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Rationale: Ivacaftor's clinical effects in the residual function mutations 3849+10kb C →T and D1152H warrant further characterization. Objectives: Evaluate ivacaftor's effect in people with cystic fibrosis aged ≥6 years with 3849+10kb C→T or D1152H residual function mutations; explore the correlation between ivacaftor-induced organoid-based cystic fibrosis transmembrane conductance regulator function measurements and clinical response to ivacaftor. Methods: Participants were randomized (1:1) in this placebo-controlled crossover study; each treatment sequence included two 8-week treatments with an 8-week washout period. The primary endpoint was absolute change in lung clearance index2.5 from baseline through Week 8. Additional endpoints included lung function, patient-reported outcomes, and in vitro intestinal organoid-based measurements of ivacaftor-induced cystic fibrosis transmembrane conductance regulator function. Results: Of 38 participants, 37 completed the study. The primary endpoint was met; the Bayesian posterior probability of improvement in lung clearance index2.5 with ivacaftor vs placebo was >99%. Additional endpoints improved with ivacaftor. Safety findings were consistent with ivacaftor's known safety profile. Dose-dependent swelling was observed in 23/25 viable organoid cultures with ivacaftor treatment. Correlations between ivacaftor-induced organoid swelling and clinical endpoints were negligible to low. Conclusions: In people with cystic fibrosis aged ≥6 years with a 3849+10kb C →T or D1152H mutation, ivacaftor treatment improved clinical endpoints vs placebo; however, there was no correlation between organoid swelling and change in clinical endpoints. The organoid assay may assist in identification of ivacaftor-responsive mutations but in this study did not predict magnitude of clinical benefit for individual people with cystic fibrosis with these two mutations. Clinical trial registered with ClinicalTrials.gov (NCT03068312).

Background: Clinical trials are all based on the assumption that patients are adherent to the study protocol. Many reports indicate that general adherence of patients with CF to their daily routine therapies is poor. However, no data exists on adherence to study drug regimens. Methods: All clinical trials carried out at the Hadassah CF Center from 2008 to 2013 were reviewed. Actual adherence as determined by counted drugs was analyzed according to drug administration mode, study lengths and number of study visits. A subset of patients answered a two-part questionnaire covering study specific and general treatment specific issues. Results: Eight studies including 118 patients, with patient numbers varying between 4 and 32 per trial were analyzed. For 7/8 studies mean adherence was between 78% to 100%. Comparison with administration mode showed that adherence decreased substantially if the drugs were not provided as "ready to be used" (63%). Study length influenced adherence, the longer the study the poorer the adherence (82% trial beginning, 44% post 36 months [two combined studies with identical drug]). A substantial decrease was noted over Holiday periods and during the summer vacation months. No correlation was found between number of study visits and adherence to study drug. Conclusion: Adherence to study drug is generally higher than that for regular treatment. Study length, mode of administration, and timing according to Holidays and vacations adversely affect adherence. (C) 2015 Wiley-Blackwell.

BackgroundAlthough cough is a commonly reported symptom, objective quantitation of cough during normal activity has not been performed in patients with CF.MethodsAn ambulatory device was used to characterize cough over 24hours. Pulmonary function and subject-reported coughing were also assessed.ResultsPatients included 19 clinically stable adults with CF (males:females=10:9; median age [range]=26 [19-57] years; median %-predicted FEV1 [range]=65 [44-106]%). Median [range] cough rate was 27 [13-66] coughs/hour, with values while awake of 41 [20-102] and while asleep of 2 [0.1-7] (p

Objectives We examined whether mitral regurgitation (MR) augments post-myocardial infarction (MI) remodeling. Background MR doubles mortality after MI, but its additive contribution to left ventricular (LV) remodeling is debated and has not been addressed in a controlled fashion. Methods Apical MIs were created in 12 sheep, and 6 had an LV-to-left atrial shunt implanted, consistently producing regurgitant fractions of ∼30%. The groups were compared at baseline, 1, and 3 months. Results Left ventricular end-systolic volume progressively increased by 190% with MR versus 90% without MR (p Conclusions In this controlled model, moderate MR worsens post-MI remodeling, with reduced contractility. Pro-hypertrophic pathways are initially upregulated but subsequently fall below infarct-only levels and baseline; with sustained caspase 3 elevation, transformation to a failure phenotype occurs. Extracellular matrix turnover increases in MR animals. Therefore, MR can precipitate an earlier onset of dilated heart failure.

