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  • br Abbreviations CRC colorectal cancer

    2020-08-12


    Abbreviations: CRC ¼ colorectal cancer; MSLN ¼ mesothelin; RIT ¼ recombinant immunotoxin therapy.
    were inoculated in the right flank with SW48 E64 on Day 0, and tumors were allowed to grow to an average volume of 100 mm3 before initiation of treatment on Day 7. LMB-100 was administered IV at 2.5 mg/kg dose every other day for 6 days for 2 cycles with 3 days in between cycles. LMB-164 has an ABD and longer half-life (194 minutes) in mouse serum. LMB-164 was administered IV at 0.3 mg/kg dose every day for 3 days for 2 cycles with 1 day between cycles. These doses were determined to be safe in prior experi-ments.13 The data in Figure 3 shows that both LMB-100 and LMB-164 produced tumor regressions. All RIT-treated tumors had regressed by 50% on Day 12. Vehicle (PBS)-treated tumors grew exponentially throughout this time. By the second dose of LMB-100 and the third dose of LMB-164, a significant difference in tumor volume between the vehicle-treated controls and RIT-treated tumors developed. This difference persisted throughout the remainder of the experiment, reaching a maximum at Day 11. In addition, there was a significant delay of 13 days for the tumor volume to reach 300 mm3 in animals treated with RIT versus vehicle.
    Combination Therapy With Actinomycin D
    Previously, we have demonstrated that co-administration of actinomycin D with RIT upregulates apoptotic proteins of both the extrinsic and intrinsic pathways and shows significant synergy in vivo in treatment of pancreatic adenocarcinoma.18 Although actinomycin D has not recently been used in the treatment of colorectal adenocarcinoma, it remains a mainstay of therapy for childhood sarcomas, with a toxicity profile that does not overlap
    Table 1 Summary of IC50 (ng/mL) Values for MSLN-targeted RITs 
    with RIT therapy. Therefore, we tested LMB-100 in combination with actinomycin D using SW48 tumors. Figures 4 and 5 show that combination treatment resulted E64 in a 95% reduction in average tu-mor volume by Day 19 in comparison with a 40% reduction in tumor size for RIT monotherapy. Actinomycin D alone caused a growth delay shortly after initial dosing, but the tumors did not regress. In addition, combination therapy resulted in complete re-gressions in 13 of 22 treated mice. Figure 5 shows a spider plot of the tumor volume of an individual mouse from a representative anti-tumor experiment with 4 of 8 complete responses in the combination treatment group. There was no significant weight loss in animals treated with LMB-100 in combination with actinomycin D, although actinomycin D alone caused some weight loss after the second dose (see Supplemental Figure 1 in the online version).
    Combination Therapy With Oxaliplatin
    Previous data has shown synergy of chemotherapeutic agents with RIT therapy in in vivo models.19,20 Furthermore, combination screening of various chemotherapeutics noted a positive trend in combination of cisplatin and anti-MSLN targeted RITs.18 Given the widespread use of oxaliplatin platinum in CRC therapy, we investigated the potential of combing it with anti-MSLN RITs. We first analyzed combination efficacy using live/dead staining flow cytometry and noted a trend toward increasing numbers of dead and apoptotic cells with higher concentrations of oxaliplatin and LMB-100 (See Supplemental Figure 2 in the online version). We then tested the efficacy of oxaliplatin combination therapy with LMB-100 in mice. Although tumor volume regressed significantly in combination-treated mice in comparison to
    Abbreviations: IC50 ¼ half maximal inhibitory concentration; MSLN ¼ mesothelin; RIT ¼ recombinant immunotoxin therapy.
    Adam Cerise et al
    Figure 3 Female Nude Mice Were Inoculated Subcutaneously
    Figure 4 Female Nude Mice Were Inoculated Subcutaneously
    With SW48 Cells in the Right Flank and Underwent
    With SW48 Cells in the Right Flank and Underwent
    Treatment Beginning 7 Days Post Inoculation (Plotted
    Treatment Beginning 10 Days Post Inoculation
    as Day 0). Each Data Point Represents the Average
    (Plotted as Day 10). Each Data Point Represents
    Tumor Volume for n [ 8 Animals Treated With the
    Average tumor Volume for n [ 22 Animals Treated
    Indicated Recombinant Immunotoxin Therapy or
    With LMB-100, actinomycin D, Combination of
    Phosphate-buffered Saline Control. There Is a
    Actinomycin D and LMB-100, or Phosphate-buffered
    Significant Difference Between Immunotoxin-treated
    Saline Control. There Is a Significant Difference
    Tumors and Control Tumors (P < .05) Beginning on
    Between Groups Treated With Recombinant