Search PubMed⌕ Search

Biomedical subjects

C Kao

Publications and source records attributed to C Kao.

At least 37 records · Page 2Linked to original sources
PubMed2000

Dynamic structure factor of diamond and LiF measured using inelastic X-Ray scattering

The dynamic structure factors S(q-->,omega) of diamond and LiF have been measured using inelastic x-ray scattering. The experimental data are compared to results of ab initio calculations, which take into account the interaction of the excited electron with the remaining hole. In diamond, the vicinity of the indirect band gap and its momentum dependence are studied. In LiF, a larger energy range, which covers the fundamental exciton, the plasmon, and several interband transitions, is investigated. Calculations and measurements agree quite well and emphasize the need to properly include the interaction of the excited electron in the conduction band with the hole in the valence band.

Journal Article↗
PubMed2000

Osteocalcin-directed gene therapy for prostate-cancer bone metastasis.

Osteocalcin (OC) is a major noncollagenous bone protein whose expression is limited almost exclusively to osteotropic tumors and mature calcified tissue (differentiated osteoblasts). The function of OC, a highly conserved gamma-carboxyglutamic acid-containing protein, relies in part on its ability to bind hydroxyapatite and act as a chemoattractant for bone-resorbing cells. Serum osteocalcin levels are used clinically as an index of active bone turnover. Research in our laboratory has revealed that OC is expressed in several solid tumors, including osteosarcoma and ovarian, lung, brain, and prostate cancers. Evidence arising from reverse-transcription polymerase chain reaction (RT-PCR; detection of OC mRNA), immunohistochemical staining (detection of OC protein), and transient transfection and reporter assay (detection of OC mRNA transcription) reveals that OC expression is up-regulated in numerous solid tumors, with its expression being further elevated in androgen-independent prostate cancers. A recombinant, replication-defective adenovirus, Ad-OC-TK (OC promoter-driven herpes-simplex-virus thymidine kinase) was constructed and, when combined with the appropriate prodrug, either ganciclovir (GCV) or acyclovir (ACV), was found to be effective at destroying prostate-cancer cell lines in vitro and prostate tumor xenografts in vivo in both subcutaneous and bone sites. Additionally, via use of the OC promoter the supporting bone stromal cells are cotargeted when the prostate cancer interdigitates with bone stroma at the metastatic skeletal sites. Thus, maximal tissue-specific cell toxicity is achieved by the interruption of cellular communication between the prostate cancer and the bone stroma. We describe herein the preclinical foundation as well as the design and implementation of an ongoing phase I clinical trial at the University of Virginia that targets androgen-independent metastatic prostate cancer using the Ad-OC-TK vector.

Adenoviridae↗
PubMed2000

Using technetium-99M dimercaptosuccinic acid renal cortex scintigraphy to differentiate acute pyelonephritis from other causes of fever in patients with spinal cord injury.

OBJECTIVES: To differentiate acute pyelonephritis (APN) from fever due to other sources in patients with spinal cord injury by using technetium-99m dimercaptosuccinic acid (DMSA) renal cortex scintigraphy (DMSA scan). METHODS: A total of 24 patients with spinal cord injury were admitted with fever. DMSA scans were performed on all patients. The final determination of the cause of the fever was based on the medical history, physical examination, laboratory evaluation, and imaging studies. RESULTS: DMSA scan accurately diagnosed APN in 12 patients with inflammation on the DMSA scan. In 5 patients with scarred lesions on the DMSA scans and in 7 patients with negative DMSA scan, the fever was attributed to other causes. The sensitivity and specificity of the DMSA scan for detecting APN were both 100%. CONCLUSIONS: The DMSA scan is a valuable adjunct in the evaluation of fever in patients with SCI.

Acute Disease↗
PubMed2000

Tissue-specific promoters in gene therapy for the treatment of prostate cancer.

