Assessing the yield and safety of endoscopy in acute graft-vs-host disease after hematopoietic stem cell transplant.

BACKGROUND: Acute gastrointestinal (GI) graft-vs-host disease (aGVHD) is the most complication of hematopoietic stem cell transplant (HSCT) in patients with hematologic malignancy. Limited data exists on endoscopic evaluation of GVHD in post-HSCT patients with differing GI symptoms. Further, the diagnostic value of gross endoscopic findings as well as the safety of endoscopy in this commonly thrombocytopenic and neutropenic patient population remains unclear. AIM: To understand the diagnostic value of symptoms and gross endoscopic findings as well as safety of endoscopy in aGVHD patients. METHODS: We analyzed 195 endoscopies performed at City of Hope in patients who underwent allogeneic HSCT less than 100 d prior for hematologic malignancy and were subsequently evaluated for aGVHD via endoscopy. The yield, sensitivity, and specificity of diagnosing aGVHD were calculated for upper and lower endoscopy, various GI tract locations, and presenting symptoms. RESULTS: Combined esophagogastroduodenoscopy (EGD) and flexible sigmoidoscopy (FS) demonstrated a greater diagnostic yield for aGVHD (83.1%) compared to EGD (66.7%) or FS (77.2%) alone with any presenting symptom. The upper and lower GI tract demonstrated similar yields regardless of whether patients presented with diarrhea (95.7% vs 99.1%) or nausea/vomiting (97.5% vs 96.8%). Normal-appearing mucosa was generally as specific (91.3%) as abnormal mucosa (58.7%-97.8%) for the presence of aGVHD. Adverse events such as bleeding (1.0%), infection (1.0%), and perforation (0.5%) only occurred in a small proportion of patients, with no significant differences in those with underlying thrombocytopenia (P = 1.000) and neutropenia (P = 0.425). CONCLUSION: Combined EGD and FS with biopsies of normal and inflamed mucosa demonstrated the greatest diagnostic yield regardless of presenting symptom and appears to be safe in this population of patients.

Diagnostic value of clusterin immunostaining in hepatocellular carcinoma.

BACKGROUND: Histologic distinction between well differentiated hepatocellular carcinoma (HCC) and benign hepatocellular mass lesions is a known challenge. Existing biomarkers are of limited diagnostic value. Our previous studies observed an enhanced canalicular expression pattern of clusterin (CLU) in HCC, which was not observed in benign hepatocellular mass lesions such as hepatocellular adenoma. The aim of this study was to further investigate its diagnostic value for HCC by examining the expression pattern of CLU in a large number of non-hepatocellular tumors, and by comparing it with two other commonly used hepatocellular markers pCEA and CD10 that also show a canalicular staining pattern in HCC. METHODS: Enhanced canalicular staining patterns of CLU, pCEA and CD10 were analyzed on 54 surgically resected well to moderately differentiated HCCs on whole tissue sections, of which 37 had surrounding regenerative nodules while the remaining 17 had a non-cirrhotic background. CLU immunostaining was also performed on tissue microarray sections that contained 74 HCCs (40 of which were also stained for pCEA and CD10), 55 normal liver tissue samples, and 1305 non-hepatocellular tumors from multiple organs. RESULTS: Enhanced CLU canalicular staining was observed in 70% (89/128) HCCs but not in regenerative nodules, normal liver tissues or any non-hepatocellular tumors. The sensitivity and specificity for enhanced canalicular staining pattern of CLU in HCCs were 0.70 and 1.00. This enhanced canalicular pattern was observed in only 26 and 23% HCCs for CD10 and pCEA, respectively. These results further demonstrate that the distinctive enhanced canalicular pattern of CLU is unique to HCC. CONCLUSIONS: CLU is superior to pCEA and CD10 as a diagnostic immunomarker in that it can help distinguish well to moderately differentiated HCC not only from non-HCC malignancies but also from benign hepatocellular mass lesions.

Association Between Spatial Heterogeneity Within Nonmetastatic Gastroesophageal Adenocarcinomas and Survival.

