Biomedical subjects
Publications and source records attributed to H A Hopkins.
Across-sectional study was performed of pediatric hematology-oncology patients who received vancomycin; use was compared to the Centers for Disease Control and Prevention (CDC) recommendations for vancomycin use. Thirty-seven patients received 308 doses of vancomycin. AR patients initially received vancomycin as empirical therapy; 100% of this use was not consistent with the CDC recommendations.
Several studies have demonstrated that cytomegalovirus (CMV) infection increases expression of the tumor necrosis factor (TNF) gene. This effect is mediated, in part, by an effect of the CMV immediate early 1 (IE1) gene product on the TNF promoter. To further analyze these interactions, we used plasmids with TNF promoter truncations to determine the site within the promoter where the CMV IE1 gene product mediates its effect. The site was localized to a 40-base pair segment that contains a cAMP response element (CREB). Deletion of the cAMP response element increased basal promoter activation but had little effect on IE1-induced activation. Additional studies demonstrated that the cAMP element is flanked 5' by a PU.1 site and 3' by an NF-kappa B site, both of which increase expression of the TNF promoter. These sequences demonstrated IE1 responsiveness. We next determined the relevance of these observations for normal human cells by infecting human alveolar macrophages with CMV. In these studies we evaluated expression of NF-kappa B, PU.1 and CREB by gel shift assay at immediate early times after infection. We found that CMV infection increased the binding activity of NF-kappa B and PU.1 and decreased the binding activity of CREB. CMV infection also increased expression of the TNF gene in alveolar macrophages. These observations suggest that CMV increases TNF gene expression, in part, by altering the binding activity of transcription factors that regulate gene expression.
Cytomegalovirus (CMV) infection in transplant patients is associated with an increased incidence of gram-negative pneumonia; the mechanism for this is unknown. Human alveolar macrophages (HAM) are an important part of the response of the lung to gram-negative bacteria. They interact with lipopolysaccharide (LPS) via the surface receptor CD14. The effect of CMV on CD14 expression by HAM was examined. HAM were obtained from normal volunteers by bronchoalveolar lavage, and some were exposed to CMV. CD14 expression was assessed by immunofluorescent microscopy and flow cytometry. CMV inhibited the surface expression of CD14 on HAM. Release of soluble CD14 was also reduced from infected cells, and Northern blot analysis revealed that CD14 mRNA was reduced in CMV-exposed cells. These findings were specific for CD14 expression. These results demonstrate that CMV inhibits the ability of HAM to express CD14.
Several studies have shown that interleukin-4 (IL-4) down-regulates synthesis of prostaglandin E2 (PGE2). We evaluated the mechanisms for this suppression in human alveolar macrophages (HAMs). Normal HAMs were obtained from healthy nonsmoking volunteers. The cells either remained unstimulated, or were exposed to 10 micrograms/ml of lipopolysaccharide (LPS) and/or various amounts of IL-4. LPS alone induced the synthesis of large amounts of PGE2 and prostaglandin H synthase-2 (PGHS-2) protein. This effect of LPS was suppressed by increasing amounts of IL-4. Expression of LPS-induced PGHS-2 mRNA was also inhibited by IL-4. In addition, IL-4 inhibited expression of CD14, which is a receptor for LPS bound to the LPS-binding protein (LBP). We conclude that IL-4 down-regulates LPS-induced release of PGE2, by reducing expression of the enzyme, PGHS-2. One potential mechanism for this effect of IL-4 is a reduced expression of CD14, which is the LPS-LBP receptor.
This study was undertaken to examine the surface expression and regulation of CD14 on human alveolar macrophages (HAM). HAM obtained by bronchoalveolar lavage were cultured with or without LPS and with or without fetal bovine serum (FBS) on plastic or Teflon surfaces. CD14 expression was assessed by immunofluorescence. The roles of protein tyrosine kinase, protein kinase A, and protein kinase C were also evaluated utilizing inhibitors of these enzymes. HAM expressed very low levels of surface CD14 at baseline. CD14 was upregulated on HAM with time in culture in the presence of FBS and/or LPS; adherence did not affect CD14 expression. Inhibitors of protein kinase C and protein tyrosine kinase prevented the upregulation of CD14 by LPS but did not effect the upregulation of CD14 by FBS. These studies show that LPS can upregulate expression of its receptor on HAM and that this effect is accentuated by serum-derived factors.
