Ac-225 and her daughters: the many faces of Shiva

Radioimmunotherapy (RIT) has evolved over a period of 20 years from the use of polyclonal antibodies conjugated to long-ranged beta emitting halogen radionuclides such as I-131 to the use of tumor specific monoclonal antibodies, their fragments, or other tumor selective peptides employing increasingly sophisticated radionuclides^1,2 such as Bi-213, At-211, Y-90, Lu-177, and Re-188 as the means to arm the targeting moiety to render it selectively lethal to the tumor cells. We have now reported methods for targeting tumor cells with Ac-225 atomic nanogenerators.³

Ac-225 is an alpha particle emitting radionuclide with a 10.0 d half-life.⁴ Ac-225 yields several daughter radionuclides in its decay scheme (Figure 1a) and a net of four alpha particle emissions. The parent and the daughters are each individually lethal to cells. Actinium therefore can serve as an atomic scale in vivo generator of alpha particle emitting elements. Alpha particles are charged helium nuclei that travel approximately 50–80 micrometers (2–4 typical cell diameters) and individually are able to kill a target cell due to their deposition of 5–8 MeV in a short ionizing track, while largely sparing surrounding cells; it is this characteristic that offers clear advantages to other known forms of radiation as a means of selective cell kill. Monoclonal antibodies that target a variety of different cancer cells and that were internalized following binding to cell surface antigenic molecules were modified to carry Ac-225 atoms stably to the tumor cell.³ This work also describes the use of these molecular-scale devices against both disseminated and localized cancer models in mice and some of the challenges that remain to be explored.

Figure 1

(A) The decay scheme of Actinium-225, showing half-lives and major emissions. (B) Differences in the radiobiology and geometry of beta and alpha particles. The left side shows the long-ranged, low energy deposition of the beta particle from an antibody targeted to a cluster of tumor cells in the vasculature. Nearly all of the energy is deposited in the normal tissue and not in the targeted cell. The right side shows a high linear energy transfer (LET) alpha particle emitted from a radiometal chelate conjugated antibody, in which most of the energy is retained within the target, yielding little bystander damage and the unique capability of single cell kill. With the several alpha emitting daughters internalized within the target cell, the effect is amplified

The field of radioimmunotherapy has benefited from increasingly better identification of and understanding of the biology of the tumor antigen–antibody systems, the availability of promising radiometals with unique characteristics, and subsequent development of radiometal–chelate chemistry necessary for their use. Appropriate radiometal–chelate chemistry is fundamental to success in this area, providing stable attachment of the potentially toxic radiometal to the targeting molecule in order to deliver it to the cell or into the cell without loss of it to metabolic or catabolic pathways which might endanger the patient or diminish the focus of the administered dose.⁵ Radiometals typically will remain inside a cell when internalized, despite cellular catabolic events. On the other hand, halogens, of which radioiodine or radioastatine are examples, are often rapidly excreted from the cell following catabolism⁶ reducing the efficacy of the dose administered and increasing doses to non-target tissues such as the thyroid, stomach and urinary tract.

The requirement for reliable sources of pure reactive radiometals, possessing emissions that are therapeutically, diagnostically and dosimetrically useful, has resulted in the development and study of a number of beta particle emitting nuclides including Y-90, Lu-177, and Re-188 and a number of alpha particle emitting radionuclides including Bi-213, At-211, and Ac-225. Alpha particles have a far shorter path length and a much higher linear energy transfer than beta particles (Figure 1b); they have been demonstrated to be significantly more selective and potent in killing targeted cells.⁷ The therapeutic potential of Ac-225 and its decay daughters was unrealized in vivo due to the varied chemical periodicity of the daughters, as no single chelating agent would bind all of them. One proposed solution to this problem was to bind the Ac-225 stably to an appropriate chelate in order to deliver it to the tumor and then rely on the antigen–antibody complex to modulate and efficiently transport the Ac-225 inside the cell. Once inside, it was demonstrated that the Ac-225 and its daughters remain internalized. Thus the daughters were also utilized therapeutically and not released systemically to accumulate in normal tissue.

Internalization of the radionuclide increases the probability that an ensuing particulate alpha emission will traverse the interior of the cell. Furthermore, the multiple alpha emissions from the Ac-225 nanogenerator source further increase the chances of a cytotoxic event occurring. In a variety of cancer models studied in vitro, a specific antibody that is labeled with Ac-225 is approximately 1000 times more potent on a mCi basis than the same antibody labeled with Bi-213 which emits only a single alpha and which has a 46 min half-life. This time is so short that only a fraction of the administered atoms will be internalized into target cells. The increased potency may be thus rationalized by both the 313-fold longer Ac-225 half-life and the four net alphas emitted.

Cellular responses to alpha particle exposure have been characterized and include gene mutation, chromosomal aberrations, cell cycle arrest, the induction of micronuclei and sister chromatid exchanges, lethality⁸ and the induction of apoptosis.⁹ The exact mechanisms by which alpha particles damage cells have not been determined. Obvious possibilities include direct alpha particle interaction with DNA and hydroxyl radical interactions, with DNA promoted by the high energy particle track through the cell yielding a significant number of ionizing events. Less obvious is the biological production of reactive oxygen species, superoxide and hydrogen peroxide, following exposure of human cells to alphas, which has been shown to mediate DNA damage indirectly in the absence of direct alpha hits on the cell.^8,10 The induction of apoptosis may be initiated in some cell types as a result of passive DNA damage following alpha exposure⁸ and contribute to the overall cytotoxicity. Microvascular endothelial apoptosis has been demonstrated to be the primary lesion leading to stem cell damage in mice following radiation exposure¹¹ and this may be a factor in vivo.

A surprising phenomena that we have observed is the resistance of a drug resistant leukemia cell line, RV+, to alpha particles. Exposure of RV+ and HL60 leukemia cells to either Bi-213 or Ac-225 labeled HuM195 (both of these cell lines express the CD33 antigen in similar amounts and both internalize the radiolabeled HuM195 antibody with similar kinetics) demonstrate a marked difference in the LD50 of radiolabeled construct necessary to kill cells. This phenomena necessitates further exploration since it may have implications for effectiveness of RIT of drug resistant cancers, commonly found in humans, and the mechanism of relative radioresistance to targeted alphas, an observation not previously described.

The extraordinary potency of alpha emissions predicts that use of Ac-225 labeled antibodies clinically in humans will utilize extremely low amounts of radioactivity, perhaps a fraction of a milliCurie (approximately 4E6 disintegrations per second). Furthermore, Ac-225 is more potent and specific than other radionuclides and may prove more lethal to tumor cells and other diseases when attached to an optimal targeting vehicle. Little is known now about the biology, chemistry or pharmacology of drugs based on Ac-225, and future efforts involving Ac-225 should include investigations into its mechanisms of action and resistance, its chemistry and daughter properties, better targeting and pretargeting vehicles for cancers and other diseases, the role of tumor vascular targeting, and intrathecal and intraperitoneal regional therapeutic approaches in patients.