Phase 1 study of JNJ-64619178, a protein arginine methyltransferase 5 inhibitor, in patients with lower-risk myelodysplastic syndromes.

Splicing factor (SF) gene mutations are frequent in myelodysplastic syndromes (MDS), and agents that modulate RNA splicing are hypothesized to provide clinical benefit. JNJ-64619178, a protein arginine methyltransferase 5 (PRMT5) inhibitor, was evaluated in patients with lower-risk (LR) MDS in a multi-part, Phase 1, multicenter study. The objectives were to determine a tolerable dose and to characterize safety, pharmacokinetics, pharmacodynamics, and preliminary clinical activity. JNJ-64619178 was administered on a 14 days on/7 days off schedule or every day on a 21-day cycle to patients with International Prognostic Scoring System (IPSS) Low or Intermediate-1 risk MDS who were red blood cell transfusion-dependent. Twenty-four patients were enrolled; 15 (62.5 %) patients had low IPSS risk score, while 18 (75.0 %) had an SF3B1 mutation. Median duration of treatment was 3.45 months (range: 0.03-6.93). No dose limiting toxicities were observed. The 0.5 mg once daily dose was considered better tolerated and chosen for dose expansion. Twenty-three (95.8 %) patients experienced treatment-emergent adverse events (TEAE). The most common TEAEs were neutropenia (15 [62.5 %]) and thrombocytopenia (14 [58.3 %]). JNJ-64619178 pharmacokinetics was dose-dependent. Target engagement as measured by plasma symmetric di-methylarginine was observed across all dose levels; however, variant allele frequency of clonal mutations in bone marrow or blood did not show sustained reductions from baseline. No patient achieved objective response or hematologic improvement per International Working Group 2006 criteria, or transfusion independence. A tolerable dose of JNJ-64619178 was identified in patients with LR MDS. However, no evidence of clinical benefit was observed.

Phase 3 SELENE study: ibrutinib plus BR/R-CHOP in previously treated patients with follicular or marginal zone lymphoma.

The phase 3 SELENE study evaluated ibrutinib + chemoimmunotherapy (CIT; bendamustine and rituximab [BR]; or rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone [R-CHOP]) for patients with relapsed/refractory (R/R) follicular lymphoma (FL) or marginal zone lymphoma (MZL). Adult patients who had received ≥1 prior line of CIT were randomized 1:1 to oral ibrutinib (560 mg) or placebo daily, plus 6 cycles of BR/R-CHOP. The primary end point was investigator-assessed progression-free survival (PFS). Overall, 403 patients were randomized to ibrutinib + CIT (n = 202) or placebo + CIT (n = 201). Most patients received BR (90.3%) and had FL (86.1%). With a median follow-up of 84 months, median PFS was 40.5 months in the ibrutinib + CIT arm and 23.8 months in the placebo + CIT arm (hazard ratio [HR], 0.806; 95% confidence interval [CI], 0.626-1.037; P = .0922). Median overall survival was not reached in either arm (HR, 0.980; 95% CI, 0.686-1.400). Grade ≥3 treatment-emergent adverse events (TEAEs) were reported in 85.6% and 75.4% of patients in the ibrutinib + CIT and placebo + CIT arms, respectively. In each arm, 13 patients had TEAEs leading to death. The addition of ibrutinib to CIT did not significantly improve PFS compared with placebo + CIT. The safety profile was consistent with known profiles of ibrutinib and CIT. This trial was registered at www.clinicaltrials.gov as #NCT01974440.

Phase 1 Study of JNJ-64619178, a Protein Arginine Methyltransferase 5 Inhibitor, in Advanced Solid Tumors.