Delivery of therapeutic toxic genes to and their expression in tumor cells through the use of tissue-specific promoters could decrease their toxic effect on neighboring normal cells when virus-mediated gene delivery results in their infection. We have demonstrated the utility of two prostate cancer-specific promoters, long PSA and osteocalcin, for tissue-specific toxic gene therapy for prostate cancer. The two promoters were highly active in both androgen-dependent and androgen-independent prostate cancer cells. We also introduce the Phase I trial of osteocalcin promoter-based toxic gene therapy for bone metastases of prostate cancer, which is in progress at the University of Virginia.

Acyclovir↗
PubMed1999

The N-terminal half of the brome mosaic virus 1a protein has RNA capping-associated activities: specificity for GTP and S-adenosylmethionine.

The N-terminal half of the brome mosaic virus (BMV) 1a replication-associated protein contains sequence motifs found in RNA methyltransferases. We demonstrate that recombinant BMV methyltransferase-like (MT) domain expressed in Escherichia coli forms an adduct with a guanine nucleotide in a reaction that requires S-adenosylmethionine (AdoMet) and divalent cations. Moieties in GTP and AdoMet required for adduct formation were determined using a competition assay and chemical analogues. In the guanine nucleotide the ribose 2' hydroxyl, the triphosphates, the base C6 keto group, and possibly the N1 imine are required. In AdoMet, the methyl group and the ability to transfer a methyl group to guanine nucleotide were demonstrated to be required for adduct formation. The effects of methyltransferase inhibitors on viral RNA synthesis was determined using an in vitro RNA synthesis assay. These results are consistent with the previously reported activities of alphaviral nsP1 methyltransferase protein and identify the chemical moieties required for the BMV methyltransferase activity.

Bromovirus↗
PubMed1999

A simple and efficient method to reduce nontemplated nucleotide addition at the 3 terminus of RNAs transcribed by T7 RNA polymerase.

DNA templates modified with C2'-methoxyls at the last two nucleotides of the 5' termini dramatically reduced nontemplated nucleotide addition by the T7 RNA polymerase from both single- and double-stranded DNA templates. This strategy was used to generate several different transcripts. Two of the transcripts were demonstrated by nuclear magnetic resonance spectroscopy to be unaffected in their sequence. Transcripts produced from the modified templates can be purified with greater ease and should be useful in a number of applications.

Base Sequence↗
PubMed1999

Cytotoxicity of adenoviral-mediated cytosine deaminase plus 5-fluorocytosine gene therapy is superior to thymidine kinase plus acyclovir in a human renal cell carcinoma model.

PURPOSE: An estimated 11,600 Americans will die of renal cell carcinoma in 1998. The lack of effective chemotherapy or radiotherapy requires the investigation of novel treatment modalities. We compared two forms of toxic gene therapy, cytosine deaminase (CD) plus 5-fluorocytosine (5-FC) and thymidine kinase (TK) plus acyclovir (ACV), in pre-clinical models of human renal cell carcinoma. MATERIALS AND METHODS: Replication-deficient recombinant adenoviral vectors containing the Rous sarcoma virus promoter driving CD (Ad-RSV-CD) or TK (Ad-RSV-TK) gene expression were constructed and tested for in vitro cell-killing assays at various viral multiplicity of infection (MOI) and in vivo for growth inhibition of a human renal cell carcinoma, SK-RC-29 models. Subcutaneous tumors of SK-RC-29 were examined by electron microscopy for presence of intercellular gap junctions. Levels of expression of the gap junctional associated connexin 43 protein in SK-RC-29, 31, 38, 42, 52 human RCC cell lines were examined by western immunoblotting. RESULTS: In vitro cell-killing assay comparing Ad-RSV-CD/5F-C and Ad-RSV-TK/ACV at a wide range of MOI (2.5 to 20) revealed superior cell-kill by Ad-RSV-CD/5-FC over Ad-RSV-TK/ACV. Consistent with these results, we observed that Ad-RSV-CD/5-FC but not Ad-RSV-TK/ACV demonstrated a significant in vivo tumor growth inhibition. These results are corroborated by the lack of gap junctions in SK-RC-29 subcutaneous tumors by the electron microscopy and the absence of connexin-43 in all five human RCC cell lines by western immunoblotting. CONCLUSION: We have demonstrated in this study that Ad-RSV-CD/5-FC is superior to Ad-RSV-TK/ACV for the treatment of human RCC in cell culture and animal models. The results are supported by the lack of gap junctional communication between RCC cells assessed by connexin-43 expression.