Importance: Intratumoral heterogeneity has been recognized as a significant barrier in successfully developing targetable biomarkers for gastroesophageal adenocarcinoma (GEA) and may affect neoadjuvant precision medicine approaches. Objective: To describe intratumoral spatial heterogeneity of tumor cell populations in nonmetastatic GEA and its association with survival. Design, Setting, and Participants: This case series retrospectively identified 41 patients with GEA who underwent up-front surgical resection at a tertiary referral cancer center from January 1, 1989, through December 31, 2013. Survival was calculated from date of surgery to date of death through June 1, 2017. Data were analyzed from June 2, 2017, to March 1, 2019. Main Outcomes and Measures: Overall survival, intratumoral clonal composition determined by genomic single-nucleotide variation array and bioinformatic analysis, and intercellular tumoral distances determined by multiprobe fluorescence in situ hybridization. Results: Among the 41 patients included in the analysis (22 men [54%]; mean [SD] age, 63 [12] years), a high proportion (19 [46%]) presented with tumors possessing high intratumoral heterogeneity. Kaplan-Meier analysis demonstrated that cases with an intratumoral clonal composition count of at least 2 exhibited worse survival compared with cases with a clonal composition count of 0 to 1 (univariate hazard ratio, 3.92; 95% CI, 1.27-12.08; P = .02). This finding remained significant on multivariate analysis controlling for stage, Lauren histologic subtype, receipt of adjuvant therapy, and age (multivariate hazard ratio, 4.55; 95% CI, 1.09-19.04; P = .04). Multiprobe fluorescence in situ hybridization demonstrated intratumoral clonal populations coexisting at submillimeter distances with differing relevant oncogenic copy number alterations, such as EGFR, JAK2, FGFR2, MET, CCND1, KRAS, MYC, PIK3CA, CD274, and PDCD1LG2. Conclusions and Relevance: This study found that spatial intratumoral heterogeneity of oncogenic copy number alterations exists before metastatic dissemination, and increased heterogeneity was associated with worse outcomes in resected GEA. Baseline heterogeneity illustrates the challenges in GEA targeted therapy. Further study may offer insight into strategies on combinatorial and/or sequential targeted and immunotherapeutic approaches.

Precision medicine and actionable alterations in lung cancer: A single institution experience.

OBJECTIVES: Oncology has become more reliant on new testing methods and a greater use of electronic medical records, which provide a plethora of information available to physicians and researchers. However, to take advantage of vital clinical and research data for precision medicine, we must initially make an effort to create an infrastructure for the collection, storage, and utilization of this information with uniquely designed disease-specific registries that could support the collection of a large number of patients. MATERIALS AND METHODS: In this study, we perform an in-depth analysis of a series of lung adenocarcinoma patients (n = 415) with genomic and clinical data in a recently created thoracic patient registry. RESULTS: Of the 415 patients with lung adenocarcinoma, 59% (n = 245) were female; the median age was 64 (range, 22-92) years with a median OS of 33.29 months (95% CI, 29.77-39.48). The most common actionable alterations were identified in EGFR (n = 177/415 [42.7%]), ALK (n = 28/377 [7.4%]), and BRAF V600E (n = 7/288 [2.4%]). There was also a discernible difference in survival for 222 patients, who had an actionable alteration, with a median OS of 39.8 months as compared to 193 wild-type patients with a median OS of 26.0 months (P<0.001). We identified an unprecedented number of actionable alterations [53.5% (222/415)], including distinct individual alteration rates, as compared with 15.0% and 22.3% in TCGA and GENIE respectively. CONCLUSION: The use of patient registries, focused genomic panels and the appropriate use of clinical guidelines in community and academic settings may influence cohort selection for clinical trials and improve survival outcomes.

Long-term Proton Pump Inhibitor Administration Caused Physiological and Microbiota Changes in Rats.

