Biomedical subjects

M S Wicha

Publications and source records attributed to M S Wicha.

At least 19 recordsLinked to original sources

PubMed2000

A novel, conditionally replicative adenovirus for the treatment of breast cancer that allows controlled replication of E1a-deleted adenoviral vectors.

The efficiency of gene therapy strategies against cancer is limited by the poor distribution of the vectors in the malignant tissues. To solve this problem, a new generation of tumor-specific, conditionally replicative adenoviruses is being developed. To direct the replication of the virus to breast cancer, we have considered one characteristic present in a great proportion of these cancers, which is the expression of estrogen receptors (ERs). On the basis of the wild-type adenovirus type 5, we have constructed a conditionally replicative adenovirus (Ad5ERE2) in which the E1a and E4 promoters have been replaced by a portion of the pS2 promoter containing two estrogen-responsive elements (EREs). This promoter induces transcriptional activation of the E1a and E4 units in response to estrogens in cells that express the ERs. Ad5ERE2 is able to kill ER(+) human breast cancer cell lines as efficiently as the wild-type virus, but has decreased capacity to affect ER(-) cells. By complementation of the E1a protein in trans, Ad5ERE2 allows restricted replication of a conventional E1a-deleted adenoviral vector. When a virus expressing the proapoptotic gene Bc1-xs (Clarke et al., Proc. Natl. Acad. Sci. U.S.A. 1995;92:11024-11028) is used in combination with Ad5ERE2, the ability of both viruses to induce cell death is dramatically increased, and the effect can be modulated by addition of the antiestrogen tamoxifen.

Adenoviridae↗

PubMed2000

Evaluating the financial impact of clinical trials in oncology: results from a pilot study from the Association of American Cancer Institutes/Northwestern University clinical trials costs and charges project.

PURPOSE: Medical care for clinical trials is often not reimbursed by insurers, primarily because of concern that medical care as part of clinical trials is expensive and not part of standard medical practice. In June 2000, President Clinton ordered Medicare to reimburse for medical care expenses incurred as part of cancer clinical trials, although many private insurers are concerned about the expense of this effort. To inform this policy debate, the costs and charges of care for patients on clinical trials are being evaluated. In this Association of American Cancer Institutes (AACI) Clinical Trials Costs and Charges pilot study, we describe the results and operational considerations of one of the first completed multisite economic analyses of clinical trials. METHODS: Our pilot effort included assessment of total direct medical charges for 6 months of care for 35 case patients who received care on phase II clinical trials and for 35 matched controls (based on age, sex, disease, stage, and treatment period) at five AACI member cancer centers. Charge data were obtained for hospital and ancillary services from automated claims files at individual study institutions. The analyses were based on the perspective of a third-party payer. RESULTS: The mean age of the phase II clinical trial patients was 58.3 years versus 57.3 years for control patients. The study population included persons with cancer of the breast (n = 24), lung (n = 18), colon (n = 16), prostate (n = 4), and lymphoma (n = 8). The ratio of male-to-female patients was 3:4, with greater than 75% of patients having stage III to IV disease. Total mean charges for treatment from the time of study enrollment through 6 months were similar: $57,542 for clinical trial patients and $63,721 for control patients (1998 US$; P =.4) CONCLUSION: Multisite economic analyses of oncology clinical trials are in progress. Strategies that are not likely to overburden data managers and clinicians are possible to devise. However, these studies require careful planning and coordination among cancer center directors, finance department personnel, economists, and health services researchers.

Aged↗

PubMed2000

Differential regulation of apoptosis in normal versus transformed mammary epithelium by lutein and retinoic acid.

