VP2 capsid domain of the H-1 parvovirus determines susceptibility of human cancer cells to H-1 viral infection.

Although H-1 parvovirus is used as an antitumor agent, not much is known about the relationship between its specific tropism and oncolytic activity. We hypothesize that VP2, a major capsid protein of H-1 virus, determines H-1-specific tropism. To assess this, we constructed chimeric H-1 viruses expressing Kilham rat virus (KRV) capsid proteins, in their complete or partial forms. Chimeric H-1 viruses (CH1, CH2 and CH3) containing the whole KRV VP2 domain could not induce cytolysis in HeLa, A549 and Panc-1 cells. However, the other chimeric H-1 viruses (CH4 and CH5) expressing a partial KRV VP2 domain induced cytolysis. Additionally, the significant cytopathic effect caused by CH4 and CH5 infection in HeLa cells resulted from preferential viral amplification via DNA replication, RNA transcription and protein synthesis. Modeling of VP2 capsid protein showed that two variable regions (VRs) (VR0 and VR2) of H-1 VP2 protein protrude outward, because of the insertion of extra amino-acid residues, as compared with those of KRV VP2 protein. This might explain the precedence of H-1 VP2 protein over KRV in determining oncolytic activity in human cancer cells. Taking these results together, we propose that the VP2 protein of oncolytic H-1 parvovirus determines its specific tropism in human cancer cells.

Cancer upregulated gene 2, a novel oncogene, confers resistance to oncolytic vesicular stomatitis virus through STAT1-OASL2 signaling.

We have recently found a novel oncogene, named cancer upregulated gene 2 (CUG2), which activates Ras and mitogen-activated protein kinases (MAPKs), including ERK, JNK and p38 MAPK. Because activation of these signaling pathways has previously been shown to enhance cancer cell susceptibility to oncolysis by certain viruses, we examined whether vesicular stomatitis virus (VSV) could function as a potential therapeutic agent by efficiently inducing cytolysis in cells transformed by CUG2. Unexpectedly, NIH3T3 cells stably expressing CUG2 (NIH-CUG2) were resistant to VSV because of the activation of signal transducers and activators of transcription 1 (STAT1). The result was supported by evidence showing that suppression of STAT1 with short interference RNA (siRNA) renders cells susceptible to VSV. Furthermore, 2'-5' oligoadenylate synthetase-like (OASL) 2 was the most affected by STAT1 expression level among anti-viral proteins and furthermore suppression of OASL2 mRNA level caused NIH-CUG2 cells to succumb to VSV as seen in NIH-CUG2 cells treated with STAT1 siRNA. In addition, Colon26L5 carcinoma cells stably expressing CUG2 (Colon26L5-CUG2) exhibited resistance to VSV, whereas Colon26L5 stably expressing a control vector yielded to VSV infection. Moreover, Colon26L5-CUG2 cells stably suppressing STAT1 succumbed to VSV infection, resulting in apoptosis. Taken together, we propose that VSV treatment combined with the selective regulation of genes such as STAT1 and OASL2 will improve therapeutic outcomes for CUG2-overexpressing tumors.

CUG2, a novel oncogene confers reoviral replication through Ras and p38 signaling pathway.

As we have recently found a novel oncogene, the cancer-upregulated gene 2 (CUG2), which was elevated in a variety of tumor tissues such as the ovary, liver, lung and pancreas, we examined whether reovirus could efficiently induce cytolysis in cancer cells expressing CUG2 and thus be used as a potential cancer therapeutic agent. In this study, we describe experiments in which we use reovirus to treat NIH3T3 cells stably expressing either CUG2 (NIH-CUG2) or vector only (NIH-Vec). NIH-CUG2 cells readily support reoviral proliferation and undergo apoptosis, whereas NIH-Vec cells are highly resistant to reoviral infection and virus-induced apoptosis. This notable result may be explained by the observation that CUG2 expression inhibits PKR activation, leading to reoviral proliferation in nonpermissive NIH3T3 cells. Furthermore, reovirus infection results in almost complete regression of tumorgenic NIH-CUG2 cells in transplanted nude mice. As we found that CUG2 enhances activation of MAPK (ERK, JNK and p38), Src kinase and Ras, we examined whether CUG2 confers reoviral replication independent of the Ras or p38 MAPK signaling pathway. From these experiments we found that either inhibition of p38 MAPK or Ras blocks reoviral proliferation even in the presence of CUG2 but inhibition of ERK, JNK and Src kinase does not, indicating that activation of p38 MAPK and Ras has critical roles in reoviral replication in CUG2-expressing tumor cells. Accordingly, we propose that reovirus can be useful in the treatment of transformed cells expressing CUG2, which is commonly detected in various tumor tissues.

