68Gallium- and 90Yttrium-/177Lutetium: “theranostic twins” for diagnosis and treatment of NETs

Abundant expression of somatostatin receptors (SSTR) is frequently identified in differentiated neuroendocrine tumors and may serve as potential target for diagnostic imaging and treatment. This article discusses the “theranostic approach” of SSTR-targeting compounds including an overview of its role for diagnosis, staging and restaging, discussing its way to being established in clinical routine, and giving an outlook about further potentially relevant developments.


Introduction
Recent developments in cancer research have resulted in a wide spectrum of therapies. Due to the large heterogeneity of patients and tumors, there is an increasing demand for personalized medicine. Introduction of the theranostic approach is based on the idea of selecting patients through a diagnostic study indicating whether a patient will benefit from a therapy or not.
The theranostic principle has been applied in the field of nuclear medicine for more than 60 years and 131 I and 89 Sr are still established in daily clinical routine. 131 I was first discovered in 1938 by Seaborg and Livingood at the University of California Berkeley. Already in 1946, Seidlin et al. [1] reported the therapeutic use of 131 I for patients with metastasized adenocarcinoma of the thyroid. Since then the combination of radiolabeled iodine for diagnostic imaging and therapy represents an established and accepted ''theranostic'' approach. In 1942, Pecher demonstrated that 89 Sr accumulates in bone tumors in animals; and later, these findings were translated to humans and 89 Sr was widely used in cancer patients [2,3]. More recently, a number of new theranostic approaches have been introduced including 123 I-/ 131 I-labeled Metaiodobenzylguanidine (MIBG) for diagnosis and treatment of neuroblastoma [4][5][6]. An additional theranostic pair consisting of 123 I-and 131 Imetomidate for diagnosis and treatment of adrenocortical carcinomas (ACC) has been established at the University Hospital of Würzburg. The diagnostic scan with 123 Imetomidate allows for the prediction whether an ACC patient will benefit from 131 I-metomidate radionuclide therapy after determination of an individualized dose by dosimetry [7,8]. First, clinical data revealed tumor control in half of the patients undergoing 131 I-metomidate therapy [7,9].
In this article, we focus on the recently introduced and clinically translated ''theranostic approach'' of somatostatin receptor (SSTR) targeting compounds for treatment of patients with neuroendocrine tumors (NETs). We will present an overview of its role for diagnosis, staging and restaging, discussing its way to being established in clinical routine, and giving an outlook about new developments.