The two primary therapeutic strategies in cancer have been to give either chemotherapy and radiation therapy together or give a complete course of one treatment modality before starting the second. Clinical studies show that toxicity has been one of the major deterrents to substantial improvements in cancer management when the two modalities are administered together. On the other hand, the prolonged time necessary to administer all of one modality followed by the other makes it likely that repopulation of the tumor during sequential treatment will diminish therapeutic effectiveness. A third strategy of giving chemotherapy and radiation therapy has been developed. This new regimen was designed to give chemotherapy initially, maintain the chemotherapy schedule to avoid any reduction in its effectiveness, and add radiation therapy as early as possible in between courses of chemotherapy to minimize the development of cross resistance. One of the primary objectives of alternating chemotherapy and radiation therapy is to increase the therapeutic index by reducing toxicity without a significant reduction in therapeutic effectiveness. Recent clinical, experimental, and theoretic results with radiation therapy and chemotherapy for cancer management emphasize the necessity of giving both modalities with the greatest intensity possible in the initial phase of induction therapy. Cancer treatment scheduling determines the toxicity and thereby limits the dose intensity that can be tolerated. Scheduling may also govern the antitumor effect directly; however, normal host tissue makes the determination of the direct effects on the tumor difficult, if not impossible, in clinical studies. Well-defined experimental solid-tumor systems provide the means for determining directly the relationship between toxicity and antitumor effects in relation to tumor burden and total therapeutic dose. In addition, its relationship to dose intensity and scheduling can be determined by the using more sophisticated research techniques, such as response surface methods. Well-defined clinical protocols to determine how to interact chemotherapy with radiation therapy more effectively hold considerable potential for rapid improvement in treatment of radiosensitive and chemosensitive cancers.
Clinical and experimental results with radiotherapy and chemotherapy for the treatment of cancer emphasize the necessity of giving treatment with both modalities with the greatest intensity possible in the initial phase of induction therapy. This can best be accomplished by utilizing new approaches, such as alternating the chemotherapy with newer methods of delivery of radiotherapy, which has the potential for greater destruction of tumor within acceptable limits of host toxicity. The clinical data from Institut Gustave Roussy, Villejuif, shows that alternating chemotherapy and radiotherapy produced an excellent complete response rate of 79% and a 4 year relapse-free survival rate of 22% in 109 patients with limited small cell lung cancer. Based on 5 year survival criteria, approximately one-fourth of the patients can be considered cured in 1 more year if no further change occurs. The results of the University of Virginia experimental studies have also demonstrated the superiority of alternating cyclophosphamide and radiotherapy in three courses of induction therapy over conventional methods of delivery of the two modalities. Using the experimental solid tumor 3924A, a cure rate of 50% or greater was obtained with this protocol with acceptable toxicity to the host, as opposed to no cures from three or more courses of either modality alone. One of the major deterrents to tumor cure with concomitant chemotherapy and radiotherapy has been excessive toxicity, which can be avoided by temporal separation (+/- 7 days) of the delivery of the two modalities without a significant loss of therapeutic effectiveness. Well-defined clinical protocols to determine how to more effectively interact chemotherapy with radiotherapy offer some of the greatest potential for a more rapid improvement in treatment.
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The experimental tumor 3924A in ACI rats metastasizes to the lungs, but this is rarely apparent unless the life span of the animal is increased by treatment of the tumor. Pulmonary metastases were prevalent among animals given radiation daily or in multiple fractions per day over periods of 3-4 weeks at total doses ranging from 3,000 to 11,250 rad. Multiple doses of cyclophosphamide, each at the level that was lethal to 10% of the animals, did not cure the primary tumor but appeared to delay the appearance of metastases. Treatment schedules that alternated the delivery of multiple 250-rad fractions of radiation during 2-day periods with cyclophosphamide at a dose of 150 mg/kg for a total of three courses were effective in curing the primary tumor and eliminating pulmonary metastases.