Down-regulation of HIF-1alpha by oncolytic reovirus infection independently of VHL and p53.

Many oncolytic viruses are currently being tested as potential cancer therapeutic agents. To be effective, these viruses must replicate and propagate efficiently through the tumor mass. However, it is possible that the hypoxia that characterizes many tumors may be an obstacle to viral therapy because of its inhibition of viral replication and propagation. We, therefore, decided to test how oncolytic reovirus and its target cells respond to hypoxia. We found that reovirus infection suppresses hypoxia inducible factor (HIF)-1alpha protein levels (but not transcript abundance) in colon cancer HCT116 cells under CoCl(2) or hypoxia. Reovirus infection was able to reduce HIF-1alpha levels in both von Hippel Lindau (VHL)-/- renal carcinoma A498 and p53-/- HCT116 cells, indicating that the decrease of HIF-1alpha mediated by reovirus requires neither VHL nor p53 proteins. However, treatment with the inhibitor MG132 restored HIF-1alpha levels, suggesting that reovirus-induced HIF-1alpha decrease needs proteosomal activity. A498 VHL-/- cells with constitutive expression of HIF-1alpha were relatively resistant to reovirus-induced apoptosis when compared with A498 VHL+/+ cells. However, we found that the use of YC-1 to target HIF-1alpha promoted reovirus-induced apoptosis in A498 VHL-/- cells. Accordingly, we propose that reovirus may be used together with YC-1 as a potential therapeutic agent against chemoresistant or radioresistant tumors that are hypoxic and show increased levels of HIF-1alpha.

Expression of HBX, an oncoprotein of hepatitis B virus, blocks reoviral oncolysis of hepatocellular carcinoma cells.

Although reovirus has been used in tests as a potential cancer therapeutic agent against a variety of cancer cells, its application to hepatocellular carcinoma cells, in which the hepatitis B virus (HBV) X (HBX) protein of HBV plays a primary role, has not yet been explored. Here, we describe experiments in which we use reovirus to treat Chang liver carcinoma cells expressing either a vector only (Chang-vec) or a vector encoding HBX protein (Chang-HBX). Although Chang-vec cells readily support reoviral proliferation and undergo apoptosis, Chang-HBX cells are highly resistant to reoviral infection and virus-induced apoptosis, even though HBX protein induces activation of Ras and inactivation of PKR, which are normally thought to enhance reoviral oncolysis. The resistance of Chang-HBX cells to reovirus may instead be explained by HBX-induced downregulation of death receptor 5 and activation of Stat1. Phosphorylated Stat1 activates interferon (IFN)-stimulated regulatory element (ISRE)- and IFN-gamma-activated sequence (GAS)-mediated transcription, leading to the production of IFN-beta, whereas the reduced expression of Stat1 with its siRNA results in a decrease in IFN-beta production, by which Chang-HBX cells eventually succumb to reovirus infection. This result further indicates that HBX induces the establishment of an antiviral state through Stat1 activation. Thus, it appears that active Ras does not override the antiviral effect mediated by the activation of Stat1. Accordingly, we report that HBX, an oncoprotein of HBV, can prevent reoviral oncolysis of hepatocellular carcinoma. This suggests there may be limits to the practical application of reovirus in the treatment of human cancers already expressing other oncoviral proteins.

Prostate blood flow characteristics in the chronic prostatitis/pelvic pain syndrome.

PURPOSE: We determine whether the chronic prostatitis/pelvic pain syndrome is associated with abnormal prostate blood flow. MATERIALS AND METHODS: We used color Doppler ultrasonography to examine 53 patients with inflammation, 80 men without inflammation and 22 healthy controls. Images were recorded and scored using standardized criteria to characterize the degree and distribution of prostatic vascularity. RESULTS: Flow was observed to the entire prostatic capsule in 77% of patients but only 18% of controls (p<0.0001). Parenchymal flow was evaluated using several criteria. On a 2-point scale flow was classified as grade 2 in 74% of patients compared to 27% of controls (p<0.0001). Similar findings were noted on a Doppler spot scale, with flow classified as grade 2 in 47% of patients compared to 14% of controls (p<0.004). Patients also had more parenchymal Doppler spots than controls (p<0.01). Diffuse blood flow throughout the prostatic parenchyma was observed in 63% of patients compared to 36% of controls (p<0.03). There was no significant difference in the amount or distribution of blood flow in patients with and without inflammation. CONCLUSIONS: The chronic prostatitis/pelvic pain syndrome was associated with increased blood flow to the prostatic capsule and diffuse flow throughout the prostatic parenchyma. Despite technical limitations, color Doppler ultrasonography may provide objective documentation of prostate blood flow abnormalities in patients with this syndrome.

