Novel Radiopharmaceuticals for Therapy
In the era of personalized medicine, “target radionuclide therapy” (TRT) is designed to damage only the cancerous cells while sparing unnecessary damage to the adjacent healthy cells/tissues. Unlike conventional external beam radiation therapy, TRT is intended to cause less or no collateral damage to normal tissues, as it aims at achieving targeted drug delivery either to a clinically diagnosed cancer not amenable to surgery or to metastatic tumor cells and tumor cell clusters, thus providing systemic therapy of cancer. Currently there are hundreds of new pathway-targeted anticancer agents undergoing phase II and phase III clinical trials. TRT is just one type within the domain of “targeted therapies.” In addition to the effective targeted radiopharmaceuticals already well validated for routine clinical use, newer radiolabeled agents are still in the phase of either preclinical or clinical validation.
This chapter describes the main physical and radiochemical characteristics of radionuclides that have potential or have already been employed to label biologically reactive molecules for the development of novel radiopharmaceuticals for therapy. Some of these agents have entered advanced clinical trials in tumor-bearing patients. Results of these clinical trials cover a wide spectrum of potential clinical usefulness.
The chapter is divided into two main parts depending on the type of particle emission (α- or β-associated or not with the emission of either γ- or β+-radiation). Within each domain, there is some exchange of experience and shift of focus in the various phases of development, depending on the modalities of ascertaining efficient tumor targeting according to the principles of theranostics. The example of a novel α-emitting radiopharmaceutical that has most recently achieved approval by regulatory agencies for clinical use (223Ra-dichloride) is presented in detail as the paradigm for an agent that is showing a survival advantage besides the original target of pain palliation from bone metastases.
KeywordsTargeted therapies α-emitters β-emitters Auger electron emitters Antibody Peptide Ligand
Bispecific monoclonal antibody
Carbonic anhydrase isoenzyme 9
Clear-cell renal carcinoma
2-(4-isothiocyanatobenzyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (macrocyclic coupling agent to label compounds of biological interest with metal radionuclides)
External beam radiation therapy
European Medicines Agency
Ethylenediamine tetra(methylene phosphonic acid)
Food and Drug Administration of the United States of America
Human anti-mouse antibody
Human epidermal growth factor receptor 2, also known as receptor tyrosine-protein kinase erbB-2, or HER2/neu
- L-19 SIP
Small immunoreactive protein, (scFv)2, derived from monoclonal antibody L19
Monoclonal antibody recognizing the EDB domain of fibronectin
Metastatic castrate-resistant prostate cancer
Metastasis-associated protein encoded by the MTA1 gene
Medullary thyroid cancer
Maximum tolerated dose
Maximum tolerated radiation dose
A monoclonal antibody recognizing the sodium dependent phosphate transport protein 2b
Sodium-dependent phosphate transport protein 2b
Proteases, members of the urokinase-type plasminogen activator family
Positron emission tomography
Peptide receptor radionuclide therapy
Prostate-specific membrane antigen
Severe combined immunodeficiency
Single-photon emission computed tomography
Targeted alpha therapy
2-(4-isothiocyanatobenzyl-1,4,7,10-tetraaza-1,4,7,10-tetra-(2-carbamonylmetyl)-cyclododecane (macrocyclic coupling agent to label compounds of biological interest with metal radionuclides)
An agent with both diagnostic and therapeutic capabilities (e.g., 131I-iodide – in low activity it is a diagnostic agent, in high activity it is a therapeutic agent)
Cell-surface glycoprotein overexpressed in adenocarcinomas, correlated with tumor aggressiveness
Targeted radionuclide therapy
Vascular endothelial growth factor
World Health Organization
Targeted radionuclide therapy (TRT), is based on the use of high-affinity molecules as carriers of radionuclides to tumor cells (see also “Principles of Molecular Targeting for Radionuclide Therapy”). After intravenous administration these radiopharmaceuticals are distributed to tissues, so that they can reach their target molecule on the surface of tumor cells.
In clinical practice, TRT is often administered for the treatment of the most radiosensitive tumors, such as leukemias and lymphomas. Whereas solid tumors are usually more radioresistant, therefore greater ionizing radiation doses are needed for therapy. Because of the combination of a short path length in tissues (50–80 μm) with a high linear energy transfer (100 keV μm−1), targeted α-particle therapy offers the potential for more specific tumor cell killing with less damage to surrounding normal tissue than therapy with β-particle emitters. These favorable physical properties, for which a routine clinical application has already been developed with the novel radiopharmaceutical 223Ra-chloride for therapy of bone metastatic disease, offer the real possibility of targeted (or pre-targeted) α-therapy suitable for clearing minimal residual or micrometastatic disease.
Main characteristics of radionuclides already used or with potential for use in alpha-targeted therapy that are discussed in this chapter (arranged according to rising mass element number)
α, 5.9 (42)
α, 5.8 (97%); β, 1.4 (100%); α/βb, 0.02
α, 5.7 (100%)
α, 5.7 (100%)
α, 6.0 (48%)
The first alpha-emitting radiopharmaceutical that has obtained registration in the USA for treatment of bone metastases from prostate cancer is 223RaCl2 (Xofigo®), as described in detail further below in this chapter.
The alpha-emitting radioisotopes 211At, 213Bi, 225Ac, and 227Th are being used to label targeting vectors such as monoclonal antibodies for specific cancer therapy indications. In this review, safety and tolerance issues are considered with respect to microdosimetry, specific energy, Monte Carlo model calculations, biodosimetry, equivalent dose, and mutagenesis. When radiolabeled antibodies against the VEGF receptors are used, the clinical efficacy of targeted alpha therapy (TAT) may be enhanced also by their anti-neoangiogenesis effects, especially in the case of solid tumors. This review emphasizes key aspects of TAT investigations with respect to the PAI2-uPAR complex and the monoclonal antibodies (mAbs) bevacizumab, C595, and J591. Clinical trial outcomes are reviewed for neuroendocrine tumors, leukemia, glioma, melanoma, non-Hodgkin’s lymphoma, and bone metastases from prostate cancer.
Radium-223 ( 223 Ra) is commercially available for therapy of patients with metastatic castration-resistant prostate cancer (mCRPC). Radium-223 has a half-life of 11.4 days and releases 94% of its energy as α-particles with very little γ-emission. It is produced from the 227Ac/227Th system and purified using actinium-resin to immobilize 227Ac and 227Th . α-emission has certain advantages over β-emission, in particular lower bone marrow toxicity due to its narrow range in tissues. Similar to cationic strontium, the radium-223 ion is a calcium analog and therefore constitutes a natural bone seeker as it complexes with hydroxyapatite crystals in osteoblastic bone metastases, inducing non-repairable DNA strand breaks in cells.
The convenient biologic characteristics for alpha-emitters have fueled investigations on the use of radium-223 (administered as the dichloride salt) in preclinical models. In particular, an early study has reported a significant antitumor effect of radium-233 on experimental skeletal metastases . The model used for bone metastases was based on laboratory studies where the human cancer cells were injected intravenously into athymic nude rats. The authors used MTA-1 cells which are an estrogen and progesterone receptor-negative human breast cancer cell line. This model results in bone and bone marrow metastases as well as in the growth of tumor lesions in the spinal cord and in soft tissues. Biodistribution studies in this model identified selective uptake of radium in bone when compared to soft tissue. Over the course of time, the relative content of radium-223 in non-bone metastatic sites (including the liver, spleen, and kidney) increased relative to content in the femur. All tumor-bearing control animals (receiving no therapy) were sacrificed between 20 and 30 days, following the onset of secondary to paralysis caused by spinal infiltration of tumor. In contrast, mice receiving radium-223 developed symptoms significantly later, and many of them (7 out of 19, or 37%) were alive beyond 67 days. Furthermore, examination of sacrificed animals showed no significant changes of the bone marrow after radium-223. In a separate report, Henriksen et al.  directly compared the biodistribution of radium-223 with that of strontium-89 in murine xenograft models. Both compounds were found to accumulate in a higher concentration at bony surfaces as compared to soft tissues. Conversely, distribution at soft tissue sites was limited with both compounds. Supporting the notion that beta-emitters may be more myelotoxic than alpha-emitters, radium-223 was found to cause less damage to normal marrow when compared with strontium-89 based on an equal dose-deposition basis.
