Pre-Clinical Evaluation of the Anti-CD22 Immunotoxin CAT-8015 in Combination with Chemotherapy Agents for Childhood B-Precursor Acute Lymphoblastic Leukemia (Pre-B ALL).

Despite advances in curative treatment, childhood ALL remains a leading cause of cancer-related mortality in pediatrics and survivors are at risk of multiple late effects. Targeted therapies that overcome chemotherapy resistance and decrease nonspecific toxicities are needed. The B-lineage differentiation antigen CD22 represents a relevant target for pre-B ALL. Monotherapy with the anti-CD22 immunotoxin RFB4(dsFv)-PE38 (CAT-3888 or BL22) has been well tolerated in a pediatric Phase I trial. To date however, only transient clinical activity has been observed with this agent in children with ALL. We conducted cytotoxicity studies to evaluate combination regimens of a higher affinity anti-CD22 immunotoxin (CAT-8015 or HA22) with standard ALL chemotherapeutic agents.

Methods: CD22+ human malignant cell samples studied included cell lines of Burkitt lymphoma (CA46) and pre-B ALL (REH and KOPN-8), as well as primary lymphoblasts from 2 children with multiply relapsed pre-B ALL. Flow cytometry was used to determine CD22 expression and anti-CD22 antibody binding using the BD Quantibrite System. Asparaginase, cytarabine, doxorubicin, methotrexate, and vincristine were added to cells individually and in various combinations with CAT-8015. Total incubation time was 2 days for cell lines and 5 days for primary patient blasts. Cell viability was assessed using WST-8. The Chou-Talalay median-effect method was used for data analysis (

Results: All samples were sensitive to CAT-8015 and to most chemotherapy agents tested. Simultaneous addition of a fixed concentration of CAT-8015 with varying concentrations of each chemotherapy drug led to a 2- to 4-fold reduction in IC₅₀ in comparison to individual agents. Assays with simultaneous addition of equipotency ratios of CAT-8015 and chemotherapy agents had synergistic or additive interactions. These interactions were sequence dependent, and the addition of CAT-8015 followed by chemotherapy led to greater positive interactions than the reverse order. Further assays with simultaneous addition of CAT-8015 and doublet chemotherapy drugs showed enhanced synergistic and additive interactions compared to assays with only doublet agents.

Conclusions: CAT-8015 combined with standard chemotherapy agents results in synergistic or additive cytotoxicity against CD22-positive Burkitt lymphoma and pre-B ALL cell lines and primary blasts from children with pre-B ALL. High affinity anti-CD22 recombinant immunotoxins have the potential to overcome chemotherapy resistance and to decrease nonspecific treatment-associated toxicities. In addition, CAT-8015 might synergistically enhance chemotherapy-induced cytotoxicity, thus improving efficacy and allowing the use of less toxic regimens in the future. Clinical trials of recombinant immunotoxins targeting CD22 in combination with standard ALL chemotherapy regimens are planned.