The doubling time (Td) for hepatoma 3924A in ACI rats is relatively constant between different treatments when growth resumes after treatment with either chemotherapy or radiotherapy. Advantage was taken of this to estimate the fraction of surviving cells (SF) from in vivo growth delay (GD) data using the expression SF = 1/2GD/Td. Survival curves were constructed for several recently published treatment schedules which employed alternating radiotherapy and chemotherapy, and for both daily and multiple fractions per day (MFD) radiation schedules. A doubling time of 5.2 days was assumed, in the range observed for control and regrowing treated tumors, which yields one surviving cell at the TCD37 (3650 cGy single dose). The single fraction, multi-target, single hit model, SF = 1 - (1 - e-D/D0)n with D0 = 406 cGy and n = 1.63 is a reasonable representation of the data over the dose range 375-2250 cGy. The D0 of 406 cGy should be considered as a relative, not absolute value which is dependent on the radiosensitivity of the tumor cells and the accuracy of the doubling time, but is useful in relating the single dose data to more complex radiation schedules. Using D0 = 406 cGy, n = 1.63, and f = number of fractions, the multi-fraction, multi-target, single hit model SF = (1 - [1 - e-D/D0]n)f closely fits our data for 30 daily fractions given at 100 to 375 cGy/day for total radiation doses of 3000 to 11,250 cGy. The fraction of surviving cells for daily radiation alternated with three courses of cyclophosphamide (CP) was in good agreement with an additive effect of the two modalities at radiation doses of 3000 to 5640 cGy. Multiple fractions per day radiation given as 250 cGy fractions was more effective than predicted by the model both when given alone or alternately with cyclophosphamide.
A series of experimental studies has been carried out to assess the relationship between the effectiveness of tumor control, normal tissue reaction and median survival following different radiation fractionation schedules on the solid tumor model 3924A in the ACI rat. The total radiation dose of 7500 rad and the dose per fraction of 250 rad were held constant. Tumor control and life expectancy improved with 250 rad given continuously as multiple fractions per day (MFD) (1, 2 and 3 fractions per day over 30-, 15- and 10-day periods, respectively). However, the maximum acute skin reaction surrounding the tumor was greater for MFD given continuously than for single daily fractions. We have also completed studies of the therapeutic effectiveness of MFD given intermittently at 11-day intervals. Tumor control following MFD given intermittently was comparable to that following the daily fraction schedule given continuously, and the normal tissue reaction was acceptable at a total radiation dose of 7500 rad. Experimental and clinical results in head and neck cancer indicate that large total radiation doses in the order of 7000 rad given as continuous MFD are associated with unacceptable normal tissue reaction. However, these clinical and experimental results also indicate that large total radiation doses can be given as MFD intermittently and effectively used in cancer management. The marked superiority of alternating chemotherapy and radiotherapy in obtaining a tumor cure rate of greater than or equal to 50% in our experimental system and the superiority of alternating chemotherapy and radiotherapy in extensive Hodgkin's disease (complete response rate--87%) over chemotherapy alone, radiotherapy alone, or the two modalities given together or as split course provides the clinical and experimental basis for the development of more effective protocols in the treatment of tumors responsive to chemotherapy and radiotherapy. A complete response rate of 78% in patients with head and neck cancer, and a complete response rate of 89% in patients with limited small cell carcinoma of the lung using alternating chemotherapy and radiotherapy indicate that it is now possible to proceed rapidly in clinical protocol design for these two additional tumors responsive to both chemotherapy and radiotherapy.