Phase I Assessment
Results of the first clinical trial with radium-223 dichloride were reported by Nilsson et al. with reference to a phase I study in which 25 patients were enrolled, including 15 with prostate cancer and 10 with breast cancer . Five dose levels of the compound were explored: 46, 93, 163, 213, and 250 kBq kg−1. At each dose level, five patients received a single dose of the agent with evaluation of pain response at weeks 1, 4, and 8. Fifty to sixty percent of patients experienced improvement in bone pain at each time point. A median overall survival of approximately 20 months was noted, and declines in alkaline phosphatase (ALP) levels were noted in 29% and 52% of breast and prostate cancer patients, respectively. Toxicities were generally mild, even at the highest dose levels explored. The incidence of grade 3 neutropenia was only 12%. In particular, imaging studies (based on the X-ray and gamma emission associated with decay of 223Ra) indicated that the majority of radium-223 accumulated in metastatic bone lesions.
A second phase I study reported by Carrasquillo et al.  evaluated a total of ten patients with mCRPC. A different array of dose levels of radium-223 than with the aforementioned experience was explored (50, 100, and 200 kBq kg−1). All patients received a single dose of the agent, while six patients (60%) received a second dose of 50 kBq kg−1. In this small cohort of patients, no dose-limiting toxicities were observed. Radium-223 was rapidly cleared from the circulation, with pharmacokinetic studies suggesting that only 0.5% of the drug remained in the circulating blood 24 h after administration. Most of the excretion occurred through the fecal route, as 52% of administered radium-223 was detected in the gastrointestinal tract at 24 h. As in the previously mentioned phase I experience, most of the observed toxicities were hematologic, with 60%, 70%, and 30% of patients developing any grade anemia, leukopenia, and thrombocytopenia, respectively. Besides grade 3–4 leukopenia observed in three patients (30%), grade 3–4 toxicities were limited.
Phase II Assessment
Three phase II studies have further assessed the activity of radium-223 in the setting of mCRPC. These studies differ slightly regarding eligibility and treatment schedule. The first study followed a randomized placebo controlled design and included 64 patients with mCRPC and either one painful bony lesion or multiple bony metastatic sites . All patients received external beam radiation therapy (EBRT) to a metastatic site and then were randomized to receive four concurrent treatment cycles with radium-223 or saline. Two primary endpoints were evaluated in this study: (1) change in bone ALP levels and (2) time to first skeletal‑related event (SRE). The study met the first of these two primary endpoints, with a more significant decline in bone ALP levels with radium-223 (−65.6% vs. +9.3%, P < 0.0001). Furthermore, there was a substantial (albeit at borderline of statistical significance) difference in time to first skeletal-related events (SRE, P = 0.065). Notably, a follow-up report from this trial suggests a survival advantage with radium-223 (65 vs. 46 weeks, P = 0.056), consistent with the subsequently noted phase III experience . A separate phase II study assessed a total of 100 patients with mCRPC and painful bony metastases . In contrast to the previous study design, patients did not receive concomitant external beam radiation therapy (EBRT) and were randomized to receive a single dose of radium-223 at one of four dose levels (5, 25, 50, or 100 kBq kg−1). The primary endpoint of the study was to characterize pain responses using two primary indices: (1) the visual analog scale (VAS) for pain and (2) the use of analgesics for pain control. The study met its primary endpoint, with an increased pain response at higher dose levels of radium-223 observed as early as week 2 (P = 0.035). At week 8, the proportions of individuals with a decrease in pain and stable analgesic use (at increasing dose levels) were 40%, 63%, 56%, and 71%, respectively (P = 0.040). As with the prior phase II experience, changes in markers of bone turnover were correlated with increasing dose levels, a significant reduction being observed in patients treated with 100 kBq kg−1.
The third and largest randomized phase II trial assessed 122 patients with mCRPC who had failed all available therapeutic modalities . While the previous two studies were designed to evaluate palliative endpoints (time to SRE, pain response, etc.), this study was powered to evaluate the proportion of individuals who achieved a prostate-specific antigen (PSA) decline of ≥50%. Patients were randomized in a double-blind fashion to receive a total of three 6-weekly doses of radium-223 at 25, 50, or 80 kBq kg−1. There was a dose-response relationship, with 0%, 6%, and 13% of the patients achieving ≥50% PSA decline in the respective cohorts (P = 0.0297). Greater declines in ALP values were also seen at higher dose levels, with a ≥50% decline in just 16% of patients treated at 25 kBq kg−1, as compared with 66% and 67% of patients treated at 50 and 80 kBq kg−1, respectively.
Phase III Assessment
Radium-223 dichloride is the first radiopharmaceutical to demonstrate a clear survival benefit, as reported by Parker and colleagues in an international phase III randomized trial (ALSYMPCA) . A total of 922 patients with mCRPC received six injections of 223Ra (50 kBq kg−1, every 4 weeks) or placebo. Eligibility criteria included symptomatic patients with progressive mCRPC with two or more bone metastases, no known visceral metastases, and castrate serum testosterone <1.7 nMol/L. In both groups, 57% of patients had received docetaxel, while the remainders did not (either because they were not good candidates for such chemotherapy or because docetaxel was not available).
Planned interim analysis found an improved median survival in the 223Ra group [14.9 vs. 11.3 months, hazard ratio (HR) for survival = 0.7; 95% CI 0.55–0.88, two-sided P = 0.002]. This study also demonstrated increased time to SREs (15.6 vs. 9.8 months, HR = 0.66; 95% CI 0.52–0.83, P < 0.001) and improved quality of life in the 223Ra-dichloride group [9, 10]. Radium-223 dichloride and placebo shared similar rates of all adverse events (93% vs. 96%, respectively), adverse events of grade 3 or higher (56% vs. 62%) and serious adverse events (47% vs. 60%). Only 16% of the patients enrolled discontinued treatment due to adverse events in the 223Ra arm compared with 21% in the placebo arm. Given the improvement in survival, excellent tolerance and ease of use, 223Ra-dichloride is likely to become the radiopharmaceutical of choice for treating patients with mCRPC and predominant skeletal metastasis. In this regard, it should be noted that other bone-seeking radiopharmaceuticals had been approved by regulatory agencies (FDA in the USA, EMEA in Europe) for “palliation of bone pain” from skeletal metastases. Whereas based on the results described above regarding the impact of 223Ra-dichloride on overall survival, this agent has been approved for “therapy” of mCRPC – a notion that has been recognized and embedded in the most recent international guidelines on treatment of mCRPC [11, 12].
Astatine-211 ( 211 At) is a cyclotron-produced α-particle emitter that has a 7.2-h half-life . It offers many potential advantages for α-particle therapy; however, its use for this aim is constrained by its limited availability.
This radionuclide is an attractive option for TAT, such as quantitative α-particle emission with each 211At decay, no long-lived α-particle-emitting daughter products, a half-life well suited for use with a diversity of chemistry applications, and possibility for conjugation with a variety of carrier molecules capable of efficient tumor targeting in vivo. Astatine-211 can be produced in reasonable yield from natural bismuth targets via the 209Bi(α,2n)211At nuclear reaction utilizing straightforward methods. There is some debate as to the best incident α-particle energy for maximizing 211At production while minimizing production of 210At; the latter occurrence is in fact problematic because of its 138.4-day half-life α-particle-emitting daughter, 210Po. The intrinsic cost for producing 211At is reasonably modest and comparable to that of commercially available 123I. The major limitation to 211At availability is the need for a medium energy α-particle beam for its production. On the other hand, there are about 30 cyclotrons in the world that have the beam characteristics required for 211At production . Considering its half-life of only 7.2 h, the current scarcity of 211At production sites limits the ability to ship it to distant sites for therapeutic applications.
Despite this drawback, there are many potential advantages of 211At as a therapeutic α-particle emitter, including cost, satisfactory experience regarding radiochemistry of conjugation, and evidence derived from proof-of-concept clinical trials. In addition, the decay of 211At also results in the emission of a 77- to 92-keV polonium X-ray, which enables noninvasive imaging of real-time biodistribution with either planar or SPECT scintigraphy . It is conceivable that the current supply bottleneck will be overcome as more clinical evidence provides further proof-of-concept for the use of 211At.
In 1989, labeling of a monoclonal antibody (mAb) with 211At using N-succinimidyl 3-(trimethylstannyl) benzoate as a synthon was reported . Studies with an internalizing mAb labeled with 211At using N-succinimidyl 5-[211At]astato-3-pyridinecarboxylate ([211At]SAPC) as a synthon showed that astatine-211 was intracellularly retained after internalization . Additional preclinical studies have recently shown that combined treatment with 211At-labeled anti-CD45 combined with bone marrow transplantation prolonged survival in a murine model of disseminated acute myeloid leukemia . Biodistribution studies showed excellent localization of the 211At-anti-murine CD45 mAb 30F11 to the bone marrow and spleen within 24 h. In syngeneic hematopoietic stem cell transplantation studies, radioimmunotherapy (RIT) with a 211At-conjugate improved the median survival of leukemic mice in a dose-dependent fashion, with minimal toxicity.