PURPOSE: In this first-in-human, Phase 1, open-label, multicenter study, we evaluated JNJ-64619178, a selective and potent PRMT5 inhibitor, in patients with advanced malignant solid tumors or non-Hodgkin lymphomas (NHL). The primary objective was to evaluate the safety and to identify a recommended Phase 2 dose (RP2D) of JNJ-64619178. PATIENTS AND METHODS: Adult patients with treatment-refractory advanced solid tumors or NHL and measurable disease received escalating doses of JNJ-64619178 following two schedules (Schedule A: 14 days on/7 days off; Schedule B: every day on a 21-day cycle). Safety, pharmacokinetics (PK), pharmacodynamics (PD), and clinical activity were evaluated. RESULTS: Ninety patients received JNJ-64619178. Thrombocytopenia was identified as the only dose-limiting toxicity. JNJ-64619178 showed dose-proportional PK and robust target engagement, as measured by plasma symmetric dimethylarginine, across all dose levels. The objective response rate was 5.6% (5 of 90). Patients with adenoid cystic carcinoma (ACC) had an ORR of 11.5% (3 of 26) and a median progression-free survival of 19.1 months. CONCLUSIONS: JNJ-64619178 demonstrated manageable dose-dependent toxicity and preliminary evidence of antitumor activity in ACC and other tumor types. Plasma exposure was dose dependent, and target inhibition was maintained with intermittent and continuous dosing. On the basis of safety, clinical activity, PK, and PD findings, two provisional RP2Ds were selected: 1.5 mg intermittently and 1.0 mg once daily. Aside from ACC, clinical benefit was limited, and biomarkers to enrich for responsiveness to PRMT5 inhibition will be needed for further development.

Association between PCV13 pneumococcal vaccination and risk of hospital admissions due to pneumonia or sepsis among patients with haematological malignancies: a single-centre retrospective cohort study in Israel.

OBJECTIVES: Patients with haematological malignancies receiving immunosuppressive therapy are at highest risk of invasive pneumococcal disease. Our goal was to investigate whether vaccination of haematological patients with pneumococcal 13-valent conjugated vaccine (PCV13) prior to therapy initiation is associated with decreased hospital admissions due to pneumonia or sepsis within 12 months. DESIGN AND SETTING: A longitudinal retrospective cohort study was conducted at the haematology unit of Carmel Medical Center, Israel. PARTICIPANTS: Information on adult patients (>18 years) who were diagnosed between 1 January 2009 and 30 December 2019 with haematological malignancies and received immunosuppressive therapy was retrieved from the electronic health records. Patients with haematological malignancies who received the PCV13 vaccination during or after initiation of the immunosuppressive therapy were excluded from the study. OUTCOME MEASURES: A multivariate logistic regression model was performed to determine whether PCV13 vaccination is associated with fewer hospital admissions due to pneumonia or sepsis. RESULTS: The cohort included 616 patients, of which 418 (67%) patients were not vaccinated and 198 (33%) were vaccinated. Within 12 months, 15.1% (n=63) of non-vaccinated patients compared with only 7.1% (n=14) of the vaccinated patients were hospitalised due to pneumonia or sepsis. The logistic regression analysis demonstrated that receiving PCV13 vaccination is associated with 45% (OR=0.45, 95% CI: 0.246 to 0.839, p=0.012) reduced odds of being hospitalised due to pneumonia or sepsis in patients with haematological malignancies receiving immunosuppressive therapy. CONCLUSION: This is the first observational study to demonstrate the association between PCV13 vaccination and hospital admissions in patients with haematological malignancies receiving immunosuppressive therapy. Patients receiving PCV13 vaccination before immunosuppressive therapy initiation had significantly reduced odds of hospitalisation due to pneumonia or sepsis compared with non-PCV13-vaccinated patients.

Adherence with direct oral anticoagulants in patients with atrial fibrillation: Trends, risk factors, and outcomes.

