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Anti-CD5 ricin A chain immunoconjugate (XZ-CD5) is an immunotoxin that inhibits proliferative and cytotoxic responses to alloantigen in vitro and has activity in the treatment of acute graft-vs.-host disease (GVHD). To determine if XZ-CD5 could be used to prevent acute GVHD, 11 adult recipients of HLA-identical allogeneic marrow received XZ-CD5 0.1 mg kg-1 day-1 intravenously with high-dose methyl-prednisolone for 10, 14 or 17 doses early post-transplant. Six additional patients received 17 doses of XZ-CD5 and cyclosporine (CSA). All patients engrafted. Severe capillary leak syndrome was the most common serious toxicity and occurred more frequently in patients receiving CSA (5/5 vs. 3/11, P = 0.03). All evaluable patients developed acute GVHD; 88% had grade II-IV GVHD. Flow cytometric analysis demonstrated a substantial number of circulating CD5+ and CD3+ lymphocytes during and early after administration of XZ-CD5. These results suggest that the immunotoxin did not eliminate alloreactive T cells in this setting.

Type-I (insulin-dependent) diabetes mellitus is an immunologically mediated disease that results in destruction of the insulin secreting beta cells of the pancreas. T cells have been implicated in the pathogenesis of this disease. One novel form of anti-T-cell therapy is the immunoconjugate CD5-Plus. This agent is composed of the murine IgG1 monoclonal antibody H65, which is directed toward the CD5+ antigen; and ricin A chain, a ribosomal inhibitor protein. We performed a pilot study to evaluate the safety of the immunoconjugate in subjects with type-I diabetes mellitus. We conducted a dose-escalation study using CD5-Plus given as an intravenous infusion for 5 consecutive days. Fifteen subjects (12 men and 3 women) with a mean age of 26 years, a mean duration of diabetes of 4.8 months, and a minimum stimulated C peptide of 0.3 pmol/mL were entered. Six subjects each were treated at the 0.1 and 0.2 mg/kg/day dosage levels, and three subjects were treated at the 0.33 mg/kg/day dose. Glycemic control was determined monthly by recording the glycohemoglobin, total daily insulin requirements, and fasting blood glucoses. Beta-cell function was measured by determining the C-peptide response to a mixed formula meal (Sustacal) at baseline and at 1,3,6,9, and 12 months after treatment. The area under the curve (AUC) of the C-peptide response was calculated and, to reduce variability, related to that of the same subject at baseline. An analysis of subjects who retained at least 80% of their baseline beta-cell function as measured by the AUC was performed.(ABSTRACT TRUNCATED AT 250 WORDS)

XOMAZYME-Mel (XMMME-001-RTA) is an immunoconjugate comprised of ricin A chain conjugated to a murine monoclonal antibody directed against high molecular weight melanoma antigens. Although not necessarily related to increased toxicity or decreased efficacy, the development of anti-immunoconjugate antibodies may limit repetitive dosing with an immunoconjugate. We evaluated the role of cyclosporine A in blocking the antibody response in patients with melanoma treated with XMMME-001-RTA. Patients received cyclosporine in divided daily doses to achieve serum levels by HPLC of 150-200 ng/ml on days 1-22. On day 3, XMMME-001-RTA was begun at dosages 0.2-0.6 mg/kg daily for 5 days. Treatment was repeated every 35 days. Three patients were treated in each dosage tier (0.2, 0.4, 0.6 mg/kg). Nine patients were entered and all nine were evaluable. Patients had histologically confirmed melanoma. Metastatic sites included skin, soft tissue, and lymph nodes (seven), lung (two), liver (one), and spleen (one). There were four men and five women aged 46-75 years. Toxicities included myalgia, arthralgia, hypoalbuminemia, fatigue, elevations in liver function tests, and increased peripheral edema. Four patients received two to five repeated dosages of XMMME-001-RTA. One wheal-and-flare reaction from an immunotoxin test dose of XMMME-001-RTA was noted after five cycles. After a test dose subsequent to one cycle, two patients experienced chest tightness without ECG changes and were removed from the study. All toxicities resolved without sequelae. One patient experienced partial lymph node remission for 9 months. A second patient had stable mediastinal disease for 20 months. XMMME-001-RTA is safe when given repeatedly with cyclosporine.

To assess the biological effect of therapeutic monoclonal antibodies (mAbs) immunoconjugates, flow cytometric assays are often performed on patients' blood samples. We report here that, using standard protocols, staining whole blood samples with mAb-fluorochromes can result in erroneous immune cell phenotyping. Blood samples were obtained from patients treated with H65-RTA, a murine IgG1 anti-CD5 mAb conjugated to ricin A chain. While a transient decrease in CD4+ and CD8+ cells was observed during the 5 days of therapy, alterations in lymphocyte phenotype were also noted, beginning around days 15-30. The most prominent effect was an apparent loss of CD4+ cells. However, if the cells were separated from plasma prior to staining, the patients' samples demonstrated their pre-treatment lymphocytic phenotypes. Co-incubation of post-treatment (day greater than or equal to 15) patients serum with lymphocytes from normal donors also resulted in artifactual staining. The effects of the post-treatment serum could be correlated with the onset of a human immune response directed against H65-RTA. Moreover, co-incubation of normal PBMC with goat anti-mouse immunoglobulin could replicate the artifactual staining patterns. Even though the human immune response was generated against a murine IgG1 mAb, there were sufficient human antibodies crossreactive with other murine IgG isotypes to induce artifactual staining with all mAb-fluorochromes tested. To minimize artifacts, cells should be separated from plasma prior to flow cytometric analysis as well as for other immunoassays involving murine mAbs.