Dose-dependent neovascularization-promoting effect of adenovirus vector CI-1023 in a rat hindlimb ischaemic model.

CI-1023 (AdGVVEGF121.10) is a replication-deficient adenovirus vector (complete E1a-, partial E1b-, partial E3-) delivering human vascular endothelial growth factor-121 gene. Previous studies from this group have established that CI-1023 can successfully transfer human vascular endothelial growth factor-121 gene resulting in local tissue expression of vascular endothelial growth factor protein. The purpose of this study was to evaluate neovascularization-promoting potency and efficacy of CI-1023 in a wide dose range. In a rat hindlimb ischaemic model, we measured neovascularization-promoting effect of CI-1023 using three end-points: post mortem angiography, immuno-histochemistry and Laser Doppler scanning of tissue blood perfusion. Neovascularization-promoting activity of CI-1023 over the dose range of 4 x 10(6) pu-4 x 10(10) pu was evaluated. Our data demonstrated an obvious dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu. The neovascularizing effect is somewhat plateaued at the levels between 4 x 10(8) pu and 4 x 10(10) pu. We conclude CI-1023 is a potent neovascularization-promoting compound, with a dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu in the rat hindlimb ischaemic model.

New chemotherapeutic agents: update of major chemoradiation trials in solid tumors.

The institution of combined modality therapy for unresected solid tumors has resulted in significant improvements in tumor control and survival benefit compared with radiotherapy (RT) alone. A number of chemotherapy agents that can enhance the effectiveness of RT, such as cisplatin and 5-fluorouracil, are now considered standard treatment for patients with a number of cancer types. There is growing interest in a number of additional agents that have also been found to have radiosensitizing ability. These include paclitaxel, docetaxel, irinotecan, gemcitabine, and vinorelbine, as well as biologic agents. Other agents may be of value because they act to counter dose-limiting toxicities associated with RT. This article provides an update of some important, recently completed and ongoing clinical trials evaluating novel chemoradiation protocols, with examples taken primarily from studies conducted by the Radiation Therapy Oncology Group (RTOG). Theoretical approaches to the development of new agents and combined modality regimens are also discussed.

The effect of vascular endothelial growth factor on a rat model of traumatic arteriogenic erectile dysfunction.

PURPOSE: We tested the hypothesis that intracavernous injection of vascular endothelial growth factor (VEGF) can restore erectile function in a rat model of traumatic arteriogenic erectile dysfunction. MATERIALS AND METHODS: Exploration of bilateral internal iliac arteries was performed in 50, 3-month-old male rats. A total of 44 rats underwent bilateral ligation of the internal iliac arteries and 6 that underwent exploration only served as the sham operated group. Minutes later intracavernous injection of phosphate buffered saline (PBS) plus bovine serum albumin in 16 rats, 2 microg. VEGF plus PBS plus BSA in 12 and 4 microg. VEGF plus PBS plus BSA in 16 was performed. At weeks 1, 2 and 6 about a third of the rats in each group underwent electrostimulation of the cavernous nerves to assess erectile function and were then sacrificed. Penile tissues were collected for histochemical and electron microscopy examinations. RESULTS: No impairment of erectile function was noted in sham operated rats. Immediately after arterial ligation all rats showed little or no erectile response to neurostimulation. In PBS treated rats modest recovery of erectile function was noted at week 6. Significant recovery of erectile function was noted in VEGF treated rats at weeks 1 and 2 in the 4 microg. group only and at week 6 in the 2 and 4 microg. groups. Neuronal nitric oxide synthase staining showed a reduction in neuronal nitric oxide synthase positive nerve fibers in the dorsal or intracavernous nerves at week 1. Moderate recovery of neuronal nitric oxide synthase positive nerve fibers was noted in the 2 and 4microg. VEGF treated groups but not in the PBS treated group. Electron microscopy revealed no pathological change in sham operated rats. In dorsal nerves the atrophy of myelinated and nonmyelinated nerve fibers was noted in ligated plus PBS treated rats. Partial recovery was observed in VEGF treated rats. Scattered atrophic smooth muscle cells were seen in PBS and occasionally in VEGF treated rats but not in the sham operated group. The most dramatic findings in VEGF treated rats were hypertrophy and hyperplasia of the endothelial cells, especially those lining the small capillaries. CONCLUSIONS: Ligation of bilateral internal iliac arteries produced a reliable animal model of traumatic arteriogenic erectile dysfunction. Intracavernous injection of VEGF minutes after arterial ligation facilitated the recovery of erectile function.