BACKGROUND: Adherence to direct oral anticoagulants (DOACs) remains a concern among non-valvular atrial fibrillation (AF) patients. We aimed to assess patterns of adherence with DOACs and examine their association with ischemic stroke and systemic embolism (SE). METHODS: This retrospective cohort study includes all adult members of Clalit Health Services, the largest healthcare provider in Israel, with newly diagnosed non-valvular AF between January 2014 and March 2019, who initiated DOACs within 90 days of AF diagnosis and used DOACs exclusively. Adherence was assessed using the proportion of days covered (PDC) over the first year of treatment, and high adherence was defined as PDC ≥80%. Regression models were used to identify predictors of high adherence to DOACs and to examine the association between adherence and stroke or SE. RESULTS: Overall 15,255 patients were included in this study. The proportion of highly adherent (PDC ≥80%) DOACs users was around 75% and decreased slightly over the years. On multivariable analyses, the likelihood of high adherence to DOACs increased with age and across higher socioeconomic classes, and was more likely among females, Jews, statins users, and patients with CHA2DS2-VASc score ≥2. Risk of stroke and SE was lower among highly adherent DOACs users; adjusted HR 0.56 (95% CI, 0.45-0.71), compared to users with PDC <80%. CONCLUSIONS: Adherence with DOACs is still sub-optimal among non-valvular AF patients, resulting in a higher risk of stroke and SE.

Novel mutation in coagulation factor VII (Carmel mutation): Identification and characterization.

BACKGROUND: Measurement of factor VII (FVII) activity does not enable prediction of bleeding tendency in individuals with inherited FVII deficiency. OBJECTIVE: To characterize the molecular and functional features of FVII in a family with FVII deficiency and correlate them with the bleeding tendency. PATIENTS/METHODS: We studied 7 family members with very low FVII activity using prothrombin time (PT), activated factor VII (FVIIa), FVII activity level, and thrombin generation. The factor 7 gene was sequenced and the mutation was analyzed by prediction software. RESULTS: The proband has very low FVII activity (0%-4%), with PT ranging between 5% to 18% depending on the tissue factor (TF) origin. Direct sequencing demonstrated a single homozygous nucleotide substitution G > A in exon 6, predicting a novel missense mutation Cys164Tyr. Three members of the family were found to be heterozygous carriers of this mutation. One of them was a compound heterozygote, carrying both the Cys164Tyr and Ala244Val mutation (linked to Arg353Gln polymorphism). Her FVII activity and antigen levels were 3%-7% and 8%, respectively. The other heterozygous carriers demonstrated FVII activity of 41%-54%, FVII antigen of 46%-66%, and FVIIa activity of 30%. FVIIa was undetectable in the homozygous and compound heterozygous subjects. Thrombin generation was normal in the presence of calcium, but no response to TF addition was observed in the homozygous proband, and a reduced response was observed in the compound heterozygous subject. CONCLUSION: The patient homozygous for the "Carmel" mutation has mild clinical manifestations despite very low FVII activity, which correlates with thrombin generation results.

Implementation of oral anticoagulation treatment guidelines in patients with newly diagnosed atrial fibrillation.

AIMS: Oral anticoagulants (OACs) are considered the mainstay in preventing stroke in atrial fibrillation (AF). OAC treatment remains suboptimal among AF patients, even after the introduction of direct oral anticoagulants (DOACs). We aimed to assess trends overtime and current implementation of OAC treatment guidelines in AF, using a large dataset of real world data from Israel. METHODS: This is a retrospective cohort study that includes all adult members of Clalit Health Services, the largest healthcare provider in Israel, with newly diagnosed nonvalvular AF between January 2014 and December 2019 with CHA2 DS2 -VASc score ≥2. OAC treatment rates were calculated and multivariate regression models were used to identify predictors of OACs initiation. RESULTS: Overall, 46 531 patients were included in the study. The 3-months cumulative OAC treatment rates increased consistently over the years: 46.9% (95% confidence interval, 46.1-47.7%), 54.9% (54.1-55.6%) and 61.7% (60.9-62.4%) during 2014-2015, 2016-2017 and 2018-2019, respectively. DOACs constituted 51.3% of prescribed OACs in 2014-2015 and increased to 95.1% during 2018-2019. On multivariate analyses, the likelihood of OACs initiation among AF patients increased across the years and across higher socioeconomic classes, and was more likely among females, Jews, statins users and patients previously screened for colorectal cancer, but less likely among smokers and patients with impaired renal function. The likelihood of treatment increased with higher CHA2 DS2 -VASc score and decreased with higher HAS-BLED score. CONCLUSION: Despite the increasing OAC treatment rates among high-risk AF patients, mainly attributed to the expanding DOAC use, OAC treatment scope is still far from optimal.

