Transcriptional changes in the aphid species Myzus cerasi under different host and environmental conditions.

Aphids feature complex life cycles, which in the case of many agriculturally important species involve primary and secondary host plant species. Whilst host alternation between primary and secondary host can occur in the field depending on host availability and the environment, aphid populations maintained as laboratory stocks generally are kept under conditions that allow asexual reproduction by parthenogenesis on secondary hosts. We used Myzus cerasi (black cherry aphid) to assess aphid transcriptional differences between populations collected from primary hosts in the field and those adapted to secondary hosts under controlled environment conditions. Transfer of M. cerasi collected from local cherry trees to reported secondary host species resulted in low survival rates. Moreover, aphids were unable to survive on the secondary host land cress, unless first adapted to another secondary host, cleavers. Transcriptome analyses of the different aphid populations (field collected and adapted) revealed extensive transcriptional plasticity to a change in environment, with predominantly genes involved in redox reactions differentially regulated. Most of the differentially expressed genes were duplicated and we found evidence for differential exon usage. Our data suggest that aphid adaptation to different environments may pose a major hurdle and leads to extensive gene expression changes.

Phosphatidylserine exposure on the surface of Leishmania amazonensis amastigotes modulates in vivo infection and dendritic cell function.

Leishmania amazonensis parasites can cause diverse forms of leishmaniasis in humans and persistent lesions in most inbred strains of mice. In both cases, the infection is characterized by a marked immunosuppression of the host. We previously showed that amastigote forms of the parasite make use of surface-exposed phosphatidylserine (PS) molecules to infect host cells and promote alternative macrophage activation, leading to uncontrolled intracellular proliferation of the parasites. In this study, we demonstrated that treatment of infected mice with a PS-targeting monoclonal antibody ameliorated parasite loads and lesion development, which correlated with increased proliferative responses by lymphocytes. In addition, we observed an enhanced dendritic cell (DC) activation and antigen presentation in vitro. Our data imply that the recognition of PS exposed on the surface of amastigotes plays a role in down-modulating DC functions, in a matter similar to that of apoptotic cell clearance. This study provides new information regarding the mechanism of immune suppression in Leishmania infection.

Investigations of mechanical vibrations for beamlines at the Canadian Light Source.

Vibration is often a problem causing poor quality of photon beams at synchrotron radiation facilities, since beamlines are quite sensitive to vibrations. Therefore, vibration analysis and control at synchrotron radiation facilities is crucial. This paper presents investigations on mechanical vibrations at four beamlines and endstations at the Canadian Light Source, i.e. the Canadian Macromolecular Crystallography Facility 08ID-1 beamline, the Hard X-ray MicroAnalysis 06ID-1 beamline, the Resonant Elastic and Inelastic Soft X-ray Scattering 10ID-2 beamline, and the Scanning Transmission X-ray Microscope endstation at the Spectromicroscopy 10ID-1 beamline. This study identifies vibration sources and investigates the influence of mechanical vibrations on beamline performance. The results show that vibrations caused by movable mechanical equipment significantly affect the data acquired from beamlines.

Matrix metalloproteinase-9 triggers the angiogenic switch during carcinogenesis.

During carcinogenesis of pancreatic islets in transgenic mice, an angiogenic switch activates the quiescent vasculature. Paradoxically, vascular endothelial growth factor (VEGF) and its receptors are expressed constitutively. Nevertheless, a synthetic inhibitor (SU5416) of VEGF signalling impairs angiogenic switching and tumour growth. Two metalloproteinases, MMP-2/gelatinase-A and MMP-9/gelatinase-B, are upregulated in angiogenic lesions. MMP-9 can render normal islets angiogenic, releasing VEGF. MMP inhibitors reduce angiogenic switching, and tumour number and growth, as does genetic ablation of MMP-9. Absence of MMP-2 does not impair induction of angiogenesis, but retards tumour growth, whereas lack of urokinase has no effect. Our results show that MMP-9 is a component of the angiogenic switch.

Emergence of immunoglobulin variants following treatment of a B cell leukemia with an immunotoxin composed of antiidiotypic antibody and saporin.

