Current Radiopharmaceuticals - Volume 1, Issue 3, 2008

In this thematic issue of Current Radiopharmaceuticals the research and development in the field of internal radionuclide therapy with alpha-particle emitters is addressed. Targeted radionuclide therapy is a viable alternative to external beam radiation therapy, at least for some cancer forms, and can deliver selectively curative doses of radiation to cancer cells. The radiopharmaceuticals in vogue for these applications are labeled with radionuclides that emit β-particles such as 131I and 90Y. Beta-particles have a relatively long range in tissues and therefore deposit their energy over several millimeters. As a result of this, the fraction of absorbed radiation dose in tumor decreases with the decreasing size of the tumor. Thus β-particle radiations are appropriate for the treatment of larger tumors and those in which the uptake of the radiopharmaceuticals is not homogeneous. But radiopharmaceuticals tagged with β-particle emitting radionuclides can be detrimental to normal tissues adjacent to tumors. On the other hand, alpha-emitters, which deliver a much more energetic and localized radiation, are ideal for the treatment of minimal residue diseases, settings in which the targeted radiotherapy has the greatest chance of success. Such diseases include micrometastatic lesions, residual tumor margins that remain after debulking the primary tumor by surgery, skeletal metastases which appear in close vicinity to radiosensitive bone-marrow cells, and tumors in circulation including lymphoma and leukemia. In addition, targeted alpha particle therapy may be ideal for the treatment of local metastases in body cavities, e.g., ovarian cancer and neoplastic meningitis which spread as thin sheets of compartmental tumor often accompanied by free-floating cells. Alpha-particles are radiations of high linear energy transfer (LET) and have a number of radiobiological advantages. These include high relative biological effectiveness, less dependence of cytotoxicity on dose rate, and the ability to treat both normoxic and hypoxic cell populations. Also, the cytotoxic effect of high-LET radiations is only modestly affected, if at all, by the cell cycle status. Another positive feature of alpha-particle emitting radionuclides is that it is easy to shield hospital staff and families of patients from receiving radiation exposure since most alpha-emitters are associated with relatively little gammas and x-rays. High LET radiations like α-particles, are in general considered more carcinogenic than beta particles and conventional external beam irradiation. However, this problem can be reversed in clinical applications of targeted alpha therapy, since the short alpha-particle range facilitates the exposure of smaller normal tissue volumes, thus minimizing the number of normal cells at risk. The translational research in this field has for many years been hampered by lack of funding and supply of α-emitting radionuclides in clinically relevant amounts, but results of four clinical trials have been reported. A phase I dose escalation study with a 213Bi-labeled anti-CD33 monoclonal antibody for the treatment of patients with recurrent acute myeloid leukemia was presented in 2002. Yet another phase I study was performed in evaluating the therapeutic efficacy of an 211At-labeled chimeric anti-tenascin antibody in the treatment of patients with recurrent brain tumors. Results from a phase I study evaluating the usefulness of dissolved [223Ra]radium dichloride for the treatment of patients with prostate and breast carcinoma and skeletal metastases have been reported. A phase II trial of this compound is completed and a large scale phase III study, estimated to include 750 patients randomized two-to-one in favor of 223Ra, has recently been initiated. If this phase III study becomes successful, it could attract the interest for alpha-emitters among larger commercial players and secure a basis for further development of this exciting field of nuclear medicine. As mentioned above, the lack of availability of alpha particle-emitting radionuclides is one of the major impediments for the successful application of this mode of therapy. Fisher has dwelt on the commercial availability of various alpha-emitters and has made several recommendations to improve their supply. Morgenstern et al. have reviewed the production aspects describing the pros and cons of various processes..........

Alpha-emitting radionuclides provide effective cell-killing properties and have been shown to be effective in cancer treatment. The number of different alpha emitters having suitable physical and chemical characteristics for applications in medicine is relatively few. Development and testing of new radiopharmaceuticals requires a reliable supply of alpha- emitters in high quality, with timely delivery, but at reasonable cost. Applications and commercially availability of the following alpha emitters are reviewed: Actinium-225, bismuth-213, astatine-211, radium-223, bismuth-212, radium- 224, radium-226, terbium-149, and thorium-227. Nations with the capability to produce or to support production of alpha emitters should be actively involved in promoting the development of production capabilities and supporting applications research. Recommendations for improving the supply of these alpha emitters include an increased United States government commitment (through funding and joint-agency cooperation), establishing new production capabilities, and strengthening federal-private partnerships with companies involved to help to meet critical needs, both in the U.S. and elsewhere.

