) assess the prognostic impact of RAS pathway mutations, including individual RAS pathway mutations. After approval by the Mayo Clinic (Rochester, MN, USA) institutional review board, patients with World Health Organization (WHO)-defined MDS/MPN overlap neoplasms were included in the study [6]. The bone marrow (BM) morphology, cytogenetics and 2016 WHO diagnoses were retrospectively reviewed and all patients underwent targeted next generation sequencing (NGS) for 29 myeloidrelevant genes, obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods [7]. The performance statistics for the NGS assay include minimum depth coverage of 250×, along with a sensitivity of 5% variant allele fraction (VAF) for single base substitutions and insertion/deletion events. Patients were included in this study at first referral only if their diagnosis was within 6 months of referral and provided they had not been treated with disease modifying agents, especially hypomethylating agents (HMA). Prior use of hydroxyurea, erythropoiesis stimulating agents and transfusions were allowed. Germline mutational status in JMML was confirmed by sequencing skin fibroblast-derived DNA. For the purposes of this study, NRAS, CBL, KRAS, and PTPN11 mutations were considered as RAS pathway mutations (given the low frequency, NF1 was not included in the NGS panel). All variants were manually curated to assess pathogenicity. Statistical analyses considered clinical and laboratory parameters obtained at the time of diagnosis or first referral. Continuous variables were presented as median (range) and categorical variables as frequency (percentage). Differences in the distribution of continuous variables * Mrinal M. Patnaik [email protected]
Systemic immune tolerance is not well-characterized in chronic myelomonocytic leukemia (CMML). Du... more Systemic immune tolerance is not well-characterized in chronic myelomonocytic leukemia (CMML). Due to the presence of clonal plasmacytoid dendritic cells (pDC) in CMML, and the established association of lymph node indoleamine 2,3-dioxygenase-1 (IDO1)-positive (+) DC populations (IDC) with systemic immune tolerance in other malignant contexts, we sought to determine the association of IDO1 expression and bone marrow (BM) DC populations with systemic T-cell compartment changes using primary CMML patient samples (BM, plasma, and peripheral blood mononuclear cells) via immunohistochemistry (IHC), liquid chromatography-mass spectrometry (LC-MS), and time-of-flight mass cytometry (CyTOF). Our results highlight that aggregate BM IDC (CD123 and/or CD11c positive) occur in 33% CMML patients at any disease time-point (IHC), correlate with accentuated tryptophan catabolism (LC-MS, increased kynurenine level, median 4.7 versus 3 microM, P=0.049*), systemic regulatory T-cell expansion (CyTOF, %...
The Runt-related transcription factor 1 (RUNX1) gene encodes a transcription factor that plays a ... more The Runt-related transcription factor 1 (RUNX1) gene encodes a transcription factor that plays a crucial role in embryogenesis and definitive hematopoiesis [1]. Germline RUNX1 mutations (RUNX1) result in a familial platelet disorder with propensity to myeloid malignancies (RUNX1FPD), a rare autosomal dominant condition characterized by mild to moderate thrombocytopenia, qualitative platelet defects, and inherent predisposition to develop hematological malignancies such as myelodysplastic syndrome (MDS), acute myelogenous leukemia (AML), and T-cell acute lymphoblastic leukemia (T-ALL) [2]. The lifetime risk of developing a myeloid malignancy in RUNX1-FPD is about 30–40%, with a median age at onset of 33 years [3]. In addition, germline pathogenic RUNX1 variants can be found in up to 29% of the families with two or more cases of AML or MDS [4]. Somatic RUNX1 have been identified as recurrent abnormalities in myeloid neoplasms, including AML (10%), MDS (12%), myeloproliferative neoplasms (MPN, 2–4%), and MDS/MPN overlap syndromes such as chronic myelomonocytic leukemia (CMML, 15%) [5–8]. In CMML, RUNX1 are associated with a higher risk of leukemic transformation (LT), especially in the context of mutations located in the C-terminal region, with a recent study also demonstrating an adverse impact on overall survival (OS) [7, 9]. In addition, RUNX1 have been identified in ~2–4% of patient with clonal cytopenias of unclear significance (CCUS), a premalignant condition associated with a probability of developing a myeloid neoplasm 14 times higher than that of patients with no evidence of clonal disease [10, 11]. We carried out this study to (i) define the genomic landscape of clonal evolution in RUNX1-FPD patients, (ii) compare and contrast germline RUNX1 with somatic RUNX1 seen in patients with MDS/MPN overlap syndromes (due to the availability of a well-defined data set) and (iii) assess phenotypic and survival differences between somatic RUNX1 and RUNX1 wild-type patients with MDS/MPN overlap syndromes. With due approval from the Mayo Clinic (Rochester, MN, USA) institutional review board, after obtaining written informed consent, families with RUNX1-FPD detected by germline sequencing of buccal swab-derived DNA or skin fibroblast-derived DNA (target capture-NGS and/or whole exome sequencing) and deletion/duplication assays (array comparative genomic hybridization assaysCGH) (Supplementary Table 1) and patients with 2016, World Health Organization (WHO)-defined MDS/MPN overlap syndromes (except juvenile myelomonocytic leukemia) were included in the study. Index patients with * Mrinal M. Patnaik [email protected]
The expression of the extracellular sulfatase SULF2 has been associated with increased hepatocell... more The expression of the extracellular sulfatase SULF2 has been associated with increased hepatocellular carcinoma (HCC) growth and poor patient survival. However, the molecular mechanisms underlying SULF2-associated tumor growth remain unclear. To address this gap, here we developed a transgenic mouse overexpressing Sulf2 in hepatocytes under the control of the transthyretin promoter. In this model, Sulf2 overexpression potentiated diethylnitrosamine-induced HCC. Further analysis indicated that the transcription factor GLI family zinc finger 1 (GLI1) mediates Sulf2 expression during HCC development. A cross of the Sulf2-overexpressing with Gli1-knockout mice revealed that Gli1 inactivation impairs SULF2-induced HCC. Transcriptomic analysis revealed that Sulf2 overexpression is associated with signal transducer and activator of transcription 3 (STAT3)-specific gene signatures. Interestingly, the Gli1 knockout abrogated SULF2-mediated induction of several STAT3 target genes, including s...
ABSTRACTChronic myelomonocytic leukemia (CMML) is an aggressive hematological malignancy with lim... more ABSTRACTChronic myelomonocytic leukemia (CMML) is an aggressive hematological malignancy with limited treatment options. Whole exome (WES) and targeted sequencing of several independent cohorts of CMML patients, comparing dysplastic (dCMML) to proliferative (pCMML) CMML, as well as paired chronic phase disease and acute leukemic transformation (LT), associate acquisition of oncogenic RAS pathway mutations, the most common being NRASG12D, with aggressive disease and with disease progression. Using patient derived progenitor colony assays and a NRASG12D-Vav-Cre mouse model, we further demonstrate the role of mutant RAS signaling in driving and maintaining pCMML phenotype. RNA-sequencing links RAS pathway mutations with an increased expression of genes encoding the mitotic checkpoint kinases PLK1 and WEE1. Further, we dmeoinstrated that non-mutated lysine methyltransferase KMT2A (MLL1) acts as mediator of NRAS-induced PLK1 and WEE1 expression. Finally, we demonstrate the translational ...