The potency and specificity of immunotoxins consisting of monoclonal antiidiotype conjugated to the ribosome-inactivating protein, saporin, have been evaluated in the treatment of guinea pig L2C B lymphocytic leukemia. The immunotoxins were therapeutically much more effective than their parent antibodies. Their specificity reflected that of their antiidiotype component. Although the leukemia emerged eventually in most animals treated with these conjugates, most of the cells showed altered Ig expression, which rendered them resistant to the therapy. Commonly, the emerging cells had lost mu heavy chain production, leaving them negative for intracellular, surface, and secreted IgM, but still positive for lambda light chain production. In addition, a minor group of L2C variants was identified in a protocol designed to detect mutants at very low frequency: here the cells were exposed in vitro to immunotoxin and, while still viable as judged by dye-exclusion, inoculated in large numbers into animals. In tumor that emerged under these circumstances, the majority of cells were again immunoglobulin-negative; however a minority exhibited IgM with an altered idiotype (Idiotope-loss variants), rendering them unreactive with immunotoxin. Immunotherapy with unmodified anti-Id antibody alone does not reveal these variants, and we suggest it is the increased selective force exerted by the highly potent immunotoxins that allow these minor nonreactive populations to emerge.

Endovascular stents for intermittent claudication.

BACKGROUND: Endovascular stents have been suggested as a means to improve the patency of arteries after angioplasty in patients with intermittent claudication. This is an update of a Cochrane review published in 2002. OBJECTIVES: The null hypothesis to be tested by this review is that for individuals with claudication the use of an endovascular stent, in addition to percutaneous transluminal angioplasty, does not improve symptoms of life-style limiting claudication when compared to percutaneous angioplasty alone. SEARCH STRATEGY: For this update the Cochrane Peripheral Vascular Diseases Group searched their Specialised Register (last searched August 2009) and the Cochrane Central Register of Controlled Trials (CENTRAL) in The Cochrane Library (last searched 2009, Issue 3). SELECTION CRITERIA: Randomised trials comparing angioplasty alone versus angioplasty with endovascular stents in patients with intermittent claudication. DATA COLLECTION AND ANALYSIS: Two authors independently assessed trial quality and extracted the data. Only published trial data were used but unpublished data were sought for the update. Effectiveness was measured by the pre-defined primary outcome measures restenosis or reocclusion rates and maximum walking distance. MAIN RESULTS: Two studies were included involving a total of 104 participants. Both studies included only individuals with femoro-popliteal disease. They compared angioplasty and stenting with the Palmaz stent against angioplasty alone. Although one study showed a slight statistical advantage in arterial patency after angioplasty alone, this was not found when the two studies were combined. No differences in the secondary outcomes were detected in either study. AUTHORS' CONCLUSIONS: The small number of relevant studies identified together with the small sample sizes and methodological weaknesses severely limit the usefulness of this review in guiding practice. The results from larger multicentre trials are needed.

Tumor infarction in mice by antibody-directed targeting of tissue factor to tumor vasculature.

Selective occlusion of tumor vasculature was tested as a therapy for solid tumors in a mouse model. The formation of blood clots (thrombosis) within the tumor vessels was initiated by targeting the cell surface domain of human tissue factor, by means of a bispecific antibody, to an experimentally induced marker on tumor vascular endothelial cells. This truncated form of tissue factor (tTF) had limited ability to initiate thrombosis when free in the circulation, but became an effective and selective thrombogen when targeted to tumor endothelial cells. Intravenous administration of the antibody-tTF complex to mice with large neuroblastomas resulted in complete tumor regressions in 38 percent of the mice.

The role of subintimal angioplasty in the management of arterial disease of the lower limb: the North-East of Scotland experience.