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This study with the rat hepatoma 3924A demonstrated the marked improvement in tumor cure rates and control of tumor growth that can be achieved by the addition of cyclophosphamide (CP) to multiple fractions of radiation per day (MFD) schedules given intermittently. MFD radiation was delivered over a 2-day period followed by CP (150 mg/kg or 0.9 g/m2) 1 day later; this combined course was repeated at 11-day intervals (to allow for gastrointestinal tract and bone marrow recovery) for a total of 3 courses over a 23-day period. Cure rates of 30, 50 and 60% were achieved with total radiation doses of 4500, 6000 and 7500 rad, respectively, when the MFD radiation was given with CP. No cures and no complete responses were realized when the same intermittent MFD schedules for radiation were employed up to 9000 rad without CP. Other groups of 10 animals each were treated with daily fractions of 100, 150, 188, 250 and 375 rad given on days 0-9, 11-20 and 22-31. A 150 mg/kg or 0.9 g/m2 dose of CP was given after each course of daily radiation on days 10, 21 and 32 in the combined treatment groups. No complete responses or tumor cures occurred with radiation alone given daily for total radiation doses, which were increased from 3000 to 11,250 rad. Only the highest radiation dose given, 375 rad per day to a total of 11,250 rad, resulted in a complete response rate and tumor cure rate of 50% when CP was added. The addition of CP to the daily fractionation schedules reduced the total dose needed to give a growth delay of 100 days by 39% (5600 rad versus 9200 rad). The addition of CP to the intermittent MFD schedules further reduced the total dose needed to give a growth delay of 100 days to 4200 rad. Major improvements in some types of cancer treatment may be realized if we can develop clinical protocols for the alternate use of chemotherapy and radiotherapy as we have done successfully in our experimental program. The finding that intermittent MFD radiation schedules are as effective as daily schedules when given alone suggests that greater flexibility of patient management in clinical radiotherapy may be possible without a major loss of therapeutic effectiveness. These alternated fractionated schedules offer the possibility of optimizing treatment in terms of patient convenience and economy as well as the potential for improving the effectiveness of the interaction of radiotherapy with radiosensitizers, radioprotectors, and hyperthermia in addition to chemotherapy.
Major increases in the time between administration of two modalities, radiation and cyclophosphamide (CP), from 1 to 7 days and in the overall time of delivery of 3 courses of combined therapy from 24 to 35 days were carried out in rats with hepatoma 3924A without major loss of therapeutic effectiveness. Cure rates of 50% or greater could be maintained even though treatment was given over much longer time periods. The radiation was given as hyperfractionated, split-course schedules which were devised by increasing the number of 250 rad fractions over a 2-day period. In one series of experiments these 2-day schedules were given at 11-day intervals for 3 courses on days 0 and 1, 11 and 12, 22 and 23; and CP (150 mg/kg) was given 1 day after each of the 3 radiation courses on days 2, 13, and 24. In the second series of experiments radiation was given on days 0 and 1, 14 and 15, 28 and 29; and this was alternated with 3 single doses of CP given 1 week after each of the 3 courses of radiation, on days 7, 21 and 35. Increasing the total radiation dose from 6000 to 7500 rad in the series given CP 1 day after each of three courses of radiation results in an increase in total tumor cure rates from 50% to 60%. The tumor cure rate in the series given CP 7 days after radiation increased from 10% to 70% when the total radiation dose was increased from 6000 to 7500 rad. Increasing the total radiation dose from 6000 to 7500 rad increased the magnitude of the acute skin reaction as well as the duration of recovery. However, the skin reactions for both the 6000 and 7500 rad were acceptable. Host toxicity and normal tissue reaction were within acceptable limits for both modalities. The results of these studies, therefore, indicate that excessive toxicity, one of the major deterrents to the effective combined utilization of these two primary means of cancer management, may be avoided by temporal separation of delivery while maintaining tumor cure rates of 50% or greater.
Tumour cords have been examined quantitatively in two rat hepatomas, 3924A and H-4-II-E, that differ in their radiobiological oxygenation status (oxygen enhancement ratio for growth delay [tumour clamped: tumor 'in air'] was 1.35 for 3924A and only 1.08 for H-4-II-E). The average thickness of tumour cords in 3924A was 118 microns and only 69 microns in H-4-II-E. The migration rates across the cords of the two tumours were approximately the same (1.7 and 1.4 micron X h-1) but for any given distance from the subtending blood vessel, the proportion of histologically-dead cells within the cord was always higher for H-4-II-E. Volume for volume, H-4-II-E contained four times as much vascular space as 3924A but it is suggested that the poor quality of this vasculature in H-4-II-E contributed to its relative radioresistance.