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Measles virus (MV) vaccine strains have shown significant preclinical antitumor activity against glioblastoma (GBM), the most lethal glioma histology. In this first in human trial (NCT00390299), a carcinoembryonic antigen-expressing oncolytic measles virus derivative (MV-CEA), was administered in recurrent GBM patients either at the resection cavity (Group A), or, intratumorally on day 1, followed by a second dose administered in the resection cavity after tumor resection on day 5 (Group B). A total of 22 patients received study treatment, 9 in Group A and 13 in Group B. Primary endpoint was safety and toxicity: treatment was well tolerated with no dose-limiting toxicity being observed up to the maximum feasible dose (2×10 7 TCID50). Median OS, a secondary endpoint, was 11.6 mo and one year survival was 45.5% comparing favorably with contemporary controls. Other secondary endpoints included assessment of viremia, MV replication and shedding, humoral and cellular immune response to the injected virus. A 22 interferon stimulated gene (ISG) diagonal linear discriminate analysis (DLDA) classification algorithm in a post-hoc analysis was found to be inversely (R = -0.6, p = 0.04) correlated with viral replication and tumor microenvironment remodeling including proinflammatory changes and CD8 + T cell infiltration in post treatment samples. This data supports that oncolytic MV derivatives warrant further clinical investigation and that an ISG-based DLDA algorithm can provide the basis for treatment personalization., (© 2024. The Author(s).)

Background: Attenuated measles virus (MV) strains are promising agents currently being tested against solid tumors or hematologic malignancies in ongoing phase I and II clinical trials; factors determining oncolytic virotherapy success remain poorly understood, however., Methods: We performed RNA sequencing and gene set enrichment analysis to identify pathways differentially activated in MV-resistant (n = 3) and -permissive (n = 2) tumors derived from resected human glioblastoma (GBM) specimens and propagated as xenografts (PDX). Using a unique gene signature we identified, we generated a diagonal linear discriminant analysis (DLDA) classification algorithm to predict MV responders and nonresponders, which was validated in additional randomly selected GBM and ovarian cancer PDX and 10 GBM patients treated with MV in a phase I trial. GBM PDX lines were also treated with the US Food and Drug Administration-approved JAK inhibitor, ruxolitinib, for 48 hours prior to MV infection and virus production, STAT1/3 signaling and interferon stimulated gene expression was assessed. All statistical tests were two-sided., Results: Constitutive interferon pathway activation, as reflected in the DLDA algorithm, was identified as the key determinant for MV replication, independent of virus receptor expression, in MV-permissive and -resistant GBM PDXs. Using these lines as the training data for the DLDA algorithm, we confirmed the accuracy of our algorithm in predicting MV response in randomly selected GBM PDX ovarian cancer PDXs. Using the DLDA prediction algorithm, we demonstrate that virus replication in patient tumors is inversely correlated with expression of this resistance gene signature (ρ = -0.717, P = .03). In vitro inhibition of the interferon response pathway with the JAK inhibitor ruxolitinib was able to overcome resistance and increase virus production (1000-fold, P = .03) in GBM PDX lines., Conclusions: These findings document a key mechanism of tumor resistance to oncolytic MV therapy and describe for the first time the development of a prediction algorithm to preselect for oncolytic treatment or combinatorial strategies.

Edmonston vaccine strains of measles virus (MV) have significant antitumor activity in mouse xenograft models of ovarian cancer. MV engineered to express the sodium iodide symporter gene (MV-NIS) facilitates localization of viral gene expression and offers a tool for tumor radiovirotherapy. Here, we report results from a clinical evaluation of MV-NIS in patients with taxol- and platinum-resistant ovarian cancer. MV-NIS was given intraperitoneally every 4 weeks for up to 6 cycles. Treatment was well tolerated and associated with promising median overall survival in these patients with heavily pretreated ovarian cancer; no dose-limiting toxicity was observed in 16 patients treated at high-dose levels (10(8)-10(9) TCID50), and their median overall survival of 26.5 months compared favorably with other contemporary series. MV receptor CD46 and nectin-4 expression was confirmed by immunohistochemistry in patient tumors. Sodium iodide symporter expression in patient tumors after treatment was confirmed in three patients by (123)I uptake on SPECT/CTs and was associated with long progression-free survival. Immune monitoring posttreatment showed an increase in effector T cells recognizing the tumor antigens IGFBP2 and FRα, indicating that MV-NIS treatment triggered cellular immunity against the patients' tumor and suggesting that an immune mechanism mediating the observed antitumor effect. Our findings support further clinical evaluation of MV-NIS as an effective immunovirotherapy., (©2014 American Association for Cancer Research.)