INTRODUCTION: The management of occlusive femoropopliteal disease continues to evolve and a definitive strategy remains to be defined. We examine the utility ofsubintimal angioplasty (SIA) in our institution. METHODS: A retrospective study with predefined end-points, including technical success and primary patency. RESULTS: 61 consecutive cases were identified (claudicants n=29 and critical ischaemia n=32). Sixty-four percent of occlusions were greater than 10 cm with poor run-off (60% with two vessels or less). Technical and physiological success was 95% and 79% respectively, with clinical improvement reported by 72%. At a mean follow-up of 20 months twelve-month primary patency (assessed clinically, with ABPI and selective duplex scanning) was 67% (subgroup analysis: claudicants 83%, criticals 53%, p=0.02) and morbidity 8% with no limb loss or procedure related mortality. CONCLUSION: SIA is an effective procedure for chronic lower limb ischaemia with acceptable outcome. Our experience correlates well with evidence in the current literature.

TAT-BH4 counteracts Abeta toxicity on capillary endothelium.

Oxidative stress is one of the factor contributing to blood brain barrier degeneration. This phenomenon is observed during pathological conditions such as Alzheimer's disease or cerebral amyloid angiopathy in which brain haemorrhages are very frequent. Both diseases are characterized by beta amyloid peptide deposition either in neurons or in vessels. Oxidative stress leads to impairment of mitochondrial functions and apoptotic cell death subsequent to caspases activation. In this paper we demonstrate that BH4 domain of Bcl-xl administrated to endothelial cells as the conjugated form with TAT peptide, reverts Abeta-induced apoptotic cell death by activating a survival programme which is Akt/endothelial nitric oxide synthase dependent.

An immunotoxin composed of monoclonal anti-Thy 1.1 antibody and a ribosome-inactivating protein from Saponaria officinalis: potent antitumor effects in vitro and in vivo.

The ribosome-inactivating protein saporin, from Saponaria officinalis, was coupled by a disulfide bond to monoclonal anti-Thy 1.1 antibody (OX7) and to its F(ab')2 fragment. The immunotoxins were at least as toxic as the plant toxin ricin to the Thy 1.1-expressing cell lines AKR-A and BW5147 in tissue culture. They reduced the rate at which the cells incorporated [3H]leucine into protein by 50% at cell concentrations of 1.5-3 X 10(-11) and 3 X 10(-12) M, respectively. The toxic effect was specific. No toxicity was seen when the immunotoxins were applied to Thy 1.2-expressing EL 4 lymphoma cells at 3 X 10(-8) M, and a control immunotoxin made from an antibody (R10) of irrelevant specificity was without effect on AKA-A cells. Further, the treatment of spleen cells from AKR mice with OX7-saporin at 10(-8) M abolished their response to the T-lymphocyte mitogen concanavalin A, without impairing their response to the B-lymphocyte mitogen lipopolysaccharide. A single iv injection of OX7-saporin into nu/nu randombred mice bearing peritoneal AKR-A lymphoma cells prolonged the survival time of the animals by an extent corresponding to that expected if 99.999% of the tumor cells had been eradicated by the immunotoxin. None of the control materials (unconjugated OX7, unconjugated saporin, OX7 plus saporin, or R10-saporin) delayed tumor growth. The OX7 F(ab')2-saporin conjugate was also highly effective as an antitumor agent, although significantly less so than the conjugate made with intact OX7. Unexpectedly, the acute toxicity of saporin to mice (median lethal dose = 6.8 mg/kg) was elevated eightfold to sixteenfold by conjugation to OX7, R10, or OX7 F(ab')2. Histologic examination of recipients of the immunotoxin revealed gross damage to hepatic parenchymal cells and to the white pulp of the spleen, neither of which was caused by unconjugated saporin. Ricin A-chain coupled to OX7 antibody was one hundredfold to one thousandfold less effective than OX7-saporin as an antitumor agent in vivo, although the two immunotoxins were equally cytotoxic to AKR-A cells in vitro.

Comparison of two anti-Thy 1.1-abrin A-chain immunotoxins prepared with different cross-linking agents: antitumor effects, in vivo fate, and tumor cell mutants.