BACKGROUND/OBJECTIVES Interplay between the Hedgehog (HH) and epidermal growth factor receptor (E... more BACKGROUND/OBJECTIVES Interplay between the Hedgehog (HH) and epidermal growth factor receptor (EGFR) pathways modulating the outcome of their signaling activity have been reported in various cancers including pancreatic ductal adenocarcinoma (PDAC). Therefore, simultaneous targeting of these pathways may be clinically beneficial. This Phase I study combined HH and EGFR inhibition in metastatic PDAC patients. METHODS Combined effects of HH and EGFR inhibition using Vismodegib and Erlotinib with or without gemcitabine in metastatic solid tumors were assessed by CT. Another cohort of patients with metastatic PDAC was evaluated by FDG-PET and tumor biopsies-derived biomarkers. RESULTS Treatment was well tolerated with the maximum tolerated dose cohort experiencing no grade 4 toxicities though 25% experienced grade 3 adverse effects. Recommended phase II dose of Vismodegib and Erlotinib were each 150 mg daily. No tumor responses were observed although 16 patients achieved stable disease for 2-7 cycles. Paired biopsy analysis before and after first cycle of therapy in PDAC patients showed reduced GLI1 mRNA, phospho-GLI1 and associated HH target genes in all cases. However, only half of the cases showed reduced levels of desmoplasia or changes in fibroblast markers. Most patients had decreased phospho-EGFR levels. CONCLUSIONS Vismodegib and Erlotinib combination was well-tolerated although overall outcome in patients with metastatic PDAC was not significantly impacted by combination treatment. Biomarker analysis suggests direct targets inhibition without significantly affecting the stromal compartment. These findings conflict with pre-clinical mouse models, and thus warrant further investigation into how upstream inhibition of these pathways is circumvented in PDAC.
Introduction: Truncating mutations in Additional Sex Combs-Like 1 (ASXL1) are associated with a h... more Introduction: Truncating mutations in Additional Sex Combs-Like 1 (ASXL1) are associated with a high-risk disease phenotype in myeloid malignancies. In chronic myelomonocytic leukemia (CMML), truncating ASXL1 mutations are known to increase transcriptional activity of leukemic driver genes and have been associated with gene body hypermethylation. We interrogated the transcriptome and methylome of patients with ASXL1-mutant (MT) and -wildtype (WT) CMML using a multi-omics approach to test the hypothesis that gene expression is mediated through gene body methylation. Methods: Bone marrow mononuclear cells from patients with ASXL1 WT (n=8) and MT (n=8) CMML were subjected to targeted NGS of DNA, whole transcriptome shotgun sequencing (RNA-seq), and immunoprecipitation of DNA methyl residues (DIP-seq). After quality control all samples were sequenced on an Illumina HiSeq 4000 before further processing and data analysis. Differential gene expression analysis was performed to identify gen...
Introduction: Additional Sex Combs-Like 1 (ASXL1) is a chromatin modifier frequently affected by ... more Introduction: Additional Sex Combs-Like 1 (ASXL1) is a chromatin modifier frequently affected by truncating mutations in myeloid malignancies. In chronic myelomonocytic leukemia (CMML), truncating ASXL1 mutations are associated with the overexpression of leukemogenic driver genes, however the molecular mechanisms underlying this transcriptional activation remain controversial. We performed single-cell chromatin accessibility studies of ASXL1-mutant (MT) and -wildtype (WT) CMML to identify cis-regulatory elements (CREs) such as distal enhancer elements that may explain the observed transcriptional activity in MT CMML. Methods: Bone marrow mononuclear cells from a patient with WT and a patient with MT CMML were subjected to single-cell DNA transposase accessibility studies (scATAC-seq). The cryopreserved cells were thawed and resuspended and approximately 100000 viable mononuclear cells per sample were subjected to transposase assays before proceeding to single-cell partitioning into ...
Anal squamous cell carcinoma (ASCC) is a rare malignancy, with most cases associated with human p... more Anal squamous cell carcinoma (ASCC) is a rare malignancy, with most cases associated with human papilloma virus and an increased incidence in immunocompromised patients. Progress in management of ASCC has been limited not only due to its rarity, but also the associated lack of research funding and social stigma. Historically, standard of care for invasive ASCC has been highly morbid surgical resection, requiring a permanent colostomy. Surgery was associated with disease recurrence in approximately half of the patients. However, the use of chemotherapy (5-fluorouracil and mitomycin C) concomitantly with radiation in the 1970s resulted in disease regression, curing a subset of patients and sparing them from morbid surgery. Validation of the use of systemic therapy in prospective trials was not achieved until approximately 20 years later. In this review, advancements and shortcomings in the use of systemic therapy in the management of ASCC will be discussed. Not only will standard-of-c...
