[Making decisions in times of a pandemic : A qualitative study on perspectives of managers of long-term care facilities]. / Entscheidungen treffen in Pandemiezeiten : Eine qualitative Studie zu Perspektiven von Leitungspersonen der stationären Langzeitpflege.

BACKGROUND: In the spring of 2020 during the first wave of the pandemic an above-average number of residents and staff in nursing homes fell ill and accounted for the highest number of incidences. Due to the pandemic, managers in nursing homes had to make new decisions on a daily basis as well as interpret and integrate decisions made by higher level authorities. AIM OF THE STUDY: The aim was to describe the decisions that had to be made by the managers of nursing homes in dealing with the COVID-19 pandemic and related consequences. MATERIAL AND METHODS: A qualitative multicentre cross-sectional design was chosen. Data collection was conducted with semi-structured telephone interviews. The recorded audio data were transcribed, analyzed using the framework analysis method and reflected in peer debriefings. RESULTS: A total of 78 interviews were conducted in 43 nursing homes and 3 main themes with 10 subthemes emerged: decisions about social participation, decisions on quarantine and isolation and staff adjustments. DISCUSSION: Clearer information and directives for the implementation of measures are needed, e.g. through standardized guidelines nationwide. Additionally, public health departments should play a stronger and more responsible role in a pandemic situation. The consequences of their decisions were hardly foreseeable for the managers and were marked by uncertainty. Responsibilities for and consequences of pandemic-related decisions should be further evaluated to empower managers in times of crises.

Efficacy of oncolytic adenovirus expressing suicide genes and interleukin-12 in preclinical model of prostate cancer.

Oncolytic adenovirus-mediated suicide gene therapy has been shown to improve local tumor control in preclinical tumor models and in the clinic. Although local tumor control is important, for most human cancers, new therapies must also target metastatic disease if they are to have an impact on survival. Here, we test the hypothesis that adding cytokine gene therapy to our multimodal platform improves both local and metastatic tumor control in a preclinical model of prostate cancer. An oncolytic adenovirus (Ad5-yCD/mutTKSR39rep-mIL12) expressing two suicide genes and mouse interleukin-12 (IL-12) was generated. Relative to an adenovirus lacking IL-12 (Ad5-yCD/mutTKSR39rep), Ad5-yCD/mutTKSR39rep-mIL12 improved local and metastatic tumor control in the TRAMP-C2 prostate adenocarcinoma model, resulting in a significant increase in survival. Ad5-yCD/mutTKSR39rep-mIL12 resulted in high levels of IL-12 and interferon gamma in serum and tumor, increased natural killer (NK) and cytotoxic T-lymphocyte lytic activities, and the development of tumor-specific antitumor immunity. Immune cell depletion studies indicated that both the innate and adaptive arms of immunity were required for maximal Ad5-yCD/mutTKSR39rep-mIL12 activity. The results demonstrate that the addition of IL-12 significantly improves the efficacy of oncolytic adenovirus-mediated suicide gene therapy and provide the scientific basis for future trials targeting locally aggressive cancers.

Efficacy of suicide gene therapy in hypoxic rat 9L glioma cells.

Viral vector mediated suicide gene therapy (SGT) involving thymidine kinase (TK) or cytosine deaminase (CD) have considerable promise in the treatment of malignant brain tumors. An unresolved issue is to what extent tumor hypoxia influences the outcome of SGT since brain tumors characterized by regions of hypoxia have potentially reduced cellular metabolism and SGT's cytotoxicity is manifest through cellular metabolism. We studied in vitro and in vivo, the effect of hypoxia on the cytotoxicity of SGT in rat 9L glioma cells. Neither acute nor chronic hypoxia affected the cell killing of SGT by TK or CD. In vivo confirmation that SGT efficacy was not adversely affected by tumor hypoxia using the hypoxic cell marker pimonidazole was shown by the absence of a change in tumor hypoxia by SGT. These studies support the use of SGT utilizing either TK or CD gene strategies even when tumors are characterized by a hypoxic microenvironment.

Polyethylene glycol (molecular weight 400 DA) vehicle improves gene expression of adenovirus mediated gene therapy.