Feasibility of 211At-RIT was reported in two clinical trials. The first study assessed 211At-RIT with an anti-tenascin mAb followed by chemotherapy in patients with glioblastoma ; the radioimmunoconjugate was injected into the resection cavity and the maximal injected activity was 347 MBq (9.4 mCi). Six patients out of 18 had reversible grade 2 neurotoxicity, but no grade 3–4 toxicities were observed. Maximal tolerated activity was not reached and the observed median survival favorably compared with that of historical control groups. In the second phase I study, 211At-MX35-F(ab′)2 was assessed in women in complete response (CR) after a second-line chemotherapy for recurrent ovarian carcinoma; MX35 is a mAb against the sodium-dependent phosphate transport protein 2b (NaPi2b) . The aim was to determine dosimetry and toxicity. MX35-F(ab′)2 was labeled with 211At using N-succinimidyl-3-(trimethylstannyl)-benzoate. Nine patients underwent laparoscopy 2–5 days before 211At-RIT. Before intraperitoneal infusion of 211At-MX35-F(ab′)2, the abdominal cavity was inspected to exclude the presence of macroscopic tumor growth or major adhesions. The radioimmunoconjugate was infused at the activity concentration of 22.4–101 MBq/L in the peritoneal dialysis solution via a transcutaneous catheter. The estimated absorbed dose to the peritoneum was 15.6 ± 1.0 mGy per MBq/L, to the red bone marrow 0.14 ± 0.04 mGy per MBq/L, and to the unblocked thyroid 24.7 ± 11.1 mGy per MBq/L (decreasing to 1.4 ± 1.6 mGy per MBq/L when the thyroid was blocked). No adverse effects were reported. This study indicates that intraperitoneal 211At-RIT delivers therapeutic absorbed doses in microscopic tumor clusters, without significant associated toxicity.
Actinium-225 ( 225 Ac) is a particularly attractive radionuclide for therapy, since each decay is associated with the emission of four α-particles; this feature holds high promise for a very potent radiobiological effect – assuming that this radionuclide is conjugated with a selective and efficient tumor-targeting carrier. Actinium-225 has a half-life of 10 days and is produced as a decay product of uranium [20, 21]. In addition to α-particle emission, 225Ac decay also results in the emission of a 440-keV gamma ray that can be used for imaging after administration of a therapeutic agent. The relatively long half-life of 225Ac allows its shipment to potential users from a centralized production or purification site. In the USA, the Oak Ridge National Laboratory currently purifies 225Ac for commercial use from uranium stockpiles. The limiting factor for 225Ac is its cost, which currently exceeds $1,000/mCi. Besides purification from a uranium source, small amounts of high purity 225Ac can also be produced employing a standard proton-beam cyclotron, although using very long irradiation times (7–50 h) . Thanks to the widely available radiochemistry based on macrocyclic bifunctional chelators, 225Ac can be easily conjugated to antibodies and peptides without damaging the in vivo targeting properties of the antibody or peptide [23, 24]. Nevertheless, in the process of 225Ac conjugation to compounds of biological interest, there is an important radiochemical challenge; in particular, production of radiotoxic daughter products (such as 213Bi) during decay of 225Ac introduces in the system elements that have chemical properties different from those of 225Ac – therefore dissociate from the targeting moiety. A solution to this problem can probably be found in a novel approach based on the so-called 225Ac nanogenerator; in this approach the delivery system is designed to be internalized into the targeted tumor cell, where the toxic daughter elements may dissociate from the targeting molecule but will be trapped inside the cell, thus adding to the therapeutic effect of 225Ac [21, 25]. Moreover, nanotechnology techniques have also been explored to induce intracellular retention of several daughter products during 225Ac-based therapies [26, 27]. In particular, the retention of 225Ac and its radioactive daughters in the targeted tumor cell after internalization, according to the concept of an “in vivo generator,” was demonstrated using a mAb derivatized with DOTA [14, 28].
Clinical studies are ongoing in the USA with an 225Ac-labeled anti-CD33 antibody in patients with acute myeloid leukemia. An ongoing phase I trial of alpha-RIT based on the 212Pb/212Bi generator concept uses instead an anti-HER2 mAb infused intraperitoneally in patients with peritoneal carcinomatosis from HER2-expressing tumors [29, 30]. Production of daughter radionuclides during 225Ac decay complicates radiation dosimetry issues to nontarget organs, such as the kidneys, as an excretion route .
An interesting approach to the use of an 225Ac-labeled ligand for TAT is based on the high selectivity of the prostate-specific membrane antigen (PSMA) and on the long tumor retention of ligands for PSMA. In particular, a ligand named PSMA-617 has been developed, characterized by optimized tumor cell internalization and reduced kidney excretion versus the PSMA-11 ligand used for PET imaging after labeling with 68Ga. In addition, the PSMA-617 ligand contains DOTA as more universal chelator that makes radiolabeling of this compound with a variety of metal nuclides possible [32, 33]. Besides initial clinical trials with 177Lu-PSMA-617 [34, 35, 36], the results of a pilot, proof-of-concept study for the therapeutic use of 225Ac-PSMA-617. This treatment was offered as salvage therapy to two patients with mCRPC who had failed all lines of treatment and had widespread metastatic disease . Therapy consisted in the administration of three cycles of 100 kBq/kg body weight 225Ac-PSMA-617 at bimonthly intervals, followed in one of the two patients by further administration of 6 MBq 2 months later for consolidation. The results achieved following this regimen were impressive both from the imaging point of view (disappearance of virtually all tumor lesions visualized at PET imaging with 68Ga-PSMA-11) and from the biochemical point of view (reduction of serum PSA to <0.1 ng/mL from the initial levels >3,000 ng/mL in one patient and about 300 ng/mL in the other patient, respectively). Clearly, this pilot experience will need to be confirmed by additional controlled clinical trials, but will most likely open new perspectives in the treatment of advanced mCRPC.
Thorium-227 ( 227 Th) is a radionulide that decays to 223Ra, a long-lived radionuclide with proven good tolerance . A recent preclinical study reported the contribution of mAb internalization in the therapeutic efficacy of intraperitoneal 212Pb-RIT for small volume carcinomatosis, compared targeting of HER2 (internalizing) or CEA (non-internalizing) [39, 40]. An advantage was observed using internalizing anti-HER2 compared with non-internalizing anti-CEA 212Pb-labeled antibodies.
Bismuth-213 ( 213 Bi) is a short-half-life (46 min) α-particle emitter that is generally supplied through an 225Ac generator. The useful radionuclide production life of such a generator is about 10 days. The main drawbacks in the use of 213Bi as a label for therapy agents are its very short half-life and a somewhat high cost linked to the use of 225Ac, the starting material for the 213Bi generator. Another technical problem with 213Bi is the possible failure of the generator due to radiation damage, causing breakthrough of contaminant radionuclides such as 225Ac . 213Bi is limited by time constraints for radiolabeling and administration into patients of the tumor-targeting carrier molecule. Nevertheless, despite these limitations, a number of groups that have used 213Bi-labeled agents have been investigated for feasibility studies both in preclinical and in clinical trials [42, 43]. The results of these studies suggest however the superior therapeutic efficacy of the 225Ac-based nanogenerator described above over 213Bi-labeled antibodies, due to its greater α-particle yield per decay; additional roles are played by the longer half-life of 225Ac and its daughter products, which match the longer biologic half-life of large-molecular-weight antibodies . On the other hand, conjugation of 213Bi with biological tumor-targeting peptides might offer advantages because of the faster clearance of these compounds.
An early trial with a 213B1-labeled anti-CD33 humanized mAb administered to 18 patients with acute myeloid leukemia showed a reduction in circulating blasts in about 80% of the patients, without detectable extramedullary toxicity . In another pilot clinical study, 213Bi-DOTA-substance P was injected intratumorally via an implanted catheter in five patients with WHO grade II–IV gliomas; radiation-induced tumor necrosis was demonstrated after this form of therapy . More recently, an anti-mouse CD138 mAb labeled with 213Bi was employed for 213Bi-RIT in a syngeneic multiple myeloma mouse model ; when administered 10 days after engraftment, this treatment significantly prolonged survival and caused only moderate and transient hematologic toxicity.