Key Points Systemic immune microenvironment signatures in CMML indicate an altered T- and natural... more Key Points Systemic immune microenvironment signatures in CMML indicate an altered T- and natural killer cell balance. CMML bone marrow dendritic cell aggregates associate with disease progression and systemic regulatory T-cell phenotypic switch.
The Hedgehog-regulated transcription factors GLI1 and GLI2 play overlapping roles in development ... more The Hedgehog-regulated transcription factors GLI1 and GLI2 play overlapping roles in development and disease; however, the mechanisms underlying their interplay remain elusive. We report for the first time that GLI1 and GLI2 physically and functionally interact in cancer cells. GLI1 and GLI2 were shown to co-immunoprecipitate in PANC1 pancreatic cancer cells and RMS13 rhabdomyosarcoma cells. Mapping analysis demonstrated that the zinc finger domains of both proteins are required for their heteromerization. RNAi knockdown of either GLI1 or GLI2 inhibited expression of many well-characterized GLI target genes (BCL2, MYCN, PTCH2, IL7 and CCND1) in PANC1 cells, whereas PTCH1 expression was only inhibited by GLI1 depletion. qPCR screening of a large set of putative canonical and non-canonical Hedgehog/GLI targets identified further genes (e.g. E2F1, BMP1, CDK2) strongly down-regulated by GLI1 and/or GLI2 depletion in PANC1 cells, and demonstrated that ANO1, AQP1 and SOCS1 are up-regulate...
Proliferative chronic myelomonocytic leukemia (pCMML), an aggressive CMML subtype, is associated ... more Proliferative chronic myelomonocytic leukemia (pCMML), an aggressive CMML subtype, is associated with dismal outcomes. RAS pathway mutations, mainly NRASG12D, define the pCMML phenotype as demonstrated by our exome sequencing, progenitor colony assays and a Vav-Cre-NrasG12D mouse model. Further, these mutations promote CMML transformation to acute myeloid leukemia. Using a multiomics platform and biochemical and molecular studies we show that in pCMML RAS pathway mutations are associated with a unique gene expression profile enriched in mitotic kinases such as polo-like kinase 1 (PLK1). PLK1 transcript levels are shown to be regulated by an unmutated lysine methyl-transferase (KMT2A) resulting in increased promoter monomethylation of lysine 4 of histone 3. Pharmacologic inhibition of PLK1 in RAS mutant patient-derived xenografts, demonstrates the utility of personalized biomarker-driven therapeutics in pCMML.
Understanding the molecular events controlling melanoma progression is of paramount importance fo... more Understanding the molecular events controlling melanoma progression is of paramount importance for the development of alternative treatment options for this devastating disease. Here we report a mechanism regulated by the oncogenic SOX2-GLI1 transcriptional complex driving melanoma invasion through the induction of the sialyltransferase ST3GAL1. Using in vitro and in vivo studies, we demonstrate that ST3GAL1 drives melanoma metastasis. Silencing of this enzyme suppresses melanoma invasion and significantly reduces the ability of aggressive melanoma cells to enter the blood stream, colonize distal organs, seed and survive in the metastatic environment. Analysis of glycosylated proteins reveals that the receptor tyrosine kinase AXL is a major effector of ST3GAL1 pro-invasive function. ST3GAL1 induces AXL dimerization and activation that, in turn, promotes melanoma invasion. Our data support a key role of the ST3GAL1-AXL axis as driver of melanoma metastasis, and highlight the therapeu...
Chromosome region maintenance protein1 (CRM1) mediates protein export from the nucleus and is a n... more Chromosome region maintenance protein1 (CRM1) mediates protein export from the nucleus and is a new target for anti-cancer therapeutics. Broader application of KPT-330 (selinexor), a first in class CRM1 inhibitor recently approved for relapsed multiple myeloma and diffuse large B-cell lymphoma, have been limited by substantial toxicity. We discovered that salicylates markedly enhance the anti-tumor activity of CRM1 inhibitors by extending the mechanisms of action beyond CRM1 inhibition. Using salicylates in combination enables targeting of a range of blood cancers with a much lower dose of selinexor, thereby potentially mitigating prohibitive clinical adverse effects. Choline salicylate (CS) with low-dose KPT-330 (K+CS) had potent, broad activity across high-risk hematological malignancies and solid organ cancers ex vivo and in vivo. The K+CS combination was not toxic to non-malignant cells as compared to malignant cells and was safe without inducing toxicity to normal organs in mic...