We examined the effects of all-trans retinoic acid (ATRA) and lutein (a nonprovitamin A carotenoid), on apoptosis and chemosensitivity in primary normal human mammary epithelial cells, SV40 transformed mammary cells, and MCF-7 human mammary carcinoma cells. ATRA and lutein selectively induced apoptosis in transformed but not normal human mammary cells. In addition, both compounds protected normal cells, but not transformed cells, from apoptosis induced by the chemotherapy agents etoposide and cisplatin. Furthermore, lutein and ATRA selectively increased the ratio of Bcl-xL:Bax protein expression in normal cells but not transformed mammary cells, suggesting a possible mechanism for selective modulation of apoptosis. The differential effects of lutein and ATRA on apoptotic pathways in normal versus transformed mammary epithelial cells may have important implications for chemoprevention and therapy.

Antineoplastic Agents↗

PubMed1999

Inhibition of protein synthesis by didemnin B is not sufficient to induce apoptosis in human mammary carcinoma (MCF7) cells.

Didemnin B (DB) is one member of a class of natural cyclic depsipeptides that display potent cytotoxicity in vitro. The detailed mechanism of action of DB is unknown, although it appears to involve the inhibition of protein biosynthesis. Additional activities of DB have established DB as a rapid and potent inducer of apoptosis in HL-60 cells. Our aim was to determine if the induction of apoptosis by DB is mediated through inhibition of protein synthesis in MCF-7 human breast carcinoma cells. Apoptosis was observed only at > or = 100 nM DB, even though inhibition of protein synthesis occurred at much lower DB concentrations (IC50 = 12 nM). DB-induced apoptosis was mediated by caspase activation, since cleavage of the caspase substrate poly(ADPribose) polymerase was observed as early as 6 hr after DB exposure. Two additional protein synthesis inhibitors, cycloheximide (CHX) and emetine (ET), failed to induce apoptosis at concentrations that completely inhibited protein synthesis. Moreover, DB-induced apoptosis was enhanced only slightly by pre- and co-treatment with CHX and ET. Thus, inhibition of protein synthesis alone was not sufficient to induce apoptosis in these cells. As a measure of antiproliferative potential, DB (1-5 nM) inhibited the colony forming ability of MCF7 cells regardless of pretreatment with CHX. In conclusion, additional effects of DB, independent of protein synthesis inhibition, are proposed to account for its ability to induce apoptosis and prevent cell proliferation.

Antineoplastic Agents↗

PubMed1999

Overexpression of Bcl-x(L) promotes chemotherapy resistance of mammary tumors in a syngeneic mouse model.

Bcl-x(L), a prosurvival member of the Bcl-2 family that is expressed in many tumors, represses apoptosis induced by chemotherapeutic drugs in vitro. However, the contribution of apoptosis and prosurvival Bcl-2-related proteins to chemotherapy resistance in vivo is unknown and has been challenged by recent results with clonogenic survival assays. To test the ability of Bcl-x(L) to provide chemotherapy resistance to tumors, we transfected the mouse bcl-x(L) gene into the tumorigenic SCK mammary cell line and assessed the response of tumor cells to chemotherapeutic drugs in clonogenic assays and in a syngeneic mouse model. Bcl-x(L) conferred protection on SCK cells against methotrexate at certain drug concentrations, but not at all against 5-fluorouracil in clonogenic survival assays in vitro. Injection of SCK cells transfected with Bcl-x(L) or control plasmid in the mammary fat pads of syngeneic recipient mice resulted in tumors of similar size. However, although the volume of control tumors regressed up to 80% after 4 to 5 days of chemotherapy, SCK tumors expressing Bcl-x(L) did not regress and continued to grow in the presence of methotrexate or 5-fluorouracil. In addition, numbers of apoptotic cells were significantly higher in control tumors as compared to Bcl-x(L)-expressing tumors in animals treated with methotrexate or 5-fluorouracil. These results provide evidence that inhibition of apoptosis through Bcl-x(L) overexpression can promote resistance to chemotherapy in tumors in vivo.

Adenocarcinoma↗

PubMed1999

Diet and risk for breast cancer recurrence and survival.