Coronary artery disease (CAD) is a highly prevalent disease with significant morbidity and mortality. Periodontal disease has been suggested to influence this disease and has been associated with CAD in some epidemiologic studies. However, this relation is still controversial. This study aimed to determine the relationship between periodontal disease measures and CAD and acute coronary syndromes (ACSs).Two hundred one patients presenting with stable angina or ACS referred for coronary angiography underwent a periodontal assessment including evaluation of periodontal pathogens. Severity of CAD was determined by the number of obstructed coronary arteries.Patients with severe CAD defined by multiple vessel disease had significantly more periodontal destruction than those with mild CAD, as shown by mean clinical attachment level, a measure of chronic periodontal disease (CAL; 5.43 +/- 1.8 versus 4.85 +/- 1.6; P = 0.02), percentage of teeth with CALor=5 mm (82.1 +/- 23.4 versus 70.4 +/- 26.9; P = 0.002), and number of missing teeth (8.75 +/- 6.6 versus 6.76 +/- 6.6; P = 0.03). Logistic regression analysis showed that percentage of teeth with CALor=5 mm was significantly associated with CAD severity. Patients with ACS had significantly higher plaque scores, gingival index, and Porphyromonas gingivalis counts than stable patients. Logistic regression analysis showed that either plaque score or percentage of P. gingivalis was significantly associated with ACS.Periodontal destruction measures are significantly correlated with CAD severity, whereas periodontal infectious measures are significantly associated with clinical cardiac status.

Clinical trials are all based on the assumption that patients are adherent to the study protocol. Many reports indicate that general adherence of patients with CF to their daily routine therapies is poor. However, no data exists on adherence to study drug regimens.All clinical trials carried out at the Hadassah CF Center from 2008 to 2013 were reviewed. Actual adherence as determined by counted drugs was analyzed according to drug administration mode, study lengths and number of study visits. A subset of patients answered a two-part questionnaire covering study specific and general treatment specific issues.Eight studies including 118 patients, with patient numbers varying between 4 and 32 per trial were analyzed. For 7/8 studies mean adherence was between 78% to 100%. Comparison with administration mode showed that adherence decreased substantially if the drugs were not provided as "ready to be used" (63%). Study length influenced adherence, the longer the study the poorer the adherence (82% trial beginning, 44% post 36 months [two combined studies with identical drug]). A substantial decrease was noted over Holiday periods and during the summer vacation months. No correlation was found between number of study visits and adherence to study drug.Adherence to study drug is generally higher than that for regular treatment. Study length, mode of administration, and timing according to Holidays and vacations adversely affect adherence.

PURPOSE To investigate if a platelet inhibitor (aurintricarboxylic acid [ATA]) and a heparin-mimicking antagonist (RG-13577) of basic fibroblast growth factor 2 (bFGF2) could be combined as a stable compound and attached to conventional bare metal stents to hinder thrombus formation and inflammatory reactions of stenting. METHODS Fifteen domestic pigs were stented with RG-13577/ATA-coated (n=6), ATA-coated (n=12), and bare metal stents (n=12) in the left anterior descending (LAD) and left circumflex (LCX) coronary arteries. All surviving pigs were evaluated with contrast angiography and intravascular ultrasonography (IVUS) after 4 weeks. Histological analysis of the stented arteries was performed after hematoxylin-eosin staining. Tissue factor (TF) staining and scanning electron microscopy (SEM) were performed in animals with acute stent thrombosis. RESULTS Five of the 6 animals receiving an RG-13577/ATA-coated stent experienced acute stent thrombosis, while no adverse events occurred in the animals of the other 2 groups. Follow-up angiography did not show significant in-stent stenosis in either bare or ATA-coated stents. However, histomorphometry revealed larger neointimal area (3.54+/-0.69 mm2 versus 1.82+/-0.27 mm2, p