In this issue, Hou and colleagues present their exciting work demonstrating that, through remodel... more In this issue, Hou and colleagues present their exciting work demonstrating that, through remodeling of the local tumor microenvironment (TME), pancreatic ductal adenocarcinoma forms a tumor-supportive niche capable of liberating cancer cells from dependence on oncogenic KRAS signaling. Through extensive experimentation both in vitro and in vivo, the authors reveal that the HDAC5-CCL2 axis drives the recruitment of tumor-associated macrophages to the TME to provide trophic signaling.See related article by Hou et al., p. 1058.
Malignant melanoma, the most aggressive form of skin cancer, is characterized by high prevalence ... more Malignant melanoma, the most aggressive form of skin cancer, is characterized by high prevalence of BRAF/NRAS mutations and hyperactivation of extracellular signal-regulated kinase 1 and 2 (ERK1/2), mitogen-activated protein kinases (MAPK), leading to uncontrolled melanoma growth. Efficacy of current targeted therapies against mutant BRAF or MEK1/2 have been hindered by existence of innate or development of acquired resistance. Therefore, a better understanding of the mechanisms controlled by MAPK pathway driving melanogenesis will help develop new treatment approaches targeting this oncogenic cascade. Here, we identify E3 ubiquitin ligase PARK2 as a direct target of ELK1, a known transcriptional effector of MAPK signaling in melanoma cells. We show that pharmacological inhibition of BRAF-V600E or ERK1/2 in melanoma cells increases PARK2 expression. PARK2 overexpression reduces melanoma cell growth in vitro and in vivo and induces apoptosis. Conversely, its genetic silencing increas...
RAS pathway mutations are driver mutations in several cancers including myeloid neoplasms. In par... more RAS pathway mutations are driver mutations in several cancers including myeloid neoplasms. In particular, in myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN) overlap syndromes, RAS pathway mutations are associated with a proliferative phenotype, increased rates of leukemic transformation (LT) and adverse outcomes [1, 2]. In fact, juvenile myelomonocytic leukemia (JMML), a pediatric MDS/MPN overlap syndrome, is a bona fide RASopathy, with germline and somatic mutations in RAS pathway genes giving rise to disease [3]. Whole genome and exome sequencing (WES) studies in MDS/ MPN overlap syndromes, including chronic myelomonocytic leukemia (CMML), have shown recurrent somatic mutations involving NRAS, CBL, KRAS, PTPN11, and NF1 [4]. In CMML, RAS pathway mutations are seen in 30% of patients and are commonly associated with MPN-CMML, increased rate of LT and shortened survival [5]. We carried out this study in patients with MDS/MPN overlap neoplasms to (i) define the prevalence and phenotypic correlates of RAS pathway mutations and (ii) assess the prognostic impact of RAS pathway mutations, including individual RAS pathway mutations. After approval by the Mayo Clinic (Rochester, MN, USA) institutional review board, patients with World Health Organization (WHO)-defined MDS/MPN overlap neoplasms were included in the study [6]. The bone marrow (BM) morphology, cytogenetics and 2016 WHO diagnoses were retrospectively reviewed and all patients underwent targeted next generation sequencing (NGS) for 29 myeloidrelevant genes, obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods [7]. The performance statistics for the NGS assay include minimum depth coverage of 250×, along with a sensitivity of 5% variant allele fraction (VAF) for single base substitutions and insertion/deletion events. Patients were included in this study at first referral only if their diagnosis was within 6 months of referral and provided they had not been treated with disease modifying agents, especially hypomethylating agents (HMA). Prior use of hydroxyurea, erythropoiesis stimulating agents and transfusions were allowed. Germline mutational status in JMML was confirmed by sequencing skin fibroblast-derived DNA. For the purposes of this study, NRAS, CBL, KRAS, and PTPN11 mutations were considered as RAS pathway mutations (given the low frequency, NF1 was not included in the NGS panel). All variants were manually curated to assess pathogenicity. Statistical analyses considered clinical and laboratory parameters obtained at the time of diagnosis or first referral. Continuous variables were presented as median (range) and categorical variables as frequency (percentage). Differences in the distribution of continuous variables * Mrinal M. Patnaik [email protected]