A hemizygous short tandem repeat polymorphism 3' to the human phosphoglycerate kinase gene.

The human phosphoglycerate kinase (PGK) gene is located within Xq11-Xq13, a region implicated in genitourinary diseases including: prostate cancer, androgen insensitivity, perineal hypospadias, and other genetic abnormalities. The PGK gene and the androgen receptor gene are in linkage disequilibrium. PGK has been mapped extensively for nuclease-sensitive sites, methylation sites, and flanking DNA sequences. A PGK-associated BstXI polymorphism has been used to determine clonality of neoplastic tissues. Using fluorescent PCR product analysis and DNA sequencing, we discovered that a short tandem repeat (STR) in the 3' flanking region of the PGK gene is polymorphic. Among 231 individuals, there were nine distinct alleles, including eight based on variations in the number of TATC repeats. The PGK STR demonstrated hemizygosity, consistent with its X-chromosomal location and with an absence of cross-hybridizing autosomal homologs. The polymorphic PGK STR shows promise for rapid investigation of neoplastic clonality, for personal identification, and for studies of inherited predisposition to urologic disorders.

Magnetic resonance imaging in hemospermia.

PURPOSE: We evaluated the prostate and seminal tract with magnetic resonance imaging (MRI) in patients with hemospermia. MATERIALS AND METHODS: To evaluate the prostate and seminal tract in 17 patients 20 to 59 years old (mean age 44) with hemospermia we performed transrectal ultrasound and MRI using an endorectal surface coil with a 1.5 tesla unit. Mean duration of hemospermia was 32 months (1 week to 16 years). RESULTS: Abnormalities were noted on transrectal ultrasound in 15 of the 17 patients (88%) and on MRI in all. Of the 12 cases of hemorrhage 10 involved the seminal vesicle and 2 involved the ejaculatory duct. There were 12 cystic lesions, including 7 in the müllerian and 5 in the ejaculatory ducts. Of 19 cases calculi were detected in the prostate in 5, seminal vesicle in 8, and ejaculatory and müllerian duct cysts in 4 and 2, respectively. There was 1 case of prostatic atrophy and 1 wolffian duct anomaly associated with an ejaculatory duct cyst, ectopic ureterocele and absence of the left kidney. CONCLUSIONS: MRI with an endorectal surface coil is a powerful modality for evaluating the seminal tracts of patients with hemospermia. It can be performed clinically when transrectal ultrasonography is not satisfactory.

Ten years of experience with various penile prosthesis in Korean.

Currently there are more than 10 types of penile prosthesis available, ranging from the very simple to the very sophisticated. We review our experiences with various penile prosthesis, with particular regard to the complication rate. From Dec. 1983 to Jul. 1993, we implanted 295 penile prosthesis of eight different types. The average age of patients was 44 years. Every patient was evaluated with various multidisplinary diagnostic approaches. The etiologies of impotence were vasculogenic 29%, diabetogenic 22%, spinal cord injury 16%, pelvic bone injury 11%, etc. The types of implanted prosthesis were AMS malleable 143, Jonas 42, Dynaflex 36, Hydroflex 8, Uni-Flate 1000 2, AMS 700 CXM 58, Ultrex 3, Mentor alpha-1 3 and the mean follow-up period was 34 months. The diameters of implanted prosthesis were from 9.5 mm to 13 mm, mostly 9.5 mm (52.9%). The length of implanted prosthesis were from 10 cm to 20 cm, mostly 16 approximately 18 cm (68.8%). Cases with uneven diameters or lengths were 20 (6.8%). The int aoperative complications were 1 corporeal rupture and 1 bladder rupture, and the postoperative complications were 2 prosthesis infections, 2 mechanical failures, and 1 prosthesis infection with mechanical failure. In those 4 patients reimplantations were successful. More than 99% (290/291) patients still have functioning prosthesis. Every prosthesis has their advantages and disadvantages. Factors to be analysed in the selection of proper prosthesis should include patients economic status, education, personality, social activity, hand dexterity, and penile size.(ABSTRACT TRUNCATED AT 250 WORDS)