Willingness and concerns of transfusion-dependent hematological patients toward the option of home transfusion therapy.

BACKGROUND: One of the main obstacles of providing home-based palliative care to transfusion-dependent hematology patients is the lack of home transfusions services. While healthcare professionals are concerned with safety and cost of home transfusions, the attitude of the patients toward home transfusions are mostly unknown. AIM: To obtain quantitative data regarding the willingness and concerns of transfusion-dependent patients with hematological diseases toward the option of home transfusions. DESIGN: A cross sectional survey including a self-administered questionnaire in one of the three main spoken languages in Israel was administered to patients in 17 hospital hematology outpatient clinics between May 2019 and March 2020. RESULTS: About 52% of 385 patients that participated in the survey preferred home transfusions to hospital transfusions. Gender, age, education, or type of disease were not associated with preference for home transfusions, nor were hospital location or its size. The likelihood to prefer home transfusions was significantly higher among the Hebrew-speakers and those who had not experienced adverse effects previously. The most significant factor associated with preference of home transfusions was a perceived negative effect of hospital-based transfusion on quality of life. The main reason to reject home transfusions was fear of possible adverse effects and concerns over losing contact with the medical staff at the treating hospital. CONCLUSION: These data suggest that a significant portion of transfusion-dependent patients in Israel view home transfusions as a preferred treatment option and that its successful implementation requires maintaining ongoing contact with the treating hospital.

Association between myelofibrosis and risk of non-hematologic malignancies: a population-based retrospective cohort study.

Controversy regarding the risk of non-hematologic malignancies in myelofibrosis patients still exists. We aimed to examine the association between myelofibrosis and non-hematologic malignancies. A cohort of 1,469,790 adults without a diagnosis of myelofibrosis was identified on 1 January 2007, from the electronic medical records of the largest healthcare provider in Israel. Participants were followed up until 31 December 2015, for the occurrence of myelofibrosis. All cases of myelofibrosis were adjudicated by reviewing patients' electronic medical files. Using risk set sampling, four randomly selected controls (without myelofibrosis) were matched to each case of myelofibrosis on age, sex, ethnicity, and index date. Patients with and without myelofibrosis were followed from the index date until 31 December 2016 for the occurrence of non-hematologic malignancies based on the data from the Israel National Cancer Registry. The study included 550 patients with myelofibrosis and 2200 matched controls. Non-hematologic cancers occurred in 35 patients with myelofibrosis and 138 patients without myelofibrosis, reflecting a crude incidence rate of 27.9 and 15.3 per 1000 person-years, respectively. Myelofibrosis was independently associated with increased risk of non-hematologic malignancies with propensity score adjusted HR of 1.85 (95% CI, 1.09-3.15). No significant association was detected between myelofibrosis and the specific sites of non-hematologic malignancies. Treatment with ruxolitinib was not significantly associated with non-hematologic malignancies HR 1.36 (0.60-3.11). In conclusion, myelofibrosis appears to be associated with increased risk of non-hematologic malignancies. However, this study raises concerns about surveillance bias, suggesting that the association might be attributed to earlier detection rather than real increased risk.

Preoperative direct oral anticoagulants treatment and all-cause mortality in elderly patients with hip fracture: A retrospective cohort study.