Targeted alpha therapy (TAT) is a promising approach for the treatment of cancer and infectious diseases. The availability of suitable alpha emitting isotopes in clinically relevant amounts is a main prerequisite for further development of TAT and its widespread clinical application. Several alpha emitting isotopes have been proven effective in preclinical studies and/or clinical trials, including 225Ac / 213Bi, 230U / 226Th, 227Th / 223Ra, 212Pb / 212Bi, 211At and 149Tb. Here we give an overview on methods for the production of these nuclides, describe advantages and limitations of the various processes and give an outlook on future availability and isotope supply.

A review of the literature has been conducted on 211At-labeled compounds, which focuses on their in vitro and in vivo stability towards deastatination. In vivo stability is very important in 211At-labeled compounds being developed for treating human disease. Instability of the 211At-carbon bond found in many labeled compounds leads to release of [211At]astatide. Therefore, the review includes information from literature reports on toxicity resulting from free [211At]astatide in animal models. The review also examines 211At-bond stability obtained when using different methods or reagents to label a variety of large and small molecules. Fortunately, in vivo stability is readily determined by conducting coinjected dual-label studies, where the study compound is labeled with [125/131I]radioiodine and separately with [211At]astatine. Radionuclide concentrations in selected tissues (i.e. lung, spleen, thyroid or neck, stomach) from several literature reports have been plotted to provide a visual indicator of relative in vivo stability obtained from the various labeling approaches. When the labeling approach involves formation of a carbon-astatine bond, a major factor determining stability appears to be the rate of metabolism of the carrier molecule. Other factors such as charge on the molecule, electronic and steric encumbrance about the astatine, and use of non-metabolizable (D-) forms of amino acids may also contribute to in vivo stability, but these could not be delineated from the literature reports. An alternate 211At-labeling approach, where a B-At bond is formed on anionic aromatic boron cage moieties, appears to increase stability to in vivo deastatination.

For the treatment of minimum residual diseases such micrometastases and residual tumor margins that remain after debulking of the primary tumor, targeted radiotherapy using radiopharmaceuticals tagged with α-particle-emitting radionuclides is very attractive. In addition to the their short range in tissue, which helps minimize harmful effects on adjacent normal tissues, α-particles, being high LET radiation, have several radiobiological advantages. The heavy halogen, astatine-211 is one of the prominent α-particle-emitting radionuclides in practice. Being a halogen, it can often be incorporated into biomolecules of interest by adapting radioiodination chemistry. A wide spectrum of compounds from the simple [211At]astatide ion to small organic molecules, peptides, and large proteins labeled with 211At have been investigated with at least two reaching the stage of clinical evaluation. The chemistry, cytotoxic advantages, biodistribution studies, and microdosimetry/pharmacokinetic modeling of some of these agents will be reviewed. In addition, potential problems such as the harmful effect of radiolysis on the synthesis, lack of sufficient in vivo stability of astatinated compounds, and possible adverse effects when they are systemically administered will be discussed.

Transfer of the gene encoding the noradrenaline transporter (NAT) into tumor cells rendered them amenable to dose dependent, [131I]meta-iodobenzylguanidine ([131I]MIBG) mediated cell kill. We have utilised the human telomerase component promoters (hTR and hTERT) to drive NAT gene expression specifically in tumor cells. Transfected cells were treated with benzylguanidine conjugated to the α-emitting radionuclide [211At]-astatine - [211At]MABG. This radiopharmaceutical was shown to be 1000 times more radiotoxic than the conventional β-emitter [131I]MIBG. Moreover the high LET emissions from 211At kill tumor cells regardless of cell cycle or oxygen status. The interactive components of this novel gene therapy strategy should provide a safe and potent form of radiation-mediated tumor cell kill. Indirect effects of ionizing radiation may contribute significantly to the effectiveness of radiotherapy by sterilizing malignant cells that are not directly hit by the radiation. However, there have been few investigations of the importance of indirect effects in targeted radionuclide treatment. We compared the induction of bystander effects by external beam γ- radiation with those resultant from exposure to three radiohaloanalogues of meta-iodobenzylguanidine (MIBG): [131I]MIBG (low linear energy transfer (LET)β-emitter), [123I]MIBG (potentially high LET Auger electron emitter), and meta-[211At]astatobenzylguanidine ([211At]MABG) (high LET α-emitter). We observed that potent toxins were generated specifically by cells which concentrated radiohalogenated MIBG. These may be LET-dependent and distinct from those elicited by conventional radiotherapy.

