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Biomedical subjects

M Awwad

Publications and source records attributed to M Awwad.

At least 55 records · Page 3Linked to original sources
PubMed1998

Depletion of anti-Gal(alpha)1-3Gal antibody in baboons by specific alpha-Gal immunoaffinity columns.

Ongoing studies at our center on facilitating transplantation of discordant xenogeneic organs are focused on tolerance induction. To abrogate hyperacute rejection, we have used adsorption methods to eliminate natural anti-Gal(alpha)1-3Gal (alphaGal) antibodies from the circulation of baboons. We have analyzed data concerning antibody removal in baboons that were 1) immunologically naive, 2) receiving conventional pharmacologic immunosuppressive therapy (IS), and 3) treated with a conditioning regimen for tolerance induction. We compared the efficiency of removing alphaGal antibody 1) by perfusion of whole blood through an alphaGal affinity column (CP; n=5) with 2) perfusion of plasma (separated from cellular components by apheresis) through an alphaGal column (CPA; n=39). Our studies demonstrate that 1) CP and CPA are equally effective in removing anti-alphaGal antibody, 2) CPA is the method of choice if multiple adsorptions are required, 3) CPA in naive animals transiently affects levels of total IgG and IgM, 4) four CPAs repeated at 2-4 day intervals in association with heavy IS reduce the pool of anti-alphaGal antibody and total Ig, and 5) splenectomy and/or IS delay the return of anti-alphaGal antibody.

Animals↗
PubMed1998

Pharmacologic immunosuppressive therapy and extracorporeal immunoadsorption in the suppression of anti-alphaGal antibody in the baboon.

The aim of this study was to deplete baboons of anti-(alpha)galactosyl (alphaGal] antibody and attempt to maintain depletion by pharmacologic immunosuppressive therapy (PI). In 12 experiments, involving nine baboons, repeated extracorporeal immunoadsorption (EIA) was carried out by plasma perfusion through immunoaffinity columns of synthetic alphaGal trisaccharide type 6. Five of the baboons were immunologically naive and four had undergone various procedures at least 6 months previously. All, however, had recovered lymphohematopoietic function and (with one exception) had levels of anti-alphaGal antibody within the normal range. Eleven protocols included continuous i.v. cyclosporine (to maintain whole blood levels of approximately 1,600 ng/ml). In addition, in ten protocols, the baboon received one or more of the following drugs: cyclophosphamide (1-20 mg/kg/day), mycophenolate mofetil (70-700 mg/ kg/day), brequinar sodium (1-12 mg/kg/day), prednisolone (1 mg/kg/day), melphalan (0.15-0.6 mg/kg/day), methylprednisolone (125 mg/day x3), and antilymphocyte globulin (ATG) (50 mg/kg/day x3). EIA was carried out on 1-9 occasions in each study and was temporarily successful in removing all antibody. When no PI was administered, antibody returned close to pre-EIA levels within 48 hr. Cyclosporine alone delayed the rate of antibody return only slightly. While EIA was continuing on a daily or alternate day schedule, antibody levels (both IgM and IgG) were maintained at 20-45% of pre-EIA levels. Once EIA was discontinued but PI maintained, IgM rose to 40-90% and IgG to 30-60% of pre-EIA levels. In vitro testing demonstrated significant cytotoxicity to pig cells at these antibody levels. We conclude that i) EIA utilizing columns of alphaGal trisaccharide is successful in temporarily depleting baboons of anti-alphaGal antibody, but ii) none of the PI regimens tested suppressed antibody production to levels which would be expected to prevent antibody-mediated rejection of pig xenografts. Additional strategies will therefore be required if xenotransplantation is to become a clinical reality.

ABO Blood-Group System↗
PubMed1994

Modification of monoclonal antibody carbohydrates by oxidation, conjugation, or deoxymannojirimycin does not interfere with antibody effector functions.