Theranostic twins in NET
The term ''theranostics twins'' includes diagnostic molecular imaging followed by a personalized treatment decision based on the predictive value of the diagnostic scan. Translated to NETs ''theranostic twins'' relate to the pairing of 111 In-/ 68 Ga-labeled diagnostic imaging and 90 Y-/ 177 Lu-labeled treatment compounds targeting the SSTR (Fig. 1).
The rationale of this theranostic approach bases on the fact that well-differentiated NETs usually overexpress SSTR on their tumor cell surface which may serve as diagnostic and therapeutic targets [10]. Five different membrane-bound receptors have been evaluated: SSTR1, SSTR2, SSTR3, SSTR4 and SSTR5. Out of these potential targets usually SSTR2 is addressed for diagnosis and therapy. Using radiolabeled somatostatin analogs for functional imaging the diagnosis of NETs can be confirmed and the tumor burden (metastases) assessed. These nuclear medicine imaging procedures provide essential information about SSTR density, which is relevant for treatment decisions, and selecting patients for treatment with peptide receptor radionuclide therapy (PRRT) [11]. In general, PRRT is recommended in inoperable, metastasized cases expressing SSTR on tumor cell surface [12]. 111 In-labeled twins 111 In-DTPA-octreotide, binding to SSTR2, was the first and most widely used radiopharmaceutical for detecting and staging NETs [13]. Planar images should be performed and, if available, single photon emission computed tomography (SPECT) after 4 and 24 h [14]. With an overall sensitivity of 80 % 111 In-DTPA-octreotide scintigraphy (including SPECT) seems to be an effective method to detect tumor burden [15]. However, reduced detection rate has been reported in small and deep-seated lesions, even when using hybrid SPECT/CT scanners [16]. Historically verified, 111 In can not only be used for diagnosis and staging of NETs, but also for treatment: treating 40 patients by using the Auger electron emitting radionuclide 111 In, partial remission (PR) was reported in one patient, minor remission in six patients and stable disease (SD) in 14 patients [17]. In another study, only two of 27 patients (8 %) showed imaging-based morphological PR [18]. Due to the short particle range and the resulting limited tissue penetration, tumor regression only rarely occurred [19].
Thus, in the previous decade, the use of 111 In was abandoned and replaced by the following new theranostic twins.  All these compounds bind with a high affinity to SSTR2 on the tumor cell surface [16]. Compared with 111 In-DTPAoctreotide, 68 Ga-DOTATATE PET/CT was found to be the superior functional imaging modality especially in small lesions with a low density of SSTRs because of its higher resolution [20]. Initial results were confirmed by Haug et al. in a larger patient cohort. Offering better imaging properties, 68 Ga-SSTR PET/CT was able to depict NETs with an accuracy of 87 % [21] (Fig. 2). In 2010, conventional imaging's such as CT or endoscopic ultrasound were compared to PET/CT in 90 patients. SSTR PET/CT led to a modification of staging in 29 % and management of therapy in 76 % of patients [22]. Additionally, PET is more patient friendly since it can be done as a single imaging session performed within 30-60 min after injection [16] as opposed to multiple day imaging required for 111 In-DTPAoctreotide scintigraphy.
Currently, the most commonly used isotopes for treatment intended radiolabeling of somatostatin analogs are the ß-emitting isotopes 90 Y with DOTATOC or ß-and y-emitting 177 Lu with DOTATATE. Otte et al. [23] reported treatment in a palliative setting of 29 patients with advanced SSTR-positive tumors by administering at least 3.7 GBq of 90 Y-DOTATOC in 127 single treatment cycles. Only three patients suffered from progressive disease. Waldherr et al. [24] injected 7.4 GBq 90 Y-DOTATOC (4 treatment cycles, time interval of 6 weeks) resulting in 5 % complete responses (CR) and 18 % PR. The most common reported side effect was renal impairment with a decline in creatinine clearance of 7.3 % per year [25].
However, 177 Lu-DOTATATE seems to be better tolerated than 90 Y [25]. Thus, it is also routinely used as the ''theranostic twin'' to 68 Ga. 310 NET patients treated with 27.8-29.6 GBq (4 treatment cycles, time interval of 6-10 weeks) showed a survival benefit of 40-72 months from diagnosis in comparison to historical controls [26]. Quality of life improved significantly after 177 Lu-DOTA-TATE [27] and even in patients undergoing repeat salvage PRRT a median PFS of 13 months was reported [28]. At least two treatment cycles should be performed because PFS seems to be determined by the injected dose [29]. The most important delayed side effects significant renal impairment and myelosuppression occur in approximately 1 % of patients [30,31]. Amino acid (AA) solutions are recommended directly prior and during PRRT to reduce renal absorbed dose and subsequent damage to the renal parenchyma. Treating physicians should be aware of potentially life-threatening acute hyperkalemia which can occur as an acute side effect, of administering AA [14,32].
Because of deeper tissue penetration the high energy 90 Y beta emitter is recommended for larger lesions, whereas the lower energy 177 Lu should be administered for smaller lesions. A combination of 90 Y-/ 177 Lu-DOTATATE demonstrates better therapy response in comparison to single-injection of 90 Y-DOTATATE by providing a significantly longer median OS [33].
The term ''theranostics'' emphasizes the inseparability of diagnosis and therapy. However, individual treatment planning determining prognosis should be part of this concept. 68 Ga-DOTATATE PET/CT can also predict progressionfree-survival (PFS) in patients undergoing PRRT: Evaluating the early prediction of clinical outcome 3 months after the initial treatment cycle, the tumor-to-spleen SUV ratio was able to independently predict the time-to-progression in patients undergoing SSTR PET/CT [34].
In summary, the diagnosis and therapy monitoring of NETs can be assessed by using functional imaging with different somatostatin analogs. 111 In-labeled twins were used in the last decade showing a good overall sensitivity and treatment effects. However, in small and deep-seated lesions, 68 Ga-labeled radiopharmaceuticals for PET are superior, even superior to stand alone contrast-enhanced multislice CT [35]. For its twin, 177 Lu or 90 Y DOTATATE/ DOTATOC, good results for overall survival, improvement of quality of life but also less side effects were demonstrated [26,27].