The A-chain of the plant toxin abrin was covalently linked to monoclonal anti-Thy 1.1 antibody (OX7) with the use of either N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP) or 2-iminothiolane hydrochloride (2IT). The SPDP reagent generates a linkage containing a disulfide bond and an amide bond, whereas the 2IT reagent generates a linkage containing a disulfide bond and an amidinium bond. The two immunotoxins were powerfully and specifically toxic to Thy 1.1-expressing murine AKR-A lymphoma cells in vitro. Both reduced the rate of protein synthesis of the cells by 50% at a concentration of 10(-11) M. However, clonogenic assays revealed that about 1% of the AKR-A cells survived treatment with high concentrations of OX7-SPDP-abrin A, whereas only about 0.1% survived treatment with similar concentrations of OX7-2IT-abrin A. Several clones of the surviving cells were isolated. Of 11 clones of cells that had survived exposure to OX7-SPDP-abrin A, 10 were resistant to further treatment with OX7-SPDP-abrin A but had normal sensitivity to OX7-2IT-abrin A. These clones expressed moderate to high levels of the Thy 1.1 antigen and were fully sensitive to abrin. In contrast, all 10 clones of cells that had survived exposure to OX7-2IT-abrin A were substantially or entirely resistant to both immunotoxins. They expressed low to high levels of the Thy 1.1 antigen and were fully sensitive to abrin. The 2IT-linked immunotoxin was much more effective than the SPDP-linked immunotoxin at protecting nu/nu mice against the growth of AKR-A lymphoma cells in the peritoneal site. A single iv injection of 0.3 nmol OX7-2IT-abrin A eradicated at least 99.99% of the tumor cells, as judged from the extension in the median survival time of the animals, whereas OX7-SPDP-abrin A eradicated only about 99% of the cells. The tumors that developed in the animals that received OX7-2IT-abrin A were Thy 1.1-negative, whereas those in the recipients of OX7-SPDP-abrin A generally expressed normal levels of the Thy 1.1 antigen. The difference in antitumor activity of the immunotoxins was not due to differences in their in vivo fate, inasmuch as they were cleared from the bloodstream at an identical rate and broke down at the same rate to release free antibody.(ABSTRACT TRUNCATED AT 400 WORDS)

Therapeutic transarterial embolisation in the management of benign and malignant renal conditions.

PURPOSE: The aim of this study was to critically appraise the efficacy and complications of Therapeutic Transarterial Embolisation (TAE) in various benign and malignant renal conditions. MATERIAL AND METHODS: The records of all patients who underwent renal embolisation procedures, at a single institution, between March 1992 and March 2004, were reviewed. The patients were identified from hospital records via the procedure coding system and the radiology department procedures book and were analysed retrospectively. Twenty-nine patients were analysed, looking at indications, clinical outcome, complications and long-term results. RESULTS: Twenty-nine patients underwent 35 embolisation procedures during this period. Fourteen patients with benign diseases underwent 17 embolisation procedures for haematuria or intractable pain. In the haematuria group, selective embolisation was used to treat bleeding post percutaneous nephrolithotomy (PCNL) (n=4), angiomyolipoma (n=2), arteriovenous (AV) malformation (n=l1), renal artery aneurysm (n=1) and renal trauma (n=2). In the renal pain group (n=3), non-selective embolisation was done. Two of these patients had recurrence of pain despite repeat embolisation and subsequently underwent nephrectomy. Fifteen patients with advanced renal malignancy, who were deemed unfit for surgery, underwent 18 embolisation procedures for symptomatic haematuria. Twelve of the 15 patients had successful outcomes with cessation of haematuria. Three patients required repeat embolisation procedures for continuing haematuria with success. There were no major embolisation-related complications. Minor complications were self-limiting and settled with conservative management. CONCLUSION: Renal artery embolisation is effective in managing haematuria in benign and malignant renal conditions where indicated, with minor and easily treatable adverse effects

The outcome of direct current cardioversion (DCC) for the treatment of atrial fibrillation (AF) in a district general hospital in Ireland.

