Program: Oral and Poster Abstracts
Session: 614. Acute Lymphoblastic Leukemia: Therapy, excluding Transplantation: Poster I
Methods: Cytotoxicity was assessed with growth inhibition assays (WST-8). ALL cells were incubated with RIT for various times, washed, re-plated for a total of 72 h, stained for apoptosis by Annexin V-PE/7-AAD, and cell death analyzed by flow cytometry. NSG mice were inoculated with 5 million ALL cells IV on day 1 and treated with RIT from day 8 either IV or by continuous infusion with ALZET osmotic pumps implanted into the peritoneal cavity. Bone marrow (BM) samples were analyzed by flow cytometry for ALL infiltration and apoptosis.
Results: In vitro cytotoxicity (IC50) of cell lines varied 210-fold (0.02 to 4.6 ng/ml). The time that cells had to be exposed to RIT to kill >90% of the cells varied greatly from <30 minutes to >4 days. We observed similar exposure time variability on 8 primary ALL samples. Despite this variability, for 57% (8/14) of all samples tested, the two toxins HA22 and LMB-11 were similar in the time to induce cell death. For 38% (5/14) of samples, HA22 was up to 10-fold more active and killed more rapidly than LMB-11. One example was KOPN-8, which was killed after less than 30 minutes exposure to HA22, but needed more than 9 hours exposure to LMB-11 for similar effects. The difference in exposure time dependent killing between the two toxins was related to intracellular RIT-processing. Using two newly developed ALL xenograft models with KOPN-8 or HAL-01, we found that response in vivo correlated directly with the time needed to kill the cells in vitro. Because RITs have a short serum half-life in mice (<30 minutes) but killing of KOPN-8 or HAL-01 cells needed 9 h or 24 h exposure to LMB-11, we replaced bolus dosing with small doses given at frequent intervals or with continuous infusion. The change in dosing strongly improved response of both KOPN-8 and HAL-01 xenografts (table 1).
Conclusion: The previously unrecognized high variability of the time a CD22-targeting immunotoxin requires to kill ALL blasts provides a potential explanation for differences between in vitrocytotoxicity and the response rates observed in clinical trials. Our data indicate that ALL patients might show improved responses to anti-CD22 immunotoxins if treated with continuous infusion rather than the currently used bolus injection.
Table 1) In vivo response to immunotoxin improved with change in dosing.
RIT = recombinant immunotoxin, QOD = every other day, IP = intraperitoneal, SD = stable disease, CR = complete response, PD = progressive disease, PR = partial response.
ALL Xenograft Cell line | RIT | Change in dosing |
Change in response |
KOPN-8 | LMB-11 | QOD → frequent injections | SD → CR |
HAL-01 | HA22 | QOD → continuous infusion | PD → PR |
Disclosures: Wayne: Medimmune: Honoraria , Other: travel support , Research Funding ; NIH: Patents & Royalties ; Kite Pharma: Honoraria , Other: travel support ; Pfizer: Honoraria ; Spectrum Pharmaceuticals: Honoraria , Other: travel support , Research Funding . Pastan: NIH: Patents & Royalties: Coinventor on NIH Patent .
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