INTRODUCTION: Hip fracture is common among the elderly and is associated with increased morbidity and mortality, particularly when surgery is delayed. Direct oral anticoagulants (DOACs) use might increase bleeding and postpone hip repair surgery. We aimed to assess the association between preoperative DOACs use and adverse outcomes in elderly patients with hip fracture. MATERIALS AND METHODS: This retrospective cohort study included all elderly patients (≥65 years), from the district of Haifa and Western Galilee, Israel, who underwent hip repair surgery for hip fracture between 2014 and 2018. Regression models with adjustment for propensity score were used to assess the association with all-cause mortality and other adverse outcomes. RESULTS: A total of 3418 patients with hip fracture were included of whom 163 (4.8%) were vitamin K antagonists (VKAs) users and 247 (7.2%) were DOCAs users. Propensity score adjusted models revealed that, compared to no anticoagulants use, DOACs use were independently associated with decreased risk of 30-day and 90-day mortality; HR 0.38 (95% CI, 0.17-0.88) and 0.47 (0.27-0.82), respectively. No significant associations were detected between VKAs use and all-cause mortality, compared to no anticoagulants use. DOACs and VKAs had significantly longer waiting time for hip repair surgery, and longer stay in hospital. DOACs and VKAs users had a non-significant higher estimated intraoperative bleeding. However, only VKAs users required a significantly higher number of blood transfusions. CONCLUSIONS: Albeit being associated with longer waiting time for surgery and longer hospitalization, DOACs use appears to be associated with reduced risk of mortality among elderly patients with hip fracture.

Association between myelofibrosis and thromboembolism: A population-based retrospective cohort study.

BACKGROUND: The risk of thromboembolism in myelofibrosis remains incompletely understood. OBJECTIVES: To examine the association between myelofibrosis and each of venous and arterial thromboembolism. METHODS: A cohort of 1 469 790 adults without a diagnosis of myelofibrosis was identified on 1 January 2007, from the electronic medical records of the largest health-care provider in Israel. Participants were followed until 31 December 2016 for the occurrence of myelofibrosis. Four randomly selected controls (without myelofibrosis) were matched to each case of myelofibrosis on age, sex, religious identification, and index date. The two groups were followed from the index date until 31 December 2017 for the occurrence of venous and arterial thromboembolism. RESULTS: The study included 642 patients with myelofibrosis and 2568 matched controls. Myelofibrosis was independently associated with increased risk of venous thromboembolism but not with arterial thromboembolism. The propensity score adjusted hazard ratios (HRs) were 6.88 (95% confidence interval [CI], 2.02-23.45) for venous thromboembolism, and 0.94 (0.49-1.77) for arterial thromboembolism. Atypical sites of venous thromboembolism occurred almost exclusively in patients with myelofibrosis (four events of Budd Chiari versus none, and two mesenteric vein thrombosis events versus one) and were more likely to occur around the time of myelofibrosis diagnosis. No significant association was found between JAK2 inhibitor treatment (ruxolitinib) and the risk of venous HR 0.97 (0.30-3.12) or arterial thromboembolism 1.68 (0.78-3.62). CONCLUSIONS: Myelofibrosis is associated with increased risk of venous thromboembolism but not of arterial thromboembolism. Atypical sites of venous thromboembolism are more frequent in myelofibrosis and are more likely to occur shortly after diagnosis.

Association of factor V activity with risk of venous thromboembolism and atherothrombotic cardiovascular events: A retrospective population-based cohort study.