Dietary factors may influence the risk for breast cancer and also the prognosis following diagnosis and treatment. The aim of this study was to assess whether self-reported prediagnosis diet or other patient factors associated with breast cancer incidence were predictive of recurrence and survival. Patients (n = 149) diagnosed with primary breast cancer between 1989 and 1991 were followed for five or more years. Total energy (hazard ratio (HR) = 1.58, 95%, confidence interval (CI) = 1.05, 2.38) as well as total (HR = 1.46, 95% CI = 1.05, 2.01), saturated (HR = 1.79, 95% CI = 1.05, 3.04), and monounsaturated (HR = 1.65, 95% CI = 1.09, 2.49) fat intakes were associated with increased risk, and energy-adjusted bread and cereal consumption (HR = 0.55, 95% CI = 0.33, 0.93) with decreased risk of recurrence. Both total energy (HR = 1.58, 95% CI = 1.03, 2.43) and polyunsaturated fat (HR = 1.84, 95% CI = 1.09, 3.13) intakes were associated with an increased risk of death. All associations between dietary fat and recurrence and survival attenuated following energy adjustment. Oral contraceptive use (HR = 1.28, 95% CI = 1.03, 1.60), lymph node positive status (HR = 2.36, 95% CI = 1.01, 5.49), and tumor stage (HR = 2.22, 95% CI = 1.02, 4.81) were associated with increased risk of recurrence. Tumor stage (HR = 4.96, 95% CI = 1.86, 13.23), lymph node positive status (HR = 3.31, 95% CI = 1.38, 7.95), and estrogen receptor negative status (HR = 2.46, 95% CI = 1.02, 5.94) were associated with increased risk, and arm muscle circumference (HR = 0.27, 95% CI = 0.09, 0.86) and mammographic utilization (HR = 0.77, 95% CI = 0.61, 0.98) with decreased risk of death. Higher levels of energy, fat intakes, and selected patient characteristics (particularly disease stage and anthropometric indicators of adiposity) appear to increase risk of recurrence and/or shortened survival following the diagnosis of breast cancer.

Adult↗

PubMed1998

A method of limited replication for the efficient in vivo delivery of adenovirus to cancer cells.

Replication-deficient viral vectors are currently being used in gene transfer strategies to treat cancer cells. Unfortunately, viruses are limited in their ability to diffuse through tissue. This makes it virtually impossible to infect the majority of tumor cells in vivo and results in inadequate gene transfer. This problem can be addressed by allowing limited viral replication. Limited viral replication facilitates greater penetration of virions into tissue and can improve gene transfer. We have developed a strategy of limited viral replication using AdRSVlaclys, a chemically modified E1-deleted adenovirus, to codeliver an exogenous plasmid encoding the adenovirus E1 region. This system allows one round of viral replication. We examined the effect of this limited adenovirus replication in vitro and in vivo. In culture, codelivery of virus and pE1 resulted in a large increase in infected cells when compared with control cells exposed to virus and pUC19. In experiments on nude mice bearing HeLa ascites tumors, intraperitoneal injection of AdRSVlaclys/pE1 resulted in a significantly higher percentage of infected HeLa cells as compared with the PBS controls (p < 0.05) or the AdRSVlaclys/pUC19 controls (p < 0.01). These data demonstrate that the transcomplementation of replication-deficient adenovirus with exogenous E1 DNA leads to limited replication, and this controlled replication enhances gene transfer efficiency of adenovirus in vivo.

Adenoviridae↗

PubMed1998

Targeting cancer cell death with a bcl-XS adenovirus.