BACKGROUND: Direct current cardioversion (DCC) is a method to control persistent AF, to facilitate a reduction in stroke risk. Although this is a frequently performed procedure, there are no available published data regarding its outcome in an Irish setting. AIMS: To determine the short- and long-term outcome of DCC, factors predicting a successful outcome, and its safety. METHODS: Data relating to each DCC were collected retrospectively from patient notes over a 6.3 year-period, and subsequently entered into a Microsoft Access database before subsequent statistical analysis. RESULTS: Forty-five consecutive unselected patients were identified, in which 59 DCCs were performed. Sinus rhythm (SR) was achieved immediately after DCC in 54/59 (91%) patients. There was a significant positive correlation between patient body weight and the energy level required to achieve SR (p=0.0001). No thromboembolic complications were noted. After a mean follow-up time of 12 +/- 13.7months, 30/45 (67%) had maintained SR. After univariate analysis, a number of important factors predictive of maintenance of SR at follow-up were identified. CONCLUSION: DCC was found to be an effective method for short- and long-term control of AF, without thromboembolic complications, and patients with a favourable long-term outcome after DCC could conceivably be predicted on the basis of a methodical history, careful examination, simple investigations and pharmacological variables.

A murine model for antibody-directed targeting of vascular endothelial cells in solid tumors.

An attractive approach to the therapy of solid tumors would be to target cytotoxic agents or coagulants to the vasculature of the tumor rather than to the tumor cells themselves. This strategy has 3 advantages: (a) it should be applicable to many types of solid tumors because all require a blood supply for survival and growth; (b) the target endothelial cells are directly accessible through the blood and are normal cells, making the outgrowth of resistant mutants unlikely; and (c) there is an in-built amplification mechanism because thousands of tumor cells are reliant on each capillary for nutrients and oxygen. Despite its theoretical attractions, the approach of tumor vascular targeting has not been testable because antibodies that recognize tumor vascular endothelial cell antigens with adequate specificity are currently not available. In this study, we developed a model system in which to investigate the antibody-directed targeting of vascular endothelial cells in solid tumors in mice. A neuroblastoma transfected with the mouse interferon-gamma gene, C1300(Mu gamma), was grown in antibiotic-treated BALB/c nude mice. The interferon-gamma secreted by the tumor induces the expression of major histocompatibility complex Class II antigens on the tumor vascular endothelium. Class II antigens are absent from the vasculature of normal tissues, although they are present on B-lymphocytes, cells of monocyte/macrophage lineage, and some epithelial cells. Anti-Class II antibody administered i.v. strongly stains the tumor vasculature, whereas an antitumor antibody directed against a major histocompatibility complex Class I antigen of the tumor allograft produces classical perivascular tumor cell staining. This model should enable the theoretical superiority of tumor vascular targeting over conventional tumor cell targeting to be tested.

Effect of chemical deglycosylation of ricin A chain on the in vivo fate and cytotoxic activity of an immunotoxin composed of ricin A chain and anti-Thy 1.1 antibody.

The carbohydrate present on ricin A chain causes ricin A chain immunotoxins to be cleared rapidly in animals by the reticuloendothelial system. In an effort to overcome this problem we destroyed the carbohydrate on ricin A chain by treating it with a mixture of sodium metaperiodate and sodium cyanoborohydride and then linked the "deglycosylated" A chain to monoclonal anti-Thy 1.1 antibody. The deglycosylation procedure did not affect the ability of the A chain component of the immunotoxin to inhibit protein synthesis in a cell-free system or the capacity of the immunotoxin to inhibit protein synthesis in Thy-1.1 positive lymphoma cells in vitro. Immunotoxins prepared with deglycosylated A chain were cleared from the bloodstream of mice more slowly than native ricin A chain immunotoxins. The difference in the blood clearance rates of the two immunotoxins could be accounted for by a decreased entrapment of the deglycosylated ricin A chain immunotoxin by the liver. Both immunotoxins broke down in vivo with the appearance of free antibody in the bloodstream. The site of cleavage of the immunotoxin was possibly the liver because immunotoxins taken up by it rapidly became unreactive with antiricin but retained reactivity with anti-mouse immunoglobulin G suggesting that dissociation of the A chain from the antibody had occurred. The immunotoxins taken up by the liver were metabolized further and the acid insoluble radioactive metabolites gradually accumulated in the stomach, thyroid, and salivary gland. The deglycosylated ricin A chain immunotoxin should be a more effective antitumor agent in vivo because it is cleared from the blood more slowly and so has greater opportunity to localize within the tumor target.