Lead-212 ( 212 Pb), one of the radionuclides generated during the decay of thorium-232, is currently being investigated to develop new antitumor treatments . There is a definite potential for the clinical use of 212Pb (10.6-h half-life). 212Pb itself is not an α-emitter, but its physical decay results in the emission of two short-lived α-particles with potent therapeutic efficacy to cellular nuclei. 212Pb decays to 212Bi via β-emission. 212Bi, with a 60-min half-life, has a split decay chain and emits an α-particle at 36% frequency with an average energy of 6.1 MeV; 212Bi decays to 212Po for the remaining 64% via β-emission. In turn, 212Po decays to stable 208Pb in microseconds by emission of an 8.8-MeV α-particle. Betas are of low energy and/or frequency such that they are not expected to contribute significantly to toxicity or efficacy. Cumulative energy from the γ-emissions is <12% of those from the α-particles, but the 238.6-keV γ-ray with a 43% yield can provide the basis for imaging with a gamma camera. In preclinical studies, 212Pb has shown significant therapeutic efficacy in both in vitro and in vivo model systems [48, 49]. With the advent of the first clinical trial that employed 212Pb as the therapeutic agent (212Pb-TCMC-trastuzumab), this radionuclide has finally reached a landmark position in the family of α-emitters validated as suitable for such applications .
Among the variety of technologies that are available to conjugate radionuclides to a monoclonal antibody (mAb), two chelating systems have been used successfully for labeling with 212Pb, namely, the macrocyclic agents 2-(4-isothiocyanatobenzyl-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (C-DOTA)) and 2-(4-isothiocyanatobenzyl-1,4,7,10-tetraaza-1,4,7,10-tetra-(2-carbamonylmetyl)-cyclododecane (TCMC)). The TCMC chelate was specifically developed for chelating Pb(II) radionuclides, while C-DOTA is more multipurpose and has been used for chelating other metals including 111In, 90Y, 177Lu, and 212Bi. Although DOTA is known to form strong complexes with both Bi and Pb, a significant portion of 212Bi (~35%) is released from the carrier following the 212Pb/212Bi radioactive decay, thus resulting in the formation of highly ionized daughter atoms. Release of 212Pb after internalization of mAb delivery to cells has been reported as a source of marrow toxicity when using a DOTA conjugate, as the dissociated 212Pb was free to be transported to the bone and subsequently decay to 212Bi . In contrast, the differences in susceptibility to acidic conditions by the Pb(II)-TCMC complex were well demonstrated by Chappell et al. The Pb(II) [4-NCS-Bz-TCMC] complex, which has been very efficiently radiolabeled with Pb(II) radionuclides, was less labile at pH 3.5 than the corresponding DOTA complex, thus conferring enhanced resistance to acid-catalyzed dissociation with the cell. More recently, the structure of the Pb(II)-TCMC complex revealed that Pb(II) is fully encapsulated by TCMC with the eight-coordinate sphere saturated by the four ring nitrogen and four amide oxygen atoms; this complex is extremely stable and has decreased acid lability for in vivo sequestration of Pb(II) [51, 52, 53].
Based on the successful 212Pb preclinical data published from Milenic et al.  and from Anderson et al. , 212Pb-TCMC-trastuzumab may have potential for an improved therapeutic ratio over β-emitters such as 90Y- and 177Lu-targeted conjugate therapy and thus have beneficial activity against patients with malignancies expressing HER2. Recently, a phase I clinical trial with i.p. 212Pb-TCMC-trastuzumab at the University of Alabama (Birmingham, Alabama, USA) was designed to determine distribution, pharmacokinetics, and safety of i.p. 212Pb-TCMC-trastuzumab in patients with HER2-expressing malignancy . In this study, 212Pb-TCMC-trastuzumab was administered i.p. to three patients with HER2-expressing cancer who had failed standard therapies. After administration of 7.4 MBq/m2, gamma camera imaging studies showed no distribution of radioactivity out of the peritoneal cavity, nor normal organ uptake. These data were consistent with preclinical studies. Cumulative urinary excretion was <6% in 2.3 half-lives. The maximum external exposure rate immediately post-infusion at skin contact over the abdomen averaged 7.67 mR/h and dropped to 0.67 mR/h by 24 h. The data points correlate closely with 212Pb physical decay. Follow-up for over 6 months showed no evidence of agent-related toxicity, thus indicating good tolerance.
In the second study using 212Pb-TCMC-trastuzumab, Meredith et al.  evaluated the safety, pharmacokinetics, immunogenicity, and tumor response of 212Pb-TCMC-trastuzumab administered i.p. in patients with HER2-expressing malignancy . Five dose levels of treatment with i.p. 212Pb-TCMC-trastuzumab resulted in little toxicity in patients with ovarian cancer. Limited redistribution of radioactivity out of peritoneal cavity to circulating blood, which cleared via urinary excretion, and no specific uptake in major organs were observed at 24 h. The highest administered activity concentration at 500 Bq/mL was needed for high therapeutic efficacy without any side effects. Therefore, the phase study I with i.p. 212Pb-TCMC-trastuzumab confirmed its clinical potential and feasibility. The data obtained are consistent with those of prior nonhuman studies showing prolonged retention of 212Pb-TCMC-trastuzumab within the peritoneal cavity, with no evidence for localization to normal organs on planar images. The relatively low radiation dose is also closely comparable to dosimetry data obtained with the use of 211At-MX35-F(ab′)2.
There are currently eight β-emitting agents approved by the FDA for therapy of cancer in humans: 131I-iodide (for differentiated thyroid cancer), 131I-metaiodobenzylguanidine (131I-MIBG, for neural-crest-derived neuroendocrine tumors), 131I-tositumomab and 90Y-ibritumomab tiuxetan (Bexxar® and Zevalin®, respectively, both for CD-20-expressing non-Hodgkin’s lymphomas), 89Sr-chloride and 153Sm-EDTMP (Metastron® and Quadramet®, respectively, both for palliation of bone pain from skeletal metastases), and finally 90Y-labeled microspheres (TheraSphere® and SIR-Spheres®, for radioembolization of primary and metastatic liver tumors). Not all these agents meet the criteria to be considered as true “tumor-targeted” therapeutic agents based on specific interaction/binding of the radiopharmaceutical with antigens/receptors on the surface of tumor cells.
On the other hand, numerous preclinical and clinical investigations have addressed a much broader spectrum both of carrier molecules targeting tumor-specific sites and of additional β-emitting radionuclides. A tentative list of the latter includes at least eight radionuclides: lutetium-177 (177Lu), holmium-166 (166Ho), rhenium-186 (186Re), rhenium-188 (188Re), copper-67 (67Cu), promethium-149 (149Pm), gold-198 (198Au), and rhodium-105 (105Rh).
Main characteristics of radionuclides already used or with potential for use in beta-targeted therapy that are discussed in this chapter (arranged according to rising mass element number)
β, 0.4 (100%)
β, 2.2 (100%)
β, 0.5 (100%)
β, 2.0 (100%)
β, 0.6 (~80%); γ, 0.2 (44%); 0.08 (18%)
β, 0.6(89%); γ, 0.364 (81%)
β, 1.854 (50%), 1.774 (48.7%); γ, 0.8 (6.7%)
β, 0.480 (86.5%), 356.2 (3.3%), 0.7 (10.2%)
β, 1.07(97%); γ, 0.28 (3%)
β, 0.96 (100%)
β, 0.56 (70%); γ, 0.25 (30%)
Although certainly not the latest emerging β-emitter for therapy, iodine-131 still fulfills an important role in the development of new tumor-seeking radiopharmaceuticals with therapeutic intent. In particular, this radionuclide is being employed in a series of clinical trials based on monoclonal antibody constructs against a family of target antigens that are essential components of the extracellular matrix (ECM). The molecular architecture of the ECM is extremely heterogeneous with wide variation in its constituents in different organs and tissue compartments as well as within the same tissue depending on different developmental and physiologic states . The main components of the ECM are laminins, collagens, fibronectins, and elastins, along with other compounds such as tenascin, thrombospondins, members of the coagulation system, neuronal guidance molecules, and specific growth factor-associated proteins [58, 59]. During oncogenesis, the molecular composition and structure of the ECM is dynamically modified .