Transformation is a complex cellular process that requires several genetic abnormalities. In many cases, one of these abnormalities is an inhibition of PCD, which provides a selective advantage for tumor cells. This has been recently shown in an in vivo model, where overexpression of Bcl-XL, is a crucial step in the progression from hyperplasia to neoplasia and is accompanied by a significant decrease in tumor apoptosis [56]. Frequently, overexpression of a member of the Bcl-2 family results in a block in cell death and appears to nullify many built-in cellular defense mechanisms against cancer. Such a block presents a problem because radiation and chemotherapy, standard cancer treatments, ultimately exert their effect by induction of apoptosis and would also be made less effective. Therefore, to better treat cancer it may be necessary to develop novel methods to overcome the effects of the Bcl-2 family. One way to approach this problem is to target the cause--the molecular machinery that allows a cancer cell to survive. Advances in our understanding of apoptosis has identified the Bcl-2 family as a mediator of most apoptosis pathways, including those initiated by oncogenes, tumor suppressor genes, growth factor withdrawal, and external damaging signals. Therefore, functional inhibition of Bcl-2 family members is lethal to many cancer cells. Using gene transfer technology, we can now deliver genes that accomplish this goal. Further investigation will reveal whether this translates to improved therapy in the future.

Adenoviridae↗

PubMed1998

Immunolymphoscintigraphy in breast cancer: evaluation using 131I-labeled monoclonal antibody B72.3.

Noninvasive axillary lymph node staging was investigated using [131I]murine monoclonal antibody B72.3 in 16 patients with breast cancer scheduled for axillary dissection. [131I]B72.3 was injected into ipsilateral finger webs or around the breast biopsy. Scintigraphy to 72 h and gamma-counting/immunohistochemistry of nodes were performed. Specific antibody uptake (%ID/g) and the ratio of specific:nonspecific antibody uptake were not significantly different in tumor-positive versus tumor-negative nodes, suggesting that [131I]B72.3 is unsuitable to discriminate axillary node tumor involvement.

Antibodies, Monoclonal↗

PubMed1996

bcl-xs gene therapy induces apoptosis of human mammary tumors in nude mice.

Bcl-xs is a dominant negative repressor of Bcl-2 and Bcl-xL, both of which inhibit apoptosis. We used a replication-deficient adenoviral vector to transiently overexpress Bcl-xs in MCF-7 human breast cancer cells, which overexpress Bcl-xL. Infection with this vector induced apoptosis in vitro. We then determined the effects of intratumoral injection of bcl-xs adenovirus on solid MCF-7 tumors in nude mice. Tumors injected four times with the bcl-xs adenovirus showed a 50% reduction in size. Using terminal transferase-mediated dUTP-digoxigenin nick end labeling, we observed apoptotic cells at sites of bcl-xs adenoviral injection. These experiments demonstrate the feasibility of using bcl-xs gene therapy to induce apoptosis in human breast tumors.

Animals↗

PubMed1995

Murine granulocytic cell adhesion to bone marrow hemonectin is mediated by mannose and galactose.

Hemonectin (HN) is a bone marrow (BM) protein that promotes specific attachment of immature granulocytes and their precursors within the BM. We report that HN is a glycoprotein containing both mannose and galactose residues, and provide evidence that these carbohydrates mediate granulocytic cell adhesion to HN. Carbohydrate structure was determined by digoxigenin-conjugated lectin binding to HN and indicated the presence of mannose, galactose, sialic acid, and the absence of fucose-linked oligosaccharides. The role of carbohydrates in mediating cell adhesion was examined by chemical and enzymatic deglycosylation. Deglycosylation of HN with trifluoromethanesulfonic acid, which cleaves N- and O-linked oligosaccharides, inhibits 66% of cell attachment to HN, and results in an apparent decrease in molecular weight from 60 to 50 kD. Enzymatic deglycosylation with endo-B-N-acetylglucosaminidase H, which hydrolyzes specific N-linked mannose residues, inhibits 30% of cell adhesion to HN. Finally, the role of these specific sugars in hemonectin-mediated cell adhesion was confirmed with neoglycoprotein blocking. Preincubation of BM cells with mannosyl- and galactosyl-BSA probes produces a dose-dependent inhibition of cell attachment to HN, whereas fucosyl-BSA does not inhibit cell adhesion to HN. These results show that mannose and galactose partially mediate adhesion of BM granulocytes to HN.