Vascular endothelial growth factor as a marker of tumor endothelium.

Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that is a primary stimulant of the vascularization of solid tumors. VEGF production is induced by oncogenic gene mutations in the tumor cells and by hypoxic conditions inside the tumor mass. Hypoxia and the locally increased concentration of VEGF lead to an up-regulation of VEGF receptor expression on tumor endothelial cells. Therefore, in the tumor microenvironment, there is an up-regulation of both VEGF and its receptor, leading to a high concentration of occupied receptor on tumor vascular endothelium. The VEGF:receptor complex presents an attractive target for the specific delivery of drugs or other effectors to tumor endothelium. In the present study, several hybridomas that secrete monoclonal antibodies against the VEGF:receptor (Flk-1) complex or against VEGF itself have been raised. Three of the antibodies (3E7, GV39M, and 11B5) bind with high affinity to the VEGF:Flk-1 complex in ELISA and to tumor endothelium in frozen sections of human tumors, rodent tumors, and human tumor xenografts. 3E7 and GV39M localize selectively to tumor endothelium after i.v. injection into mice bearing human tumor xenografts. Additionally, one antibody (2C3) was raised that blocks the interaction between VEGF and KDR/Flk-1. 2C3 inhibits VEGF-mediated growth of endothelial cells in vitro and localizes strongly to connective tissue in tumors after injection into mice bearing human tumor xenografts. These findings suggest that 3E7, GV39M, and 2C3 are candidates for targeting and imaging the vasculature or connective tissue of tumors.

An assay that predicts the ability of monoclonal antibodies to form potent ricin A chain-containing immunotoxins.

In this report, we describe an assay for screening monoclonal antibodies for their cytotoxic potential as ricin A chain-containing immunotoxins. The assay involves treating cells with dilutions of the test antibody followed by a Fab fragment of a secondary antibody coupled to ricin A chain ("indirect assay"). The cytotoxicity of the indirect assay is compared to that of the direct assay where the monoclonal antibody is coupled to ricin A chain. Indirect and direct assays were carried out using 14 antibodies and a panel of 8 human and mouse cell types. The two assays showed virtually 100% correlation. The indirect assay, therefore, predicts the potency of a given monoclonal antibody to make an effective immunotoxin and should be useful in screening monoclonal antibodies for use as immunotoxins.

Selective inhibition of vascular endothelial growth factor (VEGF) receptor 2 (KDR/Flk-1) activity by a monoclonal anti-VEGF antibody blocks tumor growth in mice.

Vascular endothelial growth factor (VEGF) is a multifunctional angiogenic growth factor that is a primary stimulant of the development and maintenance of a vascular network in embryogenesis and the vascularization of solid tumors. At the present time there are two well-characterized receptors for VEGF that are selectively expressed on endothelium. VEGF receptor 2 [VEGFR2 (KDR/Flk-1)] mediates endothelial cell mitogenesis and permeability increases, whereas the role of VEGF receptor 1 [VEGFR1 (Flt-1)] has not been clearly defined. In the present study, a monoclonal antibody, 2C3, is shown to block the interaction of VEGF with VEGFR2 but not with VEGFR1 through ELISA, receptor binding assays, and receptor activation assays. 2C3 blocks the VEGF-induced vascular permeability increase in guinea pig skin. 2C3 has potent antitumor activity, inhibiting the growth of newly injected and established human tumor xenografts in mice. These findings demonstrate the usefulness of 2C3 in dissecting the pathways that are activated by VEGF in cells that express both VEGFR1 and VEGFR2, as well as highlighting the dominant role of VEGFR2 in mediating VEGF-induced vascular permeability increase and tumor angiogenesis.

Infarction of solid Hodgkin's tumors in mice by antibody-directed targeting of tissue factor to tumor vasculature.

