Molecular Imaging Companion Diagnostics

C H A P T E R

10 Molecular Imaging Companion Diagnostics Kathryn M. Tully1,2, Nicholas B. Sobol1, Patrı´cia M.R. Pereira1 and Jason S. Lewis1,2,3,4 1

Department of Radiology, Memorial Sloan Kettering Cancer Center, New York, NY, United States 2Department of Pharmacology, Weill Cornell Medical College, New York, NY, United States 3Department of Radiology, Weill Cornell Medical College, New York, NY, United States 4Program in Molecular Pharmacology, Memorial Sloan Kettering Cancer Center, New York, NY, United States O U T L I N E 10.1 Introduction

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10.2 Nuclear Imaging

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10.3 Molecular Imaging With Small Molecules 204 10.3.1 Small Molecule Imaging for Diagnosis: 18F-fluorodeoxyglucose (18F-FDG) 204 10.3.2 Small Molecule Imaging for Patient Staging and Endoradiotherapy: 123I-Labeled Metaiodobenzylguanidine (123IMIBG) 205 10.3.3 Small Molecule Imaging to Determine Intertumor Heterogeneity and Predict Therapy Response—18Ffluoroestradiol (18F-FES) 207

Companion and Complementary Diagnostics DOI: https://doi.org/10.1016/B978-0-12-813539-6.00010-9

10.3.4 Small Molecule Imaging to Assess Pharmacodynamics of Novel Therapeutics: 18 F-fluorodihydrotestosterone (18F-FDHT) and Enzalutamide 10.3.5 Small Molecule Imaging to Assess EGFR Mutations for Targeted TKI Therapeutics 10.4 Peptides for Molecular Imaging 10.4.1 Somatostatin Receptor Targeting 10.4.2 Gastrin-Releasing Peptide Receptor Targeting 10.5 Molecular Imaging Using Antibodies 10.5.1 Antibody Fragments

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© 2019 Elsevier Inc. All rights reserved.

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10.5.2 HER Targeting Antibodies 10.5.3 Carbonic Anhydrase-IX Targeting Antibodies 10.5.4 Vascular Endothelial Growth Factor Targeting Antibodies 10.5.5 Immunopositron Emission Tomography to Detect Brain and Bone Metastases

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10.5.6 Programmed Death-Ligand 1 Targeting Antibodies 10.5.7 Shedding Targets and Pretargeting

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10.6 Conclusion

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References

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10.1 INTRODUCTION The biological heterogeneity of tumors was observed as early as 1977 [1,2]. Since then, an enormous amount of data has been collected to support this initial observation and understand the clinical implications of tumor heterogeneity. Tumoral heterogeneity can exist between patients, within the same tumor, and between a primary and metastatic lesion in the same patient. This heterogeneity manifests itself in different ways, such as immunogenicity, cell surface receptors, growth rate, and antigenic properties [3,4]. In addition to initial heterogeneity, expression of tumor biomarkers [e.g., epidermal growth factor receptor (EGFR), HER2] and biomolecular processes (e.g., glycolysis) are dynamic and change during tumor development and/or in response to therapy. Despite this, protein or genetic analyses of tumor biopsies are the primary method of diagnosis and staging for most types of cancer. Not only are tumor biopsies invasive and slightly increase the risk for neoplastic seeding but also biopsy results can be misleading because of tumor heterogeneity and dynamic processes [5]. Biopsy in patients with metastatic disease poses further complications, as trying to biopsy every lesion could be difficult and impractical. In the era of precision medicine, there is a growing need for noninvasive diagnostic assays which can determine a patient’s eligibility for a given targeted therapy, as well as monitor a patient’s response. Because of the shortcomings of tumor biopsies, molecular imaging modalities have been developed as companion techniques to noninvasively diagnose and assess disease extent. The Society of Nuclear Medicine defines molecular imaging as “the visualization, characterization, and measurement of biological processes at the molecular and cellular levels in humans and other living systems” [6]. Importantly, imaging modalities can simultaneously provide information on every lesion within a patient while remaining noninvasive. Common types of single imaging modalities include computed tomography (CT), magnetic resonance imaging (MRI), ultrasound, positron emission tomography (PET), and single-photon emission CT (SPECT). Imaging techniques can also be combined (multiplexed imaging) in order to further enhance their ability to diagnose and stage patients as well as plan targeted treatments and monitor their efficacy [7,8]. For example, PET/CT imaging combines the molecular information of a PET scan with the anatomical information of a CT scan. This chapter will focus on how nuclear imaging (PET and SPECT) with radiolabeled small molecules, peptides, and antibodies can be used to noninvasively obtain molecular information within the context of oncology, and how they have

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potential as companion diagnostics to provide additional information to conventional diagnostic methods.

