The HSV1-TK has been used as a tool in gene therapy13
. Even the notion that a viral TK might be used to target radiation therapy has been explored by others 14
. Similarly, induction of the EBV-TK with a histone deacetylase inhibitor followed by treatment with ganciclovir has been reported to induce remissions in some patients15
. Here, we model a new approach: b
argeting of r
adionuclide (BETR) therapy. We show that this BETR approach yields promising results in lymphoid and epithelial malignancies in murine xenografts of human tumors.
How do the doses administered in the murine model extrapolate to the clinical setting? Experience with 131I-labeled antibodies against lymphoma in patients demonstrates that clinical responses are achievable with 0.5 to 2 Gy to the tumor16
. The biodistribution data presented in allow estimates of the absorbed dose to tumor and vital organs, and suggests that the kidneys and red marrow are the dose limiting organs for [131
I]FIAU. As detailed in the online supplement, it should be possible to deliver 0.5 to 2 Gy to a 1 gram tumor in a 70 kg patient with critical organ doses well below toxicity.
FIAU was previously investigated as an antiviral17
. Chronic dosing of FIAU was associated with hepatic damage and in some cases fatality18-20
. In those trials, treatment for less than four weeks with a cumulative dose less than 200 mg was not associated with either clinical or biochemical evidence of toxicity. Thus, the “no-effect dose” of FIAU might be conservatively estimated to be on the order of 0.1 mg/kg per administration. [124
I]FIAU has been administered to patients for imaging without any adverse effect on hepatic function21
. Extrapolating from our xenograft experiments, a radiotherapeutic effect is likely to be achieved at doses of FIAU that are orders of magnitude times less than the no-effect FIAU dose in humans.
BETR treatment results in concentration of FIAU in lymphoma and carcinoma xenograft models and impacts on tumor growth curves for each of the tumors studied (). However, in the lymphoma models, growth stops, whereas in the gastric carcinoma model growth only appears to be slowed. This difference likely reflects two phenomena. First, epithelial tumors are more resistant to radiation than lymphoid tumors. Second, the percentage of injected radioactivity per gram of tumor achieved in the lymphomas is greater than in the carcinoma (, ). Optimization for human tumors will ultimately require further xenograft and clinical studies, but the ability to image EBV-TK activation directly in patients and thus assess the relevant pharmacokinetics of radiolabeled FIAU offers the promise that optimization can be rapid.
One concern with all 131I-labeled radiopharmaceuticals is deiodination. In the case of tositumomab, dehalogenases decouple the radioisotope from the antibody moiety, resulting in free, circulating 131I. In order to block this accumulation, standard procedure involves pretreatment of patients with thyroid protective agents (potassium iodide) beginning before drug administration and continuing until 2 weeks after administration. In comparison, biotransformation of FIAU is limited because of the high chemical and metabolic stability of the N
-glycosyl linkage in pyrimidine nucleosides that contain the 2'-fluoro substituent in the arabinosyl (“up”) configuration22
. Studies in serum and whole blood over a 24 h period indicate excellent stability and low susceptibility to deiodination. FIAU is also cleared more rapidly from the plasma than tositumomab (3.9 versus 48 hours). Thus with less deiodination and more rapid clearance, it may be expected to have a better safety profile with respect to radiation exposure in normal tissues than agents already in the clinic.
BETR therapy also offers possible advantages over other lytic induction-suicide prodrug approaches. All lytic induction approaches are limited by the tendency of gammaherpesviruses to latency. When ganciclovir or a similar agent mediates cell killing, a therapeutic effect will likely require that a large fraction of tumor cells express viral enzymes. Although the phenomenon of bystander killing attributed to the exchange of phosphorylated nucleotides and nucleotide analogues between cells via gap junctions has been recognized23
, such killing is limited. In the gene therapy setting attempts have been made to engineer increased expression of the connexin protein in the gene therapy vector so as to increase such killing24
. However, this approach to increasing bystander killing is not readily applicable in approaches that do not involve gene therapy. Indeed, some of the therapeutic agents used to treat tumors may interfere with such bystander killing25
I]FIAU bystander killing is not limited by connexin expression or gap junctions but is a function of beta emissions with a maximum energy of 0.61 MeV that deposit 90% of their energy in a sphere of radius 0.7 mm26
. This “cross-fire” effect is likely an important contributor to the efficacy of radioimmunotherapy in lymphoma27
. “Cross-fire” with [131
I]FIAU may be inferred from where the tumor growth rate is substantially slowed when even 10% of tumor cells harbor the EBV-TK. Cell level dosimetry modeling is presented in the online supplementary material
A second limitation of the lytic induction-suicide prodrug approaches is that ganciclovir-mediated killing is cell cycle dependent. As reported by several groups of investigators, activation of lytic infection in EBV and KSHV infected cells leads to cell cycle arrest28-30
, thus likely compromising cell cycle-dependent tumor kill. In contrast, radiation effects are not similarly diminished by lytic induction.
BETR therapy may also offer advantages over radioimmunotherapeutic approaches. Internalization of radioimmunoconjugates may lead to degradation and release of free isotope with consequent loss of specificity. In contrast, we observed that following injection of FIAU, isotope is stably associated with EBV-TK expressing tumor (). Enzymatic phosphorylation and incorporation into DNA (as has been reported in HSV1-TK expressing cells 6
) offers the possibility of achieving high concentrations in tumor tissue not limited by concentration gradients. Finally, both FIAU and bortezomib are small molecules and thus better tumor penetration would be anticipated in comparison with antibody conjugates. FIAU does not penetrate the blood-brain barrier but does reach brain tumors reflecting disruption of the blood brain barrier22
. More broadly, the use of small molecules to activate tissue or tumor-specific metabolic pathways suggests the possibility that metabolic targeting of radiation may have applications beyond thyroid cancer.