Successful engraftment of hematopoietic stem cells requires a supportive hematopoietic stromal microenvironment (HSM). Defects in the HSM associated with aplastic anemia, myelofibrosis, or caused by intensive ionizing radiation and chemotherapy generally result in failure of bone marrow (BM) engraftment. Transplantation of donor BM within donor HSM may therefore provide optimal conditions for allogeneic BM transplantation. We have transplanted donor hematopoietic cells together with their own HSM to improve acceptance of allogeneic or xenogeneic BM. The nonmyeloablative treatment used induced tolerance to murine allografts and provided conditions for the lifelong acceptance of allogeneic HSM. Allogeneic BM transplanted within it’s own HSM under the kidney capsule caused less graft-versus-host disease than BM transplanted i.v. Tolerance in mice to xenogeneic (rat) HSM was less complete. Ectopic ossicles were small and contained fewer hematopoietic cells. However, simultaneous transplantation of rat BM and HSM to preconditioned mice improved engraftment of rat BM compared with transplantation of BM alone. Donor hematopoietic cells survived longer on their own HSM than on HSM of recipients. Over the last 30 years allogeneic bone marrow transplantation (BMT*) has become an accepted curative therapy for a variety of malignant hematopoietic diseases, immunodeficiencies, and diseases caused by genetically abnormal hematopoietic progenitor cells (1). Stem cells infused in the course of BMT seed in the host hematopoietic stromal microenvironment (HSM) and a direct interaction of the latter with the stem cells is probably mandatory for differentiation and maturation of hematopoietic progenitor cells (2, 3). In diseases associated with primary or secondary defects of HSM, namely certain forms of aplastic anemia or myelofibrosis, and disorders resulting from intensive ionizing radiation or chemotherapy, graft failure, distinguishable from graft rejection, can occur (4). Long-term hematopoietic engraftment of transplanted bone marrow (BM) may therefore ultimately require transplantation of donor HSM.

In the course of allogeneic bone marrow transplantation (BMT), avoiding graft-versus-host disease (GVHD) while retaining the antileukemic effects of the T cells remains a major challenge. T-cell depletion (TCD) reduces the incidence of GVHD but increases the relapse rate after allogeneic BMT. We attempted to develop a regimen that would retain or increase the graft-versus-leukemia effect induced by donor T cells while preventing GVHD. Immunosuppressed mice were given immunocompetent donor cells, i.e., fresh lymphocytes or lymphokine-activated killer (LAK) cells differing from the host in major (MHC) or minor (MiHC) histocompatibility antigens. Engraftment of donor cells was documented by polymerase chain reaction analysis. Administration of MHC- and MiHC-incompatible allogeneic LAK cells, especially in conjunction with recombinant interleukin-2 (rIL-2), increased disease manifestations and mortality associated with GVHD. On the other hand, irradiated LAK cells or TCD-LAK cells prevented GVHD in both mice models studied. Phenotypic analysis of LAK cells demonstrated that CD8 + -equivalent (Lyt-2) T cells are of significance in aggravation of GVHD. The in vitro cytotoxic capacity of LAK cells against MHC-nonrestricted target cells was not reduced by either irradiation or TCD. These results provide the background for designing improved protocols for immunotherapy of residual disease after BMT. In addition, the data imply that antitumor effects may be retained by irradiated rIL-2–activated allogeneic cells without causing GVHD. Whereas unmodified allogeneic LAK cells can induce more effective graft-versus-leukemia effects at the cost of GVHD, irradiated allogeneic donor LAK cells might play some role in eradication of minimal residual disease following autologous or allogeneic BMT without causing GVHD.

