β−
and future perspectives
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1. Introduction
Due to a lack of awareness and misconceptions regarding toxicity and costs radionuclide therapy of bone metastases is an underutilized treatment modality [1–5]. However, the targeted treatment with phosphonate-based radiopharmaceuticals in advanced cancer patients with a positive bone scintigram is an appealing example of theranostics with a high probabili- ty of improving quality of life [6–8]. Rhenium-188-HEDP (188Re-HEDP) might be a preferable compound for the targeted treatment of painful bone metastases. It can be prepared in a typical hospital pharmacy department, enabling fast and tailored therapy upon presentation of the patient [9,10].
As explicated in the General introduction (chapter 1), the central goal of the research described in this thesis was the bench-to-bedside development of 188Re-HEDP. We performed the phar- maceutical development, preclinical studies and clinical evaluation of this bone-seeking thera- peutic radiopharmaceutical. In this chapter the main outcomes of our investigations are sum- marized and discussed. Furthermore, some future perspectives are provided.
1.1 Phosphonate-based therapeutic radiopharmaceuticals
Chapter 2 is a review of the development of therapeutic phosphonate-based radiopharma- ceuticals and their investigation for the targeted treatment of painful bone metastases across several decades. We performed a systematic literature search to unlock research data on the pharmaceutical development, preclinical research and early human studies of these radio- pharmaceuticals. A total of 91 phosphonate-based therapeutic radiopharmaceuticals were identified, from which only six agents have been clinically used. Only 186Re-HEDP, 188Re-HEDP and 153Sm-EDTMP have been investigated in extensive clinical studies. 153Sm-EDTMP is the only compound that received marketing authorization in many countries. 188Re-HEDP is genera- tor-based, enabling on demand preparation. In India, 177Lu-EDTMP has recently been approved for clinical use.
The topics we explored in reviewing the literature on the development of these theranostic radiopharmaceuticals included their energy and range in tissue, physical half-life, suitabili- ty for imaging, precursor availability and quality, preparation and quality control aspects, stability, characterization and biodistribution.
To obtain a comparable radiation dose in a shorter time period, a high-energetic radionuclide (e.g. 188Re) is preferable. In view of availability, continuity of patient care and healthcare costs generator-based radionuclides, like 188Re, are advantageous. In a recent review comparing 11 radionuclides for palliative treatment of metastatic bone pain, 188Re is proclaimed the best option for medium to large sized metastases [11].
The composition and preparation conditions may be crucial for product quality and biodis- tribution of phosphonate-based radiopharmaceuticals. Although the molecular structure is important for the in vivo behaviour of these radiopharmaceuticals, characterization has been
the subject of only very few studies. Complexes of rhenium with phosphonates like 188Re- HEDP consist of different species of which the exact molecular structures and spatial orienta- tion are not known. Therefore, extensive research on the preparation and stability is required in each centre that prepares this radiopharmaceutical.
In order to exploit the full potential of therapeutic phosphonate-based radiopharmaceuticals, simple and reproducible preparation and quality control methods are essential. Proper un- derstanding of the radiochemistry of these agents is indispensable to design these methods.
Moreover, extensive biodistribution and dosimetry studies are required before these products can be administered to humans. The knowledge provided in this review should guide research on the development and potential approval of new promising agents.
This review demonstrates the difficulty of introducing newly developed radiopharmaceuti- cals into clinical trials. Legal and economical hurdles may contribute to the challenging route to marketing authorization [12].
1.2 Regulations for the preparation of radiopharmaceuticals
To enable clinical studies and introducing new pharmaceuticals adherence to appropriate regulations is essential. In Chapter 3 European legislation and guidelines on the preparation of radiopharmaceuticals are reviewed. The strict regulations on production and radiation safety may constitute a hurdle for their development and clinical use. Moreover, they may have a large budget impact and are not always appropriate for their production, due to some particularities of radiopharmaceuticals.
Therefore, we propose a risk-based approach for the implementation of these regulations and provide guidance on interpreting and applying Good Manufacturing Practice (GMP) for the small-scale production of radiopharmaceuticals, including cleanroom classification, air pressure regime, cleanroom qualification and microbiological monitoring. We present an algorithm to assess the risk of microbiological contamination of a radiopharmaceutical preparation process and propose corresponding GMP classification levels. We argue that the risk of carry-over of radiopharmaceuticals cannot be contained by pressure differences and that a simple cleanroom design is sufficient in most radiopharmacy departments. We propose a sterility assurance level of 10-2 for the preparation of radiopharmaceuticals that are administered within a working day, and suggest adopting less strict limits for environmental monitoring of microbial contamination than those provided in GMP Annex 1. Recent regula- tory documents seem to be more liberal than current legislation and appear to recognize the use of risk-assessment [13]. We state risk-assessment as the gold standard for implementation of appropriate quality assurance standards for the preparation of unlicensed radiopharma- ceuticals.