Animals↗

PubMed1995

Overexpression of Bcl-XS sensitizes MCF-7 cells to chemotherapy-induced apoptosis.

Resistance to apoptosis plays an important role in tumors that are refractory to chemotherapy. We report that Bcl-XL, which functions like Bcl-2 to inhibit apoptosis, is highly expressed in MCF-7 human breast carcinoma cells. We used Bcl-XS, a dominant negative inhibitor of Bcl-2 and Bcl-XL, to demonstrate the role of these genes in modulating chemotherapy-induced apoptosis. Bcl-XS overexpressed in MCF-7 cells by stable transfection does not affect viability by itself but induces a marked increase in chemosensitivity to VP-16 or taxol. Using an ELISA assay which quantitates DNA damage, we demonstrate that this sensitization is due to apoptosis, suggesting the therapeutic utility of targeting this pathway.

Antineoplastic Agents↗

PubMed1992

cDNA cloning and molecular characterization of MSE55, a novel human serum constituent protein that displays bone marrow stromal/endothelial cell-specific expression.

Hemonectin is a lineage-specific cytoadhesive protein that may be involved in the developmentally regulated adhesion of granulocytic cells to bone marrow stroma. Immunoblot analysis using an anti-hemonectin antibody recognizes two distinct immunoreactive species in endothelial cell lysates (approximately M(r) 65,000) and human serum (approximately M(r) 55,000). Initial characterization of the 55-kDa protein has now been completed by isolating the cDNA from a human endothelial cell expression library. Sequence analysis of overlapping clones identifies a composite sequence spanning 2030 nucleotides with an open reading frame of 1173 base pairs. No significant sequence similarity was observed on analysis of current GenBank databases. The open reading frame was expressed as a recombinant protein in Escherichia coli and used as an immunogen for the production of a specific polyclonal antibody. Immunoblotting with this antibody identifies a single immunoreactive species of apparent M(r) 55,000 in HUVEC lysates and human serum, confirming that a secreted form normally circulates as a serum constituent protein. This antibody fails to recognize purified hemonectin, suggesting that the M(r) 55,000 protein is not hemonectin. Cross-species Southern blot analysis reveals persistent hybridizing fragments in all species tested, suggestive of a developmentally conserved function. Northern blot analysis demonstrates expression limited to endothelial and bone marrow stromal cells, but not poly(A) RNA from monkey liver, spleen, brain, lung, and kidney. On this basis, we have designated this novel protein MSE55, for marrow stromal/endothelial cell protein with a molecular mass of 55,000 daltons. Its tissue-specific expression may suggest a functional role in hematopoiesis.

Adaptor Proteins, Signal Transducing↗

PubMed1992

cDNA cloning and genomic analysis of a new multigene family sharing common phylogenetic and expression profiles with the laminin receptor gene.

We have isolated a human laminin receptor (LR) cDNA which bears a different sequence in its 5' end with regard to the corresponding region in the regular LR mRNA. This different sequence hybridizes to a 1 kb mRNA. We have cloned a 740 bp cDNA for this transcript (cDNA 48-1). Search on sequence data bases revealed no sequence homology with known messengers or proteins. Using cDNA 48-1 in a simplified version of the protocol with which we had previously characterized the LR gene as a member of a retrogene family in mammals, we show in the present paper that the gene of this new transcript exhibits phylogenic, expression and amplification features that strickingly recall those of the LR gene.

Amino Acid Sequence↗

PubMed1992

Posttranscriptional regulation of alpha-casein mRNA accumulation by laminin.

We have utilized primary cultures of rat mammary epithelial cells to study mechanisms by which laminin regulates the prolactin-dependent accumulation of alpha-casein mRNA. Mammary cells accumulate approximately fivefold more alpha-casein mRNA when cultured on laminin than when cultured on tissue plastic and the accumulation of alpha-casein mRNA is prolactin dependent. On the basis of transcription assays there is approximately a twofold increase in the alpha-casein mRNA transcription rate in cells cultured on laminin over that of tissue culture plastic. Measurements on the turnover of alpha-casein mRNA show that this mRNA is stabilized fourfold more on laminin than on tissue culture plastic, while there was no significant difference in the turnover of poly(A) RNA on either substratum. These data indicate that laminin regulates the cytoplasmic levels of alpha-casein mRNA accumulation primarily at the post-transcriptional level by increasing the stabilization of this mRNA.