PURPOSE: A significant limitation of adenoviral mediated suicide gene therapy is poor gene distribution in vivo. The choice of vehicle has been demonstrated to affect the level of adenoviral delivered gene transduction. We examined the hypotheses that 1) adenovirus suspended in PEG400 improves gene expression in the naïve canine prostate model, 2) improved transgene expression with PEG400 results in improved tumor control and 3) vehicle affects the initial adenoviral spread from a single intratumor injection. MATERIALS AND METHODS: The magnitude and volume of gene expression were measured 24 hours following intraprostatic injection of adenovirus suspended in PEG400 (12.5% weight per volume) or saline as vehicle. Tumor growth delay was measured in mice bearing human tumor xenografts following the injection of adenovirus in PEG400 and saline. The initial spread of adenovirus was measured by confocal microscopy following a single injection of fluorescently labeled adenoviral particles in human tumor xenografts using each vehicle. RESULTS: Adenovirus suspended in PEG400 provided an average of twice the level of gene expression in the canine prostate and significantly better tumor control relative to saline in preclinical tumor models (p = 0.046 and 0.036, respectively). The initial spread of adenovirus with PEG400 was superior to that of adenovirus in saline and the latter was largely limited to the needle tract. CONCLUSIONS: Adenoviral gene therapy vectors suspended in PEG400 results in improved tumor control because of greater initial adenoviral spread, and the increased volume and magnitude of gene expression in vivo.

[Allocating and scheduling operating room time]. / Zuweisung und Planung von Operationsraumkapazitäten Maximierung der Operationsraumeffizienz am Beispiel einer deutschen Universitätsklinik.

During the past decade many scientific advances have been made concerning the development of methodologies to maximize efficiency of surgical facilities through allocating and scheduling of operating rooms. In this article such a methodology is described. Using the analysis of historical data of surgical activity in a facility, future demand is predicted and planned. Part of the methodology includes principles and rules needed for the daily organization and operative management of surgical facilities. They are also derived from the same science and therefore the basis for rational and structured decision making. Medical aspects such as patient safety and free choice of day for surgery have higher priority than the economic goal of maximizing operating room efficiency.

Cyclandelate in the prophylaxis of migraine: a placebo-controlled study.

The prophylactic action of cyclandelate was investigated in a multicentre, randomized, placebo-controlled, parallel group study. A 4-week baseline period was followed by a 4-week placebo phase and a 16-week treatment period with either 1600 mg cyclandelate or placebo. Patients (n = 251) with two to six migraine attacks/month were randomized. Neither the primary study endpoint (reduction of migraine days from baseline to the last 28 days) nor most of the secondary endpoints (reduction in the number of migraine attacks, severity or duration of attacks, frequency of autonomic disturbances, medication for treatment of attacks) showed a difference between cyclandelate and placebo. Cyclandelate, however, was superior to placebo in a global impression of efficacy rated by the patients and the treating physicians. Both treatments were well tolerated. In conclusion, cyclandelate was not superior to placebo in the prophylaxis of migraine with regard to parameters usually used in migraine prophylaxis trials.

Antisense GLUT-1 protects mesangial cells from glucose induction of GLUT-1 and fibronectin expression.

A stable clone of rat mesangial cells expressing antisense GLUT-1 (i.e., MCGT1AS cells) was developed to protect them from high glucose exposure. GLUT-1 protein was reduced 50%, and the 2-deoxy-[(3)H]glucose uptake rate was reduced 33% in MCGT1AS. MCLacZ control cells and MCGT1 GLUT-1-overexpressing cells were used for comparisons. In MCLacZ, 20 mM D-glucose increased GLUT-1 transcription 90% vs. no increase in MCGT1AS. Glucose (8 mM) and 12 mM xylitol [a hexose monophosphate (HMP) shunt substrate] did not stimulate GLUT-1 transcription. An 87% replacement of the standard 8 mM D-glucose with 3-O-methylglucose reduced GLUT-1 transcription 80%. D-Glucose (20 mM) increased fibronectin mRNA and protein by 47 and 100%, respectively, in MCLacZ vs. no increases in MCGT1AS. Fibronectin synthesis was elevated 48% in MCGT1 and reduced 44% in MCGT1AS. We conclude that 1) transcription of GLUT-1 in response to D-glucose depends on glucose metabolism, although not through the HMP shunt, and 2) antisense GLUT-1 treatment of mesangial cells blocks D-glucose-induced GLUT-1 and fibronectin expression, thereby demonstrating a protective effect that could be beneficial in the setting of diabetes.