Acyclovir↗
PubMed1998

Establishing human prostate cancer cell xenografts in bone: induction of osteoblastic reaction by prostate-specific antigen-producing tumors in athymic and SCID/bg mice using LNCaP and lineage-derived metastatic sublines.

LNCaP lineage-derived human prostate cancer cell lines C4-2 and C4-2B4 acquire androgen independence and osseous metastatic potential in vivo. Using C4-2 and C4-2B4 the goals of the current investigation were 1) to establish an ideal bone xenograft model for prostate cancer cells in intact athymic or SCID/bg mice using an intraosseous route of tumor cell administration and 2) to compare prostate cancer metastasis by administering cells either through intravenous (i.v.) or intracardiac administration in athymic or SCID/bg mice. Subsequent to tumor cell administration, prostate cancer growth in the skeleton was assessed by radiographic bone density, serum prostate-specific antigen (PSA) levels, presence of hematogenous prostate cancer cells and histopathologic evaluation of tumor specimens in the lymph node and skeleton. Our results show that whereas LNCaP cells injected intracardially failed to develop metastasis, C4-2 cells injected similarly had the highest metastatic capability in SCID/bg mice. Retroperitoneal and mediastinal lymph node metastases were noted in 3/7 animals, whereas 2/7 animals developed osteoblastic spine metastases. Intracardiac injection of C4-2 in athymic hosts produced spinal metastases in 1/5 animals at 8-12 weeks post-injection; PC-3 injected intracardially also metastasized to the bone but yielded osteolytic responses. Intravenous injection of either LNCaP or C4-2 failed to establish tumor colonies. Intrailiac injection of C4-2 but not LNCaP nor C4-2B4 cells in athymic mice established rapidly growing tumors in 4/8 animals at 2-7 weeks after inoculation. Intrafemoral injection of C4-2 (9/16) and C4-2B4 (5/18) but not LNCaP (0/13) cells resulted in the development of osteoblastic bone lesions in athymic mice (mean: 6 weeks, range: 3-12 weeks). In SCID/bg mice, intrafemoral injection of LNCaP (6/8), C4-2 (8/8) and C4-2B4 (8/8) cells formed PSA-producing, osteoblastic tumors in the bone marrow space within 3-5 weeks after tumor cell inoculation. A stepwise increase of serum PSA was detected in all animals. Reverse transcription-polymerase chain reaction (RT-PCR) to detect hematogenously disseminated prostate cancer cells could not be correlated to either serum PSA level or histological evidence of tumor cells in the marrow space. We have thus established a PSA-producing and osteoblastic human prostate cancer xenograft model in mice.

Animals↗
PubMed1998

Overcoming cellular senescence in human cancer pathogenesis.