10.2 NUCLEAR IMAGING PET and SPECT are widely used methods in molecular imaging of cancer due to their high sensitivity, allowing detection of imaging agents in the nanomolar range. Briefly, PET imaging works by administration of a targeting vector bound to a positron-emitting radionuclide (e.g., fluorine-18, gallium-68, zirconium-89, etc.) After positron emission, the positron will travel until it collides with an electron in the surrounding tissue and annihilates, converting the positron into two photons (511 keV) that are emitted 180 degrees apart [9]. When the photons reach the PET detectors simultaneously, the PET scanner registers this as a “coincidence event” which is then converted into a tomographic image. SPECT imaging is similar to PET, but instead of a pair of photons, the detector registers signals from single photons of specific energy (100
200 keV) to reconstruct a tomographic image [9]. In the tomographic image, the intensity of the signal will be proportional to the amount of the targeted receptor and/or biochemical process. Designing an imaging agent requires a thoughtful combination of a targeting vector with an appropriate radionuclide. The targeting vector provides specificity for a molecular pathway or target that is overexpressed, selectively expressed, or mutated in the disease of interest, and types of targeting vectors include small molecules, peptides, antibodies, antibody fragments, nanoparticles, etc. The localization of the cellular target helps determine what type of vector should be selected, as an extracellular target might utilize an antibody while an intracellular target might require a small molecule. After the targeting vector is determined, the appropriate radionuclide can be selected. The choice of radionuclide is crucial, as the nuclear half-life of the tracer should match the pharmacokinetic profile and biological half-life of the targeting vector. For example, a small molecule with a short biological half-life and fast pharmacokinetics might utilize the PET radionuclide fluorine-18, while an antibody that remains in circulation longer and takes more time to accumulate at the tumor site might require a longer lived radionuclide, such as zirconium89. Depending on the radionuclide and the chemical composition of the targeting vector, a chelator may need to be incorporated into the imaging agent to bind the radionuclide to the targeting vector. The chemical structure of some small molecules can be directly altered to incorporate the radionuclide [e.g., 18F-fluorodeoxyglucose (18F-FDG)], while peptides or antibodies usually require a chelator [e.g., thedesferrioxamine (DFO) chelator for zirconium-89]. By binding a PET or SPECT agent to a targeting vector, the location of the imaging agent can be tracked over time, providing insight into the expression of the cellular target that the vector is binding, uptake of the vector, and changes in the cellular target over time. Within oncology, understanding the characteristics of a cellular target can help provide a more specific cancer diagnosis, evaluate inter- and intratumor heterogeneity, select patients for target-specific therapies, aid in drug discovery and development, and more quickly follow disease progression in response to therapy [10,11]. Molecular imaging has enormous potential in a variety of disease types, particularly within oncology as personalized medicine continues to revolutionize the field.