Sustained improvements in patients with dementia of Alzheimer's type (DAT) 6 months after termination of Cerebrolysin therapy.

The present study is an extension of the work of Rüther et al. (1994). 101 patients suffering from DAT were evaluated 6 months after completion of a 4 week (5 days per week) therapy with either 30 ml Cerebrolysin or placebo. The significant and clinically relevant improvements in the global rating (CGI), clinical symptomatology (SCAG), cognitive performance (ZVT-G) as primary efficacy variables, as well as the improvements in the secondary efficacy variables activities of daily living (NAI) and wellbeing (Bf-S), achieved in the Cerebrolysin group after only 4 weeks of active therapy, were maintained to a large extent during the follow-up period. Although there was a moderate tendency in the drug group towards loss of improvement, the differences between baseline and follow-up examination, as well as the differences between the verum and the placebo group, clearly document a sustained improvement in patients treated with Cerebrolysin in the first 4 weeks of the study period. It can be speculated that relatively short treatment courses with Cerebrolysin in patients suffering from neurodegenerative dementia can lead to long term influence on disease progression, which is in accordance with the proposed neurotrophic - nerve growth factor like - mode of action.

Evaluation of the biodistribution, persistence, toxicity, and potential of germ-line transmission of a replication-competent human adenovirus following intraprostatic administration in the mouse.

Adenovirus-mediated gene transfer may hold much promise in the treatment of human cancer. However, concerns regarding vector dissemination beyond the target tissue, particularly with replication-competent viruses, require an evaluation of the persistence of viral infection in collateral tissue and vector-associated toxicities. In addition, for indications such as prostate cancer, the proximity of the point of viral administration to organs of the male reproductive system raises concerns regarding inadvertent germ-line transmission of genes carried by the virus. To address these concerns, the biodistribution, persistence, toxicity, and potential of germ-line transmission of a replication-competent adenovirus (Ad5-CD/TKrep) following intraprostatic administration in the mouse was examined. Ad5-CD/TKrep (10(10) vp, 5 x 10(11) vp/kg) was injected intraprostatically on Day 1 of the study and its presence in the major organs of the male urogenital tract (prostate, testes, seminal vesicles, and urinary bladder) and liver was determined on Days 8 and 29. For comparison, a parallel group of animals was injected with the same dose of a related replication-defective Ad5-FGNR virus. To evaluate germ-line transmission, Ad5-CD/TKrep-injected males were mated to females on Days 8 and 29 and resulting embryos were examined for AdS-CD/TKrep viral DNA. Ad5-CD/TKrep viral DNA was detected in all major organs of the adult male urogenital tract and liver 7 and 28 Days postinjection. Interestingly, relative to the replication-defective Ad5-FGNR adenovirus, the replication-competent Ad5-CD/TKrep virus accumulated to a much greater level (approximately 300-fold) and persisted for a longer period of time in prostate, testes, and liver. This difference could not be explained on the basis of differences in viral infectivity, suggesting that the AdS-CD/TKrep virus may be capable of replicating in mouse tissues in vivo. In vitro infection of six mouse cell lines representing prostate, testes, and liver demonstrated that the Ad5-CD/TKrep virus was indeed capable of replicating in these mouse cell types, albeit with reduced efficiencies relative to human cells. Despite the fact that the Ad5-CD/TKrep vector persisted in the adult male gonads and may have replicated in vivo, we observed no evidence of germ-line transmission in 149 offspring examined. To evaluate the toxicity of combining Ad5-CD/TKrep viral therapy with CD/5-FC and HSV-1 TK/GCV suicide gene therapies as a prerequisite for a human trial, an escalating dose (10(8), 10(9), 10(10) vp) of Ad5-CD/TKrep was administered intraprostatically followed by 7 days of 5-FC and GCV double prodrug therapy. Although the virus persisted in the mouse urogenital tract and liver for up to 28 days postinjection, most of the toxicities observed were expected, minimal, and self-limiting. These results lead us to believe that intraprostatic administration of the Ad5-CD/TKrep virus to humans concomitant with double suicide gene therapy will be associated with acceptable toxicities and will not result in vertical transmission of viral-encoded genes through the germ line.