Elevation of p16, the CDKN2/p16 tumor suppressor gene (TSG) product, occurs at senescence in normal human uroepithelial cells (HUC). Immortal HUCs and bladder cancer cell lines show either alteration of p16 or pRb, the product of the retinoblastoma (RB) TSG. In addition, many human cancers show p16 or pRb alteration along with other genetic alterations that we associated with immortalization, including +20q and -3p. These observations led us to hypothesize that p16 elevation plays a critical role in senescence cell cycle arrest and that overcoming this block is an important step in tumorigenesis in vivo, as well as immortalization in vitro. Using a novel approach, we tested these hypotheses in the present study by examining p16 and pRb status in primary culture (P0) and after passage in vitro of transitional cell carcinoma (TCC) biopsies that represented both superficial bladder tumors and invasive bladder cancers. We demonstrated that all superficial TCCs showed elevated p16 after limited passage in vitro and then senesced, like normal HUCs. In contrast, all muscle invasive TCCs contained either a p16 or a pRb alteration at P0 and all spontaneously bypassed senescence (P = 0.001). Comparative genomic hybridization (CGH) was used to identify regions of chromosome loss or gain in all TCC samples. The application of a statistical model to the CGH data showed a high probability of elevated alteration rates of +20q11-q12 (0.99) and +8p22-pter (0.94) in the immortal muscle invasive TCCs, and of -9q (0.99) in the superficial TCCs. Three myoinvasive TCCs lost 3p13-p14. In this study, four of six myoinvasive TCCs also showed TP53 mutation that associated well with genome instability (P = 0.001), as previously hypothesized. Notably, TP53 mutation, which has been used as a marker of tumor progression in many human cancers, was less significant in associating with progression in this study (P = 0.04) than was p16 or pRb alteration (P = 0.001). Thus, these data support a new model in which overcoming senescence plays a critical role in human cancer pathogenesis and requires at least two genetic changes that occur in several combinations that can include either p16 or pRb loss and at least one additional alteration, such as +20q11-q12, -3p13-p14, or -8p21-pter.

Adult↗
PubMed1998

Reconstitution of estrogen-dependent transcriptional activation of an adenoviral target gene in select regions of the rat mammary gland.

Estrogen regulates proliferation and morphogenesis of mammary ductal epithelium by interacting with a specific intracellular estrogen receptor (ER) that acts as a hormone-dependent transcriptional regulator of gene expression. The mechanisms by which ER regulates transcription in response to estrogen have been analyzed extensively in tissue culture and in cell-free systems. These studies have demonstrated that the transcriptional activity of ER is strongly influenced by cellular context and highlight the need to address ER transcriptional activity in an appropriate cellular background. Thus, to gain insight into the mechanistic role of ER in mammary epithelial morphogenesis, we have used an adenoviral gene delivery strategy to introduce an estrogen-responsive reporter gene into the mammary epithelium and to monitor the activity of endogenous ERs in their natural environment where cellular context including stromal-epithelial interactions can be taken into account. Using this approach, we first demonstrated highly efficient adenoviral delivery throughout the mammary epithelium using a beta-galactosidase (betagal) reporter gene under the control of the constitutively active cytomegalovirus (CMV) promoter. Next, we constructed an adenoviral vector by substituting the CMV promoter with an estrogen-dependent promoter fragment-linked betagal (Ad-ERE-tk-betagal). This adenoviral reporter system provides evidence that ER positive mammary epithelial cells display a differential sensitivity in a region-specific manner toward estrogen induction. Our data suggest that the availability of factor(s) other than ER is necessary for ER-mediated gene activation and may be important in modulating the differential responses of mammary epithelial cells to estrogen.

Adenoviridae↗
PubMed1998

Development of prostate-specific antigen promoter-based gene therapy for androgen-independent human prostate cancer.

PURPOSE: The goal of this study is to develop a tissue-specific toxic gene therapy utilizing the prostate specific antigen (PSA) promoter for both androgen-dependent (AD) and androgen-independent (AI) PSA-secreting prostate cancer cells. Ideally this gene therapy would be effective without the necessity of exposing the target cells to circulating androgens. MATERIALS AND METHODS: An AI subline of LNCaP, an AD PSA-secreting human prostate cancer cell line, C4-2, was used in this study. Castrated mice bearing C4-2 tumors secrete PSA. A transient expression experiment was used to analyze the activity of two PSA promoters, a 5837 bp long PSA promoter and a 642 bp short PSA promoter, in C4-2 cells. A recombinant adenovirus (Ad-PSA-TK) carrying thymidine kinase under control of the long PSA promoter was generated. The tissue-specific activity of Ad-PSA-TK was tested in vitro and in vivo. RESULTS: The long PSA promoter had superior activity over short PSA promoter, and higher activity in C4-2 cells than in LNCaP cells. High activity of Ad-PSA-TK was observed in C4-2 cells in an androgen deprived condition. In vitro, Ad-PSA-TK was further demonstrated to induce marked C4-2 cell-kill by acyclovir in medium containing 5% FBS. No cell-kill was observed in control WH cells (a human bladder cancer cell line). In vivo, Ad-PSA-P-TK with acyclovir significantly inhibited subcutaneous C4-2 tumor growth and PSA production in castrated animals. CONCLUSION: The 5837 bp long PSA promoter was active in the androgen free environment and could be used to target both androgen-dependent and independent PSA-producing prostate cancer cells in vitro, and prostate tumors in castrated hosts.