Site-specific attachment of metal chelators or cytotoxic agents to the carbohydrate region of monoclonal antibodies results in clinically useful immunoconjugates [Doerr et al. (1991) Ann Surg 214: 118, Wynant et al. (1991) Prostate 18: 229]. Since the capacity of monoclonal antibodies (mAb) to mediate tumor cell lysis via antibody-dependent cellular cytotoxicity (ADCC) or complement-dependent cytotoxicity (CDC) may accentuate the therapeutic effectiveness of immunoconjugates, we determined whether site-specific modification of mAb carbohydrates interfered with these functions. The chemical modifications examined consisted of periodate oxidation and subsequent conjugation to either a peptide linker/chelator (GYK-DTPA) or a cytotoxic drug (doxorubicin adipic dihydrazide). mAb-associated carbohydrates were also modified metabolically by incubating hybridoma cells in the presence of a glucosidase inhibitor deoxymannojirimycin to produce high-mannose antibody. All four forms (unaltered, oxidized, conjugated and high-mannose) of murine mAb OVB-3 mediated tumor cell lysis via CDC. Similarly, equivalent ADCC was observed with native and conjugated forms of mAb OVB-3 and EGFR.1. ADCC was achieved with different murine effector cells such as naive (NS), poly (I*C)- and lipopolysaccharide-stimulated (SS) spleen cells, or Corynebacterium-parvum-elicited peritoneal cells (PEC). All murine effector cell types mediated tumor cell lysis but differed in potency such that PEC > SS > NS. Excellent ADCC activity was also demonstrable by human peripheral blood mononuclear cells with OVB-3-GYK-DTPA and high-mannose OVB-3 mAb. ADCC activity was detectable in vivo: both native and conjugated OVB-3 inhibited growth of OVCAR-3 xenografts in nude mice primed with C. parvum. In conclusion, modification of mAb carbohydrates did not compromise their in vivo or in vitro biological functions. Therefore, combination therapy using immunomodulators to enhance the effector functions of site-specific immunoconjugates could be seriously contemplated.

1-Deoxynojirimycin↗
PubMed1990

Radiosensitive barrier to T-cell-mediated adoptive immunotherapy of established tumors.

This study shows that it is not possible to cause regression of the immunogenic SA-1 sarcoma by adoptive immunotherapy with tumor-sensitized T-cells, unless the tumor-bearing recipient is exposed to a sublethal dose of gamma-irradiation to remove a barrier that prevents adoptive immunity from being expressed. This barrier to adoptive immunotherapy was found to be regenerated between 2 and 4 weeks following irradiation, and its regeneration was associated with general repopulation of host T-cells. However, it was not regenerated in the absence of the thymus, thus showing that it is T-cell dependent. Evidence that it is caused by the presence of CD4+ suppressor T-cells was shown by the finding that it can be removed by depleting mice of CD4+ T-cells with anti-L3T4 monoclonal antibodies, but not by depleting them of CD8+ T-cells with anti-Lyt-2 monoclonal antibodies. Again, the barrier could be restored to irradiated recipients by infusing them with CD4+ T-cells, but not with CD8+ T-cells, from tumor-bearing donors. The barrier to adoptive immunotherapy was found to be tumor induced and to be paradoxically generated in concert with host concomitant immunity.

Animals↗
PubMed1990

Elimination of cycling CD4+ suppressor T cells with an anti-mitotic drug releases non-cycling CD8+ T cells to cause regression of an advanced lymphoma.

This paper describes a model of successful immunotherapy of advanced lymphoma based on the selective elimination of cycling tumour-induced suppressor T cells. It shows that a single injection of the anti-mitotic drug, vinblasting (Vb), results in complete regression of a large L5178Y lymphoma and its metastases, but not if it is growing in an immunocompetent host. Vb-induced, immunologically mediated tumour regression was dependent on the anti-tumour function of CD8+ T cells, because regression was prevented by depleting the host of this subset of T cells 24 hr after Vb was given. Regression was also prevented by infusing the host with Vb-sensitive, CD4+ T cells from a tumour-bearing donor. These and other results are in keeping with the interpretation that Vb-induced regression of the L5178Y lymphoma depends on the ability of the drug to eliminate CD4+ suppressor T cells that are replicating, and to spare non-replicating CD8+ effector cells. It is suggested that at an advanced stage of growth of the L5178Y lymphoma the host possesses an acquired population of antigen-primed CD8+ effector T cells that are unable to become activated in response to abundant tumour antigen because of the dominant influence of CD4+ suppressor cells. Activation of these CD8+ T cells was indicated by the finding that they were rapidly converted from being cyclophosphamide (Cy) resistant to being highly Cy sensitive within 48 hr of giving Vb.

