Anti-ceramide single-chain variable fragment mitigates radiation GI syndrome mortality independent of DNA repair
Jimmy A. Rotolo, Chii Shyang Fong, Sahra Bodo, Prashanth K.B. Nagesh, John Fuller, Thivashnee Sharma, Alessandra Piersigilli, Zhigang Zhang, Zvi Fuks, Vijay K. Singh, Richard Kolesnick
Jimmy A. Rotolo, Chii Shyang Fong, Sahra Bodo, Prashanth K.B. Nagesh, John Fuller, Thivashnee Sharma, Alessandra Piersigilli, Zhigang Zhang, Zvi Fuks, Vijay K. Singh, Richard Kolesnick
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Research Article Stem cells Vascular biology

Anti-ceramide single-chain variable fragment mitigates radiation GI syndrome mortality independent of DNA repair

Abstract

After 9/11, threat of nuclear attack on American urban centers prompted government agencies to develop medical radiation countermeasures to mitigate hematopoietic acute radiation syndrome (H-ARS) and higher-dose gastrointestinal acute radiation syndrome (GI-ARS) lethality. While repurposing leukemia drugs that enhance bone marrow repopulation successfully treats H-ARS in preclinical models, no mitigator potentially deliverable under mass casualty conditions preserves GI tract. Here, we report generation of an anti-ceramide 6B5 single-chain variable fragment (scFv) and show that s.c. 6B5 scFv delivery at 24 hours after a 90% lethal GI-ARS dose of 15 Gy mitigated mouse lethality, despite administration after DNA repair was complete. We defined an alternate target to DNA repair, an evolving pattern of ceramide-mediated endothelial apoptosis after radiation, which when disrupted by 6B5 scFv, initiates a durable program of tissue repair, permitting crypt, organ, and mouse survival. We posit that successful preclinical development will render anti-ceramide 6B5 scFv a candidate for inclusion in the Strategic National Stockpile for distribution after a radiation catastrophe.

Authors

Jimmy A. Rotolo, Chii Shyang Fong, Sahra Bodo, Prashanth K.B. Nagesh, John Fuller, Thivashnee Sharma, Alessandra Piersigilli, Zhigang Zhang, Zvi Fuks, Vijay K. Singh, Richard Kolesnick

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Figure 2

Anti-ceramide 6B5 scFv protects against radiation-induced crypt lethality.

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Anti-ceramide 6B5 scFv protects against radiation-induced crypt lethalit...
(A) Anti-ceramide 6B5 scFv inhibits Jurkat cell apoptosis when administered to cell culture medium before 10 Gy exposure, as quantified in Figure 1B. **P < 0.01 vs. control, Wilcoxon’s rank sum test with continuity correction. Data are compiled from 3 independent experiments. (B) Anti-ceramide 6B5 scFv inhibits crypt lethality, determined by the microcolony assay, as in Figure 1C, when administered by i.v. infusion to C57BL/6 mice 15 minutes before 15 Gy WBR. ***P < 0.001 vs. control, Wilcoxon’s rank sum test with continuity correction. (C) Anti-ceramide 6B5 scFv (100 μg/25 g mouse) delivers equal or greater protection from crypt lethality when administered by the i.p., s.c., or i.m. route compared with i.v. injection. ***P < 0.001 vs. i.p. h2A2 IgG (1 mg/25 g mouse), Wilcoxon’s rank sum test with continuity correction. Data from microcolony assays show 1 representative experiment of 3 experiments, with 2 mice each, analyzing 10–20 intestinal circumferences per mouse. Only continuous, fully intact circumferences were counted.