Proton pump inhibitors (PPIs) are used for the long-term treatment of gastroesophageal disorders and the non-prescription medicines for acid reflux. However, there is growing concerns about PPI misuse, overuse and abuse. This study aimed to develop an animal model to examine the effects of long-term use of PPI in vivo. Twenty one Wistar rats were given omeprazole orally or intravenously for 30 days, and caerulein as a positive control. After euthanization, the serum and stool were collected to perform MS-based quantitative analysis of metabolites. We carried out 16S-based profiling of fecal microbiota, assessed the expression of bile acid metabolism regulators and examined the immunopathological characteristics of bile ducts. After long-term PPI exposure, the fecal microbial profile was altered and showed similarity to those observed in high-fat diet studies. The concentrations of several metabolites were also changed in various specimens. Surprisingly, morphological changes were observed in the bile duct, including ductal epithelial proliferation, micropapillary growth of biliary epithelium, focal bile duct stricture formation and bile duct obstruction. These are characteristics of precancerous lesions of bile duct. FXR and RXRα expressions were significantly reduced, which were similar to that observed in cholangiocarcinoma in TCGA and Oncomine databases. We established a novel animal model to examine the effects of long-term use of omeprazole. The gut microbes and metabolic change are consequences of long-term PPI exposure. And the results showed the environment in vivo tends to a high-fat diet. More importantly, we observed biliary epithelial hyperplasia, which is an indicator of a high-fat diet.

Integrin α6 signaling induces STAT3-TET3-mediated hydroxymethylation of genes critical for maintenance of glioma stem cells.

Both the extracellular matrix (ECM) and DNA epigenetic regulation are critical for maintaining stem cell phenotype and cancer progression. Whether and how ECM regulates epigenetic alterations to influence cancer stem cells (CSCs) remain to be explored. Here we report that ECM through laminin-integrin α6 upregulates ten-eleven translocation enzyme 3 (TET3) dioxygenase. TET3 in turn mediates DNA cytosine 5'-hydroxymethylation (5hmC) and upregulates genes critical for maintenance of glioma stem cells (GSCs). Activating integrin α6-FAK pathway increases STAT3 activity, TET3 expression and 5hmC levels in GSCs. Moreover, targeting STAT3 disrupts integrin α6-FAK signaling and inhibits TET3+ GSC maturation in vivo. STAT3 directly regulates TET3 expression and the two proteins are co-localized with 5hmC in GSC clusters. 5hmC is upregulated by STAT3 at the promoters of several tumorigenic genes, including c-Myc, known to be critical for GSCs. In vivo silencing of TET3 in GSC-enriched tumors reduces 5hmC accumulation and expression of the GSC critical genes, leading to tumor growth inhibition. TET3 expression and 5hmC accumulation also co-segregate with integrin α6 in patient malignant glioma. Thus, ECM- integrin α6-STAT3-TET3 axis regulates hydroxymethylation of genes important for GSCs, thereby increasing GSC tumorigenicity and resistance to therapies.

STAT3 Activation-Induced Fatty Acid Oxidation in CD8+ T Effector Cells Is Critical for Obesity-Promoted Breast Tumor Growth.

Although obesity is known to be critical for cancer development, how obesity negatively impacts antitumor immune responses remains largely unknown. Here, we show that increased fatty acid oxidation (FAO) driven by activated STAT3 in CD8+ T effector cells is critical for obesity-associated breast tumor progression. Ablating T cell Stat3 or treatment with an FAO inhibitor in obese mice spontaneously developing breast tumor reduces FAO, increases glycolysis and CD8+ T effector cell functions, leading to inhibition of breast tumor development. Moreover, PD-1 ligation in CD8+ T cells activates STAT3 to increase FAO, inhibiting CD8+ T effector cell glycolysis and functions. Finally, leptin enriched in mammary adipocytes and fat tissues downregulates CD8+ T cell effector functions through activating STAT3-FAO and inhibiting glycolysis. We identify a critical role of increased oxidation of fatty acids driven by leptin and PD-1 through STAT3 in inhibiting CD8+ T effector cell glycolysis and in promoting obesity-associated breast tumorigenesis.

Monitoring and Determining Mitochondrial Network Parameters in Live Lung Cancer Cells.