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1.3 Development of GMP-grade rhenium-188-HEDP
As 188Re-HEDP might be a preferable therapeutic radiopharmaceutical for the treatment of painful bone metastases, we developed a simple method for the preparation and quality control of GMP-grade 188Re-HEDP for its application in routine clinical practice and for support of clinical studies. In chapter 4 the entire bench-to-bedside development is presented. When we started to develop 188Re-HEDP in our hospital, no standardized preparation method was available. Standardization is crucial, since drug composition and preparation conditions are known to affect product quality, stability and efficacy. Also the production of sterile non-radioactive starting materials is described. These materials constitute a cold kit (con- taining the ligand HEDP, the reductant stannous chloride and the antioxidant gentisic acid), an ammonium perrhenate solution (carrier rhenium) and a sodium acetate solution (for pH correction). Furthermore, validation, stability and microbiological data are provided. Finally, hydroxyapatite binding assay results and biodistribution data of 188Re-HEDP in mice and in patients with bone metastases are presented.
We showed that our composition and preparation process result in a product with requisite quality, stability and biodistribution in mice. Scintigrams acquired 3 hours after adminis- tration of 188Re-HEDP to patients demonstrated only uptake in the predicted localizations.
Product quality review during the first year of routine use of 188Re-HEDP proved that the preparation method was robust. This completely validated method to prepare GMP-grade
188Re-HEDP may be transferred to other centres with sufficient numbers of patients who are eligible for bone-targeted radionuclide therapy.
1.4 Influence of carrier concentration
The above-mentioned development of 188Re-HEDP included investigations on the influence of non-radioactive (carrier) rhenium concentration. The presence of carrier rhenium in the reaction mixture is known to be required for adequate product quality and bone affinity of 188Re-HEDP, but the optimal carrier concentration was unknown when we started our research. In chapter 5 investigations on the influence of carrier concentration on radiochem- ical purity, in vitro hydroxyapatite affinity and in vivo bone accumulation of 188Re-HEDP in mice are presented.
The carrier concentration proved to influence both hydroxyapatite binding in vitro and bone accumulation in vivo. Hydroxyapatite binding was around 95% at carrier amounts between 0.1 and 20 µmol. The radiochemical purity of the complex was sufficient for clinical use at carrier amounts from 0.01 µmol up to 20 µmol. In the in vivo experiments in mice no bone accumula- tion was observed for the 188Re-HEDP formulation without carrier, and only soft tissue uptake was visible. A carrier amount of 0.01 µmol showed both bone and soft tissue uptake. With 10 µmol of carrier only bone uptake was detected and soft tissue uptake was absent.
Since sufficient radiochemical purity and hydroxyapatite affinity not always resulted in
acceptable in vivo bone accumulation of 188Re-HEDP, we hypothezise that the 188Re-HEDP-com- plex is only stable in vivo at higher carrier concentrations. We conclude that human adminis- tration of phosphonate-based radiopharmaceuticals should only be initiated after evaluation of animal biodistribution experiments.
1.5 Quality by design of the preparation method
Since preparation conditions may influence the product quality and in vivo behaviour of
188Re-HEDP, we investigated the effect of critical process parameters on product quality and stability. The methods and results are described in chapter 6. We adopted the quality by design (QbD) concept of the ICH Q8 (Pharmaceutical Development) guideline and followed a stepwise approach.
First potential critical process conditions were identified. Subsequently, these conditions were varied, investigating the impact of each variable on radiochemical purity and stability of the product upon dilution and storage. The acceptable ranges of the process conditions were established by boundary testing.
Only two boundary tests: starting with a larger eluate volume than in the standard prepara- tion method and a reaction time of 10 minutes resulted in inadequate radiochemical purity and stability. Furthermore, diluting the end product and storage at elevated temperature had a strong negative influence on product stability.
Our standard preparation method proved to fall well within the acceptable ranges, demon- strating the intrinsic robustness of both composition and preparation method. We showed that applying QbD principles is feasible for the small-scale preparation of radiopharmaceuti- cals. Mapping the acceptable ranges by validating the preparation process for each parameter while keeping all other variables constant is meaningful for assuring product quality.
1.6 Combination with taxanes
In order to investigate the potential added value of combining chemotherapy and 188Re, exploiting the radiosensitization concept, we explored the combined treatment of the taxanes docetaxel and cabazitaxel with 188Re in prostate carcinoma cell lines. In chapter 7 we report on the methods and results of this research. We investigated the cytotoxic effects of single and combined treatment with taxanes and 188Re in three human prostate carcinoma cell lines, using the colony-forming assay (CFA).