Immunocompetent donor-derived T lymphocytes play a crucial role in the elimination of residual leukemic cells post allogeneic bone marrow transplantation. Because this graft versus leukemia (GVL) effects is absent after autologous stem cell transplantation (ASCT), a high rate of relapse ensues. We introduced cell-mediated immunotherapy at the stage of minimal residual disease in lymphoma patients to help effect a GVL-like reaction by adoptive transfer of immunocompetent human leukocyte antigen-matched donor peripheral blood lymphocytes (PBL). Thirteen consecutive patients with high-risk lymphoma were treated with allogeneic cell therapy (AlloCT) after having undergone ASCT. In the absence of graft-versus-host disease, cell therapy-induced graft-versus-lymphoma reaction was amplified by human recombinant interleukin 2 (rIL-2) during 3 days to activate donor PBL in vivo, followed by infusion of in vitro rIL-2 activated donor lymphocytes combined with 3-day rIL-2 therapy. Nine of the patients underwent the treatment protocol well. In the four other patients, in whom the AlloCT resulted in marrow aplasia due to elimination of host hematopoietic cells, treatment with donor marrow cell infusion without further conditioning was performed. Adoptive cell therapy in the form of AlloCT may turn out to be an effective therapeutic modality for the treatment of resistant residual disease in lymphoma patients.

PURPOSE Donor lymphocyte infusion (DLI) was used to reverse relapse after allogeneic bone marrow transplantation (BMT) in a patient with beta-thalassemia major. PATIENTS AND METHODS The patient with unstable mixed chimerism after BMT was treated with graded increments of donor lymphocytes (10(5) T cells/kg to 5 x 10(7) T cells/kg) to displace residual hematopoietic host cells. RESULTS DLI resulted in complete donor-derived reconstitution of the hematopoietic compartment. The patient developed mild graft-versus-host disease (GVHD) that could be controlled by steroid treatment. CONCLUSIONS This case report shows that DLI can effectively eradicate host stem cells in mixed chimeras after BMT in nonmalignant hematopoietic diseases.

Immunity to murine B-cell leukemia/lymphoma (BCL 1 ) induced by multiple injections with irradiated tumor cells, prevented leukemia development in primary and adoptive transfer recipients despite long-lasting persistence of residual tumor cells. Detection of dormant BCL, cells was carried out by PCR analysis using the V H -rearranged DNA sequence as a BCL 1 clonal marker. Dormant tumor cells were detected >250 days following immunity induction in 40% of spleens from healthy immune mice having no detectable symptoms of disease. Tumor dormancy was not abrogated by adoptive transfer of BCL 1 -containing splenocytes into syngeneic recipients, indicating that cell-mediated anti-tumor immunity contributes to maintenance of the tumor dormant state and prevents renewed tumor-cell growth. Splenocytes but not sera from immune mice conferred specific radiosensitive protection from a lethal dose of BCL, cells included in cell mixtures transferred to secondary recipients. A therapeutic effect of transferred immune splenocytes was shown in BCL 1 -bearing mice, which remained disease-free for >200 days after inoculation ; nevertheless, dormant BCL 1 cells were detected by PCR analysis in some of the surviving mice. Our results suggest that an efficient tumor-cell vaccine can lead to induction of tumor dormancy that can be maintained by a cell-derived mechanism for a long period of time.

Smokers hospitalized with acute coronary syndrome (ACS) are at high risk for subsequent ischemic events. Nevertheless, over two-thirds of patients continue to smoke after an acute myocardial infarction. Bupropion hydrochloride has proven efficacy as a smoking cessation aid, but data regarding its safety and efficacy in ACS patients are limited.In a double-blind, randomized controlled trial, we compared the safety and efficacy of 8 weeks of treatment with bupropion slow-release (SR) or placebo for smokers hospitalized with ACS as an adjunct to nurse-led hospital- and telephone-based support. Primary efficacy outcome was smoking abstinence at 1 year. Primary safety outcome was clinical events at 1 year.A total of 151 patients were enrolled; all but 2 completed follow-up. Abstinence rates at 3 months were 45% and 44% in the bupropion SR and placebo groups, respectively (P = .99); 37% vs 42% (P = .61) at 6 months; and 31% vs 33% (P = .86) at 1 year. On multivariate analysis, an invasive procedure performed during index hospitalization was an independent predictor for smoking abstinence at 1 year (odds ratio [OR], 4.2; 95% confidence interval [CI], 1.22-14.19). Presence of adverse effects attributed to treatment was a negative predictor for smoking cessation (OR, 0.23; 95% CI, 0.07-0.78). Treatment with bupropion SR was not associated with an increase in clinical events or change in blood pressure or body mass index, but dizziness was more common compared with placebo (14% vs 1.4%; P = .005).In hospitalized patients with ACS who received continuous, intensive nurse counseling about smoking cessation, bupropion did not increase the rates of smoking abstinence.