Adenoviridae↗
PubMed1997

Cytotoxic effects of recombinant adenovirus p53 and cell cycle regulator genes (p21 WAF1/CIP1 and p16CDKN4) in human prostate cancers.

PURPOSE: Overexpressing or restoring the basal levels of tumor suppressor genes in cancer cells can suppress tumorigenicity of cancer cells. In this communication, we compared tumor suppressive activities of three well-defined tumor suppressive genes (p53, p21WAF1/CIP1, and p16CDKN2) delivered individually to prostate cancer cells with adenoviral vector (Ad). MATERIALS AND METHODS: Efficacy of growth inhibition by recombinant adenoviruses bearing p53, p21WAF1/CIP1, or p16CDKN2 (Ad5CMV-p53, Ad5CMV-p21, Ad5CMV-p16) genes were tested in vitro on androgen-dependent (LNCaP) and androgen-independent (C4-2, DU-145, and PC-3) human prostate cancer cells, ex vivo and in vivo on PC-3 tumor. RESULTS: Ad5CMV-p53 was observed to exert the greatest growth inhibitory action on all of the cell lines tested; inhibition appeared to be cytolytic. In comparison to control Ad5CMV-PA added samples, the growth inhibitory action of Ad5CMV-p21 and Ad5CMV-p16 appeared to be cytostatic. Ad5CMV-p53 is more effective than Ad5CMV-p16 and Ad5CMV-p21 in inhibiting the tumor "take" rate. A similar order of antitumor activity was observed when recombinant adenoviruses were injected intratumorily to previously established PC-3 tumors in vivo. CONCLUSION: p53 is the most effective tumor suppressor gene to target human prostate cancer.

Adenoviruses, Human↗
PubMed1997

Human prostate cancer progression models and therapeutic intervention.

Our laboratory has developed two cellular models of human prostate cancer progression. The LNCaP prostate cancer progression model is based upon the well-known cellular interaction between human prostate or bone stromal cells and LNCaP cells in vivo. The marginally tumorigenic LNCaP cells acquired tumorigenic and metastatic potential upon cellular interaction with either prostate or bone fibroblasts. A subline termed C4-2 was observed to grow readily in castrated animals and acquired metastatic potential spreading from the primary tumor site to the lymph node, the seminal vesicles, and the axial skeleton, resulting in an intense osteoblastic reaction. The second model is ARCaP, where prostate cancer cells derived from the ascites fluid of a man with metastatic disease exhibited an Androgen- and estrogen-Repressed Prostate Cancer cell growth and tumor formation in either a hormone-deficient or a castrated environment. However, the growth of either the tumor cells in vitro or the tumors in vivo was suppressed by both estrogen and androgen. While the tumor cells expressed low levels of androgen receptor and prostate-specific antigen (PSA), they were highly metastatic when inoculated orthotopically. Distant metastases to a number of organs were detected, including the liver, lung, kidney, and bone. We have employed a human prostate cancer progression model as a system to study the efficacy of gene therapy. Results of the study show that whereas universal promoters, such as Cytomegalovirus (CMV) and Rous Sarcoma Virus (RSV) promoter-driven tumor suppressors (e.g. p53, p21, and p16), were effective in inhibiting prostate tumor growth, the advantages of driving the expression of therapeutic toxic genes using a tissue-specific promoter prostate-specific antigen (PSA) and a tumor--but not tissue-specific promoter, osteocalcin (OC), are preferred. In the case of the PSA promoter, we can achieve cell-kill in PSA-producing human prostate cancer cells. To circumvent the supporting role of bone stroma for prostate cancer epithelial growth, we have recently developed a novel concept where the expression of therapeutic toxic genes is driven by a tumor--but not a tissue-specific OC promoter. Osteocalcin-thymidine kinase (OC-TK) was found to efficiently eradicate the growth of osteosarcoma, prostate, and brain tumors both in vitro and in vivo. We observed that androgen-independent human prostate cancer cells lines expressed OC-TK at higher levels than androgen-dependent human prostate cancer cell lines. We have obtained data to suggest that Ad-OC-TK plus a pro-drug acyclovir (ACV) may be used as an effective therapy to treat prostate cancer bone metastasis in models where the growth of androgen-independent PC-3 and C4-2 tumors in the bone has occurred.