Systemic immune tolerance is not well-characterized in chronic myelomonocytic leukemia (CMML). Du... more Systemic immune tolerance is not well-characterized in chronic myelomonocytic leukemia (CMML). Due to the presence of clonal plasmacytoid dendritic cells (pDC) in CMML, and the established association of lymph node indoleamine 2,3-dioxygenase-1 (IDO1)-positive (+) DC populations (IDC) with systemic immune tolerance in other malignant contexts, we sought to determine the association of IDO1 expression and bone marrow (BM) DC populations with systemic T-cell compartment changes using primary CMML patient samples (BM, plasma, and peripheral blood mononuclear cells) via immunohistochemistry (IHC), liquid chromatography-mass spectrometry (LC-MS), and time-of-flight mass cytometry (CyTOF). Our results highlight that aggregate BM IDC (CD123 and/or CD11c positive) occur in 33% CMML patients at any disease time-point (IHC), correlate with accentuated tryptophan catabolism (LC-MS, increased kynurenine level, median 4.7 versus 3 microM, P=0.049*), systemic regulatory T-cell expansion (CyTOF, %...
The Runt-related transcription factor 1 (RUNX1) gene encodes a transcription factor that plays a ... more The Runt-related transcription factor 1 (RUNX1) gene encodes a transcription factor that plays a crucial role in embryogenesis and definitive hematopoiesis [1]. Germline RUNX1 mutations (RUNX1) result in a familial platelet disorder with propensity to myeloid malignancies (RUNX1FPD), a rare autosomal dominant condition characterized by mild to moderate thrombocytopenia, qualitative platelet defects, and inherent predisposition to develop hematological malignancies such as myelodysplastic syndrome (MDS), acute myelogenous leukemia (AML), and T-cell acute lymphoblastic leukemia (T-ALL) [2]. The lifetime risk of developing a myeloid malignancy in RUNX1-FPD is about 30–40%, with a median age at onset of 33 years [3]. In addition, germline pathogenic RUNX1 variants can be found in up to 29% of the families with two or more cases of AML or MDS [4]. Somatic RUNX1 have been identified as recurrent abnormalities in myeloid neoplasms, including AML (10%), MDS (12%), myeloproliferative neoplasms (MPN, 2–4%), and MDS/MPN overlap syndromes such as chronic myelomonocytic leukemia (CMML, 15%) [5–8]. In CMML, RUNX1 are associated with a higher risk of leukemic transformation (LT), especially in the context of mutations located in the C-terminal region, with a recent study also demonstrating an adverse impact on overall survival (OS) [7, 9]. In addition, RUNX1 have been identified in ~2–4% of patient with clonal cytopenias of unclear significance (CCUS), a premalignant condition associated with a probability of developing a myeloid neoplasm 14 times higher than that of patients with no evidence of clonal disease [10, 11]. We carried out this study to (i) define the genomic landscape of clonal evolution in RUNX1-FPD patients, (ii) compare and contrast germline RUNX1 with somatic RUNX1 seen in patients with MDS/MPN overlap syndromes (due to the availability of a well-defined data set) and (iii) assess phenotypic and survival differences between somatic RUNX1 and RUNX1 wild-type patients with MDS/MPN overlap syndromes. With due approval from the Mayo Clinic (Rochester, MN, USA) institutional review board, after obtaining written informed consent, families with RUNX1-FPD detected by germline sequencing of buccal swab-derived DNA or skin fibroblast-derived DNA (target capture-NGS and/or whole exome sequencing) and deletion/duplication assays (array comparative genomic hybridization assaysCGH) (Supplementary Table 1) and patients with 2016, World Health Organization (WHO)-defined MDS/MPN overlap syndromes (except juvenile myelomonocytic leukemia) were included in the study. Index patients with * Mrinal M. Patnaik [email protected]
The expression of the extracellular sulfatase SULF2 has been associated with increased hepatocell... more The expression of the extracellular sulfatase SULF2 has been associated with increased hepatocellular carcinoma (HCC) growth and poor patient survival. However, the molecular mechanisms underlying SULF2-associated tumor growth remain unclear. To address this gap, here we developed a transgenic mouse overexpressing Sulf2 in hepatocytes under the control of the transthyretin promoter. In this model, Sulf2 overexpression potentiated diethylnitrosamine-induced HCC. Further analysis indicated that the transcription factor GLI family zinc finger 1 (GLI1) mediates Sulf2 expression during HCC development. A cross of the Sulf2-overexpressing with Gli1-knockout mice revealed that Gli1 inactivation impairs SULF2-induced HCC. Transcriptomic analysis revealed that Sulf2 overexpression is associated with signal transducer and activator of transcription 3 (STAT3)-specific gene signatures. Interestingly, the Gli1 knockout abrogated SULF2-mediated induction of several STAT3 target genes, including s...