The radiobiological and radiochemical properties of radium-223 (223Ra, T1/2 = 11.4 d) render this alpha-emitting radionuclide promising for targeted cancer therapy. Together with its short-lived daughters, each 223Ra decay produces four alpha-particle emissions which enhance therapy effectiveness at the cellular level. In this paper, we review the recently published data reported for pre-clinical and clinical use of 223Ra in cancer treatment. We have evaluated two distinct chemical forms of 223Ra in vivo: 1) cationic 223Ra as dissolved RaCl2, and 2) liposome-encapsulated 223Ra. Cationic 223Ra seeks metabolically active osteoblastic bone and tumor lesions with high uptake and strong binding affinity based on its similarities to calcium. Based on these properties, we have advanced the clinical use of 223Ra for treating bone metastases from breast and prostate cancer. The results show impressive anti-tumor activity and improved overall survival in hormone-refractory prostate cancer patients with bone metastases. In other studies, we have evaluated the biodistribution and tumor uptake of liposomally encapsulated 223Ra in mice with human osteosarcoma xenografts, and in dogs with spontaneous osteosarcoma and associated soft tissue metastases. Results indicate excellent biodistributions in both species. In dogs, we found considerable uptake of liposomal 223Ra in cancer metastases in multiple organs, resulting in favorable tumor- to-normal soft tissue ratios. Collectively, these findings show an outstanding potential for 223Ra as a therapeutic agent.

A novel technology for preparing low dose rate alpha-radioimmunoconjugates has recently been developed based on the actinide radionuclide 227Th (T1/2=18.7 days). Thorium-227 has interesting properties that can be exploited in radioimmunotherapy: (1) it can be conjugated in stable fashion to antibodies using bifunctional DOTA; (2) it can be produced in virtually unlimited amounts with current technology; (3) it has a half-life suitable for central production and long distance shipment; (4) the alpha-particle released during decay has an average energy of 5.9 MeV, which is well suited for inactivation of tumor cells; (5) the in vivo fate of its radioactive daughter product in terms of distribution and toxicity is well known in animals as well as in humans. This review presents in vitro and in vivo preclinical results obtained using 227Th conjugated to the monoclonal antibodies rituximab and trastuzumab.

Poor prognosis of gastric cancer patients is based on early tumor cell dissemination into the peritoneal cavity. Therefore, efficient therapies for disseminated tumor cells are urgently needed. Patients suffering from diffuse-type gastric cancer occasionally express a mutant variant of E-cadherin lacking exon 9 (d9-E-cad). A monoclonal antibody (d9MAb) labelled with 213Bi specifically targeting d9- E-cad, therefore is a promising agent for the treatment of disseminated tumor cells. Incubation of HSC45-M2 gastric cancer cells expressing d9-E-cad with 213Bi-d9MAb induced formation of micronuclei and severe chromosomal aberrations finally causing cell death. Since inhibitors of apoptosis could not preserve cells from dying, 213Bi-d9MAb most likely induces necrotic cell death. This has been confirmed by analysis of 213Bi-d9MAb induced expression of genes involved in regulation of cell death. 213Bi-d9MAb treatment arrested HSC45-M2 cells in G2 phase as revealed by BrdU incorporation and by downregulation of genes involved in mitosis and cytokinesis. Therapeutic efficacy of 213Bi-d9MAb was investigated in a nude mouse model following intraperitoneal inoculation of HSC45-M2 cells. Therapy with 1.85 MBq 213Bi-d9MAb on day one after tumor cell inoculation defeated disease in 87% of all cases without any signs of toxicity. Therapeutic efficacy could be visualised non-invasively via bioluminescence imaging. Though higher activities of 213Bi-d9MAb also significantly prolonged survival, they caused long-term kidney toxicity. Therapy at day eight after tumor cell inoculation was less efficient, however fractionated 213Bi-immunotherapy could improve survival. Thus, locoregional radioimmunotherapy with 213Bi-d9MAb is a promising concept for treatment of early peritoneal carcinomatosis caused by intraperitoneal dissemination of gastric cancer cells.

This article is based on results from four published experimental studies. I: Three HER2 overexpressing cell lines were exposed to α-particles from the HER2 binding affibody molecule 211At- (ZHER2:4)2. The sensitivity differed; SKOV3 was resistant, SKBR3 intermediate and BT474 sensitive. The differences were unexpected since it is assumed that different types of cells should have a similar sensitivity to high-LET radiation. II: Effects of 211At-EGF in combination with the lysosomotropic base ammonium chloride were studied in A431 cells. The therapy effects of 211At-EGF increased by co-treatment with ammonium chloride, which was expected. III: Therapy effects of combined 211At-EGF and gefitinib treatment were studied using gefitinib sensitive A431 and resistant U343MG cells. The combined treatment reduced the survival of the resistant cells but in the gefitinib sensitive cells, the combined treatment increased survival, which was highly unexpected. IV: SKBR3 cells were exposed to the antibody trastuzumab that was labeled with 211At. A limited number of astatine decays per cell gave low survival, which was expected. Conclusion: The well-known high effectiveness of alpha particles to kill tumor cells was confirmed in all four studies. However, the dose-effect relations were complicated and more research is needed to evaluate factors of importance, such as internalization rate of the targeting agent, subcellular distribution and retention time of 211At.