Mitochondria are dynamic organelles that constantly fuse and divide, forming dynamic tubular networks. Abnormalities in mitochondrial dynamics and morphology are linked to diverse pathological states, including cancer. Thus, alterations in mitochondrial parameters could indicate early events of disease manifestation or progression. However, finding reliable and quantitative tools for monitoring mitochondria and determining the network parameters, particularly in live cells, has proven challenging. Here, we present a 2D confocal imaging-based approach that combines automatic mitochondrial morphology and dynamics analysis with fractal analysis in live small cell lung cancer (SCLC) cells. We chose SCLC cells as a test case since they typically have very little cytoplasm, but an abundance of smaller mitochondria compared to many of the commonly used cell types. The 2D confocal images provide a robust approach to quantitatively measure mitochondrial dynamics and morphology in live cells. Furthermore, we performed 3D reconstruction of electron microscopic images and show that the 3D reconstruction of the electron microscopic images complements this approach to yield better resolution. The data also suggest that the parameters of mitochondrial dynamics and fractal dimensions are sensitive indicators of cellular response to subtle perturbations, and hence, may serve as potential markers of drug response in lung cancer.

Phenotypic Switching of Naïve T Cells to Immune-Suppressive Treg-Like Cells by Mutant KRAS.

Oncogenic (mutant) Ras protein Kirsten rat sarcoma viral oncogene homolog (KRAS) promotes uncontrolled proliferation, altered metabolism, and loss of genome integrity in a cell-intrinsic manner. Here, we demonstrate that CD4+ T cells when incubated with tumor-derived exosomes from mutant (MT) KRAS non-small-cell lung cancer (NSCLC) cells, patient sera, or a mouse xenograft model, induce phenotypic conversion to FOXP3+ Treg-like cells that are immune-suppressive. Furthermore, transfecting T cells with MT KRAS cDNA alone induced phenotypic switching and mathematical modeling supported this conclusion. Single-cell sequencing identified the interferon pathway as the mechanism underlying the phenotypic switch. These observations highlight a novel cytokine-independent, cell-extrinsic role for KRAS in T cell phenotypic switching. Thus, targeting this new class of Tregs represents a unique therapeutic approach for NSCLC. Since KRAS is the most frequently mutated oncogene in a wide variety of cancers, the findings of this investigation are likely to be of broad interest and have a large scientific impact.

LKB1 deficiency promotes proliferation and invasion of glioblastoma through activation of mTOR and focal adhesion kinase signaling pathways.

Liver kinase B1 (LKB1), a serine/threonine kinase, is frequently inactivated in several types of human cancers. To date, inactivation of LKB1 tumor suppressor has rarely been reported in glioblastoma. In this study, we investigated LKB1 status, biological significance, and therapeutic implications in glioblastoma. Loss of LKB1 immunostaining was identified in 8.6% (5/58), while decrease of LKB1 immunostaining was found in 29.3% (17/58) of glioblastoma tissues. Notably, mining TCGA database of LKB1 expression in glioblastoma revealed that lower mRNA level of LKB1 was associated with shorter survival in glioblastoma. We found that knockdown of LKB1 significantly promoted in vitro proliferation, adhesion, invasion, and metformin-induced apoptosis, and simultaneously enhanced activation of ERK and mammalian-target of rapamycin (mTOR) signaling pathways in LKB1-compenent U87 and T98 glioblastoma cells. Moreover, global transcriptional profiling revealed that adhesion and cytoskeletal proteins such as Vinculin, Talin and signaling pathways including focal adhesion kinase (FAK), extracellular martrix (ECM) receptor interaction, and cellular motility were significantly enriched in U87 and T98 glioblastoma cells upon LKB1 knockdown. Additionally, we demonstrated that the enhanced activation of FAK by LKB1 knockdown was dependent on differentially expressed cytoskeletal proteins in these glioblastoma cells. Importantly, we further found that mTOR1 inhibitor rapamycin dominantly inhibited in vitro cellular proliferation, while FAK inhibitor PF-573288 drastically decreased invasion of LKB1-attenuated glioblastoma cells. Therefore, downregulation of LKB1 may contribute to the pathogenesis and malignancy of glioblastoma and may have potential implications for stratification and treatment of glioblastoma patients.

Differential gene expression and AKT targeting in triple negative breast cancer.