Background: Hepatocellular carcinoma (HCC) is the second most frequent cause of cancer death worldwide. Sulfatase 1 (SULF1) functions as a tumor suppressor in HCC cell lines in vitro , but also has an oncogenic effect in some HCCs in vivo ., Aim: To examine the mechanisms regulating SULF1 and its function in HCC., Methods: First, SULF1 mRNA and protein expression were examined. Second, we examined SULF1 gene copy number in HCC cells. Third, we assessed whether DNA methylation or methylation and/or acetylation of histone marks on the promoter regulate SULF1 expression. Finally, we examined the effect of 5-Aza-dC on sulfatase activity and drug-induced apoptosis., Results: SULF1 mRNA was down-regulated in 9/11 HCC cell lines but only 6/10 primary tumors. SULF1 mRNA correlated with protein expression. Gene copy number assessment by fluorescence in situ hybridization showed intact SULF1 alleles in low SULF1 expressing cell lines. CpG island methylation in the SULF1 promoter and two downstream CpG islands did not show an inverse correlation between DNA methylation and SULF1 expression. However, chromatin immunoprecipitation showed that the SULF1 promoter acquires a silenced chromatin state in low SULF1-expressing cells through an increase in di/trimethyl-K9H3 and trimethyl-K27H3 and a concomitant loss of activating acetyl K9, K14H3 marks. 5-Aza-dC restored SULF1 mRNA expression in SULF1-negative cell lines, with an associated increase in sulfatase activity and sensitization of HCC cells to cisplatin-induced apoptosis., Conclusion: SULF1 gene silencing in HCC occurs through histone modifications on the SULF1 promoter. Restoration of SULF1 mRNA expression by 5-Aza-dC sensitized HCC cells to drug-induced apoptosis.

Engineered measles virus (MV) strains deriving from the vaccine lineage represent a promising oncolytic platform and are currently being tested in phase I trials. In this study, we have demonstrated that MV strains genetically engineered to express the human sodium iodide symporter (NIS) have significant antitumor activity against glioma lines and orthotopic xenografts; this compares favorably with the MV strain expressing the human carcinoembryonic antigen, which is currently in clinical testing. Expression of NIS protein in infected cells results in effective concentration of radioactive iodine, which allows for in vivo monitoring of localization of MV-NIS infection by measuring uptake of (123)I or (99m)Tc. In addition, radiovirotherapy with MV-NIS followed by (131)I administration resulted in significant increase of MV-NIS antitumor activity as compared with virus alone in both subcutaneous (p=0.0003) and orthotopic (p=0.004) glioblastoma models. In conclusion, MV-NIS-based radiovirotherapy has significant antitumor activity against glioblastoma multiforme and represents a promising candidate for clinical translation.

Edmonston vaccine strains of measles virus (MV) have shown significant antitumor activity in preclinical models of ovarian cancer. We engineered MV to express the marker peptide carcinoembryonic antigen (MV-CEA virus) to also permit real-time monitoring of viral gene expression in tumors in the clinical setting. Patients with Taxol and platinum-refractory recurrent ovarian cancer and normal CEA levels were eligible for this phase I trial. Twenty-one patients were treated with MV-CEA i.p. every 4 weeks for up to 6 cycles at seven different dose levels (10(3)-10(9) TCID(50)). We observed no dose-limiting toxicity, treatment-induced immunosuppression, development of anti-CEA antibodies, increase in anti-MV antibody titers, or virus shedding in urine or saliva. Dose-dependent CEA elevation in peritoneal fluid and serum was observed. Immunohistochemical analysis of patient tumor specimens revealed overexpression of measles receptor CD46 in 13 of 15 patients. Best objective response was dose-dependent disease stabilization in 14 of 21 patients with a median duration of 92.5 days (range, 54-277 days). Five patients had significant decreases in CA-125 levels. Median survival of patients on study was 12.15 months (range, 1.3-38.4 months), comparing favorably to an expected median survival of 6 months in this patient population. Our findings indicate that i.p. administration of MV-CEA is well tolerated and results in dose-dependent biological activity in a cohort of heavily pretreated recurrent ovarian cancer patients.