Animals↗

PubMed1991

Genomic analysis of the 67-kDa laminin receptor in normal and pathological tissues: circumstantial evidence for retroposon features.

We have cloned two cDNAs for the human 67-kDa laminin receptor (LR). In the present report we show that these clones hybridize to many restriction fragments in Southern experiments in human. This particular pattern is accounted for by the presence of up to 16 and 21 copies of the laminin receptor gene per haploid genome in human and mouse, respectively. In contrast, a single gene copy is found in chicken. Chromosomal localization reveals four main loci: LAMRP1, laminin receptor pseudogene 1 (Chr 3); LAMRP2, laminin receptor pseudogene 2 (Chr 12); LAMRP3, laminin receptor pseudogene 3 (Chr 14); LAMRP4, laminin receptor pseudogene 4 (Chr X). Comparison of our experimental results to the known features of processed retropseudogenes enabled us to conclude that the LR gene belongs to a retroposon family in mammals.

Animals↗

PubMed1990

Developmental regulation of granulocytic cell binding to hemonectin.

Hemonectin (HN), a component of the bone marrow (BM) extracellular matrix which promotes adhesion of cells in the granulocytic lineage, was purified to near homogeneity and tested for its ability to mediate attachment of normal and leukemic cells of granulocytic lineage. Purified HN immobilized on plastic substrates promoted serum-free attachment of normal granulocyte/macrophage progenitor cells (CFC-GM), using an in situ attachment assay in which cell attachment is inhibited by specific polyclonal antisera. When unfractionated BM cells were allowed to attach to purified HN and stained in situ, HN preferentially bound cells at earlier stages of granulocytic differentiation. These observations were confirmed using cells of the HL-60 progranulocytic cell line which mirrored this differentiation-stage specific binding to HN. HN promoted attachment of 60% of uninduced HL-60 cells which were arrested at the progranulocyte stage, whereas only 15% of uninduced HL-60 cells attached to uncoated plastic and 4% to attached plastic coated with equal microgram quantities of bovine serum albumin (BSA). When HL-60 cells were induced to differentiate along the granulocytic pathway by incubation with dimethylsulfoxide (DMSO), attachment to hemonectin was reduced. Thus, both primary BM granulocytic cells and a granulocytic cell line show preferential attachment of those cells at earlier stages of differentiation. This developmentally regulated binding suggests a mechanism for release of maturing BM into the peripheral circulation.

Adolescent↗

PubMed1990

Fetal expression of hemonectin: an extracellular matrix hematopoietic cytoadhesion molecule.

Hemonectin, a component of bone marrow extracellular matrix, is a lineage- and organ-specific attachment molecule for cells of the granulocytic lineage. We hypothesized that hemonectin is an important marker of fetal granulopoiesis that is developmentally regulated during the ontogeny of the hematopoietic system. Murine hematopoiesis originates in the yolk sac and subsequently appears in the liver, spleen, and bone marrow. Using an affinity-purified polyclonal antibody to purified hemonectin as a probe of developing hematopoietic organs, we observe that hemonectin is coordinately expressed at developmental stages of the mouse in those tissues that are supporting hematopoiesis. Multiparameter flow cytometric analysis reveals that approximately 7% of fetal liver cells express hemonectin by day 13 of gestation, and that 32% of the cells are positive by day 19. Additionally, restricted hemonectin expression is noted in other tissues (cartilage, skin, developing bone, and capillary endothelial cells), suggesting that this molecule subserves other developmental functions and/or belongs to a previously unrecognized family of molecules.

Animals↗