Background: Metastatic triple negative breast cancer (mTNBC) is a heterogeneous disease with poor prognosis. Molecular evolution of TNBC through chemotherapy selection pressure is well recognized but poorly understood. PI3K/AKT/mTOR is one of the most commonly identified oncogenic-driver pathways in breast cancer. The current study is designed to understand the genomic and transcriptomic changes, focusing on the PI3K/AKT/mTOR pathway alterations in paired primary and metastatic TNBCs. Results: Genomic analysis of 7 paired specimens identified 67 known mutations including those from the following signaling pathways: cell cycle, p53, PI3K/AKT/mTOR, RAS/MAPK, and RTK/GF. Principle coordinate analysis (PCoA) identified 4 distinctive molecular groups based on the gene expression patterns of PI3K/AKT/mTOR pathway. Key differentially-expressed genes included AKT3, GSK3B, GNA11, PI3KR1, and GNAQ. Importantly, AKT-targeted therapy showed efficacy in a patient-derived xenograft (PDX) model of TNBC in vivo. Conclusion: Genomic discordance of paired primary and metastatic TNBCs was identified, with significant increase in tumor proliferation pathways seen in metastases. Among the differentially expressed genes, AKT3 can potentially serve as a target for novel combination therapy for treatment of metastatic TNBC. Methods: Paired specimens from 10 patients with TNBCs were identified through an IRB-approved protocol (2002-2015). FoundationOneTM sequencing was performed for genomic profiling, and Affymetrix Human Genechip 2.0st was used for mRNA expression profiling. The similarity among samples was calculated based on Pearson correlation coefficients, which were used to construct hierarchical clustering and heat maps.

Intraoperative bile spillage is associated with worse survival in gallbladder adenocarcinoma.

BACKGROUND: Gallbladder adenocarcinoma is often incidentally identified following cholecystectomy. We hypothesized that intraoperative bile spillage would be a negative prognostic factor. METHODS: A retrospective review of patients treated at a cancer center with histologically confirmed gallbladder adenocarcinoma, 2009-2017, was performed. Patient, disease, and treatment factors were analyzed in terms of progression-free survival (PFS) and overall survival (OS). RESULTS: Sixty-six patients were identified. Tumor stage was T1 (n = 8, 12%), T2 (n = 23, 35%), T3 (n = 35, 53%). Node stage was N0 (n = 22, 33%), N1+ (n = 26, 39%), Nx (n = 18, 27%). Operations included cholecystectomy alone (n = 27, 36%), cholecystectomy and partial hepatectomy (n = 30, 45%), or hepaticojejunostomy (n = 9, 14%). Median PFS was 7 months (interquartile range [IQR], 2-19); median OS was 16 months (IQR, 10-31). Subset multivariate proportional hazards regression of 41 patients who underwent initial cholecystectomy showed decreased PFS was associated with intraoperative spillage (n = 12, 29%; hazard ratio [HR], 5.5; P = .0014); decreased OS was associated with drain placement (n = 21, 51%; HR, 8.1; P = .006). CONCLUSIONS: Intraoperative bile spillage and surgical drain placement at initial cholecystectomy are negatively associated with PFS and OS in gallbladder adenocarcinoma. Explicit documentation of spillage and drain placement rationale is critical, possibly indicating locally advanced disease and prompting stronger consideration of systemic therapy before definitive resection.

Variation in the Thoroughness of Pathologic Assessment and Response Rates of Locally Advanced Rectal Cancers After Chemoradiation.

