Biomedical subjects
Publications and source records attributed to I A Cree.
Many of the white dot syndromes are considered to have a granulomatous pathogenesis. The histopathologic characteristics of this case of multifocal choroiditis seen within 15 months of apparent clinical onset show that the white dot lesions were nongranulomatous perivascular choroidal infiltrates, consisting mainly of B lymphocytes. Early choroidal neovascularization was also seen.
Chemotherapy for recurrent ovarian carcinoma (ROC) produces response rates of 10-80% depending on the prevalence of platinum resistance. Most patients relapse within 1 year and median progression-free survival (PFS) is generally no more than 6 months. Previous pretherapeutic chemosensitivity assays mostly failed to improve the outcome of patients with ROC. Newly developed ATP assays show promising retrospective correlation with clinical outcome. We report here the first results of ATP assay-directed chemotherapy in patients with ROC. Therapy was selected by the ATP tumor chemosensitivity assay (ATP-TCA) in a prospective open-label pilot trial for ROC. Objective response rate (ORR), PFS and overall survival (OAS) of the first 25 evaluable patients were retrospectively compared with those of 30 others having similar characteristics who were treated empirically within the same period. The actuarial median observation times were 80 weeks for the ATP-TCA group and 83.5 weeks for the control group, respectively. In the control group, a 37% ORR [two complete responses (CR) and nine partial responses (PR)] was followed by a median PFS of 20 weeks and a median OAS of 69 weeks, mainly related to the use of single-agent chemotherapy. The ORR in the ATP-TCA group was 64% (eight CR and eight PR) (p=0.04) with the majority of responses (11 of 16) achieved with novel combinations. The median PFS in this group was 50 weeks (p=0.003) and the median OAS was 97 weeks (p=0.145). Survival of responding patients was similar in both groups. Chemotherapy guided by the ATP-TCA produced a greater benefit with regard to both ORR and PFS in platinum-refractory patients. ATP-TCA-directed chemotherapy for ROC compares favorably with chemotherapy chosen by a clinician and often leads to the choice of novel drug combinations. These promising results now warrant confirmation by prospective randomized trials.
The role of the c-myc oncogene has been little investigated in uveal melanoma. In this study an analysis of c-myc oncoprotein expression was undertaken using flow cytometry in 71 patients with posterior uveal melanoma. Nuclear c-myc oncoprotein was detected in all of the tumours, and survival analysis revealed a significant association between high oncoprotein positivity and improved survival (log rank test: chi2 = 6.47, P = 0.01). Multifactorial analysis using Cox's proportional hazards model revealed nuclear c-myc oncoprotein to be an independent prognostic marker more accurate than other clinicopathological parameters (log rank test: chi2 = 6.61, P = 0.01). However, this result of high oncoprotein expression correlating with improved outcome is surprising and in contrast to our previous studies using the same method on cutaneous melanoma, where high levels of nuclear c-myc expression have been found to correlate with poor outcome both in primary and secondary disease. This study suggests that the pattern of oncogene expression in uveal melanoma is distinct from cutaneous melanoma and that the underlying biology of these tumours is different.
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PURPOSE: The aim of this study was to survey the histopathological and immunohistochemical features of rejected human corneal allografts. METHODS: Following graft failure in each case due to rejection, paraffin-embedded specimens of 17 corneal transplants which had been replaced were examined by light microscopy and immunohistochemistry. Specimens were either first (n = 9), second (n = 4) or third (n = 4) grafts and were removed at varying intervals from 4 weeks following documented rejection. RESULTS: Those grafts which were removed earliest following onset of rejection had the most intense graft inflammatory infiltrates. Immunohistochemical staining showed a high proportion of graft stroma-infiltrating cells expressing leucocyte common antigen, and many of these cells also bore T cell or macrophage markers. Leucocyte-keratocyte apposition and regional loss of keratocytes were observed in all rejection specimens, but not in non-rejected control grafts. In situ end-labelling of DNA double-strand breaks and morphological features identified keratocyte apoptosis in 5 of 12 specimens examined for this phenomenon. Corneal endothelial cells were absent in 7 specimens and present in reduced numbers in the remaining 10 specimens. CONCLUSIONS: Endothelial cell monolayer attenuation and keratocyte loss are consistent findings in grafts removed subsequent to clinically observed endothelial rejection. Death of donor corneal cells is mediated, at least in part, by apoptosis. The stromal inflammatory infiltrate consists mainly of T lymphocytes and macrophages, which may be responsible for induction of keratocyte apoptosis.