In vivo antitumor activity of ONYX-015 is influenced by p53 status and is augmented by radiotherapy.

The E1B-deleted, replication-competent ONYX-015 (dl1520) adenovirus was originally described as being able to selectively kill p53-deficient cells due to a requirement of p53 inactivation for efficient viral replication. This hypothesis has become controversial because subsequent in vitro studies have demonstrated that the host range specificity of ONYX-015 is independent of p53 gene status. Using a pair of isogenic cell lines that differ only in their p53 status, we demonstrate here that although ONYX-015 can replicate in both p53 wild-type and mutant cells in vitro, the virus demonstrates significantly greater antitumor activity against mutant p53 tumors in vivo. Moreover, ONYX-015 viral therapy can be combined with radiation to improve tumor control beyond that of either monotherapy. The results demonstrate that ONYX-015 can discern in vivo between tumors having a different p53 status and that it may be an effective neoadjuvant to radiation therapy.

Double suicide gene therapy augments the antitumor activity of a replication-competent lytic adenovirus through enhanced cytotoxicity and radiosensitization.

Replication-competent adenoviruses may provide a highly efficient means of delivering therapeutic genes to tumors. Previously, we evaluated in vitro a replication-competent adenovirus (Ad5-CD/TKrep) containing a cytosine deaminase (CD)/herpes simplex type 1 thymidine kinase (HSV-1 TK) fusion gene that allows lytic viral therapy to be combined with double suicide gene therapy. Both the CD/5-FC and HSV-1 TK/GCV enzyme/prodrug systems enhanced the tumor cell-specific cytopathic effects of the Ad5-CD/TKrep virus in vitro and sensitized cells to radiation. To extend these in vitro findings in vivo, we evaluated the antitumor activity of the Ad5-CD/TKrep virus in combination with double prodrug therapy and radiation therapy. The Ad5-CD/TKrep virus independently demonstrated significant antitumor activity against C33A cervical carcinoma xenografts. Therapeutic outcome was dramatically improved with systemic administration of double, but not single, prodrug (5-FC + GCV) therapy. When used in a neoadjuvant setting, Ad5-CD/TKrep-mediated double suicide gene therapy dramatically potentiated the effectiveness of radiation therapy. The trimodal approach of Ad5-CD/TKrep viral, double suicide gene, and radiotherapies produced significant tumor regression and ultimately 100% tumor cure. The results demonstrate the high therapeutic potential of the trimodal approach and provide a solid foundation for future clinical trials.

[Australian Refined Diagnosis Related Groups. Formal and inherent problems of grouping with the example of stroke care]. / Australian Refined Diagnosis Related Groups. Formale und inhaltliche Problematik der Anwendung am Beispiel der Schlaganfallversorgung.

BACKGROUND AND OBJECTIVES: With the Health Reform 2000, the Australian Refined Diagnosis Related Groups (AR-DRG), Version 4.1 have been chosen as the basis for the future German costing system for hospitals. With regard to Stroke Severity (Barthel Index [BI]) we investigated to what extent the grouping according to AR-DRGs can reproduce healthcare expenditures for such patients. Options to adapt and optimize the system are discussed. PATIENTS AND METHODS: 632 patients who had suffered a cerebrovascular accident and were discharged from conservative acute care in 1999, were classified according to the AR-DRGs. For the grouping we alternatively used data from the current hospital information system and a stroke database for quality assurance. The results were also compared with the clinical profiles for the public hospital sector of the corresponding DRGs in Australia (1997-98). RESULTS: On average 0.99 additional diagnoses per case were documented in the hospital information system, compared to 3.65 in the stroke database. In the stroke database 177 cases (36.8%) were assigned to the DRG with the highest cost weight. 53.7% of these patients suffered a serious stroke (BI < 30). Grouping on the basis of hospital information system data led only to 14 cases (2.8%) assigned to the DRG with the highest cost weight. CONCLUSIONS: Type and extent of additional diagnoses are crucial for the grouping process. From a clinical and economic point of view, measures of disability and impairment should be assigned to the grouping process to improve homogeneity under both aspects. Scores can also serve for determining reliable outcome parameters. For the development of an outcome related reimbursement system, procedures must be included in the definition of medical DRGs. In future, DRGs, which cover overlapping healthcare sectors, should be developed for patients with poststroke rehabilitation.