INTRODUCTION: Factor V (FV) deficiency is a rare inherited coagulation disorder associated with bleeding tendency. As a result, it has been postulated that decreased FV activity may confer protection against venous thromboembolism and atherothrombotic cardiovascular events. MATERIALS AND METHODS: Using the electronic database of the largest health care provider in Israel, we identified all adult individuals who were tested for FV activity between January 2004 and June 2017. Subjects with liver cirrhosis or FV Leiden mutation were excluded. FV activity was classified into three predefined categories; FV activity >50%, FV activity 30-50%, and FV activity ≤30%. Patients were followed from January 2004 to June 2017 for new atherothrombotic cardiovascular events (composite of myocardial infarction, stroke, and TIA) and venous thromboembolism (VTE). RESULTS: Overall 2021 individuals were included; 83.2% had FV activity >50%, 9.6% FV activity 30-50%, and 7.2% had FV activity ≤30%. Compared to individuals with FV activity >50% the adjusted HR for atherothrombotic cardiovascular events was 1.10 (95% CI, 0.63-1.90) in those with FV activity 30-50%, and 0.95 (0.49-1.8) in those with FV activity ≤30%. None of the patients with FV activity 30-50% had VTE during follow-up; therefore those with FV activity ≤50% were classified into one group. VTE incidence was lower in those with FV activity ≤50% compared to those with FV >50% activity; adjusted HR = 0.28 (0.09-0.91). CONCLUSION: This study suggests that decreased FV activity might be associated with decreased incidence of VTE. No significant association appears to exist between FV activity and atherothrombotic cardiovascular events.

Factor XI deficiency is associated with lower risk for cardiovascular and venous thromboembolism events.

Factor XI deficiency is one of the rare inherited coagulation factor deficiencies. However, its incidence is high within the Ashkenazi Jewish community. Because factor XI displays both procoagulant and antifibrinolytic activities, it has been postulated that an underlying cardiovascular benefit may exist with factor XI deficiency. This historical cohort study was performed using the electronic database of Clalit Health Services, the largest health care provider in Israel. All adults tested for factor XI activity between 2002 and 2014 were included in the study. Factor XI activity was classified into 3 categories: normal (activity >50%), mild deficiency (activity = 30%-50%), and moderate-severe deficiency (activity ≤30%). The cohort was followed until 31 December 2015 for incidence of cardiovascular events (composite of myocardial infarction, stroke, and transient ischemic attack) and venous thromboembolism (VTE). Of the 10 193 included patients, 8958 (88.9%) had normal factor XI activity, 690 (6.8%) had mild deficiency, and 542 (5.3%) had moderate-severe deficiency. Compared with individuals with normal activity, the adjusted hazard ratio (HR) for cardiovascular events was 0.52 (95% confidence interval [CI], 0.31-0.87) in those with mild deficiency, and 0.57 (95% CI, 0.35-0.93) in those with moderate-severe factor XI deficiency. The incidence of VTE was lower in those with factor XI deficiency (activity <50%) compared with those with normal activity; adjusted HR = 0.26 (95% CI, 0.08-0.84). In summary, factor XI deficiency is associated with decreased incidence of cardiovascular events and VTE.

Laboratory tests for paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is a rare hematological disorder that is often suspected in a patient presenting with non-immune hemolytic anemia associated with pancytopenia or venous thrombosis. This disorder is a consequence of acquired somatic mutations in the phosphatidylinositol glycan class A (PIG-A) gene in the hematopoietic stem cells (HSC) of patients. The presence of these mutations leads to production of blood cells with decreased glycosyl phosphatidylinositol-anchored cell surface proteins, making red blood cells derived from the clone more sensitive to complement mediated hemolysis. The diagnosis of PNH may be difficult in some cases due a low proportion of PNH cells in the blood and occasionally due to difficulties in selecting the most appropriate diagnostic studies. The latest generation of tests allow for detection of very small populations of PNH cells, for following the natural course and response to therapy of the disease, and for helping to decide when to initiate therapy with monoclonal antibody targeting the terminal complement protein C5 (Eculizumab), anticoagulation, and in some cases allogeneic HSC transplant. In this article, we review the different diagnostic tests available to clinicians for PNH diagnosis.