Prostate cancer cells overexpress the measles virus (MV) receptor CD46. Herein, we evaluated the antitumor activity of an oncolytic derivative of the MV Edmonston (MV-Edm) vaccine strain engineered to express the human sodium iodide symporter (NIS; MV-NIS virus). MV-NIS showed significant cytopathic effect (CPE) against prostate cancer cell lines in vitro. Infected cells effectively concentrated radioiodide isotopes as measured in vitro by Iodide-125 ((125)I) uptake assays. Virus localization and spread in vivo could be effectively followed by imaging of (123)I uptake. In vivo administration of MV-NIS either locally or systemically (total dose of 9 x 10(6) TCID(50)) resulted in significant tumor regression (P < 0.05) and prolongation of survival (P < 0.01). Administration of (131)I further enhanced the antitumor effect of MV-NIS virotherapy (P < 0.05). In conclusion, MV-NIS is an oncolytic vector with significant antitumor activity against prostate cancer, which can be further enhanced by (131)I administration. The NIS transgene allows viral localization and monitoring by noninvasive imaging which can facilitate dose optimization in a clinical setting.

Purpose: MicroRNA (miRNA) is a new class of small, noncoding RNA. The purpose of this study was to determine if miRNAs are differentially expressed in hepatocellular carcinoma (HCC)., Experimental Design: More than 200 precursor and mature miRNAs were profiled by real-time PCR in 43 and 28 pairs of HCC and adjacent benign liver, respectively, and in normal liver specimens., Results: Several miRNAs including miR-199a, miR-21, and miR-301 were differentially expressed in the tumor compared with adjacent benign liver. A large number of mature and precursor miRNAs were up-regulated in the adjacent benign liver specimens that were both cirrhotic and hepatitis-positive compared with the uninfected, noncirrhotic specimens (P < 0.01). Interestingly, all of the miRNAs in this comparison had increased expression and none were decreased. The expression of 95 randomly selected mRNAs was not significantly altered in the cirrhotic and hepatitis-positive specimens, suggesting a preferential increase in the transcription of miRNA. Comparing the miRNA expression in the HCC tumors with patient's survival time revealed two groups of patients; those with predominantly lower miRNA expression and poor survival and those with predominantly higher miRNA expression and good survival (P < 0.05). A set of 19 miRNAs significantly correlated with disease outcome. A number of biological processes including cell division, mitosis, and G(1)-S transition were predicted to be targets of the 19 miRNAs in this group., Conclusion: We show that a global increase in the transcription of miRNA genes occurs in cirrhotic and hepatitis-positive livers and that miRNA expression may prognosticate disease outcome in HCC.

Background & Aims: The percentage of Lens culinaris agglutinin-reactive (alpha)-fetoprotein (AFP-L3%) is proposed as a diagnostic and prognostic marker for hepatocellular carcinoma (HCC). We evaluated the utility of AFP-L3% for diagnosis of HCC in a US referral population., Methods: This retrospective study included 272 patients: 166 with HCC and 106 with benign liver disease (chronic liver disease, 77; benign liver mass, 29). The AFP-L3% was measured using a clinical auto-analyzer., Results: The AFP-L3% is not reported for a total alpha-fetoprotein (AFP) less than 10 ng/mL, and all patients with an AFP greater than 200 ng/mL had HCC; thus the AFP-L3% was noninformative for these patients. In patients with a total AFP of 10-200 ng/mL, an AFP-L3% greater than 10% had a sensitivity of 71% and a specificity of 63% for diagnosis of HCC. An AFP-L3% greater than 35% had a reduced sensitivity of 33%, but an increased specificity of 100%. The high specificity of the AFP-L3% cut-off of 35% allowed the confident diagnosis of an additional 10% of HCCs not diagnosed using an AFP cut-off of 200 ng/mL. After adjustment for AFP level, no association was observed between AFP-L3% and tumor size, stage, vascular invasion, grade, or survival., Conclusions: Patients with indeterminate total AFP values of 10-200 ng/mL present a diagnostic dilemma. We found that an AFP-L3% greater than 35% has 100% specificity for HCC in these patients. AFP-L3%, used in combination with AFP, may be a clinically useful adjunct marker for the diagnosis of HCC.