One of the principal constituents of ECM is fibronectin (FN), a large protein that exists in different isoforms (240–270 kDa) and different molecular forms . FN usually exists as a dimer of two nearly identical ~250 kDa (≈2,500 amino acids) subunits, covalently linked by a pair of disulfide bonds near the C-terminal end of the subunits. Although FNs are the product of a single FN gene, the resulting protein can exist in multiple forms, which – apart from posttranslational modifications – arise from alternative splicing of its primary RNA transcript. This polymorphism leads to the production of as many as 20 different isoforms for human FN. The 3D structure of the alternatively spliced-in EDB domain, a 91-amino acid sequence, is identical in the mouse, rat, rabbit, dog, monkey, and human. EDB has a restricted pattern of expression: undetectable in normal adult tissues and in mature blood vessels, but highly expressed in regenerating tissues and around new blood vessels [60, 61]. The FN isoform containing the ED-B domain is an ideal tumor target since it is widely expressed in the ECM of all solid tumors and is constantly associated with angiogenic processes (i.e., wound healing) , but is otherwise undetectable in normal adult tissues . The EDB-FN selective expression pattern has been extensively demonstrated in many different tumor types, in particular on invasive ductal carcinoma , brain tumors , lymphomas , and prostate cancer . High EDB expression is found around the newly forming blood vessels, thus suggesting that EDB is an important target for tumor-associated angiogenesis [67, 68, 69, 70].
After the identification of the EDB domain, monoclonal antibodies specific for EDB-FN were raised. A series of molecular biology experiments finally yielded a high-affinity clone with KD = 54 pM for EDB, termed clone L19 [71, 72]. The enhanced binding affinity of L19 as well as valence (monomeric scFv vs. dimeric scFv) lead also to improved targeting of tumoral angiogenesis, as was shown in biodistribution studies in F9 murine teratocarcinoma-bearing nude mice, where L19 labeled with 125I demonstrated rapid specific localization around tumor blood vessels . Biodistribution experiments in F9 tumor-bearing mice using radioiodinated L19 (scFv)2, L19-SIP (“small immunoreactive protein” format), and L19 IgG1 showed that L19-SIP offers the best compromise between molecular stability, clearance rate, and tumor accumulation [74, 75, 76]. In addition to the established therapeutic benefit, the dosimetric estimates obtained with 131I-L19SIP in murine models of cancer are among the most promising reported so far for radioimmunotherapy (RIT) .
The first immunoscintigraphy study in patients with primary tumors and metastatic lesions using 123I-L19 showed selective targeting in aggressive types of lung cancer and colorectal cancer . Following this proof-of-concept study, phase I and phase I/II dose-finding and efficacy studies with 131I-L19SIP in patients with a variety of cancers were initiated. 131I-L19SIP showed an excellent tolerability at radioactive amounts as high as 7,400 MBq (200 mCi) in patients with solid tumors, associated with therapeutic benefit for some patients . During the phase I study, 50 patients received a single dose of 4 mg of 131I-L19SIP radiolabeled with up to 185 MBq (5 mCi) for dosimetric purposes (“dosimetric phase” of the study). Whenever the radiation dose in at least one tumor lesion was found to be at least tenfold higher compared to the dose delivered to the bone marrow (the rate-limiting organ), the patient became eligible for RIT with a single dose of 5 mg 131I-L19SIP radiolabeled with 3,700 MBq (100 mCi) or 5,550 MBq (150 mCi), which constituted the “therapeutic phase” of the study. In an extension of the study, repeated treatments were performed, and patients with hematologic malignancies were included, based on the evidence that EDB is strongly expressed in almost all malignant lymphomas and in certain leukemias. The most impressive therapeutic effect in this study was demonstrated in a young female patient suffering from Hodgkin’s lymphoma . In general, uptake of 131I-L19SIP was observed both in primary tumor lesions and in distant metastasis, irrespective of cancer type and location of the lesions. RIT caused G3/G4 hematologic toxicity, but this was not predictable from the dosimetric estimates. Nevertheless, the treatment showed an excellent tolerability profile. Side effects were mostly hematological in nature and all were easily manageable and recovered spontaneously. Concerning in particular patients with hematologic malignancies, 18 patients with either relapsed, progressing lymphoma or multiple myeloma were enrolled as candidates for RIT with 131I-L19SIP. Tumor uptake of sufficient degree to justify RIT was observed in 11/15 patients with lymphoma and in 3/3 myeloma patients. Following a single course of RIT with 131I-L19SIP (standard activity of 3.7 GBq, reduced in some cases to 1.85 GBq because of low body weight), complete response was achieved in two patients with Hodgkin’s lymphoma and one patient with large B-cell lymphoma, while partial response was observed in one patient with Hodgkin’s lymphoma and patients with multiple myeloma experienced stabilization of disease. The objective response rate was therefore 40%, a highly promising outcome for these patients who had failed all lines of treatment, also considering that RIT with 131I-L19SIP was associated with low toxicity .
Labeling of L19SIP with 124I rather than with 131I during the dosimetric phase of the study allows serial PET imaging, with more accurate dosimetric estimates . Further advances in this field include proof of the therapeutic potential in prostate cancer patients .
Lutetium-177 ( 177 Lu) is a very promising β-emitter for targeted radiotherapy. Its β-emission, with a maximum energy of 498 keV, is useful in destroying small tumors and metastasis (optimal size 1.2–3.0 mm), while at the same time reducing damage to normal tissues. From this point of view, 177Lu can be seen as a complement to yttrium-90 or to iodine-131 . 177Lu also emits low-energy gamma rays (208 and 113 keV with 10% and 6% abundance, respectively) , which allow direct imaging of the activity distribution in the patients’ body for dosimetry calculations following administration of the therapeutic activity.
With its 6.71-day half-life, lutetium-177 lends itself to the use of more sophisticated procedures to synthesize and purify radiopharmaceuticals and to transport them to more distant nuclear medicine departments. As to the potential applications of lutetium-177 for RIT, prostate carcinoma (PCa) seems to be a favorable solid tumor because of its radiosensitivity and the typical metastatic diffusion to sites where there is potentially a high exposure to circulating mAbs (such as bone marrow and lymph nodes). In preclinical and clinical PCa therapy studies, different antibodies or peptides with affinity to mucin, ganglioside (L6), Lewis Y (Ley), adenocarcinoma-associated antigens, the TROP-2 pancarcinoma marker, and prostate-specific membrane antigen (PSMA) have been linked to radionuclides; PSMA appears the most specific of these targets [83, 84].
PSMA is a non-secreted, integral, type II membrane protein with abundant and nearly universal expression on epithelial prostate cells that is strongly upregulated in PCa [85, 86]. PSMA expressed in normal prostatic cells (brush border/luminal location) is not typically exposed to a circulating mAb, and the PMSA’s level of expression in non-prostate tissues is 100–1,000-fold less than in prostate tissue. The de-immunized J591 mAb, which links the PSMA’s external domain, seems to be the best clinical candidate for clinical RIT in patients with PCa .
In a 177Lu-J591 phase I trial, 35 patients were enrolled . The maximum tolerated dose (MTD) was determined to be 2,590 MBq/m2. In addition, it was shown that repeated dosing (up to three doses of 1,110 MBq/m2 each) could safely be injected. In all patients, the sites of metastatic disease were successfully imaged with gamma camera scintigraphy after administration of 177Lu-J591. As to the biochemical PSA response, the median duration of posttreatment PSA stabilization was 60 days (range between 28 and 601 days).
A phase II 177Lu-J591 clinical trial has been conducted in mCRPC patients . Patients were enrolled in two cohorts: 2,405 MBq/m2 in cohort 1 versus 2,590 MBq/m2 in cohort 2. Fifteen patients were enrolled in cohort 1, while cohort 2 was expanded to enroll 17 patients instead of the 15 patients as originally planned; targeting of 93.6% of the known sites of metastasis was detected by scintigraphy. Hematologic toxicity consisted in reversible thrombocytopenia and neutropenia and observed in 46.8% and 25.5% of the patients, respectively. The cohort 2 patients exhibited a higher PSA responses (46.9% vs. 13.3%, P = 0.048) associated with a longer survival (21.8 vs. 11.9 months, P = 0.03), but also more frequent hematologic toxicity. This trial shows that radiolabeled de-immunized J591 is well tolerated and non-immunogenic. Radiolabeled J591 effectively targets PCa metastases and induces PSA reduction with a dose-effect relation.
Among several alternative possibilities, pre-targeting may be achieved using a first injection of an unlabeled bispecific monoclonal antibody (BsMAb), followed by subsequent injection of a radiolabeled bivalent hapten-peptide [90, 91]. In this approach, the radiolabeled bivalent peptide binds avidly to the BsMAb attached to the antigen at the cell surface, whereas non-targeted hapten-peptide in the circulation clears rapidly through the kidneys. Medullary thyroid cancer (MTC) cells express high amounts of CEA, and encouraging therapeutic results have been obtained using anti-CEA pre-targeted 131I-di-DTPA peptide in two phase I/II and one phase II clinical trials [90, 92].