Theranostic approach in clinical routine
Nowadays, the most commonly used ''theranostic twins'' for treatment of NETs are 68 Ga and 90 Y-/ 177 Lu-DOTA-TATE/DOTATOC showing higher response rates and improvement of life quality [14].

Labeling of the twins
Preparation of radiopharmaceuticals should be performed according to the current regulations on radiation protection and guidelines on Good Radiopharmacy Practice ensuring strict hygiene requirements [36]. 68 Ga PET is usually used for pre-therapeutic proof of target expression, for staging and follow-up of patients. The half-life of the 68 Ga isotope is favorable with 68 min. The use of a 68 Ge/ 68 Ga-generator ensures a continuous cyclotron-independent supply with 68 Ga [37]. The only stable chemical form of Ga at physiological conditions is Ga 3? which still needs a bifunctional chelating agent (BCA) to bind to a target vector such as peptides or proteins [38]. Since the labeling process can be performed just prior to the PET/CT examination it is possible to inject an optimal amount of radioactivity. 177 Lu is produced by a nuclear reactor in two different ways: via ''direct pathway'' (irradiation of 176 Lu) or via ''indirect pathway'' (irradiation of 176 Yb producing 177 Yb decaying to 177 Lu). A specific activity of 37-74 MBq 177 Lu per microgram is generally recommended [14]. The method of synthesis is widely described in literature [39][40][41].
Therapeutic procedure Like any other nuclear medicine therapy, PRRT has to be performed according to the local legal and ethical requirements and a recommendation of a multidisciplinary tumor board is desirable. The ward of the nuclear medicine department must provide trained staff including physicians, radiochemists and medical physicist experts. General radiation safety arrangements are mandatory. However, official regulations with special focus on radionuclides used for PRRT do not exist; in summary, these requirements differentiate from one country to another [14].

Toward intercontinental patient care
In Japan and other non-European countries, PRRT using effective SSTR-targeting compounds like 177 Lu-DOTA-TOC are currently not available. To implement the entire ''theranostic'' concept, the use of the ''theranostic twins'' but also individual tailored treatment planning prior to PRRT is indispensable. To overcome this shortcoming, the Würzburg department of nuclear medicine among others provides special medical services for patients which do not have access to PRRT in their home countries including patient preparation at their home institution (e.g. renal scintigraphy, 111 In-DTPA-octreotide scintigraphy). Additionally, management of transfer to Germany is provided. If required also in-house renal scintigraphy, measurement of blood values (with special focus on renal/hematological parameters) but also the more sensitive 68 Ga-DOTATOC PET/CT scan proving receptor expression on tumor cell surface can be provided. To avoid environmental radiation exposure after PRRT as well as to guarantee a better posttherapy monitoring, the patients are constrained to stay hospitalized for a total of 3-5 days. After being discharged, patients are encouraged to avoid contact to small children and pregnant persons for the next 5-7 days; special paperwork to allow for a return flight will be provided.
Dose estimation as part of ''theranostic'' concept to minimize toxicity The term ''theranostic twins'' does not only refer to the use of peptides labeled with diagnostic or therapeutic radionuclides, but also emphasizes the personalized patient preparation prior to and after PRRT. The kidneys are the dose-limiting organ in PRRT and therefore kidney function has to be assessed prior to therapy by laboratory tests, 24 h urine collection and/or renal scintigraphy [14]. Van Binnebeek et al. reported in a case presentation as well as in a prospective phase II trial on the role of individualized dosimetry-based activity reduction of 90 Y-DOTATOC to prevent severe and rapid kidney function deterioration by maximizing the delivered tumor dose and minimizing the biological effective dose (BED) to potential risk organs (kidney, bone marrow). The authors used 111 In-pentetreotide for dose estimation and reported a maximal tolerable kidney dose of 37 Gy BED as the threshold to avoid severe loss of kidney function in 90 Y-DOTATOC [42,43].
Helisch et al. [44] demonstrated that both 86 Y-DOTATOC (chemically identical to 90 Y) and 111 In-pentetreotide are feasible to pre-therapeutically calculate the cumulative organ and tumor doses identifying patients with high radiation burden to the kidneys. Concerning post-therapeutic dosimetric approaches, the following data has to be collected up to 3 days after PRRT: urine, blood, whole-body-scans, planar images but also SPECT alone/ fused-SPECT/CT [14]. Software tools like OLINDA/ EXM using this equation provide internal dose information and especially dose calculations [45]. However, a general recommendation for performing pre-/post-therapeutic dosimetry is not given. Due to its y-emission, posttherapeutic scintigraphy of 177 Lu-DOTATATE allows staging/imaging and dosimetry by using the same radiolabeled tracer. Thus, during the first treatment cylce of 177 Lu-DOTATATE, a post-therapeutic dosimetry might be useful [14,46].