An update on angiogenesis therapy.

Angiogenesis is a novel approach for the therapy of various ischemia-related pathophysiologic conditions. Proangiogenic growth factors have shown promising results in preclinical studies using protein- and gene-based therapies. However, their success in clinical trials is hindered by the lack of an optimal delivery strategy that would provide sustained and localized levels of the growth factors in the diseased tissue. Targeted delivery of proangiogenic agents is expected to demonstrate therapeutic efficacy of growth factors at relatively lower doses, without the risk of systemic toxicity in terms of unwanted angiogenesis. To achieve the above objectives, various drug delivery systems are under investigation. This review describes the basic mechanism of action of growth factors, their current status in preclinical and clinical studies, and the issue of drug delivery.

Healing of a free tracheal autograft is enhanced by topical vascular endothelial growth factor in an experimental rabbit model.

OBJECTIVE: In 1996, we introduced the free tracheal autograft technique for repair of congenital tracheal stenosis from complete tracheal rings in infants and children. Sources of possible concern with this procedure include the potential for autograft ischemia, patch dehiscence, and recurrent stenosis. Vascular endothelial growth factor is a potent angiogenic inducer (particularly in the setting of ischemia, hypoxia, or both) and is postulated to promote tissue healing. The purpose of this study was to test the hypothesis that pretreatment of tracheal autografts with topical vascular endothelial growth factor would enhance tracheal healing. METHODS: In a rabbit model of tracheal reconstruction (n = 32), an elliptically shaped portion of the anterior tracheal wall was excised. The excised portion of trachea was one third of the tracheal circumference and 2 cm in length (6 tracheal rings). This portion of trachea (the autograft) was soaked in either vascular endothelial growth factor (5 microg/mL, n = 16) or normal saline solution (n = 16) for 15 minutes before being reimplanted in the resultant tracheal opening. Animals were killed and autografts were examined at 2 weeks, 1 month, and 2 months postoperatively for gross and microscopic characteristics. RESULTS: By 2 weeks, and progressing through 1 and 2 months, autografts treated with vascular endothelial growth factor, as compared with control autografts, had reduced luminal stenosis, submucosal fibrosis, and inflammatory infiltrate (P <.05). The autografts tended to become malaligned in control animals, whereas the tracheal architecture was preserved in rabbits treated with vascular endothelial growth factor. Microvascular vessel density was significantly greater in all vascular endothelial growth factor groups (P <.05) at all time intervals. CONCLUSIONS: Topical treatment of free tracheal autografts with vascular endothelial growth factor in a rabbit tracheal reconstruction model enhanced healing, as evidenced by accelerated autograft revascularization, reduced submucosal fibrosis and inflammation, and preservation of the normal tracheal architecture. Topical vascular endothelial growth factor may improve future results of tracheal reconstruction.

Evaluation of the effects of intramyocardial injection of DNA expressing vascular endothelial growth factor (VEGF) in a myocardial infarction model in the rat--angiogenesis and angioma formation.