New-generation BsMAb and bivalent hapten-peptides are currently available. Humanized, recombinant, trivalent BsMAb and the histamine-succinyl-glutamine (HSG) hapten and bivalent HSG hapten-peptides have been created [93, 94]. The use of humanized BsMAb should reduce immunogenicity and the dock-and-lock procedure allows large-scale production. Accordingly, a series of bivalent HSG haptens have been created, offering the possibility of labeling with many radionuclides, including yttrium-90 and lutetium-177 for therapy purposes . The first clinical study evaluating the anti-CEA anti-HSG BsMAb TF2 and the radiolabeled hapten-peptide, 177Lu-IMP288, in patients with mCRPC has been published recently . Different approaches were explored in four cohorts of five patients each, to identify the optimal molar doses of TF2 and IMP-288 and the optimal holdup between the two infusions: (1) shortening the interval between the BsMAb and peptide administration (1–5 days), (2) escalating the TF2 dose (from 75 to 150 mg), and (3) reducing the peptide dose (from 100 to 25 μg). Rapid and selective cancer uptake was observed within 1 h after the peptide injection, with high tumor-to-tissue ratios at 24 h. The best tumor targeting was obtained with a 1-day pre-targeting interval and with the 25-μg peptide amount. High activities of 177Lu-IMP288 (2.5–7.4 GBq) were well tolerated. Some controllable reactions during the TF2 infusions occurred, and transient grades 3–4 thrombocytopenia was reported in 10% of the patients. Dosimetric analysis showed that red bone marrow and renal uptake of 177Lu-IMP288 were relatively low, although marrow doses increased in subsequent cohorts as the TF2/Lu-IMP288 ratio was improved . The predicted kidney-absorbed doses (<0.50 mGy/MBq) did not restrict the maximum activity that could be injected. None of the patients would exceed the limit of 15 Gy to the kidneys with four cycles of 7.4 GBq 177Lu-IMP288. These data indicate that pre-targeting using dock-and-lock bispecific BsMAb would be most practical to deliver short-half-life radionuclides.
Bound with methylene phosphonic acid (EDTMP), 177Lu can be used as a bone pain palliation radiopharmaceutical. Preclinical studies and initial clinical trials with 177Lu-EDTMP have resulted in encouraging data . Results of clinical studies show that 177Lu-EDTMP is an effective radiopharmaceutical for palliation of metastatic skeletal pain in patients with breast or prostate cancer. An amount of 1,295 MBq (35 mCi) is sufficient for bone pain palliation therapy, but doses as high as 2,590 MBq (70 mCi) have been shown to be safe.
Holmium-166 ( 166 Ho) decays by β-emission to 166Er (stable) with Eβmax of 1.854 MeV (50.0%) and 1.774 MeV (48.7%). It decays with a half-life of 26.83 h. It emits 81-keV gamma photons (6.7%), which are suitable for external imaging to track the in vivo distribution and uptake of the radionuclide. The maximum soft tissue penetration of the β-particles is ~8.5 mm and hence is suitable for certain specific therapeutic applications. Holmium-166 is a very attractive candidate for therapeutic purposes, as this radionuclide emits gamma rays in addition to high-energy beta particles, thus allowing both nuclear imaging and radionuclide therapy, according to the principles of “theranostics.” Moreover, holmium can be visualized by computed tomography and magnetic resonance imaging owing to its high attenuation coefficient and paramagnetic properties .
Also favorable is its half-life of 26.9 h, which is long enough to overcome logistic problems encountered with the short-lived radionuclides and is sufficient to provide a high radiation dose rate. Its nuclear reaction cross section is comparable with that of rhenium, but 165Ho has a natural abundance of 100%; thus, only one radioisotope, 166Ho, is formed by neutron bombardment. Taking these characteristics into consideration, 166Ho is therefore an attractive candidate for use to tailor the future needs for treatments. The first study with this radionuclide was for the development of radiosynovectomy agents for treatment of rheumatoid arthritis [98, 99]. Development of 166Ho-labeled polylactic acid microspheres was also reported [100, 101]. Hydroxyapatite (HA) particles were selected as they are biodegradable and hence thought to be good as a matrix for radiolabeling with 166Ho and developing a radiosynovectomy agent. An experimental knee joint lesion that closely resembles rheumatoid arthritis was developed in rabbits by injecting ovalbumin. Bioevaluation of the 166Ho-HA particles was carried out by injecting ~2 mCi (74 MBq) of the radiotracer in 200 μL of saline solution which contained ~8 mg of 166Ho-HA particles into the knee joints of New Zealand white rabbits. The 166Ho-HA particles were injected into the arthritis-induced knee joints as well as into knee joints of healthy rabbits. Images of the injected joints of the animals recorded at regular intervals using a gamma camera showed good retention of the radiopharmaceutical. Blood samples were collected from the animals and the blood activity was assayed in a scintillation detector. Experiments were also carried out under identical conditions in normal rabbits. In both the cases, it was observed that there was no significant extra-articular leakage of the injected activity over the study period of 96 h postinjection and the particles are retained within the synovial joints . About 185 MBq (5 mCi) of the labeled preparation in sterile saline was injected intra-articularly under fluoroscopic guidance to the affected knee joints of the patients. The knee is flexed three to four times after injection and the patients were kept under observation. Gamma camera imaging was recorded to ascertain the localization of the injected dose. More than 100 joints were studied in the clinical trial. Patients suffering from rheumatoid arthritis and hemophilia showed good response from the treatment. Seventy-five percent of the 50 patients treated with the new radiopharmaceutical showed significant reduction in pain.
Although not strictly related to oncological applications, the results of the above study are nevertheless interesting because they constitute the rationale for applying to cancer therapy some of the concepts of using 166Ho-labeled particles and of locoregional administration described above. In this regard, 166Ho-microspheres have recently been introduced as an alternative radiopharmaceutical for intra-arterial therapy in patients with liver tumors. The efficacy of radioembolization for the treatment of liver tumors depends on the selective distribution of radioactive microspheres to tumor tissue. The distribution of 166Ho-poly(L-lactic acid) microspheres can be visualized in vivo both by SPECT and by MRI. In a phase I clinical trial, Smits et al.  evaluated the safety and the maximum tolerated radiation dose (MTRD) of 166Ho-radioembolization in patients with liver metastases. Patients were treated with intra-arterial 166Ho-radioembolization in cohorts of three patients, with escalating aimed whole-liver absorbed doses of 20, 40, 60, and 80 Gy. Cohorts were extended to a maximum of six patients if dose-limiting toxicity occurred. Patients were assigned a dose in the order of study entry, with dose escalation until dose-limiting toxicity was encountered in at least two patients of a dose cohort. The most frequently encountered toxicities (including grade 1) were lymphocytopenia, hypoalbuminemia, raised alkaline phosphatase, raised aspartate aminotransferase, and raised gamma-glutamyltransferase, which were all noted in 12 of 15 patients. Stable disease or partial response regarding target lesions was achieved in 14/15 patients (93%, 95% CI 70–99) at 6 weeks and in 9/14 patients (64%, 95% CI 39–84) at 12 weeks after radioembolization. Compared with baseline, the average global health status and quality of life score at 6 weeks after treatment had decreased by 13 points (P = 0.053) and by 14 points at 12 weeks (P = 0.048). In all patients, 99mTc-macroaggregated albumin SPECT, 166Ho scout dose SPECT, and 166Ho treatment dose SPECT showed similar patterns of the presence or absence of extrahepatic deposition of activity. The authors concluded by stating that 166Ho-radioembolization is feasible and safe for the treatment of patients with unresectable and chemorefractory liver metastases, and it enables image-guided treatment. Clinical 166Ho-radioembolization should be performed aiming at achieving a whole-liver absorbed dose of 60 Gy .
Ytterbium-175 ( 175 Yb) is another low-energy β-emitter , which has been proposed for labeling bone-seeking agents to be employed for bone pain palliation. The low β-energy of 175Yb (480 keV) will induce minimum radiation dose to bone marrow and hence will allow administration of higher levels of radioactivity, to potentially obtain even a therapeutic efficacy similar to that demonstrated for 153Sm and 188Re. Similar to 153Sm, 175Yb can also be chelated to polyamino-poly phosphonate ligands .
Thulium-170 ( 170 Tm) conjugated with EDTMP is considered as an alternative to 89SrCl2 in palliative therapy of bone metastases. The use of 170Tm would induce low myelosuppression, since it emits lower-energy β-particles (Eβmax = 968 keV) than those emitted by 89Sr, and the gamma photons emitted can be used for scintigraphy to detect the accumulated activity and biokinetics at the target sites . The long half-life of 128.6 days could be advantageous in an approach based on mixed radionuclide therapy using 170Tm together with 153Sm, 177Lu, or 175Yb. Such combination could provide both early and sustained long-term pain relief to patients in early stage of disease.