BACKGROUND: Pathologic complete response (pCR) is associated with better prognosis and guides management for patients with advanced rectal cancer. Response rates vary between series for unclear reasons. We examine whether the thoroughness of pathologic assessment explains differences in pCR rates. METHODS: We retrospectively reviewed pathology reports from patients with stage II/III rectal cancer who underwent chemoradiation and resection in a prospective, multicenter trial. We utilized a novel measure for the thoroughness of pathologic assessment by dividing residual tumor size by the number of cassettes evaluated (tumor size to cassette ratio, TSCR), and evaluated whether TSCR is associated with pCR. We validated our findings using a separate cohort. RESULTS: From the trial cohort, 71 of 247 (29%) patients achieved pCR. The pCR rate ranged from 0 to 45% and mean TSCR ranged 0.29 to 0.87 across 12 institutions. Within each institution, a lower TSCR was associated with pCR, demonstrating a higher degree of thoroughness used for tumors that achieved pCR. Moreover, across all samples, low TSCR was independently associated with pCR on multivariable analysis. This finding was corroborated in a separate cohort of 201 tumors evaluated by five pathologists; each pathologist had a lower mean TSCR for pCR calls compared with non-pCR calls. However, the mean TSCR for an institution was not associated with its overall pCR rate. CONCLUSIONS: Pathologists assess rectal cancers that have responded significantly to neoadjuvant therapy more thoroughly. Thoroughness does not appear to explain differences in pCR rates between institutions. Our results suggest pCR is not a sampling artifact.

Identification of Tissue-Specific DNA Methylation Signatures for Thyroid Nodule Diagnostics.

PURPOSE: Thyroid cancer is frequently difficult to diagnose due to an overlap of cytologic features between malignant and benign nodules. This overlap leads to unnecessary removal of the thyroid in patients without cancer. While providing some improvement over cytopathologic diagnostics, molecular methods frequently fail to provide a correct diagnosis for thyroid nodules. These approaches are based on the difference between cancer and adjacent thyroid tissue and assume that adjacent tissues are the same as benign nodules. However, in contrast to adjacent tissues, benign thyroid nodules can contain genetic alterations that can be found in cancer.Experimental Design: For the development of a new molecular diagnostic test for thyroid cancer, we evaluated DNA methylation in 109 thyroid tissues by using genome-wide single-base resolution DNA methylation analysis. The test was validated in a retrospective cohort containing 65 thyroid nodules. RESULTS: By conducting reduced representation bisulfite sequencing in 109 thyroid specimens, we found significant differences between adjacent tissue, benign nodules, and cancer. These tissue-specific signatures are strongly linked to active enhancers and cancer-associated genes. Based on these signatures, we developed a new epigenetic approach for thyroid diagnostics. According to the validation cohort, our test has an estimated specificity of 97% [95% confidence interval (CI), 81-100], sensitivity of 100% (95% CI, 87-100), positive predictive value of 97% (95% CI, 83-100), and negative predictive value of 100% (95% CI, 86-100). CONCLUSIONS: These data show that epigenetic testing can provide outstanding diagnostic accuracy for thyroid nodules.See related commentary by Mitmaker et al., p. 457.

Arginine starvation kills tumor cells through aspartate exhaustion and mitochondrial dysfunction.

Defective arginine synthesis, due to the silencing of argininosuccinate synthase 1 (ASS1), is a common metabolic vulnerability in cancer, known as arginine auxotrophy. Understanding how arginine depletion kills arginine-auxotrophic cancer cells will facilitate the development of anti-cancer therapeutic strategies. Here we show that depletion of extracellular arginine in arginine-auxotrophic cancer cells causes mitochondrial distress and transcriptional reprogramming. Mechanistically, arginine starvation induces asparagine synthetase (ASNS), depleting these cancer cells of aspartate, and disrupting their malate-aspartate shuttle. Supplementation of aspartate, depletion of mitochondria, and knockdown of ASNS all protect the arginine-starved cells, establishing the causal effects of aspartate depletion and mitochondrial dysfunction on the arginine starvation-induced cell death. Furthermore, dietary arginine restriction reduced tumor growth in a xenograft model of ASS1-deficient breast cancer. Our data challenge the view that ASNS promotes homeostasis, arguing instead that ASNS-induced aspartate depletion promotes cytotoxicity, which can be exploited for anti-cancer therapies.

Autophagic reliance promotes metabolic reprogramming in oncogenic KRAS-driven tumorigenesis.