OBJECTIVES: The effects of direct intramyocardial injection of the plasmid encoding vascular endothelial growth factor (phVEGF165) in the border zone of myocardial infarct tissue in rat hearts were investigated. BACKGROUND: Controversy exists concerning the ability of VEGF to induce angiogenesis and enhance coronary flow in the myocardium. METHODS: Sprague-Dawley rats received a ligation of the left coronary artery to induce myocardial infarction (MI). At 33.1 +/- 6.5 days, the rats were injected with phVEGF165 at one location and control plasmid at a second location (500 microg DNA, n = 24) or saline (n = 16). After 33.1 +/- 5.7 days, the hearts were excised for macroscopic and histologic analysis. Regional blood flow ratios were measured in 18 rats by radioactive microspheres. RESULTS: phVEGF165-treated sites showed macroscopic angioma-like structures at the injection site while control DNA and saline injection sites did not. By histology, 21/24 phVEGF165-treated hearts showed increased focal epicardial blood vessel density and angioma-like formation. Quantitative morphometric evaluation in 20 phVEGF165-treated hearts revealed 44.4 +/- 10.5 vascular structures per field in phVEGF165-treated hearts versus 21.4 +/- 4.7 in control DNA injection sites (p < 0.05). Regional myocardial blood flow ratios between the injection site and noninfarcted area did not demonstrate any difference between phVEGF,165-treated hearts (0.9 +/- 0.2) and saline-treated hearts (0.7 +/- 0.1). CONCLUSIONS: Injection of DNA for VEGF in the border zone of MI in rat hearts induced angiogenesis. Angioma formation at the injection sites did not appear to contribute to regional myocardial blood flow, which may be a limitation of gene therapy for this application.

Therapeutic efficacy of recombinant human leukemia inhibitory factor in a primate model of radiation-induced marrow aplasia.

The therapeutic efficacy of recombinant human leukemia inhibitory factor (LIF) was examined in a nonhuman primate model of radiation-induced marrow aplasia. Rhesus monkeys received 450 cGy of total-body, 1:1 mixed neutron:gamma radiation. For 23 days thereafter, each monkey received a daily subcutaneous injection of LIF or human serum albumin (HSA) at a dose of 15 micrograms/kg body weight. Complete blood counts and white blood cell differentials were monitored for 60 days postirradiation. Administration of LIF significantly decreased (P < or = .05) the duration of thrombocytopenia (platelet count < 30,000 or 20,000/microL), ie, 9.3 days or 6.3 days, respectively, versus the HSA-treated control monkeys, 12.2 days or 10.2 days, respectively. Treatment with LIF did not alter the duration of neutropenia (absolute neutrophil count < 1,000/microL) as compared with the HSA-treated control monkeys. Cytokine administration did not exacerbate the radiation-induced anemia observed in the HSA-treated control monkeys.

Prospects for gene therapy and lymphokine therapy for metastatic melanoma.

Conventional treatment of cancer, especially for patients with metastatic melanoma tumor, is often ineffective. Immunotherapy and recently introduced gene therapy have revolutionized the treatments of patients with metastatic melanoma tumor. Use of biological response modifiers, such as interleukins and interferons, have been found to enhance therapeutic benefits to patients with malignant melanoma. Initial studies with a high-dose interleukin-2 (IL-2) therapy have proved effective in patients with melanoma tumor, although a variety of systemic toxicities were observed. A low-dose IL-2 continuous infusion has shown a similar response in patients with melanoma tumor, but produced lesser toxicity. The low-dose IL-2 therapy has been studied with an adoptive transfer combined with either autologous lymphokine activated killer cells or autologous tumor infiltrating lymphocytes (TIL). IL-2 in combination with chemotherapeutic agents such as flavone acetic acid, dacarbazine, and cyclophosphamide have also been studied in patients with metastatic melanoma. Results have shown a moderate response in patients with metastatic melanoma. TIL therapy, however, has been shown to result in higher objective regression due to potent tumor-specific killing and tumor-specific targeting characters of the TIL. The tumor targeting nature of the TIL creates the possibility of using TIL as a vehicle to deliver gene product specifically to tumor tissue. Safety and toxicity of gene-transduced TIL were addressed by the use of neomycin-resistant, gene-transduced TIL in patients with metastatic melanoma. We also investigated the use of vaccinia oncolysate therapy by using the viral oncolysate prepared with IL-2 gene encoded vaccinia virus. Preliminary studies with murine hepatic metastases colon model have shown encouraging results.

[Experimental lymphokine treatment of the wound process]. / Limfokinoterapiia ranevogo protsessa v éksperimente.

It is shown in experiment on the rabbits that the autologous supernatants of the culture of stimulating lymphocytes and the fractions of lymphokines with mol. weight 40000-20000 and 60000-70000 considerably reduced the periods of wound healing.