Small proportions of leukemic cells escaping chemo-radiotherapy and/or dormant leukemic cells present in undetectable amounts in patients post therapy, i.e. minimal residual disease (MRD), are a source for relapse. The study of MRD and its treatment in a murine model for human B-cell leukemia/lymphoma, (BCL1), should lead to an improved understanding of the human disease. The standard assay for MRD in experimental mice is the adoptive transfer of spleen cells from experimental animals into naive secondary syngeneic recipients. We describe here the detection of MRD in BCL1-carrying BALB/c mice using the polymerase chain reaction (PCR). A BCL1 specific sequence from the rearranged VH-region was amplified yielding a 456 bp long fragment. PCR products hybridized to the cloned BCL1 sequence allowed the detection of a single BCL1 cell. This assay, therefore, reveals the presence of very small numbers of leukemic cells without sacrificing experimental animals.

Atherosclerosis is a chronic inflammatory process resulting in coronary artery disease.To determine the relationship between inflammatory markers and the angiographic severity of CAD.We measured inflammatory markers in consecutive patients undergoing coronary angiography. This included C-reactive protein, fibrinogen, serum cytokines (interleukin-1 beta, IL-1 receptor antagonist, IL-6, IL-8, IL-10) and tumor necrosis factor-alpha), all measured by high sensitivity enzyme-linked immunoabsorbent assay.There was a significant correlation between TNFalpha and the severity of CAD as assessed by the number of obstructed coronary vessels and the Gensini severity score, which is based on the proximity and severity of the lesions. Patients had more coronary vessel disease (70% stenosis) with increasing tertiles of serum TNFalpha; the mean number of vessels affected was 1.15, 1.33, and 2.00 respectively (P0.001). IL-6 correlated with the Gensini severity score and coronary vessel disease (70% stenosis). A weaker correlation was present with IL-1 receptor antagonist. A significant correlation was not found with the other inflammatory markers. After adjustment for major risk factors, multivariate analyses showed that significant independent predictors of CAD vessel disease were TNFalpha (P0.05) and combined levels of TNFalpha and IL-6 (P0.05). IL-6 levels were independently predictive of Gensini coronary score (P0.05).TNFalpha and IL-6 are significant predictors of the severity of coronary artery disease. This association is likely an indicator of the chronic inflammatory burden and an important marker of increased atherosclerosis risk.

Eimeria maxima gametocytes contain two major antigens with molecular masses of 56 and 82 kilodaltons (kDa) which are recognized by convalescent sera from immune chickens. Preparations enriched in these two antigens were used to immunize mice, and several monoclonal antibodies which specifically reacted with the 56-kDa antigen were produced. One of these monoclonal antibodies of the immunoglobulin M subclass, along with immune chicken sera raised against affinity-purified 56- and 82-kDa antigens, was used to passively immunize chicks. On the basis of the parameter of total oocyst output, it was found that these antibodies provided partial protection (40 to 50% inhibition) against E. maxima challenge infections.

Cannabidiol (CBD) is a major, nonpsychoactive Cannabis constituent with anti-inflammatory activity mediated by enhancing adenosine signaling. Inasmuch as adenosine receptors are promising pharmaceutical targets for ischemic heart diseases, we tested the effect of CBD on ischemic rat hearts. For the in vivo studies, the left anterior descending coronary artery was transiently ligated for 30 min, and the rats were treated for 7 days with CBD (5 mg/kg ip) or vehicle. Cardiac function was studied by echocardiography. Infarcts were examined morphometrically and histologically. For ex vivo evaluation, CBD was administered 24 and 1 h before the animals were killed, and hearts were harvested for physiological measurements. In vivo studies showed preservation of shortening fraction in CBD-treated animals: from 48 ± 8 to 39 ± 8% and from 44 ± 5 to 32 ± 9% in CBD-treated and control rats, respectively ( n = 14, P < 0.05). Infarct size was reduced by 66% in CBD-treated animals, despite nearly identical areas at risk (9.6 ± 3.9 and 28.2 ± 7.0% in CBD and controls, respectively, P < 0.001) and granulation tissue proportion as assessed qualitatively. Infarcts in CBD-treated animals were associated with reduced myocardial inflammation and reduced IL-6 levels (254 ± 22 and 2,812 ± 500 pg/ml in CBD and control rats, respectively, P < 0.01). In isolated hearts, no significant difference in infarct size, left ventricular developed pressures during ischemia and reperfusion, or coronary flow could be detected between CBD-treated and control hearts. Our study shows that CBD induces a substantial in vivo cardioprotective effect from ischemia that is not observed ex vivo. Inasmuch as CBD has previously been administered to humans without causing side effects, it may represent a promising novel treatment for myocardial ischemia.