Copper-67 ( 67 Cu) has favorable radiophysical characteristics for RIT, with a half-life of 3.4 days (well suited to pharmacokinetics of whole antibodies) and an emission of β-particles with energy comparable to that of iodine-131 and lutetium-177. Moreover, it emits gamma rays with energy suitable for imaging and a relatively weak abundance, thus reducing irradiation of patients and medical staff as that occurring with iodine-131. The production capacity of copper-67 is limited by a low cross section of the nuclear reaction, which needs high-intensity and high-energy proton beams. Only a limited number of accelerators in the world have such high energy and high intensity. This situation explains the quite limited number of clinical studies performed in the last 30 years.
Two groups of investigators carried out clinical studies in the 1990s in a limited number of patients. De Nardo and coworkers pioneered this field and compared clinical results obtained with the Lym-1 mAb against non-Hodgkin’s lymphoma (NHL) labeled with 67Cu, 131I, and 90Y, respectively [105, 106]. The authors concluded that the therapeutic indices (ratio of radiation doses to tumor relative to those to normal tissues) for 67Cu-2IT-BAT-Lym-1 and, to a lesser extent, for 90Y-2IT-BAD-Lym-1 were more favorable than those observed for 131I-Lym-1. The same conclusions were drawn in a clinical study comparing the anti-CEA mAb35 antibody labeled with 67Cu and 125I in patients with colorectal cancer . Copper-67-labeled mAb35 was more favorable than its radioiodine-labeled counterpart due to higher tumor-to-blood ratio; however, the authors observed a potential problem with 67Cu, i.e., nonspecific liver and bowel uptake.
In conclusion, copper-67 seems to be more suitable than iodine-131 and possibly than yttrium-90 for labeling mAbs for therapy, but currently lutetium-177 is a radionuclide of choice for RIT. Thus in the future, copper-67 should be compared to lutetium-177 in clinical studies when the production of large activities of copper-67 will be made possible by new high-energy/high-intensity accelerators.
Yttrium-90 ( 90 Y) undergoes ß-decay to zirconium-90 with decay energy of 2.28 MeV (average energy of 0.94 MeV) that allows for high-dose deposition with an average and maximum soft tissue penetration of 2.5 mm and 11 mm, respectively [108, 109, 110]. Yttrium-90 has a physical half-life of 64.1 h  which makes it amenable for a variety of targeted radiotherapy applications including 90Y-labeled colloid [112, 113], somatostatin receptor-targeting peptides [114, 115], tumor-targeting antibodies [116, 117], and resin/glass microspheres for selective/super-selective embolization of hepatic malignancies and metastases [118, 119, 120]. Regardless of the targeted delivery agent used, the selection of 90Y and its use for radiotherapy are complex and necessitate close collaboration among various medical specialties including nuclear medicine, interventional radiology, medical oncology, and radiation oncology . 90Y can be administered via direct injection into a space or cavity (e.g., radiosynovectomy), intravenously for peptide receptor radionuclide therapy (PRRT) and radioimmunotherapy (RIT), and intra-arterially for radioembolization (RE) therapy. Yttrium-90 emits pure β-particles, but lacks gamma photons, and therefore conventional scintigraphic imaging and assessment of the post-therapy distribution cannot be performed. This lack of gamma photons led to the development and use of surrogate gamma-emitting agents such as indium-111-labeled peptides to be used before PRRT and RIT or 99mTc-labeled macroaggregated albumin (MAA) used as a surrogate radiotracer for planning 90Y-microsphere radioembolization. Yet, using surrogate tracers may still be problematic as to the accuracy of predicting 90Y radiotheraphy effects in vivo and the precise post-therapeutic distribution [122, 123, 124, 125]. However, β-particle emission from 90Y produces bremsstrahlung photons which can also be imaged scintigraphically, although with poor-quality images . The 90Y bremsstrahlung photons are generated when the high-energy β-particle is emitted from the 90Y nucleus and then loses its kinetic energy while interacting with adjacent atoms. As the electron slows down, its kinetic energy is converted into the continuous energy spectrum of both primary and scattered photons with no dominant energy photopeak for conventional scintigraphic imaging (i.e., bremsstrahlung radiation).
However, it is important to note that, even if in an extremely small proportion (32 per million), some 90Y decay occurs through internal pair production that generates 511-keV annihilation photons that can be imaged in vivo using positron emission tomography (PET). This emission is in general too small to allow for efficient PET imaging within a reasonable acquisition time when a therapeutic 90Y-radiopharmaceutical is administered systemically and distributes throughout the body (as in the case of RIT or PRRT). Instead, when the therapeutic agent is administered loco-regionally and is retained at the site of administration (as in the case of radioembolization of liver tumors with 90Y-microspheres), then even this extremely small fraction of 90Y decay occurring through internal pair production is sufficient to acquire clinically useful PET images that are useful for dosimetric purposes .
Gold-198 ( 198 Au) is a β-emitter (β-max = 0.96 MeV) with a relatively short half-life (2.7 days). Thanks to its characteristics, 198Au has been used as a permanent implant either alone or as an adjunct to external beam radiation therapy [127, 128].
Recently, due to the development of nanotechnologies, a paradigm shift in the way diagnostic and therapeutic drugs are designed is occurring. This paradigm is based on achieving target specificity and increasing retention for significant improvement in the treatment of various tumors, including prostatic cancer [129, 130, 131, 132, 133]. Nanosized particles have extraordinary capabilities to image or treat cancers at the cellular and molecular levels [134, 135, 136, 137, 138, 139]. In this regard, gold nanoparticles (AuNPs) exhibit very high tumor uptake, due to their natural affinity to the leaky tumor vasculature, which is particularly pronounced during tumor’s neoangiogenesis [140, 141].
Chanda et al. have reported promising results in a preclinical model of prostatic cancer using gum arabic glycoprotein (GA)-functionalized gold nanoparticles (12–18 nm core diameter, 85 nm hydrodynamic diameter) loaded with 198Au (GA-198AuNPs). This agent exhibited high tumor affinity in severely compromised immunodeficient (SCID) mice bearing human prostate tumor xenografts. In this experiment, a single intratumoral administration of the β-emitting GA-198AuNPs (70 Gy) was performed, to achieve a calculated radiation dose of 70 Gy. This procedure resulted in clinically important cancer regression and successful control in the growth of prostate tumors over 30 days . This observation might constitute the basis for possible clinical translation of this agent in patients with newly diagnosed PCa who are candidate to surgery, in order to reduce tumor size before surgical resection. The study by Chanda et al. revealed no hematologic toxicity, thus confirming that the therapeutic efficacy of this treatment is associated with high in vivo tolerance.
Promethium-149 ( 149 Pm) is a radiolanthanide recently made available characterized by a moderate-energy β-emission (1.07 MeV, 95.9%) and a half-life of 2.21 days. 149Pm also emits a low abundance of an imageable γ-ray (286 keV, 3%) that might permit in vivo imaging after therapy. In a preclinical trial, three 149Pm-complexes with the DO3A-amide chelator with zero and three carbon spacers to the bombesin peptide analog BBN(7–14)NH2 were synthesized and characterized. The biological properties of the 149Pm-DO3A-amide-βAla-BBN complexes were compared with biodistribution studies in normal mice. Its in vivo behavior as the 149Pm-DO3A-amide-Ala-BBN conjugate is virtually identical to that of the 153Sm- and 177Lu-analogs. This supports the hypothesis that, given the appropriate chelate, the radiolanthanides are similar allowing fine-tuning of the nuclear properties necessary for a particular disease target. No in vivo tumor-targeting trials have as yet been carried out with 149Pm-DO3A-amide-Ala-BBN in patients; however, the fast biological clearance of this conjugate suggests that a longer spacer (five or eight carbon) may be necessary to increase the lipophilicity of the complex and reduce its clearance rate .
Rhodium-105 ( 105 Rh) is a reactor-produced radionuclide with properties suitable for radiotherapeutic applications. It is a moderate low-energy β-emitter (0.560 MeV [70%], 0.250 MeV [30%]) with a 36-h half-life, and it is available in “no-carrier-added” (NCA) concentrations. 105Rh also emits a low abundance of imageable γ-rays (306 keV [5%], 319 keV [19%]) that would allow in vivo tracking of the therapeutic dose. 105Rh is produced by a neutron capture reaction of an enriched 104Ru target to produce 105Ru, which decays to 105Rh with a 4.4-h half-life . The cis-[Rh(III)Cl2(2,5,8,11-tetrathiadodecane-1,12-dicarboxylic acid)]+ complex shows the most promising in vivo characteristics on which to base the development of a potential therapeutic radiopharmaceutical . For example, 105Rh can be simply conjugated with EDTMP, by heating for 30 min in boiling water, with >99% radiochemical yield. Ando et al. have tested this radiopharmaceutical in animals, and observed that 105Rh-EDTMP has fast blood clearance and selective uptake in the bone, with rapid elimination from irrelevant tissues; thus, it appears to have suitable features for palliative treatment of pain of bone metastases.