Bismuth-213 (213Bi) (physical half-life 46 min) is a beta-emitter (97%) and an alpha-emitter (3%) which decays to short lived alpha-emitter Polonium-213 and could therefore be used as a generator of alpha particles with the energy of around 8 MeV. 213Bi has been successfully used during the last decade in both clinical and pre-clinical work for radioimmunotherapy (RIT) of cancer with 213Bi-labeled monoclonal antibodies (mAbs). RIT has been proposed as a novel techonology for treatment of infectious diseases. 213Bi-labeled mAbs have been successfully used for treatment of experimental fungal, bacterial and viral infections with transient or none hematologic toxicity. The mechanisms of RIT of infection with 213Bi-labeled mAbs include “direct” killing of cells and induction of apoptosis. RIT results in decrease of inflammation in infected organs. Among the delivery vehicles for RIT of infection whole IgG1 mAbs seem to be the most suitable in terms of the highest uptake in the target organs and the lowest - in normal tissues. RIT with alpha-emitter 213Bi involves the application of established technology developed for the treatment of malignancies to infectious diseases. The development of RIT for infectious diseases is potentially easier than its application to tumor therapy given antigenic and tissue perfusion differences between sites of microbial infection and tumor infiltration. Nevertheless, considerable preclinical and clinical development work is likely to be required to learn how to use RIT for infection optimally.

This paper reviews the journey taken by the Centre for Experimental Radiation Oncology from bench to bedside in the development of targeted alpha therapy (TAT) for metastatic melanoma. The alpha-immunoconjugate (AIC) 213Bi-cDTPA-9.2.27 (AIC) has been prepared and investigated for stability, labelling yield, toxicity, cytotoxicity and preclinical and clinical response. Preclinical studies gave high labelling efficiency and high cytotoxicity, translating to clinical trials where the conjugate was found to be very effective in regressing tumours while sparing the normal tissues. These trials provided significant information regarding pharmacokinetics, antigen expression and tumour response. Intralesional TAT (16 patients) was found to be non-toxic up to 1350 μCi and locally efficacious at doses of 600 μCi. In a systemic trial, 38 patients received activities up to 27 mCi (1000 MBq), for which there was no evidence of adverse events at any level. However efficacy was indicated as one patient showed a near complete response, 9% of patients had partial responses and 30% had stable disease for at least 8 weeks. These responses were dose independent, implying that other parameters such as antigen expression and vascular permeability of tumour capillaries play a role, and were explained by the development of tumour anti-vascular alpha therapy (TAVAT). The above clinical trials paved way for a phase 1 bridging study to optimise key parameters so as to establish a dose response relationship, to be followed by a phase 2 trial.

Among radionuclides usable for tumor therapy, α-particle emitters are characterized by a very high linear energy transfer (LET) resulting in a larger number of ionizations in a range corresponding to a cell diameter. Therefore, they can determine a stronger therapeutic effect compared to low LET β-particle emitters, producing their ionizations in a range up to many millimeters. In fact, because the distance between the two strands of DNA is almost the same as the distance between two ionizations of α-particles, DNA double strand breaks are induced with a high probability that finally cause cell death due to inefficient repair. Conversely, no therapeutic effect can be determined outside of concentrating sites. Therefore, the short range of α-emitters makes them powerful tools mainly when a therapeutic effect has to be reached in a restricted area, as in the elimination of minimal residual disease or in micro-metastases. Therapeutic efficacy of α-emitter radionuclides has been proven in numerous pre-clinical studies, but up to today only three main human studies are reported, including the treatment of myeloid leukemia by an anti-CD33 monoclonal antibody labelled by bismuth- 213 (213Bi), the therapy of patients with bone metastases from hormone-refractory prostate cancer by radium-223 (223Ra) and the loco-regional targeted radiotherapy with astatine-211(211At)-labelled anti-tenascin monoclonal antibody in patients with recurrent malignant brain tumours. The authors reviewed these human reported studies, evaluating perspectives, advantages and limitations of the targeted α-particle therapy.