Outlook
In general, PRRT is recommended for inoperable, metastasized NETs expressing SSTR2 on the tumor cell surface [12].

Radiosensitizing, SST receptor expression increase and neoadjuvant treatment strategies
The synergism of radiolabeled compounds with chemotherapy may lead to higher efficacies. Claringbold et al. [47] reported that the addition of the radiosensitizing chemotherapeutic drug capecitabine resulted in tumor control in 94 % of patients without severe toxicity. The same research group administered 177 Lu-octreotate in combination with capecitabine and temzolomid in advanced low-grade NETs demonstrating 2-year survival rate of 90 % [48].
Furthermore, receptor expression on tumour cells could be increased by addressing different molecular targets prior to PRRT. This might guarantee a higher saturation of SST receptors expressed on the tumor cell surface which could lead to higher BED. A Swedish research group reported a higher uptake of 177 Lu-DOTA-Tyr 3 -octreotate in externalbeam irradiated small cell lung cancer cells which might be caused by an up-regulation of SSTR2 [49]. These findings may constitute a promising treatment option in the future.
On a final note, the use of PRRT in a neoadjuvant setting to downstage/downsize inoperable patients could be beneficial but has not been studied sufficiently to date.
Locoregional procedures in case of discontinuing PRRT Radioembolization with 90 Y microspheres has been described as a safe and effective treatment option in unresectable cancers of the liver [50]. Due to their hypervascularity liver-dominant NET metastases are well-suited for locoregional therapeutic procedures such as radioembolization (selective internal radiation therapy, SIRT) but also transarterial chemoembolization (TACE) combining the cytotoxic effect of intra-arterial chemotherapy with an embolizing ischemic approach [51]. Injection of 90 Yttriummicrospheres into liver metastases as salvage therapy after PRRT resulted in a median OS of 29 months [52]. TACE and transarterial embolization (TAE) demonstrated the same PFS of 36 months in patients suffering from advanced NET. However, less side effects are described for TAE [53].

''High dose'' approach in PRRT
Ongoing studies are trying to determine the role of dosimetry for minimizing kidney and bone marrow damage by potentially offering the possibility of increasing the administered activities. Currently, up to a maximum of 7.4 GBq per treatment cycle is regarded as safe [54]. Using the absorbed dose to ''risk organs'' as the limiting factor, treatment with 177 Lu-DOTATATE could be individualized Ann Nucl Med (2015) 29: 1-7 5 to avoid under-/overtreatment, minimize side effects and to be able to administer the maximum activity per treatment cycle. Forrer et al. [55] reported high inter-patient variability of bone marrow absorbed doses. Thus, individual dosimetry and a personalized ''high dose'' approach appear feasible. Results of standard PRRT (by using 7.4 GBq 177 Lu-DOTATOC) according to practical guidance [14] could be compared to a tailored ''high dose'' treatment by injecting the maximum tolerable dose per cycle. Pre-therapeutic dosimetry assesses the individual kinetic behavior in every patient which might be helpful in maximizing the cumulative dose but also optimizing the BED.

Conclusion
In summary, PRRT is an effective and safe treatment option for patients suffering from advanced NET. The ''theranostic twins'' 177 Lu-and 68 Ga-DOTATATE are routinely used in clinical practice with well-established response rates. Ongoing prospective multicenter trials such as NETTER-1 will hopefully confirm these results in larger patient populations awarding PRRT a higher clinical acceptance [56]. The importance of dose estimation as part of the ''theranostic'' concept and also pre-therapeutic upregulation of SSTR expression should be kept in mind as potential innovations of PRRT [42,43,49].