The burgeoning understanding of the molecular basis of carcinogenesis and tumor drug resistance is matched by an appreciation of the complexity of individual tumors. This complexity underlies the heterogeneity of response to treatment and is a major barrier to improving the outcome of solid tumor chemotherapy. While individualization of chemotherapy is theoretically attractive, past attempts to provide such information have produced many papers and little progress. However, the disparate molecular make-up of tumors of the same clinicopathologic type suggests that there may be no alternative and recent progress suggests that individualization of cancer therapy could have considerable benefits. In this review, we consider the alternative methods which might be employed and the requirements which need to be met before they are introduced. It will be some time before molecular analysis can predict chemosensitivity, although this may prove feasible for more specific agents than those currently in use. However, newly developed cellular chemosensitivity assays such as the ATP-tumor chemosensitivity assay allied to selected molecular measurement may already have the potential to select optimal therapy for patients. We need to develop a diverse series of acceptable and biologically logical regimens for each of the common tumor types, all of which can be tested in vitro.
Uveal melanoma has a high mortality rate and responds poorly to existing chemotherapy. We have therefore examined the sensitivity of uveal melanoma to cytotoxic agents using an ex vivo chemosensitivity assay to determine whether some agents which have not been used for this tumor might have activity. An ATP-based tumor chemosensitivity assay (ATP-TCA) was used to determine the effect of nine cytotoxic drugs at multiple dilutions in 28 primary uveal melanoma specimens. Evaluable assay results from up to 16 tumors with each drug showed variable sensitivity to alkylating agents (three of nine with mitomycin C, one of 15 with cisplatin and seven of 15 with treosulfan), cytosine arabinoside (seven of 16), paclitaxel (one of five) and doxorubicin (two of 16). No tumors were sensitive to temozolomide, or 5-fluorouracil, and only one of 14 to vincristine. The combination of treosulfan with cytosine arabinoside resulted in enhanced tumor cell inhibition in three of five tumors tested. Combinations containing paclitaxel combined with doxorubicin or cisplatin showed some activity, but none achieved 100% inhibition and the results were similar to those obtained with paclitaxel alone. Uveal melanoma shows considerable heterogeneity of sensitivity to cytotoxic drugs, with considerable resistance to most agents, matching clinical experience. The results suggest that cytosine arabinoside or gemcitabine plus treosulfan may be an active combination. Clinical trials will commence shortly. The use of the ATP-TCA provides a method of testing multiple agents and combinations in a way which would be otherwise impossible in rare tumors such as uveal melanoma.
BACKGROUND: Previous work has highlighted the prognostic importance of patterns of periodic acid Schiff (PAS) staining (the Folberg patterns) in uveal melanoma. These patterns have been ascribed to blood vessels but the patterns are different from those seen with other staining techniques for blood vessels. It has recently been shown that microvessel density is the dominant prognostic factor in uveal melanoma. This study reinvestigates the nature and significance of the PAS patterns. METHODS: The PAS patterns were compared with the patterns seen with conventional connective tissue stains and with the patterns seen in sections stained for the presence of blood vessels (by immunohistochemistry for factor VIII related antigen). The PAS patterns were determined on a panel of 117 cases of uveal melanoma. The prognostic significance of each of these patterns was determined and, as more than one pattern can exist in a tumour, principal components analysis was performed to determine the number of underlying factors. RESULTS: Comparison of the PAS patterns with other stains demonstrates that they are based on connective tissue including fibrovascular tissue. Five of the nine PAS patterns carried prognostic significance on univariate analysis. Principal components analysis suggested that these patterns represented three underlying factors, which were tentatively identified as representing disordered growth (factor 1), emergence of rapidly growing subclones (factor 2), and section orientation (factor 3). CONCLUSIONS: The PAS patterns are based on fibrovascular tissue and can be ascribed to three underlying factors. The first two of these factors carried prognostic significance and the first (disordered growth) retained independent prognostic significance in a multivariate Cox model which included microvessel density and tumour size.