Acyclovir↗
PubMed1996

Androgen-repressed phenotype in human prostate cancer.

An androgen-repressed human prostate cancer cell line, ARCaP, was established and characterized. This cell line was derived from the ascites fluid of a patient with advanced metastatic disease. In contrast to the behavior of androgen-dependent LNCaP and its androgen-independent C4-2 subline, androgen and estrogen suppress the growth of ARCaP cells in a dose-dependent manner in vivo and in vitro. ARCaP is tumorigenic and highly metastatic. It metastasizes to the lymph node, lung, pancreas, liver, kidney, and bone, and forms ascites fluid in athymic hosts. ARCaP cells express low levels of androgen receptor mRNA and prostate-specific antigen mRNA and protein. Immunohistochemical staining shows that ARCaP cells stain intensely for epidermal growth factor receptor, c-erb B2/neu, and c-erb B3. Staining is negative for chromogranin A and positive for bombesin, serotonin, neuron-specific enolase, and the c-met protooncogene (a hepatic growth factor/scatter factor receptor). ARCaP cells also secrete high levels of gelatinase A and B and some stromelysin, which suggests that this cell line may contain markers representing invasive adenocarcinoma with selective neuronendocrine phenotypes. Along with its repression of growth, androgen is also found to repress the expression of prostate-specific antigen in ARCaP cells as detected by a prostate-specific antigen promoter-beta-galactosidase reporter assay. Our results suggest that the androgen-repressed state may be central to prostate cancer progression and that advanced prostate cancer can progress from an androgen-independent to an androgen-repressed state.

Aged↗
PubMed1996

Osteocalcin promoter-based toxic gene therapy for the treatment of osteosarcoma in experimental models.

Osteocalcin (OC), a noncollagenous bone matrix protein, is expressed in high levels by osteoblasts. To determine whether the OC promoter mediates cell-specific gene expression in cells of osteoblast lineage, we constructed a recombinant adenovirus, Ad-OC-TK, which contains the OC promoter that drives the expression of herpes simplex virus thymidine kinase (TK). We tested the expression of TK by this virus in osteoblast cell lines as well as in non-osteoblastic cell lines by assessing the enzyme activity of TK in vitro. Whereas the OC promoter failed to drive the expression of the TK gene in several non-osteoblastic cell lines such as WH, a human bladder transitional carcinoma, and NIH 3T3, an embryonic mouse fibroblast cell line, the OC promoter mediated high levels of expression in osteoblast cell lines including murine ROS and human MG-63 cells. The addition of acyclovir (ACV), a pro-drug for the inhibition of cell proliferation, resulted in the induction of osteoblast-specific cell death in vitro. Intratumoral injection of Ad-OC-TK into murine ROS osteosarcoma abolished tumor growth in a host treated with subsequent i.p. ACV injection in vivo. The Ad-OC-TK virus plus ACV treatment appears to be highly selective in blocking the growth of both murine and human osteosarcoma cell lines in vitro and murine osteosarcoma in vivo.

3T3 Cells↗