Improvement by affinity chromatography on antiidiotypic monoclonal antibodies (MAbs) of immunoreactivity and in vivo targeting of radiolabelled anti-HMW-MAA MAb TP61.5 in nude mice bearing human melanoma lesions

The human high-molecular-weight melanoma-associated antigen (HMW-MAA) represents a useful marker for immunoscintigraphy in patients with melanoma. Since injection of a radiolabelled anti-HMW-MAA monoclonal antibody (MAb) visualizes only about 60% of melanoma lesions, approaches are being developed to increase the sensitivity of immunoscintigraphy. One of them aims at improving the immunoreactivity of radiolabelled anti-HMW-MAA MAbs, since this approach may improve the targeting of radiolabelled MAbs to melanoma lesions. We have previously shown that affinity chromatography on insolubilized anti-idiotypic MAbs is a useful method for purifying immunoreactive anti-HMW-MAA MAb TP61.5 from 125I-labelled MAb preparations and that not all the anti-idiotypic MAbs are useful for this purpose. Since the increasing number of available anti-idiotypic MAbs is likely to facilitate the application of this procedure in many antigenic systems, we have now tested criteria to select anti-idiotypic MAbs suitable for the purification procedure. Furthermore, we have investigated the effect of the increase in immunoreactivity of 125I-MAb TP61.5 on its in vivo targeting to human melanoma lesions transplanted into nude mice. Among the 3 anti-idiotypic MAbs tested, the most effective in purifying immunoreactive MAb TP61.5 molecules following radiolabelling is MAb TK7-110, with which 125I-MAb TP61.5 displays an immunoreactivity similar to that displayed with melanoma cells. This parameter may represent a useful criterion to identify anti-idiotypic MAbs suitable for the purification procedure, if the present results are confirmed with a large number of anti-idiotypic MAbs in different antigenic systems. We have also shown that an incubation time for up to 4 hr of 125I-MAb TP61.5 with insolubilized MAb TK7-110 is the most effective in increasing immunoreactivity and in recovering immunoreactive MAb applied to the affinity matrix. The increase in the immunoreactive fraction of 125I-MAb TP61.5 significantly increases its specific localization in human melanoma lesions transplanted into nude mice. These results suggest that purification of radiolabelled immunoreactive anti-HMW-MAA MAb TP61.5 by affinity chromatography using anti-idiotypic MAb TK7-110 represents a useful approach to increasing the sensitivity of immunoscintigraphy in patients with melanoma.     

Use of new heterobifunctional reagent for radiometallic labeling of antibodies

A new heterobifunctional reagent, 2,6-dioxo-N-(carboxymethyl)morpholine (DCM), was synthesized, reacted with a monoclonal antibody (MAb) to human high-molecular-weight, melanoma-associated antigen, and then chelated with 111In. The same MAb also was chelated with 111In with the use of the bicyclic anhydride of pentetic acid (BADTPA), a homobifunctional reagent, for comparative studies. The labeling efficiencies were similar: an average of 79% for 111In-iminodiacetic acid-MAb and an average of 75% for 111In-DTPA-MAb. However, the loss of immunoreactivity was 32.6% and 58.7% for both radiolabeled antibodies, respectively. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrated a high-molecular-weight polymer band only for the 111In-DTPA-MAb preparation. The results indicate that DCM likely causes less denaturation of antibody by eliminating the cross-linking reaction that is known to occur with homobifunctional chelating reagents.     

Permanent rat cardiac allograft survival induced by ultraviolet B-irradiated donor lymphocytes and peritransplant cyclosporine