Defects in basal autophagy limit the nutrient supply from recycling of intracellular constituents. Despite our understanding of the prosurvival role of macroautophagy/autophagy, how nutrient deprivation, caused by compromised autophagy, affects oncogenic KRAS-driven tumor progression is poorly understood. Here, we demonstrate that conditional impairment of the autophagy gene Atg5 (atg5-KO) extends the survival of KRASG12V-driven tumor-bearing mice by 38%. atg5-KO tumors spread more slowly during late tumorigenesis, despite a faster onset. atg5-KO tumor cells displayed reduced mitochondrial function and increased mitochondrial fragmentation. Metabolite profiles indicated a deficiency in the nonessential amino acid asparagine despite a compensatory overexpression of ASNS (asparagine synthetase), key enzyme for de novo asparagine synthesis. Inhibition of either autophagy or ASNS reduced KRASG12V-driven tumor cell proliferation, migration, and invasion, which was rescued by asparagine supplementation or knockdown of MFF (mitochondrial fission factor). Finally, these observations were reflected in human cancer-derived data, linking ASNS overexpression with poor clinical outcome in multiple cancers. Together, our data document a widespread yet specific asparagine homeostasis control by autophagy and ASNS, highlighting the previously unrecognized role of autophagy in suppressing the metabolic barriers of low asparagine and excessive mitochondrial fragmentation to permit malignant KRAS-driven tumor progression.

JMJD1B Demethylates H4R3me2s and H3K9me2 to Facilitate Gene Expression for Development of Hematopoietic Stem and Progenitor Cells.

The arginine methylation status of histones dynamically changes during many cellular processes, including hematopoietic stem/progenitor cell (HSPC) development. The arginine methyltransferases and the readers that transduce the histone codes have been defined. However, whether arginine demethylation actively occurs in cells and what enzyme demethylates the methylarginine residues during various cellular processes are unknown. We report that JMJD1B, previously identified as a lysine demethylase for H3K9me2, mediates arginine demethylation of H4R3me2s and its intermediate, H4R3me1. We show that demethylation of H4R3me2s and H3K9me2s in promoter regions is correlated with active gene expression. Furthermore, knockout of JMJD1B blocks demethylation of H4R3me2s and/or H3K9me2 at distinct clusters of genes and impairs the activation of genes important for HSPC differentiation and development. Consequently, JMJD1B-/- mice show defects in hematopoiesis. Altogether, our study demonstrates that demethylase-mediated active arginine demethylation process exists in eukaryotes and that JMJD1B demethylates both H4R3me2s and H3K9me2 for epigenetic programming during hematopoiesis.

Chemotherapy Induces Breast Cancer Stemness in Association with Dysregulated Monocytosis.

Purpose: Preoperative or neoadjuvant therapy (NT) is increasingly used in patients with locally advanced or inflammatory breast cancer to allow optimal surgery and aim for pathologic response. However, many breast cancers are resistant or relapse after treatment. Here, we investigated conjunctive chemotherapy-triggered events occurring systemically and locally, potentially promoting a cancer stem-like cell (CSC) phenotype and contributing to tumor relapse.Experimental Design: We started by comparing the effect of paired pre- and post-NT patient sera on the CSC properties of breast cancer cells. Using cell lines, patient-derived xenograft models, and primary tumors, we investigated the regulation of CSCs and tumor progression by chemotherapy-induced factors.Results: In human patients and mice, we detected a therapy-induced CSC-stimulatory activity in serum, which was attributed to therapy-associated monocytosis leading to systemic elevation of monocyte chemoattractant proteins (MCP). The post-NT hematopoietic regeneration in the bone marrow highlighted both altered monocyte-macrophage differentiation and biased commitment of stimulated hematopoietic stem cells toward monocytosis. Chemotherapeutic agents also induce monocyte expression of MCPs through a JNK-dependent mechanism. Genetic and pharmacologic inhibitions of the MCP-CCR2 pathway blocked chemotherapy's adverse effect on CSCs. Levels of nuclear Notch and ALDH1 were significantly elevated in primary breast cancers following NT, whereas higher levels of CCR2 in pre-NT tumors were associated with a poor response to NT.Conclusions: Our data establish a mechanism of chemotherapy-induced cancer stemness by linking the cellular events in the bone marrow and tumors, and suggest pharmacologic inhibition of CCR2 as a potential cotreatment during conventional chemotherapy in neoadjuvant and adjuvant settings. Clin Cancer Res; 24(10); 2370-82. ©2018 AACR.