Efficacy of adenovirus-mediated CD/5-FC and HSV-1 thymidine kinase/ganciclovir suicide gene therapies concomitant with p53 gene therapy.

Recent evidence has suggested that tumor cells having a wild-type p53 status are more sensitive to chemotherapeutic agents and radiation than cells that lack functional p53. The heightened sensitivity of wild-type p53 cells is thought to be attributable to their propensity to undergo p53-mediated apoptosis after insult. Given that suicide gene therapy is essentially tumor-targeted chemotherapy, we examined the hypothesis that coexpression of wild-type p53 could enhance the efficacy of adenovirus-mediated suicide gene therapy. Human Hep3B and SK-OV-3 cells, which are null for p53, were infected with a pair of replication-deficient adenoviruses that expressed a cytosine deaminase/herpes simplex virus thymidine kinase (CD/HSV-1 TK) fusion gene without (fusion gene nonreplicative adenovirus, FGNR) or with (FGNRp53) the wild-type human p53 gene. The sensitivity of cells to the CD/5-fluorocytosine (CD/5-FC) and HSV-1 TK/ ganciclovir (GCV) enzyme/prodrug systems was determined in vitro and in vivo. Coexpression of p53 did not enhance the cytotoxicity of either the CD/5-FC or HSV-1 TK/GCV system in vitro. The failure to observe an effect of p53 could not be explained on the basis of insufficient or transient p53 expression, because FGNRp53-infected cells growth arrested in G1, induced Bax, and underwent apoptosis at an increased rate after prodrug treatment, particularly when the adenovirus E1A protein was present. Intratumoral injection of FGNRp53 concomitant with single or double pro-drug therapy resulted in a tumor growth delay that was equal to or less than that observed with the FGNR virus. Our results indicate that coexpression of p53 may not necessarily improve the efficacy of adenovirus-mediated CD/ 5-FC and HSV-1 TK/GCV suicide gene therapies in vivo.

Selective radiosensitization of 9L glioma in the brain transduced with double suicide fusion gene.

PURPOSE: Suicide gene therapy has proved to be successful in enhancing the therapeutic index by sensitizing genetically modified tumor cells to prodrugs. Two of the most widely studied suicide genes, herpes simplex virus type 1 thymidine kinase and Escherichia coli cytosine deaminase, have proved effective at selectively eliminating malignant tumor cells. We previously demonstrated that transduced 9L glioma cells expressing E. coli cytosine deaminase and herpes simplex virus type 1 thymidine kinase concomitantly as a fusion protein exhibited greater levels of targeted cytotoxicity and radiosensitization than could be achieved by single suicide gene therapy. The present in vivo studies were carried out to determine whether double suicide gene therapy would enhance the tumor control rate of orthotopically implanted malignant glioma growing in the brain when coupled with radiotherapy. MATERIALS AND METHODS: Rat 9L gliosarcoma cells were transfected with retroviral vectors containing an E. coli cytosine deaminase and herpes simplex virus type 1 thymidine kinase fusion gene and maintained in Dulbecco's modified Eagle's medium. The antitumor response of 9L E. coli cytosine deaminase and herpes simplex virus type 1 thymidine kinase tumors growing in the brain of Fischer rats was evaluated with small tumors (6-day-old tumors) versus large tumors (14-day-old tumors) against single versus double prodrug treatments. In the large brain tumors, the therapeutic efficacy of the combined single and double prodrugs coupled with radiotherapy was evaluated. RESULTS: Double suicide gene therapy using two prodrugs, 5-fluorocytosine (500 mg/kg) and ganciclovir (30 mg/kg), was effective in achieving long-term tumor control (50% survival) against early-stage brain tumors (6 days after implantation) but was only marginally effective against advanced stage tumors (14 days old). However, when these prodrugs were combined with radiotherapy and double suicide gene therapy against advanced-stage tumors, more than 70% of the animals were cured, whereas radiotherapy alone (20 Gy) failed to achieve any cure at all. Combined radiotherapy and single prodrug therapy showed a moderate increase in the animal survival rate (17% and 40% for 5-fluorocytosine and ganciclovir, respectively) but was inferior to the combination therapy of radiation and double prodrugs. CONCLUSION: The present in vivo results indicate that double suicide gene therapy combined with radiotherapy may represent a new, effective approach to achieve a high tumor cure rate without producing any excessive normal tissue damage.