We demonstrated previously that selective thrombosis of the blood vessels of solid tumors in mice can be achieved by targeting the extracellular domain of tissue factor by means of an antibody to an experimentally induced marker on tumor vascular endothelium. In the present study, we extend this finding to a naturally occurring marker of tumor vascular endothelium, vascular cell adhesion molecule-1 (VCAM-1). VCAM-1 is expressed by vascular endothelial cells in Hodgkin's disease and various solid tumors in mice and humans. It is absent from vascular endothelial cells in normal tissues in mice, with the exception of the heart and lungs, where it is present on venules. A monoclonal antibody to murine VCAM-1 was covalently linked to the extracellular domain of human tissue factor to create a "coaguligand." After i.v. administration to severe combined immunodeficient mice bearing human Hodgkin's tumors, the coaguligand localized selectively to VCAM-1-expressing vessels, caused thrombosis of those vessels, and retarded tumor growth. The coaguligand also localized to VCAM-1-expressing vessels in the heart and lungs of the mice but did not induce thrombosis in these sites. An immunohistochemical evaluation of the distribution of a monoclonal anti-phosphatidylserine (PS) antibody in the mice showed that the VCAM-1-expressing vessels in the tumor expressed PS, whereas the VCAM-1-expressing vessels in the heart and lungs lacked PS. The lack of thrombotic effect of the coaguligand on heart and lung vessels may be because PS is needed to provide the procoagulant surface upon which coagulation complexes can assemble. The requirement for coincident expression of the targeted marker and PS on tumor endothelium probably contributes to the selectivity of thrombotic action and the safety of coaguligands.

Antitumor effects of ricin A chain immunotoxins prepared from intact antibodies and Fab' fragments on solid human Hodgkin's disease tumors in mice.

Three monoclonal antibodies which strongly bind to Hodgkin and Reed-Sternberg cells and two corresponding Fab' fragments were linked to deglycosylated ricin A chain (dg A) to evaluate their potential as immunotoxins for the treatment of Hodgkin's disease. Two of the antibodies, Ber-H2 and HRS-3, were shown to bind to the same epitope on the CD30 antigen, whereas the third antibody, IRac, bound to a different antigen. None of the antibodies significantly cross-reacted with normal human tissues as judged by indirect immunofluorescence and immunoperoxidase analyses on frozen sections from 28 normal tissues. All three antibodies formed potent and specific immunotoxins. They inhibited protein synthesis of the L540 Hodgkin's disease cell line in vitro by 50% at concentrations of 1 x 10(-11) M for IRac.dgA, 9 x 10(-11) M for HRS-3.dgA, and 2 x 10(-10) M for Ber-H2.dgA. HRS-3 Fab' and IRac Fab' immunotoxins were 7.8- and 60-fold less cytotoxic, respectively, than their intact counterparts in vitro. In vivo, a single i.v. injection of a dose of Ber-H2.dgA, HRS-3.dgA, or IRac.dgA corresponding to 40% of the LD50 induced lasting complete remissions in 38, 44, and 50%, respectively, of mice with solid s.c. L540 tumors of 60 to 80 mm3 size (0.5-cm diameter). At equivalent dosage (40% of the LD50), the HRS-3 Fab'.dgA and the IRac Fab'.dgA both induced lasting complete remissions in 25% of the mice, although the HRS-3 Fab'.dgA was significantly superior to IRac Fab'.dgA at retarding tumor growth in the remaining animals. The effectiveness of the immunotoxins depended on the size of the tumor at the time of injection, since IRac.dgA treatment induced complete remissions in 100% of mice with small tumors (10 to 20 mm3, approximately 0.3 cm in diameter) but only 13% of mice with larger tumors of 400 to 600 mm3 (approximately 1 cm in diameter). Tumors which regrew after IRac.dgA treatment mainly consisted of antigen-deficient mutants having reduced sensitivity to IRac.dgA but normal sensitivity to HRS-3.dgA. It is concluded that HRS-3.dgA, HRS-3 Fab'.dgA, and IRac.dgA are candidates for the treatment of Hodgkin's disease in humans.