We examined whether mitral regurgitation (MR) augments post-myocardial infarction (MI) remodeling.MR doubles mortality after MI, but its additive contribution to left ventricular (LV) remodeling is debated and has not been addressed in a controlled fashion.Apical MIs were created in 12 sheep, and 6 had an LV-to-left atrial shunt implanted, consistently producing regurgitant fractions of approximately 30%. The groups were compared at baseline, 1, and 3 months.Left ventricular end-systolic volume progressively increased by 190% with MR versus 90% without MR (p0.02). Pre-load-recruitable stroke work declined by 82 +/- 13% versus 25 +/- 16% (p0.01) with MR, with decreased remote-zone sarcoplasmic reticulum Ca(2+)-ATPase levels (0.56 +/- 0.03 vs. 0.76 +/- 0.02, p0.001), and decreased isolated myocyte contractility. In remote zones, pro-hypertrophic Akt and gp130 were upregulated in both groups at 1 month, but significantly lower and below baseline in the MR group at 3 months. Pro-apoptotic caspase 3 remained high in both groups. Matrix metalloproteinase (MMP)-13 and membrane-type MMP-1 were increased in remote zones of MR versus infarct-only animals at 1 month, then fell below baseline. The MMP tissue inhibitors rose from baseline to 3 months in all animals, rising higher in the MI + MR-group border zone.In this controlled model, moderate MR worsens post-MI remodeling, with reduced contractility. Pro-hypertrophic pathways are initially upregulated but subsequently fall below infarct-only levels and baseline; with sustained caspase 3 elevation, transformation to a failure phenotype occurs. Extracellular matrix turnover increases in MR animals. Therefore, MR can precipitate an earlier onset of dilated heart failure.

BACKGROUND We evaluated the usefulness of intravascular ultrasound (IVUS) in the non-uniform distribution of in-stent neointimal hyperplasia, comparing macroscopic measurements with IVUS-guided histomorphometry. METHODS Coronary stenting was performed in 45 left coronaries of 39 pigs, using 18 Tenax (Biotronik Gmbh and Co., Berlin, Germany), 11 bare Genius (Eurocor, Bonn, Germany), 10 polymer-coated Genius (Eurocor) and six Biodivysio Matrix LO (Biodivysio Ltd, Farnham, Surrey, UK) stents. After 4 weeks, coronary angiography and IVUS with automatic pullback were performed. IVUS images were analysed using three-dimensional analysis (EchoPlaque 2; INDEC Systems Inc., Mountain View, California, USA). The stented segments were formalin fixed, embedded in Technovit 9100 and cut to 4-8 microm thick slides. The most diseased in-stent segment was 4.49 +/- 4.54 mm away from the distal stent edge assessed by IVUS. Sections of these segments were stained for histomorphometry. RESULTS A significant correlation was found between IVUS-guided histomorphometry and three-dimensional IVUS measurements of maximal intimal thickness (r = 0.6985, P < 0.005) and area (r = 0.7736, P < 0.001). Macroscopic measurements resulted in comparable maximal intimal thickness (0.83 +/- 0.43 mm compared with 0.81 +/- 0.46 mm) and area (4.44 +/- 1.73 mm2 compared with 3.45 +/- 1.55 mm2) by IVUS and histomorphometry, respectively. Although stent length, diameter, nominal inflation pressure and time and injury score did not differ between the stents, bare Genius stents resulted in significant smaller neointimal volume compared to Tenax, polymer-coated Genius and Biodivysio stents: 24.46 +/- 4.98 mm3 compared with 59.18 +/- 26.41, 60.46 +/- 10.03 and 61.41 +/- 16.27 mm3, respectively (P < 0.05). CONCLUSION The significant correlation between IVUS-guided histomorphometry and IVUS measurements confirms the usefulness of IVUS in evaluation of experimental in-stent restenosis. Implantation of bare Genius stents resulted in significant lower neointimal hyperplasia compared to Tenax, polymer-coated Genius or phosphorylcholine-coated Biodivysio stents.