Recent advances in treatment have achieved a large drop in the prevalence of active leprosy cases, but the incidence is at best decreasing slowly. Most people within leprosy-endemic populations have been exposed to Mycobacterium leprae, but few develop disease and it seems likely that the majority of the population develops protective immunity. If the site of initial infection is in the nose, dissemination of bacilli around the body to skin and nerve implies that the initial infection is bacilliferous and it has been shown that nasal M. leprae are detectable by polymerase chain reaction (PCR) of nasal swabs. Since salivary anti-M. leprae IgA (sMLIgA) levels are correlated with protection, we have surveyed groups of leprosy patients, contacts and the general population for both their sMLIgA and nasal PCR positivity. A total of 304 subjects were enrolled in the study: PCR and mucosal challenge tests were performed in 204 of these individuals. sMLIgA was present in 66% of treated patients, 76% of leprosy workers and 72% of healthy contacts. However, only 33% of indigenous subjects were sMLIgA+, in contrast to the earlier studies showing 74% positivity. PCR for M. leprae was present in both household contacts (2%) and indigenous controls (5%). In a subsequent follow-up study, nasal swabs were taken from 97 of those studied in the first series: three PCR+ individuals followed up after one year became negative, while of the remaining 94 PCR- individuals retested, 2 became positive. Of 112 subjects retested with the mucosal challenge test for sMLIgA: 22 converted from positive to negative and 12 from negative to positive. These results suggest that there is widespread subclinical transmission of M. leprae with transient infection of the nose resulting in the development of a mucosal immune response, despite the fact that few individuals will develop clinical disease. This may explain the current lack of effect of multidrug therapy (MDT) control programmes on incidence, although the reduction in general population immunity is consistent with some effect of MDT on transmission.
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Apart from clinical trials, mitoxantrone (MX) is rarely used in breast cancer (BC) due to the anticipated anthracycline cross-resistance. We have examined this drug versus doxorubicin (DOX) using data obtained from in vitro microplate ATP tumor chemosensitivity assays (ATP-TCA) of BC cells which were derived from 55 chemotherapy-naive patients at time of primary surgery. Both drugs were tested at 6 different concentrations ranging from 6.25% to 200% peak plasma concentration in vivo (PPC). Differences between DOX and MX observed for mean IC50, IC90, and a sensitivity index (SI) were not statistically significant. In vitro response rates were 44% for DOX and 52% for MX. 34 of 52 eligible assays (65%) showed comparable activity of both drugs whereas a lack of cross-resistance was observed in the remaining 18 (35%) tumors as indicated by differences for SI. Cumulative concentration-response plots of tumors responding in vitro with a > or = 50 percent or > or = 90 percent tumor cell inhibition showed a strong dose-dependence for both DOX and MX at concentrations which normally can be achieved within clinical tumors (i.e. 6.25%-50% PPC). At higher concentrations, however, cytotoxicity of DOX and MX could not be improved by further in vitro dose escalation. Moreover, a substantial proportion of BC specimens (DOX: 48.1%; MX: 40.4%) did not experience a > or = 90 tumor cell inhibition at 200% PPC. In conclusion, in vitro results obtained by ATP-TCA indicate that there is no cross-resistance between MX and DOX in a substantial proportion of BC patients. This may be clinically useful and suggests that combinations including MX should be tested in patients clinically resistant to DOX containing regimens. Since both drugs produced sigmoidal concentration-response curves, dose escalation beyond a certain point may not produce increased sensitivity.
Intact phagocyte function is a pre-requisite for successful defence against infection, but paradoxically, these cells may also play a major role in the pathogenesis of the infant respiratory distress syndrome. Phagocyte function is known to be deficient in pre-term infants, who are at risk of infection as a result, but these infants are also at risk of respiratory distress syndrome as a result of surfactant deficiency. Despite this, few longitudinal studies of phagocyte function have been performed in pre-term infants. We have used lucigenin-enhanced chemiluminescence to examine the respiratory burst of mixed samples containing polymorphonuclear leucocytes and monocytes of 100 pre-term infants at 48- to 72-h intervals during their admission to a neonatal care unit. Increased polymorphonuclear leucocyte chemiluminescence was associated with respiratory distress syndrome and the use of intermittent positive pressure ventilation. Multiple linear regression analysis revealed a slight, but significant depression of chemiluminescence in association with the use of gentamicin and penicillin when stronger influencing factors such as the presence of respiratory distress syndrome were taken into consideration. Measurement of phagocyte function by sensitive luminescence assays requires very little blood and may be useful in pre-term infants to follow the severity of respiratory distress syndrome. However, it is probable that other factors such as antioxidant capacity also have an important influence on the degree of tissue damage.