Animals↗
PubMed1989

Elimination of CD4+ suppressor T cells from susceptible BALB/c mice releases CD8+ T lymphocytes to mediate protective immunity against Leishmania.

This study examined the capacity of BALB/c mice that had been depleted of T cell subpopulations to generate a protective immune response to Leishmania major. Thymectomized mice were depleted of either L3T4+ (CD4+) T lymphocytes, Ly2+ (CD8+) T lymphocytes, or both, by treatment with appropriate mAbs. It was found that susceptible mice were rendered resistant to Leishmania by an intravenous infusion of anti-L3T4 mAb. These mice generated an immune response that destroyed the parasite in the primary lesion and in visceral metastatic foci. CD4+ cell-depleted mice also acquired a capacity to mount a sustained delayed-type hypersensitivity (DTH) response to parasite antigens, indicating that DTH, per se, is not a disease-promoting mechanism in the susceptible murine host as has been suggested. Depleting BALB/c mice of CD8+, as well as CD4+ T cells, left them highly susceptible to Leishmania infection, thereby indicating that CD8+ lymphocytes are key protective cells. Our results can be interpreted as showing that the susceptibility of BALB/c mice is due to the generation of CD4+ cells that suppress either the generation or expression of CD8+ T cell-mediated antiLeishmania immunity.

Animals↗
PubMed1989

Cyclophosphamide-induced immunologically mediated regression of a cyclophosphamide-resistant murine tumor: a consequence of eliminating precursor L3T4+ suppressor T-cells.

It was shown that it is possible to use cyclophosphamide (Cy) to cause immunologically mediated regression of the immunogenic, Cy-resistant L5178Y lymphoma in syngeneic and semisyngeneic mice. In order to cause tumor regression it was necessary to give Cy shortly before or shortly after tumor implantation. However, regardless of whether Cy was given before or after tumor implantation, tumor regression did not commence until 10 days of progressive tumor growth, by which time the tumor was 1 cm in diameter. Tumor regression was associated with the presence in the spleen of an increased number of Lyt-2+ T-cells capable of passively transferring immunity to tumor-bearing recipients. This augmented level of immunity was sustained throughout the period of tumor regression. In contrast, a lower level of concomitant immunity generated by control tumor bearers decayed after Day 12 of tumor growth. Because the therapeutic effect of Cy could be inhibited by passive transfer of L3T4+ T-cells from normal donor mice it is apparent that the therapeutic effect of Cy is based on its ability to preferentially destroy L3T4+ suppressor T-cells. These putative precursor suppressor T-cells were regenerated 4 days after being destroyed by Cy. Taken together the results represent a striking example of the negative regulatory influence of suppressor T-cells on the immune response to an immunogenic tumor.

Animals↗
PubMed1988

Immunologically mediated regression of a murine lymphoma after treatment with anti-L3T4 antibody. A consequence of removing L3T4+ suppressor T cells from a host generating predominantly Lyt-2+ T cell-mediated immunity.

This study shows that intravenous injection of 1 mg of anti-L3T4 mAb (GK1.5) into thymectomized mice bearing the syngeneic L5178Y lymphoma results, after a delay of 2-3 d, in complete regression of this tumor and in long-term host survival. A flow cytofluorometric examination of the spleen cells of mAb-treated mice revealed that antibody treatment resulted in the elimination of greater than 98% of L3T4+ T cells, but had no effect on the Lyt-2+ T cells subset. Tumor regression was immunologically mediated, because L5178Y lymphoma cells were shown to be L3T4-, and regression of the tumor failed to occur in mice that had been lethally irradiated before anti-L3T4 mAb was given. Tumor regression was mediated by tumor-sensitized Lyt2+ T cells, as evidenced by the finding that treatment of tumor-bearing mice with anti-Lyt-2 mAb alone, or in combination with anti-L3T4 mAb, resulted in enhancement of tumor growth and a significant decrease in host survival time. Moreover, the spleens of mice whose tumors were undergoing regression in response to anti-L3T4 mAb treatment contained Lyt-2+ T cells capable, on passive transfer, of causing regression of a tumor in recipient mice. These results can be interpreted as showing that removal of tumor-induced L3T4+ suppressor T cells results in the release of Lyt-2+ effector T cells from suppression, and consequently in the generation of enough Lyt-2+ T cell-mediated immunity to cause tumor regression. This can only be achieved, however, if immunity to the tumor is mediated exclusively by Lyt-2+ T cells, as is the case for the L5178Y lymphoma. In the case of the P815 mastocytoma, treatment with anti-L3T4 mAb was without a therapeutic effect, and this was in keeping with the finding that immunity to this tumor is mediated by L3T4+, as well by Lyt-2+ T cells.