All CFA experiments showed significant dose-dependent cell growth inhibition for both the taxanes and 188Re. The half maximum effective concentration (EC50) values of the individual agents were similar to results from earlier studies. However, the EC50 values of 188Re were not reported before.
The combined treatment was studied at 0.25, 0.5, 1, 2 and 4 times the EC50 value of the agents.
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The interaction was investigated with a regression model, which was well capable of explain- ing the data. The model confirmed large negative effects on cell growth for the individual agents and proved significant additive effects of combination of the two taxanes and 188Re.
Although synergistic action might have been expected, this was not demonstrated. However, additivity is a good basis for further research. The type and extent of interaction may be less important for optimizing the therapeutic strategy due to other factors contributing to the final treatment effectiveness.
The outcome of this proof-of-mechanism study encourages the design of (pre)clinical in vivo studies (e.g. in an animal bone metastasis model) and may contribute to the optimization of the treatment of advanced prostate carcinoma.
1.7 Clinical benefit in routine clinical care
188Re-HEDP is an unlicensed radiopharmaceutical. Therefore, after introduction in the clinic, we evaluated the clinical benefit of treatment of prostate or breast cancer patients with painful bone metastases with this radiopharmaceutical in routine clinical care. This prospec- tive, observational practice study is discussed in chapter 8.
Clinical benefit was defined as a response in pain palliation (using visual analogue scale (VAS)-scores and corrected for opioid intake) or quality of life (QoL, using the EORTC QLQ-C30 questionnaire), along with acceptable haematological toxicity. In this study, 45 patients were evaluable for pain palliation and 47 for QoL.
A single injection of 188Re-HEDP resulted in an overall pain response rate of 69% and a signif- icant and relevant decrease of mean VAS-scores. The pain palliation efficacy of 188Re-HEDP in this study is comparable to that of external beam radiotherapy (EBRT).
This study is the first to evaluate the quality of life upon treatment with 188Re-HEDP. The overall QoL response rate was 68%, and a significant and relevant increase of the mean Global health status/QoL-scores was demonstrated.
Since haematological side effects were mild and transient, this study proves the clinical benefit of treatment of painful bone metastases with 188Re-HEDP and supports its application in routine clinical care.
The results in this ‘real world’ study may be influenced by the relatively large proportion of missing data. However, the measured pain palliation efficacy nicely corresponds to previous research. Moreover, a sensitivity analysis showed a pain response between 51% and 77%.
This satisfactory outcome may be predictive of the value of this treatment in routine clinical practice.
2. Future perspectives
As we have shown in chapter 2, a bone-targeting therapeutic radiopharmaceutical that is ideal in every aspect still does not exist. For future research, development of radiophar- maceuticals delivering their radiation dose over a short period of time, reducing possible myelotoxicity and allowing repeated administration and serial or even concurrent treatment with other cytotoxic drugs, should be stimulated. To obtain a comparable radiation dose in a shorter time period, use of a high-energetic radionuclide is preferable.
Although 188Re-HEDP has several advantages over other agents, the complexation of rhenium with phosphonates may result in different species with unknown molecular structures and spatial orientation. The species present in the final product may exhibit different biodis- tribution patterns. Future research should include characterization of complexes that are formed in vitro and their in vivo behaviour. These investigations may involve preparation of the non-radioactive complexes, followed by chromatographic comparison with the radio- active preparations. If the chromatogram is identical for both preparations, the non-radio- active species can be characterized by various methods, providing information regarding the molecular species formed during the radioactive preparation. In addition, the stability and fate of the administered complexes in vivo have only rarely been explored. Chromatographic analysis of the species present in blood, tissues, urine and stool at different time points after administration would provide useful information on the pharmacokinetics of targeting and non-targeting species.
With respect to the clinical application of 188Re-HEDP, some recommendations for optimizing the use and for future research can be made.
First of all, dose individualization could be explored. Up to now, fixed doses or doses based on total body weight are applied. However, it is known that renal function may impact elimina- tion of 188Re-HEDP, as it is mainly renally excreted [14] and it is unlikely that in obese patients total body weight correlates well with muscle mass. It might be worthwhile to investigate a dosing algorithm that is based on the number of lesions [15]. Moreover, a higher dose may be feasible and safe in patients with adequate platelet levels. In their dose finding study Palmedo et al. showed that the maximum tolerated dose (MTD) might be 4.4 GBq in patients with a thrombocyte count above 200•109/l [16]. A higher administered dose leads to a higher radiation dose to the metastases, which might improve the clinical outcome.