ABSTRACTChronic myelomonocytic leukemia (CMML) is an aggressive hematological malignancy with lim... more ABSTRACTChronic myelomonocytic leukemia (CMML) is an aggressive hematological malignancy with limited treatment options. Whole exome (WES) and targeted sequencing of several independent cohorts of CMML patients, comparing dysplastic (dCMML) to proliferative (pCMML) CMML, as well as paired chronic phase disease and acute leukemic transformation (LT), associate acquisition of oncogenic RAS pathway mutations, the most common being NRASG12D, with aggressive disease and with disease progression. Using patient derived progenitor colony assays and a NRASG12D-Vav-Cre mouse model, we further demonstrate the role of mutant RAS signaling in driving and maintaining pCMML phenotype. RNA-sequencing links RAS pathway mutations with an increased expression of genes encoding the mitotic checkpoint kinases PLK1 and WEE1. Further, we dmeoinstrated that non-mutated lysine methyltransferase KMT2A (MLL1) acts as mediator of NRAS-induced PLK1 and WEE1 expression. Finally, we demonstrate the translational ...
BACKGROUND/OBJECTIVES Interplay between the Hedgehog (HH) and epidermal growth factor receptor (E... more BACKGROUND/OBJECTIVES Interplay between the Hedgehog (HH) and epidermal growth factor receptor (EGFR) pathways modulating the outcome of their signaling activity have been reported in various cancers including pancreatic ductal adenocarcinoma (PDAC). Therefore, simultaneous targeting of these pathways may be clinically beneficial. This Phase I study combined HH and EGFR inhibition in metastatic PDAC patients. METHODS Combined effects of HH and EGFR inhibition using Vismodegib and Erlotinib with or without gemcitabine in metastatic solid tumors were assessed by CT. Another cohort of patients with metastatic PDAC was evaluated by FDG-PET and tumor biopsies-derived biomarkers. RESULTS Treatment was well tolerated with the maximum tolerated dose cohort experiencing no grade 4 toxicities though 25% experienced grade 3 adverse effects. Recommended phase II dose of Vismodegib and Erlotinib were each 150 mg daily. No tumor responses were observed although 16 patients achieved stable disease for 2-7 cycles. Paired biopsy analysis before and after first cycle of therapy in PDAC patients showed reduced GLI1 mRNA, phospho-GLI1 and associated HH target genes in all cases. However, only half of the cases showed reduced levels of desmoplasia or changes in fibroblast markers. Most patients had decreased phospho-EGFR levels. CONCLUSIONS Vismodegib and Erlotinib combination was well-tolerated although overall outcome in patients with metastatic PDAC was not significantly impacted by combination treatment. Biomarker analysis suggests direct targets inhibition without significantly affecting the stromal compartment. These findings conflict with pre-clinical mouse models, and thus warrant further investigation into how upstream inhibition of these pathways is circumvented in PDAC.
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Papers by Ryan M . Carr