Multiple injections of intact irradiated BCL1 cells, a murine B-cell leukemia/lymphoma can trigger a dose-dependent anti-tumor immune response in naive syngeneic mice. The ability to induce anti-BCL1 immunity and the effect of various cell-modifications on BCL1 tumorigenicity and immunogenicity was evaluated. Newcastle disease virus (NDV) infection or transfer of cytokine genes by both retroviral and Adeno 5 vectors affect neither tumorigenicity nor immunogenicity of BCL1 cells given as a non-immunogenic cell-dose. New ways will have to be developed to elicit a reliable and reproducible anti-tumor effect in spontaneously arising and non-immunogenic hematological malignancies.

We have attempted to induce immune-mediated graft-vs.-tumor (GVT) effects against solid tumors, using a murine model of mammary adenocarcinoma derived from BALB/c(H-2d) mice. A cell line (4TI) isolated from this tumor model was highly tumorigenic in syngeneic (BALB/c) or haplo-identical (BALB/c × C57B1/6)F1 mice (F1), was only partially tumorigenic in an H-2d congenic strain of mice (DBA/2) and was non-tumorigenic in a major histocompatible (MHC)-unrelated (H-2b) strain of mice (C57B1/6). 4T1 cells express class I MHC antigens and adhesion molecules but do not express MHC class II antigens or B7-I co-stimulatory molecules. Female BALB/c (H-2d) or F1 (H-2d/b) mice were reconstituted with male bone marrow (BM) cells derived from minor histocompatible (MiHC)-mismatched DBA (H-2d) donors or with MHC-mismatched C57B1/6 (H-2b) BM cells, respectively, 24 hr following lethal total body irradiation. Recipient mice carrying MiHC- or MHC-mismatched donor cells were inoculated with 4T1 cells 2–3 months following BM reconstitution. Chimeras reconstituted with allogeneic donor cells that were MiHC- or MHC-incompatible with tumor cells were able to down-regulate the development of the primary tumour expressing host-type MHC alloantigens. Tumor size in BM chimeras across MiHC or MHC antigens was significantly smaller than tumor size observed in normal BALB/c or F1 controls. The GVT effect might be of help in improving immunotherapy for solid tumors in humans. Int. J. Cancer, 71:59–63, 1997. © 1997 Wiley-Liss, Inc.

Background: Simian virus-40 (SV40) is an efficient vector for gene therapy; however, it has not yet been used in the context of myocardial gene delivery. In contrast to adenovirus, which has generally been the vector of choice for many heart trials, SV40 does not elicit an immune response allowing repeated administrations, if necessary, to prolong the effect of the delivered gene. Furthermore, SV40 vectors are capable of infecting dividing or non-dividing cells. Our unique vector, prepared in vitro from recombinant SV40 capsid proteins and plasmid DNA, has no viral sequences, providing unique critical safety advantages. Therapeutic DNA of considerable size can be readily packaged into these vectors.

A 2.5-year-old girl with neurogenic Gaucher's disease was transplanted with donor bone marrow from her HLA-compatible 12-year-old brother whose marrow was harvested 30 min post-mortem, after he suffered a severe head and neck injury. The marrow was stored in liquid nitrogen for 30 days prior to infusion. The post-transplantation period was uneventful with good engraftment and no signs of graft-versus-host disease. Currently, 6 months post-allogeneic bone marrow transplantation (alloBMT), analysis of both bone marrow and blood samples by PCR documented only cells of donor origin. This case demonstrates the feasibility of cadaveric marrow as a source of donor cells. To our knowledge, this patient is the only survivor of alloBMT from a cadaveric donor.