Animals↗
PubMed1988

Sublethal, whole-body ionizing irradiation can be tumor promotive or tumor destructive depending on the stage of development of underlying antitumor immunity.

It was shown that sublethal (500 rads), whole-body gamma-irradiation of mice bearing an established i.d. immunogenic tumor can result, after several days delay, in complete tumor regression and long-term survival, but only if radiation is given after the tumor is established and growing progressively. Exposing mice to the same dose of radiation several hours after tumor cells were implanted resulted, in contrast, in enhanced growth of the primary tumor and in earlier death from systemic disease. Irradiation-induced tumor regression failed to occur in mice that were incapable of generating antitumor immunity, because of having been made T cell deficient by thymectomy and irradiation. Again, irradiation-induced tumor regression could be blocked by infusion of spleen cells from donor mice bearing a well-established tumor. These and previously published results support the view that sublethal, whole-body ionizing irradiation causes tumor regression by preferentially destroying radiosensitive suppressor T cells, thereby enabling the host to generate a therapeutic level of concomitant immunity. It is suggested that the preferential destruction of suppressor cells by irradiation depends on the acquisition, during immunologic induction, of radioresistance by antigen-activated effector T cells, and that this is the reason irradiation causes regression only of established tumors. Not all tumors tested were immunogenic enough to undergo regression in response to gamma-irradiation.

Animals↗
PubMed1988

Cyclophosphamide (Cy)-facilitated adoptive immunotherapy of a Cy-resistant tumour. Evidence that Cy permits the expression of adoptive T-cell mediated immunity by removing suppressor T cells rather than by reducing tumour burden.

A cyclophosphamide (Cy)-resistant immunogenic tumour, the L5178Y lymphoma, was used to demonstrate that Cy-treatment of a host bearing this tumour enables passively transferred tumour-sensitized T cells to cause complete tumour regression without any need for Cy to cause a reduction in tumour burden. It was shown that whereas infusion of tumour-sensitized T cells from immune donors had very little effect on growth of the tumour, and whereas treatment with 150 mg/kg of Cy caused appreciable enhancement of tumour growth, combination therapy with Cy plus immune T cells caused complete tumour regression and resulted in long-term survival. Evidence that Cy treatment facilitated the expression of adoptive immunity against the L5178Y lymphoma by eliminating tumour-induced suppressor T cells consisted of the demonstration that tumour regression caused by combination treatment with Cy and immune T cells could be inhibited by infusing the recipient with Cy-sensitive, L3T4+ T cells from tumour-bearing but not from normal donors.

Animals↗
PubMedDate not supplied

Potential of aspirin to inhibit thrombotic microangiopathy in alpha1,3-galactosyltransferase gene-knockout pig hearts after transplantation in baboons.

Hearts from alpha1,3-Galactosyltransferase gene-knockout (GaIT-KO) pigs were transplanted heterotopically into 8 baboons that received an anti-CD154 monoclonal antibody (mAb)-based immunosuppressive regimen and heparin. Three baboons died or were euthanized with beating grafts on 16, 23, and 56 days, respectively, and the remaining 5 grafts functioned for 59-179 days. Hyperacute rejection did not occur, and classical features of acute humoral xenograft or acute cellular rejection were rare. However, thrombotic microangiopathy (TM) developed in all cases; its onset was delayed in 2 baboons that received aspirin. Function of a pig organ in a baboon for a period approaching 6 months has not been reported previously and lends encouragement that the barriers to xenotransplantation will be overcome, but TM requires investigation.

Animals↗