Signaling pathways in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a deadly malignancy characterized by a plethora of molecular alterations that include major and minor driving mutations, the presence of intense desmoplasia exhibiting numerous proliferating pancreatic stellate cells (PSC) and cancer-associated fibroblasts that produce fibronectin and collagens, and foci of inflammatory cells that produce mitogenic cytokines. This review will focus on signaling by tyrosine kinase receptors, and the role of transforming growth factor beta in this malignancy is described briefly. Potential for therapeutic interventions will be discussed in relation to specific pathways.

MicroRNA-10b expression correlates with response to neoadjuvant therapy and survival in pancreatic ductal adenocarcinoma.

INTRODUCTION: Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy. Diagnosis and management of PDAC are hampered by the absence of sensitive and specific disease biomarkers. MicroRNAs (miRNA) are noncoding regulatory RNAs involved in initiation and progression of human cancers. In this study, we sought to determine whether miR-10b could serve as a biomarker for PDAC. EXPERIMENTAL DESIGN: miRNA expression was characterized by fluorescence-based in situ hybridization using locked nucleic acid-modified DNA probes against miR-10b, miR-21, miR-155, miR-196a, and miR-210, followed by codetection of proteins by immunohistochemistry on the same tissue sections. miRNA expression in surgically resected PDAC tissues and in endoscopic ultrasonography (EUS)-guided fine-needle aspirate (EUS-FNA) samples was analyzed in cytokeratin 19 (CK19)-positive epithelial cells using optical intensity analysis. RESULTS: In 10 resected PDAC samples, miR-10b was the most frequently and consistently overexpressed miRNA among characterized miRNAs, exhibiting a four-fold increase in the cancer cells (P = 0.012). Given this preferential overexpression of miR-10b, we sought to determine whether miR-10b expression was clinically relevant. Accordingly, miR-10b expression was examined in 106 EUS-FNA samples obtained from pancreatic lesions. miR-10b expression was increased in cancer cells compared with CK19-positive epithelial cells in benign lesions (P = 0.0001). In patients with PDACs, lower levels of miR-10b were associated with improved response to multimodality neoadjuvant therapy, likelihood of surgical resection, delayed time to metastasis, and increased survival. CONCLUSION: miR-10b is a novel diagnostic biomarker for PDACs when assessing pancreatic lesions. Expression of miR-10b is predictive of response to neoadjuvant therapy and outcome in this disease.

Fluorescence-based codetection with protein markers reveals distinct cellular compartments for altered MicroRNA expression in solid tumors.

PURPOSE: High-throughput profiling experiments have linked altered expression of microRNAs (miRNA) to different types of cancer. Tumor tissues are a heterogeneous mixture of not only cancer cells, but also supportive and reactive tumor microenvironment elements. To clarify the clinical significance of altered miRNA expression in solid tumors, we developed a sensitive fluorescence-based in situ hybridization (ISH) method to visualize miRNA accumulation within individual cells in formalin-fixed, paraffin-embedded tissue specimens. This ISH method was implemented to be compatible with routine clinical immunohistochemical (IHC) assays to enable the detection of miRNAs and protein markers in the same tissue section for colocalization and functional studies. EXPERIMENTAL DESIGN: We used this combined ISH/IHC assay to study a subset of cancer-associated miRNAs, including miRNAs frequently detected at low (miR-34a and miR-126) and high (miR-21 and miR-155) levels, in a panel of breast, colorectal, lung, pancreas, and prostate carcinomas. RESULTS: Despite the distinct histopathologic alterations of each particular cancer type, general trends emerged that pinpointed distinct source cells of altered miRNA expression. Although altered expressions of miR-21 and miR-34a were manifested within cancer cells, those of miR-126 and miR-155 were predominantly confined to endothelial cells and immune cells, respectively. These results suggest a heterogeneous participation of miRNAs in carcinogenesis by intrinsically affecting cancer cell biology or by modulating stromal, vascular, and immune responses. CONCLUSIONS: We described a rapid and sensitive multicolor ISH/IHC assay and showed that it could be broadly applied as an investigational tool to better understand the etiologic relevance of altered miRNA expression in cancer.