Introduction Cardiac dysfunction is among the serious side effects of therapy with recombinant humanized anti-erbB2 monoclonal antibody. The antibody blocks ErbB-2, a receptor tyrosine kinase and co-receptor for other members of the ErbB and epidermal growth factor families, which is over-expressed on the surface of many malignant cells. ErbB-2 and its ligands neuregulin and ErbB-3/ErbB-4 are involved in survival and growth of cardiomyocytes in both postnatal and adult hearts, and therefore the drug may interrupt the correct functioning of the ErbB-2 pathway. Methods The effect of the rat-anti-erbB2 monoclonal antibody B-10 was studied in spontaneously beating primary myocyte cultures from rat neonatal hearts. Gene expression was determined by RT-PCR (reverse transcription polymerase chain reaction) and by rat stress-specific microarray analysis, protein levels by Western blot, cell contractility by video motion analysis, calcium transients by the FURA fluorescent method, and apoptosis using the TUNEL (terminal uridine nick-end labelling) assay. Results B-10 treatment induces significant changes in expression of 24 out of 207 stress genes analyzed using the microarray technique. Protein levels of ErbB-2, ErbB-3, ErbB-4 and neuregulin decreased after 1 day. However, both transcription and protein levels of ErbB-4 and gp130 increased several fold. Calreticulin and calsequestrin were overexpressed after three days, inducing a decrease in calcium transients, thereby influencing cell contractility. Apoptosis was induced in 20% cells after 24 hours. Conclusion Blocking ErbB-2 in cultured rat cardiomyocytes leads to changes that may influence the cell cycle and affects genes involved in heart functions. B-10 inhibits pro-survival pathways and reduces cellular contractility. Thus, it is conceivable that this process may impair the stress response of the heart.

The antigenicity of Eimeria maxima gametocyte proteins during the course of an infection and when injected into mice and rabbits was demonstrated using the Western blotting technique. Serum taken from chickens at various times postinfection reacted to a few gametocyte proteins, with the strongest reactivity seen with serum taken 14-days postinfection. Two major antigens having molecular weights of 56,000 and 82,000 were consistently detected by these sera. Using immune rabbit or mouse sera to whole gametocyte detergent extracts, the 56,000 and 82,000 molecular weight proteins were again the immunodominant antigens, despite their representing only a small proportion of the extract which was used to immunize the animals. These results, together with those obtained by Rose ((1971)) using recovered chicken serum to passively immunize chickens, indicate that these two gametocyte antigens may play a role in protective immunity to E. maxima.

A restriction endonuclease, BstPI, was purified from a strain of B. stearothermophilus, and its cleavage specificity was determined. The enzyme cleaves at palindromic sites of the general structure: (Formula: see text) where N.N' can be any base pair. It produces phosphorylated 5'-termini which are single stranded over a length of 5 nucleotides. Ends generated by cleavage with BstPI can be rejoined by DNA ligase.

Eimeria maxima gametocytes were isolated from infected chicken intestinal tissue by treatment with hyaluronidase and subsequent filtration through polymon filters. The isolated gametocytes were analyzed by microscopical and biochemical methods and shown to be highly enriched. The antigenicity of the gametocytes was analyzed in mice, rabbits, and chickens by ELISA and indirect immunofluorescence. Contrary to published results, we have found gametocytes to be highly immunogenic in all animals tested.

RNA was extracted from isolated Eimeria maxima gametocytes and translated in a rabbit reticulocyte cell-free protein synthesis system. The major cell-free translation products from E. maxima gametocyte RNA ranged from 225 to 50 kDa, distinct and different from uninfected chicken intestine cell-free translation products. Rabbit antiserum to E. maxima gametocytes as well as recovered chicken sera specifically precipitated some of the major gametocyte cell-free products. A time course of infected intestine RNA indicated that these cell-free synthesized gametocyte antigens appear at 130 to 138 hr postinfection.