Protocol design for lymphokine testing in clinical studies of human cancer.

Sequential clinical studies have been conducted over the last seven years in the Surgery Branch, NCI utilizing transferred cells and the administration of interleukin-2 (IL-2). Each study was based on preclinical testing in experimental models or on in vitro evidence suggesting clinical application. Although clinical studies are difficult to conduct as rigorously as those in experimental models, an identical approach is required to obtain meaningful information on which to base subsequent trials. Preclinical testing of lymphokines requires general safety testing in animals, identification of the components with and without biologic activity within the material to be administered, evidence of stability, sterility, lack of pyrogenicity and demonstration of potency. The major clinical goal is to obtain evidence of tumor regression with the added goals of determining other clinical parameters (such as toxicity, half life, endocrine changes and trafficking information). Biologic and immunologic parameters other than those detailed above are routinely obtained to guide further efforts. A sequential evaluation of clinical trials performed in the evaluation of adoptive transfer of activated lymphocytes and the administration of IL-2 is presented.

Lymphokines, monoclonal antibodies, and other biological response modifiers in the treatment of cancer.

Biologicals and biological response modifiers (BRMs) represent a new class of agents for cancer therapy. Historically, there have been many attempts to stimulate the immune response with nonspecific immunomodulators in the form of bacterial extracts, viruses, and chemicals. Although these approaches have occasionally proven useful under defined conditions in experimental models, their extension to the clinic has been largely unsuccessful. Recent advances in molecular biology and hybridoma technology have made available genetically engineered lymphokines and cytokines, as well as monoclonal antibodies, as highly purified biologicals for cancer treatment. These agents may act directly on tumor cells and/or may act on the patient's own biological responses to induce an antitumor response. Selective defects in T-cell function have recently been identified in cancer patients and in patients with acquired immunodeficiency syndrome (AIDS). Simultaneously, the availability of gamma interferon (gamma-IF) and interleukin-2 (IL-2) may allow for the selective correction of these T-cell deficits, leading to restoration of the patient's immune responses and perhaps correction of the clinical syndromes. Preliminary data suggest that gamma-IF and IL-2 have in vitro activity on these T-cell defects, and the preliminary evidence that these agents have activity in vivo will be reviewed. Extensive trials are being conducted at the National Cancer Institute with monoclonal antibodies as anticancer agents. Animal model experiments have demonstrated considerable antitumor activity of immunoconjugates using monoclonal antibodies tied to toxins. Preliminary clinical results suggest that T-101 in leukemia and lymphoma and 9.2.27 in malignant melanoma may prove useful as specific reagents in the treatment of these disorders. While the antitumor effects with these antibodies have not been dramatic, our preliminary data in approximately 30 patients with leukemia, lymphoma, and melanoma clearly demonstrate the ability of intravenous monoclonal antibody to locate and specifically label tumor cells bearing the target antigens. It has been possible to localize antibody on the tumor cells in melanoma deposits that are barely visible in the skin. These data and radioimaging data suggest a future role for immunoconjugates as anticancer agents.

Biological response modifiers.

It is clear that the rate of change in biology is rapidly accelerating. This is most apparent in the BRM area with respect to the development of new cancer therapies. New surgical approaches are being developed, and new forms of delivery for therapeutic radiation are now available. Radiosensitizers are now also being tested. Similarly, new chemotherapeutic agents and new methods of utilizing existing drugs are being developed. However, the use of BRM either alone or in combination with these other modalities is the area of greatest change and can be expected to be a major factor in achieving a greater understanding of cancer biology and a greater therapeutic specificity in the treatment of cancer patients in this decade (2, 65). Given the new technology of the 1980's, this is an exciting time. It is hoped that sufficient resources will be available to support continuing efforts to develop biologicals into effective anticancer agents. The rapidly advancing technologies of computers, cell fusion, and genetic engineering are all interrelated through the underpinnings of molecular biology to offer us unparalleled opportunities in biological research. These opportunities should translate into more effective treatments for cancer in this decade.