Selective in vivo radiosensitization by 5-fluorocytosine of human colorectal carcinoma cells transduced with the E. coli cytosine deaminase (CD) gene.

PURPOSE: The E. coli cytosine deaminase (CD) gene encodes an enzyme capable of converting the nontoxic prodrug 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU), a known radiosensitizer. Having previously shown that combined CD suicide gene therapy and radiation (RT) results in pronounced radiosensitization in vitro, we progressed to in vivo studies of combined therapy. METHODS AND MATERIALS: WiDr human colon cancer cells were transduced in vitro with the CD gene and cells expressing CD were selected for use as xenografts in a nude mouse model. After administration of 5-FC, tumors received 10-30 Gy local field radiation (RT) and tumor growth delay was compared to control animals receiving either 5-FU, 5-FC, or RT alone. RESULTS: Maximal growth delay was seen in mice treated with 5-FC for 6 consecutive days prior to RT. Combined treatment with 15 Gy radiation resulted in a dose-modifying factor (DMF) of 1.50, and a greater DMF was observed with higher doses of radiation. There was no appreciable toxicity using this new approach. In contrast, a similar treatment of combined 5-FU and radiation resulted in considerable toxicity and no appreciable radiosensitization. CONCLUSION: The present results show that combined suicide gene therapy and RT results in pronounced antitumor effect without any notable toxicity. This indicates that the CD gene may be useful in the development of novel treatment strategies combining radiation and gene therapy in the treatment of locally advanced cancers.

A novel three-pronged approach to kill cancer cells selectively: concomitant viral, double suicide gene, and radiotherapy.

Two obstacles limiting the efficacy of nearly all cancer gene therapy trials are low gene transduction efficiencies and the lack of tumor specificity. Recently, a replication-competent, E1B-attenuated adenovirus (ONYX-015) was developed that could overcome these limitations, because it was capable of efficiently and selectively destroying tumor cells lacking functional p53. In an attempt to improve both the efficacy and safety of this approach, we constructed a similar adenovirus (FGR) containing a cytosine deaminase (CD)/herpes simplex virus type-1 thymidine kinase (HSV-1 TK) fusion gene, thereby allowing for the utilization of double-suicide gene therapy, which has previously been demonstrated to produce significant antitumor effects and potentiate the therapeutic effects of radiation. The FGR virus exhibited the same tumor cell specificity and replication kinetics as the ONYX-015 virus in vitro. Importantly, both the CD/5-FC and HSV-1 TK/GCV suicide gene systems markedly enhanced the tumor cell-specific cytopathic effect of the virus, and, as expected, sensitized tumor cells to radiation. By contrast, neither the FGR virus nor either suicide gene system showed significant toxicity to normal human cells. Both suicide gene systems could be used to suppress viral replication effectively, thereby providing a means to control viral spread. The results support the thesis that the three-pronged approach of viral therapy, suicide gene therapy, and radiotherapy may represent a powerful and safe means of selectively destroying tumor cells in vivo.

Human translocation liposarcoma-CCAAT/enhancer binding protein (C/EBP) homologous protein (TLS-CHOP) oncoprotein prevents adipocyte differentiation by directly interfering with C/EBPbeta function.