Cefodizime has previously been shown to possess a number of immunomodulating properties in vivo and in vitro using several different test systems. Since most in vitro studies have been performed with cells from normal individuals, we first investigated whether cells from chronic renal failure patients would respond in vitro to cefodizime in the same way as healthy subjects. Subsequently, we investigated the effect of cefodizime (10 g over 10 days in 2-g doses) on phagocyte function ex vivo in an open study of 26 chronic renal failure patients and 16 healthy subjects. Polymorphonuclear leukocytes were tested for their ability to polarize in response to cefodizime and/or f-met-leu-phe peptide. Polymorphonuclear leukocytes and monocytes were tested for their ability to produce chemiluminescence on stimulation with either phagocytic (zymosan) or soluble phorbol myristate acetate stimuli. Phagocyte and lymphocyte membrane receptor expression was compared after exposure to cefodizime. Exposure to cefodizime in vitro causes a significant increase in polarization of polymorphonuclear leukocytes from both normal individuals and renal failure patients (both P < 0.001). It also caused increased chemotaxis and chemokinesis in a modified Boyden chamber assay. Cefodizime did not affect lucigenin-enhanced chemiluminescence and there were only minor effects on cell membrane antigen levels. In the ex vivo study there was a significant increase in polymorphonuclear leukocyte polarization (P < 0.001) attributable to cefodizime, but other investigations showed no significant differences. The results suggest that cefodizime may act as a mild priming agent for some functions, particularly chemotaxis.
Chemotherapy for breast cancer is given on the basis of empirical information from clinical trials, an approach which falls to take into account the known heterogeneity of chemosensitivity between patients. Previous attempts to determine chemosensitivity ex vivo have been disappointing, but in this study results from a newly developed tumor chemosensitivity assay (TCA) have been correlated prospectively with patient response. In this study, we have used heterogeneity data for standard regimens obtained from 116 breast TCAs to set sensitivity/resistance thresholds which were then used to interpret the results from those with known clinical responses. Assay evaluability was 97% in surgical biopsies. Clinical follow-up of stage III/ IV assessable disease was obtained from 27 breast tumors which were successfully tested for chemosensitivity, including 13 needle biopsies. The ATP-TCA assay predicted response correctly in 22 out of 29 (76%) tumors with clinically evaluable disease, suggesting that it is capable of predicting outcome in individual patients. Assays were performed in seven patients before and after chemotherapy using residual or recurrent tumor tissue. Four cases with initial sensitivity showed a decrease in sensitivity within 6 months of starting chemotherapy, while two others without clinical resistance were still sensitive by TCA. All nine courses of therapy given on the basis of TCA sensitivity resulted in partial or complete responses. Controlled trials of TCA-directed treatment against standardized empirical therapy should be conducted before this technology is widely adopted to assess its impact on rates of response, survival and the cost of treatment.
Melanin bleaching prior to the application of primary antibody can alter the immunoreactivity of a number of antigens. The effect of melanin bleaching subsequent to antigen visualisation by four different chromogens used with an immunoalkaline phosphatase technique was investigated. Vector black was the only final reaction product that withstood exposure to the permanganate-oxalate sequence for a time sufficient to bleach melanin.
There is little information about the mucosal immune response in leprosy. We have developed a nasal provocation test with leprosin A which will be used to investigate mucosal immunity to Mycobacterium leprae. Initial studies were performed with increasing doses of leprosin A (1.0 pg/ml-10 micrograms/ml) to determine the optimal safe dose of leprosin A. Anti-M. leprae IgA antibody and normal IgA concentrations were measured in the saliva of leprosy contacts and controls before and after instillation of leprosin A. Nasal leprosin A was well tolerated up to a concentration of 10 micrograms/ml without side effects. None of the six subjects who had not been exposed to leprosy had salivary IgA against whole M. leprae, whereas IgA was detected from 64 h to 140 h following instillation of leprosin A in all of the leprosy hospital workers and in 15 out of 18 healthy household contacts tested. There was no correlation between serum and salivary anti-M. leprae IgA levels before and after testing. Salivary IgA anti-lipoarabinomannan responses were seen in 12 out of 20 household contacts. Normal salivary IgA concentrations varied from 8 to 240 mg/l. The leprosin A nasal provocation test appears to be a safe method for the investigation of the role of mucosal immunity in the pathogenesis of leprosy.