A few other radionuclides could be of interest for RIT. For example, the use of scandium-47 has been suggested by Pietrelli et al. . This radionuclide can be produced carrier-free by neutron irradiation of titanium-47. It decays into stable titanium by emitting low-energy β− with a half-life of 3.35 days, with the additional emission of a 159 keV γ-ray (68%) that is suitable for imaging. Very little has been published on scandium-47 because of a poor availability, but interest in scandium-44, a positron emitter, has revived interest in scandium-47 according to the β+-/β-radionuclide pair concept. Similarly as scandium-44, it may be efficiently complexed by several chelating agent, including DOTA .
Terbium-161 has also been proposed recently as a β-emitter of interest in targeted radionuclide therapy . It decays with a half-life of 6.88 days into stable dysprosium-161 by emitting low-energy β-particles. A very interesting feature of the element terbium is the possibility of using other terbium radionuclides, such as terbium-149 (which is an α-particle emitter), terbium-152 (a β+-emitter), and terbium-155 (a γ-emitter). All these radionuclides possess manageable half-life and decay properties, thus making it possible to perform β- and α-targeted radionuclide therapy, as well as PET and SPECT imaging with isotopes of the same element – according to the principles of theranostics.
It should be emphasized that the development of novel radiopharmaceuticals for therapy often includes several steps that imply the use of different radionuclides before identifying the most suitable ligand-chelator-radionuclide conjugate for clinical use. In general, proof-of-principle studies are based on the demonstration – obtained by imaging studies – that the candidate compound of interest accumulates efficiently and is retained at the tumor site sufficiently so as to predict high radiation dosimetry with a potential therapeutic effect. This demonstration can be achieved using the compound labeled with either a γ-emitter (for single-photon imaging) or a β+-emitter (for PET imaging). Some of the radionuclides described above decay by emitting both γ-radiation and β-particles (e.g., 131I, 153Sm, 177Lu), a feature that facilitates the studies on tumor targeting. When instead the therapeutic radionuclide is a pure α- or β-emitter, a surrogate γ- or β+-emitter must be used for imaging in the exploratory phase.
The example described here below will serve to illustrate the multifaceted approach that is being followed for developing a novel therapeutic radiopharmaceutical starting from a monoclonal antibody that has initially been employed primarily for diagnostic purposes in patients with clear-cell renal carcinoma. The murine G250 mAb was raised against the carbonic anhydrase isoenzyme 9 (CAIX), a transmembrane glycoprotein [150, 151, 152] that is expressed on the cell surface of the majority (>95%) of these tumors, which constitute approximately 80% of all renal cell carcinomas . The murine mAb G250 labeled with 131I was initially tested to explore its tumor-targeting potential. However, due to the formation of human anti-mouse antibodies (HAMA) , a chimeric variant of G250 (cG250) was constructed, with high affinity for the G250 antigen (Ka = 4 × 109 L/mol). Immunoreactivity of mAb cG250 with normal human tissues is limited to the upper gastrointestinal mucosa and gastrointestinal-related structures (bile ducts, pancreas) [155, 156]. Studies on the therapeutic potential of mAb G250 can be divided into studies with the “naked” antibody (administered alone or in combination with cytokines) and radioimmunotherapy (RIT) trials, respectively. In the first dose-escalating RIT study, 131I-G250 was administered to patients with progressing metastatic renal cell carcinoma (mRCC) . Hepatic toxicity occurred, mostly as the result of specific hepatic accumulation of the murine G250 mAb, which declined with increasing doses (thus suggesting saturation of the G250 binding sites). Nevertheless, hepatic toxicity was mild and not dose limiting, whereas dose-limiting toxicity was hematologic. Overall survival of patients treated with 131I-G250 appeared to be improved versus historic control patients (17/33 stable disease and two minor responses). However, high HAMA levels developed in virtually all patients, thus precluding retreatment, a problem that was overcome with the development of cG250.
Following the protein dose-escalation trial with 131I-cG250, which established the most favorable amount of protein for administration, a phase I 131I-cG250 activity escalation study was performed to establish dose-limiting toxicity . A pre-therapy imaging phase with 131I-cG250 (administered as a tracer activity of 222 MBq per 5 mg protein) was included with the purpose of selecting CAIX-positive patients for the therapeutic administration of 1.665–2.775 GBq 131I-cG250 (still with 5 mg protein). No hepatic toxicity was observed in this study, probably because the liver compartment was saturated. Dose-limiting toxicity of 131I-cG250 was established at 2.775 GBq/m2; the maximum tolerated dose was therefore set at 2.220 GBq/m2. An antitumor response was observed in two out of eight patients: one stable disease for 3–6 months and one partial response >9 months.
A fractionated dose RIT trial was subsequently designed, based on whole-body radiation-absorbed dose , under the rationale that in animal models fractionated RIT is more successful than a single administration schedule and is associated with lower toxicity. The primary target of this study was to determine the maximum tolerated whole-body radiation-absorbed dose. Fifteen patients with measurable mRCC were enrolled for this trial, during which it was found that dose-limiting toxicity was hematologic. Moreover, the total dose that could be delivered was relatively low.
In a subsequent study , two sequential high doses of 131I-cG250 (2.22 GBq/m2) were administered after a tracer activity of 131I-cG250 (185 MBq/m2) to ascertain effective tumor targeting; 29 patients with sufficient cG250 uptake received a first therapeutic dose of 131I-cG250. Patients who did not show a grade 4 hematologic toxicity received a second therapeutic administration 3 months later, with dose escalation from 1.110 to 1.665 GBq/m2. None of the 16 patients who completed the protocol (out of the 29 enrolled) showed an objective response, but five had stabilization of disease for 3–12 months. The low therapeutic efficacy observed in this trial was probably due to the quite large tumor burden present in these patients. In this regard, dosimetric estimates proved that therapeutic radiation doses (>50 Gy) could be achieved only for lesions smaller than 5 g .
Subsequent clinical studies have adopted 90Y or 177Lu to label cG250 for RIT trials. The results of a phase I/II trial with 177Lu-cG250 were recently published , a study that was conducted in parallel with a trial based on the use of 90Y-cG250 at Memorial Sloan Kettering Cancer Center (www.clinicaltrials.gov/NCT00199875). In the 177Lu-cG250 trial, 23 patients with progressive mRCC received a diagnostic activity of 111In-cG250 (185 MBq), to ascertain adequate tumor uptake; this was followed by administration of 177Lu-cG250 1 week later. The staring dose of 177Lu-cG250 was 1.110 GB/m2, with increments of 370 MBq/m2 (three patients/dose level). In the absence of grade 4 toxicity, patients were suitable for receiving a second (13/23) and a third cycle (4/23), at 75% of the dose level of the previous administration. Hematologic toxicity was dose limiting, and the maximum tolerated dose was 2.405 GBq/m2 . The majority of patients responded with stabilization of disease, while a partial response that lasted for 9 months was observed in one patient at the 1.850 GBq/m2 dose level. The tumor-to-red marrow dose ratio was higher for 177Lu-cG250 than for 90Y-cG250, indicating a wider therapeutic window for the 177Lu-conjugate. Stillebroer et al. concluded that RIT with 177Lu-cG250 may stabilize previously progressive metastatic ccRCC .
More evidence is needed to prove that cG250 imaging can be used to guide clinical management. As suggested by Divgi et al. , the role of cG250 imaging in influencing outcome would perhaps be best assessed in a clinical trial carried out in patients with small renal tumor mass and fewer associated comorbidities. Another issue that needs further investigation is choice of the radionuclide for imaging. 111In-cG250 SPECT may provide the same information as 124I-cG250 PET, with the advantage that it can be made available as an off-the-shelf product to be used on site. Other radionuclides (e.g., 89Zr) might provide a useful alternative for PET imaging. In summary, the cG250 mAb agent has shown significant tumor-targeting capacity; although the main clinical value of the compound seems at present to be its potential as a diagnostic agent, its use as a delivery vehicle for RIT requires further investigations – possibly regarding also choice of the best radionuclide for therapy.
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