Human translocation liposarcoma (TLS)-CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) is a fusion oncoprotein found specifically in a malignant tumor of adipose tissue and results from a t(12;16) translocation that fuses the amino-terminal part of TLS to the entire coding region of CHOP. Being that CHOP is a member of the C/EBP transcription factor family, proteins that comprise part of the adipocyte differentiation machinery, we examined whether TLS-CHOP blocked adipocyte differentiation by directly interfering with C/EBP function. Using a single-step retroviral infection protocol, either wild-type or mutant TLS-CHOP were co-expressed along with C/EBPbeta in naïve NIH3T3 cells, and their ability to inhibit C/EBPbeta-driven adipogenesis was determined. TLS-CHOP was extremely effective at blocking adipocyte differentiation when expressed at a level comparable to that observed in human myxoid liposarcoma. This effect of TLS-CHOP required a functional leucine zipper domain and correlated with its ability to heterodimerize with C/EBPbeta and inhibit C/EBPbeta DNA binding and transactivation activity in situ. In contrast, the TLS-CHOP basic region was dispensable, making it unlikely that the inhibitory effect of TLS-CHOP is attributable to unscheduled gene expression resulting from TLS-CHOP's putative transactivation activity. Another adipogenic transcription factor, PPARgamma2, was able to rescue TLS-CHOP-inhibited cells, indicating that TLS-CHOP interferes primarily with C/EBPbeta-driven adipogenesis and not with other requisite events of the adipocyte differentiation program. Together, the results demonstrate that TLS-CHOP blocks adipocyte differentiation by directly preventing C/EBPbeta from binding to and transactivating its target genes. Moreover, they provide strong support for the thesis that a blockade to normal differentiation is an important aspect of the cancer process.

Antileukemic activity of TNF-alpha gene therapy with myeloid progenitor cells against minimal leukemia.

Tumor necrosis factor-alpha (TNF-alpha) has exhibited antitumor activity against a variety of tumors in rodents and human tumor xenografts in nude mice, but it has been only marginally effective in cancer patients because of dose-limiting toxicity associated with systemic TNF-alpha therapy. To circumvent toxicity and to test the antileukemic activity against quantitated minimal leukemia, we have cloned human TNF-alpha (HuTNF-alpha) gene in an advanced myeloid progenitor cell line. 32Dcl3 myeloid progenitor cells transfected with HuTNF-alpha cDNA by the retroviral supernatant infection method stably express HuTNF-alpha gene and secrete substantial amounts of HuTNF-alpha. When injected i.v. into irradiated mice, transduced cells could be detected in the marrow but not in spleen or liver 10-12 days later. Injection of 5 x 10(6) transduced cells produced no obvious symptoms of TNF-alpha toxicity (i.e., weight loss, cachexia, or fever) suggesting that TNF-alpha producing cells are well tolerated by the recipient mice. Coinjection of 5 x 10(6) transduced cells and 10(2) or 10(3) 32Dp210 leukemia (BCR/ABL+) cells resulted in inhibition of leukemia development by 10(2) but not 10(3) 32Dp210 cells. An equal dose of nontransduced 32Dcl3 cells was ineffective in inhibiting leukemia progression by 10(2) 32Dp210 cells. Mice that rejected leukemia were BCR/ABL oncogene negative 8 weeks after leukemia cell injection. These data demonstrate the potential for TNF-alpha gene therapy for destroying residual leukemia, without the toxicity of systemic TNF-alpha therapy, following cytoreductive therapy and bone marrow transplant.

Preferential radiosensitization of 9L glioma cells transduced with HSV-tk gene by acyclovir.

The antiviral drug acyclovir, an analogue of purine, was found to selectively enhance the radiosensitivity of rodent tumor cells which were transduced with the herpes simplex virus thymidine kinase gene (HSV-tk). 9L rat glioma cells transduced with HSV-tk and treated with acyclovir (20 micrograms/ml) for 24 hr before or after irradiation were highly sensitive to radiation, as compared with non-transduced glioma cells. When 9L cells transduced with HSV-tk gene were exposed to acyclovir and radiation, the sensitizer enhancement ratio (SER) was 1.6. In vivo, a significant increase in the median survival time of rats with 9L-tk tumors was observed when acyclovir was administered before and after single-dose irradiation, relative to the survival time of similar rats receiving radiation alone. The results show that an antiviral agent can selectively enhance cell killing by radiation in cells transduced with the HSV-tk, and suggest that the addition of HSV-tk gene therapy to standard radiation therapy will improve the effectiveness of treatment for brain tumors.