Pretargeting CWR22 prostate tumor in mice with MORF-B72.3 antibody and radiolabeled cMORF

Abstract

Purpose

We have now applied our MORF/cMORF pretargeting technology to the targeting of CWR22 prostate tumor in nude mice.

Methods

The antiTAG-72 antibody B72.3 was conjugated with an 18 mer MORF while the cMORF was radiolabeled with ^99mTc. The specific binding of the antibody to the CWR22 cells was first confirmed in an assay placing the radiolabeled B72.3 antibody in competition with increasing concentrations of native B72.3. Thereafter, a group of four CWR22 tumored mice intravenously received the MORF-B72.3 and, 3 days later, the ^99mTc-cMORF, and were killed at 3 h postradioactivity injection. The dosage of the labeled cMORF was selected on the basis of previous experience in LS174T tumored mice. As controls, four animals received only the radiolabeled cMORF and another four received only the ¹¹¹In-B72.3. The maximum percent tumor accumulation (MPTA) of the labeled cMORF was subsequently determined by a dosage study of labeled cMORF. Both a multipinhole SPECT image and a planar gamma camera image were obtained of a representative mouse.

Results

The CWR22 tumor was confirmed to be TAG-72-positive. The MPTA of the labeled cMORF in the CWR22 tumor was 2.22%ID/g compared to only 0.12%ID/g in control mice without pretargeting. Both the planar and tomographic images confirmed the success of the CWR22 pretargeting.

Conclusions

The MORF/cMORF pretargeting approach has been successfully applied to tumor targeting of the prostate xenograft CWR22. However, the MPTA in this tumor model is lower than that in the LS174T tumor model investigated earlier, possibly due to a lower tumor blood supply.

Appendix

If we define a space containing the whole tumor, the amount of labeled effector dQ (ng) accumulated in tumor in an infinitely short period of time dt is:

$$ dQ\,{\left( {{\text{ng}}} \right)} = F{\left( {{\text{g}} \mathord{\left/ {\vphantom {{\text{g}} {\text{s}}}} \right. \kern-\nulldelimiterspace} {\text{s}}} \right)} \times f \times C{\left( {{{\text{ng}}} \mathord{\left/ {\vphantom {{{\text{ng}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} \times E \times dt $$

where F is the cardiac output in grams of blood per second (g/s), f is the blood fraction flowing into tumor, E is the trapping efficiency, i.e., the fraction of the effector reaching the tumor that is retained, and C is the blood concentration of the effector in nanograms of effector per gram of blood (ng/g). Given that the F and f are constants for a given tumor model (i.e., animal, tumor type, location, and tumor size), the total tumor accumulation Q over the entire period from administration to the complete clearance of free effector will be:

$$ Q\,{\left( {{\text{ng}}} \right)} = F \times f \times {\int\limits_{t = 0}^{t = \infty } {E \times C{\left( {{{\text{ng}}} \mathord{\left/ {\vphantom {{{\text{ng}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} } } \times dt $$

Dividing both sides of the equation by the injected dosage of effector (ng) and the tumor weight W and multiplying by 100, the accumulation of effector now in percent of injected dosage per gram of tumor (%ID/g) becomes:

$$ Q{\left( {{{\text{\% ID}}} \mathord{\left/ {\vphantom {{{\text{\% ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)} = F \times f \times W^{{ - 1}} {\int\limits_{t = 0}^{t = \infty } {E \times C{\left( {{{\text{\% ID}}} \mathord{\left/ {\vphantom {{{\text{\% ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} } } \times dt $$

As described earlier and verified experimentally [12, 13], Q is a constant when the dosage of effector is below that required to saturate the MORF-antibody in the tumor. Because the product F × f × W ⁻¹ is also constant, the integral ${\int\limits_{t = 0}^{t = \infty } {E \times C{\left( {{{\text{\% ID}}} \mathord{\left/ {\vphantom {{{\text{\% ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} } } \times dt$ must also be a constant under these conditions. Furthermore, ${\int\limits_{t = 0}^{t = \infty } {C{\left( {{{\text{\% ID}}} \mathord{\left/ {\vphantom {{{\text{\% ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} } } \times dt$ is a constant for the free labeled effector alone. In pretargeted mice, if the antibody concentration in normal tissues is sufficiently low, to a first approximation, the integral will be unchanged from the effector alone condition. Therefore, E must be a constant and can also be put outside of the integral. However, at dosages of effector greater than required to saturate the MORF antibody in tumor, E will become zero at some point when there is no longer MORF in tumor available to retain the effector, resulting in a smaller percent accumulation. Therefore, the accumulation before saturation of pretargeting agent in tumor will be the MPTA:

$$ {\text{MPTA}}\;{\left( {{\% {\text{ID}}} \mathord{\left/ {\vphantom {{\% {\text{ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)} = F \times f \times W^{{ - 1}} \times E{\int\limits_{t = 0}^{t = \infty } {\,C{\left( {{{\text{\% ID}}} \mathord{\left/ {\vphantom {{{\text{\% ID}}} {\text{g}}}} \right. \kern-\nulldelimiterspace} {\text{g}}} \right)}_{{{\text{blood}}}} } } \times dt $$

This equation shows that the MPTA is independent of the pretargeting agent and dependent only on the tumor and the effector.