This study examines the effect of pretreatment with 10(8) ultraviolet B-irradiated donor leukocytes (UV-DL) with or without peritransplant cyclosporine (CyA) treatment (20 mg/kg on days 0, +1, and +2 relative to transplantation) on rat cardiac allograft survival across major histocompatibility loci. A single UV-DL pretreatment on day -3 or -7 (before transplantation) significantly prolonged survival of heart allografts from Wistar-Furth rats (W/F) in Lewis recipients from 6.8 +/- 0.8 days to 18.4 +/- 2.1 and 17.6 +/- 1.5 days (p less than 0.001), respectively. Multiple UV-DL infusions on days -14 and -7 increased the mean survival time to 20.0 +/- 0.9 days (p less than 0.001). Similarly, UV-DL infusion on day -3 or -7 significantly prolonged the mean survival time of heart allografts from ACI rats in Lewis rats. A single or multiple UV-DL infusions combined with peritransplant CyA led specifically to permanent W/F cardiac allograft survival (more than 200 days) in all recipients. Similarly, UV-DL infusion combined with peritransplant CyA led to indefinite survival of ACI cardiac allografts in two thirds of Lewis recipients. Adoptive transfer of splenocytes from long-term recipients of cardiac allografts, which specifically prolonged donor test grafts in syngeneic hosts, suggests that unresponsiveness to cardiac allografts is, in part, dependent on suppressor cells. This study emphasizes the importance of UV irradiation of DLs in the modulation of alloreactivity and the induction of donor-specific unresponsiveness in adult animals.     

A comparison of the cyclic anhydride and mixed anhydride methods for 111In-DTPA chelation to monoclonal antibodies

The cyclic anhydride (CA) and the mixed anhydride (MA) of DTPA were synthesized and used to chelate 111In to an antimelanoma monoclonal antibody. The CA and MA methods showed mean labeling efficiencies of 25.7 and 20.5%, respectively (p = NS). The binding efficiency of labeled antibody to human melanoma cells in tissue culture also was similar (means = 52 and 50%, respectively, p = NS), as was tumor uptake in nude mice at 96 hrs post-injection (16%-CA vs 12%-MA). The method required less complicated chemical syntheses, much less preparation time, and the product was stable over a much longer period. The results suggest that the CA method is preferable for bifunctional chelate labeling of monoclonal antibodies with 111In-DTPA.     

The Goldston Syndrome: Report of a Case

We present a case of the Goldston syndrome—that is, cystic renal dysplasia and the Dandy- Walker malformation. The condition was diagnosed by ultrasound in a 635 g fetus in the seventeenth week of gestation. Ultrasound studies showed the fetal head to be somewhat enlarged with slight dilatation of the lateral ventricles and marked dilatation of the fourth ventricle. The kidneys were symmetrically enlarged and multicystic. Autopsy revealed evidence of the oligohydramnios syndrome. The kidneys were typical of the Goldston syndrome as were the microscopic lesions in the liver. To our knowledge this is the first reported case in which this diagnosis was made during intrauterine life.     

Selective lymphoid irradiation. VI. Effect of palladium-109-labeled lymphocytes on rat cardiac allograft survival

We have previously shown that pretransplant treatment with palladium-109 hematoporphyrin (Pd-H) and two small doses of antilymphocyte globulin (ALG) leads to donor-specific permanent acceptance of cardiac allografts in weakly histoincompatible rat combinations and significant prolongation of hearts in the ACI-to-Lewis strains. Pd-H also concentrates in nonlymphoid organs to a significant degree that is undesirable, so we have examined, in the present study, the efficacy of Pd-109 attached to lymphocytes in producing selective lymphoid irradiation and compared it with the immunosuppressive effect of Pd-H. Adoptive transfer of syngeneic lymphocytes labeled with palladium-109 (Pd-L) led to significant concentration of radioactivity in the peripheral lymphoid organs relative to the bone marrow (BM), intestine, thymus, and endocrine organs. The concentration of radioactivity in the spleen relative to BM and intestinal mucosa was 23:1 and 58:1, respectively. The immunosuppressive efficacy of a suboptimal dose of 3 mCi (one-third of the dose used in our earlier reports with Pd-H) administered via Pd-L (6 X 10(9) cells) at 4 days combined with 5 mg ALG at 2 days and 1 day prior to transplantation was compared in its effect on cardiac allografts with a similar dose of Pd-H (3 mCi) combined with ALG. The mean survival time (MST) of 6.5 +/- 0.4 days in untreated recipients was moderately prolonged to 14.6 +/- 3.0 days by Pd-H and ALG, and was not significantly different from an MST of 14.1 +/- 0.3 days achieved with 2 doses of ALG alone. Pretransplant treatment of Lewis rats with Pd-L and ALG produced a significant prolongation of ACI heart allografts to 30.5 +/- 3.1 days (P less than 0.001). These results suggest that Pd-L is more effective in prolonging rat cardiac allografts than a similar dose of Pd-H, and thus it may be a new method of selective lymphoid irradiation prior to transplantation.