Secondly, repeated administration, e.g. every 8 weeks, should be pursued as long as this is feasible for the patient. The limited and transient side effects of 188Re-HEDP, maybe due to the short half-life of 188Re, make 188Re-HEDP suitable for iterated treatment. Furthermore, since the pain response is decreasing after 8 weeks, repeated injections could extend palli- ation. In our practice study, we observed similar pain responses in patients with repeated administration compared to the first treatment effect. Whether repeated treatment is able to extend survival, which is suggested in previous studies in relatively small numbers of
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patients [17,18], should be investigated in large prospective studies. These studies should prefe- rably be carried out in a head-to-head fashion compared to 223Ra-chloride, since this is the only radiopharmaceutical with proven overall survival benefit in a large phase III trial [19]. Although pain palliation was demonstrated in some studies with 223Ra-chloride, this was not investigated as one of the endpoints in the registration study. Therefore, this indication is not included in the registration label. Furthermore, a slight decline in quality of life was observed in the treatment period. We are currently planning to conduct a large randomized multicentre study comparing 188Re-HEDP and 223Ra-chloride at two endpoints: palliative effec- tiveness (pain palliation and quality of life) and overall survival. This study should provide a major contribution to determining the real value of 188Re-HEDP and to its positioning in the arsenal of radiopharmaceuticals for the treatment of bone metastases.
Thirdly, potential integration with other modalities should be explored. Combining 188Re-HEDP with chemotherapy (e.g. taxanes) may augment the treatment efficacy, which was demon- strated previously for the combination of 153Sm-EDTMP and docetaxel [20,21]. In a phase I study, the feasibility and safety of sequential therapy with docetaxel and 186Re-HEDP was showed
[22]. Results of phase II studies of the administration of docetaxel and cabazitaxel alternated with 188Re-HEDP are pending. However, no in vivo data on concurrent application of taxanes and 188Re-HEDP are available. Since significant additivity was clearly demonstrated in our study in prostate carcinoma cell lines, the design of (pre)clinical in vivo studies is warranted to explore the potential added value of concomitant treatment of taxanes and 188Re-HEDP.
These studies could be carried out using 188Re-HEDP in an animal bone metastasis model.
However, as satisfactory results have been demonstrated in phase I and II studies combining
153Sm-EDTMP and docetaxel using different schedules [20,21,23,24],the design of phase I/II studies exploring the safety and efficacy of concurrent administration of 188Re-HEDP and taxanes may be considered as well. Also combination with EBRT, treating the largest lesions selectively with EBRT and smaller or invisible lesions with 188Re-HEDP, may be worthwhile. In order to optimize treatment effectiveness, even combination with other radiopharmaceuticals (e.g.
223Ra-chloride) might be considered and investigated.
Fourthly, next to examining pain palliation, in future studies additional (surrogate) response parameters should be monitored, like prostate specific antigen (PSA), alkaline phosphatase (ALP) and bone markers, which may provide valuable information on the treatment effec- tiveness of 188Re-HEDP. The use of recently introduced PET-tracers, like 18F-methylcholine, 18F- PSMA and 68Ga-PSMA, may be of added value for treatment response monitoring as well [25–28].
Lastly, treatment in an earlier stage of the disease should be studied [3]. Usually, radionuclide therapy is applied as a last resort in the end stage of advanced cancer. From studies with
89Sr-chloride it is known that early treatment may result in a better outcome than a later start of the therapy [15,29]. This could be explained from a dosimetric point of view: small metas- tases are ablated better than larger lesions [30,31]. Complete eradication of small metastases
or prevention of lesions to grow might improve survival [31]. Moreover, patients may sustain (multiple cycles) of radionuclide therapy better in an earlier phase of their disease. Due to the short half-life of 188Re-HEDP, early administration does not preclude other treatment options in later stages of the disease. Since treatment with 188Re-HEDP is very safe, this opens pos- sibilities to extend its range of use and to investigate its application earlier in the course of the disease.
Next to the optimization of the clinical use of 188Re-HEDP, it would be interesting to compare its application with the use of therapeutic radiopharmaceuticals based on the PSMA- motif. 177Lu-PSMA has already been applied in clinical studies [32]. Although the labelling of PSMA with 188Re has been published [33], no clinical use of this agent has been reported up hitherto. Again, rapid preparation of this radiopharmaceutical would be possible, when a
188W/188Re-generator is already present in the radiopharmacy department. No literature on the labelling of PSMA with the alpha emitting radionuclide 223Ra has been identified up to now, but a case report on treatment with the alpha emitter 225Ac-PSMA was published recently [34]. When PSMA-based therapeutical radiopharmaceuticals become available for clinical use, they might be combined with bone-targeting agents as well, since they are directed to additional targets. This might further improve the clinical benefit of radionuclide therapy in patients with metastatic bone disease.
References are listed after the Dutch summary (page 222).
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