Background & Aims: Hepatocellular carcinoma (HCC) is the third most common cause of cancer death worldwide. Improved treatments for advanced HCC are urgently needed. The recently identified human sulfatase 1 enzyme (SULF1) desulfates cell surface heparan sulfate glycosaminoglycans and down-regulates cell growth signaling in HCC cells in vitro. While investigating the epigenetic regulation of SULF1, we discovered that histone H4 acetylation is up-regulated by SULF1 in HCC cells. Histone deacetylase (HDAC) inhibitors reprogram cellular gene expression through the acetylation of nucleosomal histones and promote cell growth arrest and apoptosis. Hence, they are a promising modality for cancer treatment., Methods: To explore the interaction between SULF1 expression and HDAC inhibitor action, we examined the effects of SULF1 expression on HCC cells and xenografts treated with HDAC inhibitors., Results: (1) Forced expression of SULF1 significantly delayed the growth of Huh7 and Hep3B xenografts in nude mice in vivo. (2) SULF1 increased histone H4 acetylation by modulation of cellular HDAC and histone acetyltransferase activities. (3) SULF1 enhanced the induction of apoptosis by the HDAC inhibitors apicidin and scriptaid. (4) SULF1 enhanced the inhibition of tumor growth, migration, and angiogenesis by HDAC inhibitors. We also demonstrate that knockdown of SULF1 with shRNA constructs up-regulates phosphorylation of AKT and Erk and attenuates apicidin-induced apoptosis. The interaction between SULF1 and apicidin was confirmed in vivo in Huh7 and Hep3B xenografts., Conclusions: These results show that SULF1 promotes histone H4 acetylation, potentiates the effects of HDAC inhibitors, and inhibits HCC tumorigenesis.

Purpose: Production of reactive oxygen species (ROS) during chronic inflammation has been implicated in the progression of liver diseases and carcinogenesis. Subjects with inflammatory liver disease and one non-functional allele of the base excision repair gene, MYH, may be more susceptible to progression to cancer due to MYH haploinsufficiency in repairing oxidative damage caused by ROS. Here, we investigated the association of two common germline MYH mutations in patients with hepatocellular carcinoma (HCC) and cholangiocarcinoma., Methods: DNA from patients with HCC (n=48) or cholangiocarcinoma (n=84) compared to non-cancerous controls (n=308) were genotyped for the Y165C and G382D mutations in MYH., Results: There was no significant difference in MYH mutation carrier status between patients with HCC (1/48), cholangiocarcinoma (3/84), and non-cancerous controls (4/308)., Conclusions: Patients with HCC or cholangiocarcinoma do not have an increased incidence of monoallelic MYH mutations pre-disposing them to disease.

Background and Aims: The heparin-binding growth factors fibroblast growth factor (FGF) and hepatocyte growth factor (HGF) are potent mitogens for hepatocellular carcinomas (HCCs). Heparin-binding growth factor signaling is regulated by sulfation of cell-surface heparan sulfate proteoglycans (HSPGs). We hypothesized that hSulf1, a recently described sulfatase, regulates growth signaling in HCCs., Methods: Expression of hSulf1 in human HCC tumors was determined by real-time PCR. Down-regulation of hSulf1 expression was investigated by analyzing loss of heterozygosity (LOH) at the hSulf1 locus and the effect of the DNA methylation inhibitor 5-aza-deoxycytidine on hSulf1 expression. The subcellular location of hSulf1 and sulfation state of cell-surface HSPGs were assessed by immunocytochemistry. FGF and HGF signaling was examined by phospho-specific immunoblot analysis. Cell growth was measured by trypan blue exclusion, and the MTT assay and apoptosis were quantitated by fluorescence microscopy., Results: hSulf1 expression was decreased in 29% of HCCs and 82% of HCC cell lines. There was LOH at the hSulf1 locus in 42% of HCCs. Treatment with 5-aza-deoxycytidine reactivated hSulf1 expression in hSulf1-negative cell lines. Low hSulf1-expressing cells showed increased sulfation of cell-surface HSPGs, enhanced FGF and HGF-mediated signaling, and increased HCC cell growth. Conversely, forced expression of hSulf1 decreased sulfation of cell-surface HSPGs and abrogated growth signaling. HCC cells with high-level hSulf1 expression were sensitive to staurosporine- or cisplatin-induced apoptosis, whereas low expressing cells were resistant. Transfection of hSulf1 into hSulf1-negative cells restored staurosporine and cisplatin sensitivity., Conclusions: Down-regulation of hSulf1 contributes to hepatocarcinogenesis by enhancing heparin-binding growth factor signaling and resistance to apoptosis.