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10.3 MOLECULAR IMAGING WITH SMALL MOLECULES 10.3.1 Small Molecule Imaging for Diagnosis: 18F-fluorodeoxyglucose (18F-FDG) Often referred to as the “gold standard” of molecular imaging, perhaps the most widely used small molecule in nuclear imaging is 18F-FDG. 18F-FDG is utilized within oncology for a wide variety of cancer types and applications, such as tumor staging, detecting tumor recurrence, presurgical and/or external beam therapy planning, and evaluating response to therapy [12,13]. 18F-FDG also has numerous applications in other fields, such as infection and various types of inflammation [14]. As a radioactive analog of glucose, 18F-FDG is transported into cells by glucose transport protein 1 and phosphorylated to 18F-FDGP by hexokinase. Like glucose-6-phosphate, this phosphorylation inhibits 18F-FDGP from being transported back out of the cell. However, the fluorine that replaces the hydroxyl group prevents further metabolism of the 18F-FDGP, leading to an accumulation in highly glycolytic cells. Utilizing 18F-FDG for cancer imaging relies on the Warburg effect, in that many types of malignant cells are much more glycolytically active than healthy cells and therefore will take up more 18F-FDG than healthy tissue [12]. This increased uptake produces a much higher PET signal in malignant tissue relative to most healthy cells (Fig. 10.1). While imaging with 18F-FDG is a mainstay of radiology and is extremely valuable, there are a number of limitations with using 18F-FDG. One issue with PET imaging using 18FFDG is that not all primary tumor types or metastatic sites exhibit increased glycolytic activity, so an 18F-FDG PET scan would not identify the malignant tissue as such [15]. Even within the same tumor, this glycolytic phenotype is highly heterogeneous. An additional issue with 18F-FDG imaging is that high glycolytic rates are also seen in various healthy organs (e.g., the brain) making it difficult to discriminate malignant from nonmalignant tissue in this context [12]. Inflamed or infected tissue can also exhibit high glycolytic activity, so an 18F-FDG scan might not be able to discern between a tumor and inflammation. An example of this is in pancreatic cancer, where there are conflicting findings about 18F-FDG’s ability to differentiate between inflammation due to pancreatitis versus pancreatic malignancy [16,17]. Even in the setting of a diagnosed cancer, 18F-FDG is only informative of cancer’s metabolic state; it does not provide information on specific protein expression, which can be informative of vulnerabilities within cancer and can be important in patient selection for targeted therapies. Despite 18F-FDG being a valuable tool in cancer imaging, it has considerable shortcomings in the molecular information it can provide and thus is often best combined with other types of imaging. Like 18F-FDG, many other small molecule imaging agents are radioactive analogs to endogenous small molecules. To avoid large-scale screening libraries, molecular imaging agents can be designed to mimic natural substrates in order to exhibit similar binding affinity and specificity to receptors or targets of interest. In addition, small molecule tracers are often less expensive to produce than biological tracers, making them a costeffective alternative for a molecular imaging companion diagnostic. Small molecule tracers can be utilized for a variety of applications, and while this review is not exhaustive, the following examples have been selected for cancer staging, patient selection for endoradiotherapy, determining inter- and intratumor heterogeneity, predicting therapy response, and earlier detection of tumor recurrence.

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FIGURE 10.1

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F-FDG PET images of a patient with lung adenocarcinoma before treatment [12]. PET, Positron emission tomography; 18F-FDG, 18F-fluorodeoxyglucose. Source: Zhu A, Lee D, Shim H. Metabolic positron emission tomography imaging in cancer detection and therapy response. Semin Oncol 2011;38(1):55
69. Figure 1D.

10.3.2 Small Molecule Imaging for Patient Staging and Endoradiotherapy: 123 I-Labeled Metaiodobenzylguanidine (123I-MIBG) Molecular imaging plays an integral role in the staging of neuroblastoma, a type of neuroendocrine extracranial solid cancer usually seen in early childhood. Neuroblastoma is diagnosed using the International Neuroblastoma Staging System criteria with immunohistochemistry of biopsied tumor tissue, increased serum or urine catecholamines, or identification of circulating tumor cells in bone marrow samples [18]. In addition to CT and/or MRI, 123I-metaiodobenzylguanidine (123I-MIBG) SPECT imaging is Food and Drug Administration (FDA) approved and recommended before surgical intervention to evaluate metastatic disease and risk group [19]. 123I-MIBG is a radioactive analog of catecholamines, which are preferentially taken up in neuroendocrine tumors (NETs), and 123I-MIBG accumulates in 90% of neuroblastomas (Fig. 10.2).

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Comparison of anterior and posterior (A) 123I-MIBG SPECT images and (B) 18F-MFBG PET images in a patient with neuroblastoma, illustrating that the higher resolution obtained by PET imaging allows for the delineation of metastatic sites (indicated by red arrows) not visible in the SPECT scans. CT, Computed tomography; PET, positron emission tomography; SPECT, single-photon emission CT; MIBG, metaiodobenzylguanidine; 18F-MFBG, 18F-metafluorobenzylguanidine. Source: This research was originally published in Pandit-Taskar N, Zanzonico P, Staton KD, et al. Biodistribution and dosimetry of 18F-meta-fluorobenzylguanidine: a first-in-human PET/CT imaging study of patients with neuroendocrine malignancies. J Nucl Med 2018;59(1):147
53. Figure 4. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [20
22].

FIGURE 10.2

In addition to staging, MIBG radiolabeled with iodine-131 is used for endoradiotherapy treatment in neuroblastoma patients. Because the tumor uptake and pharmacokinetic profiles of 123I-MIBG and 131I-MIBG are identical, imaging with 123I-MIBG can help determine if a patient should undergo endoradiotherapy with 131I-MIBG. 131I-MIBG has been evaluated in numerous clinical studies for use as monotherapy or in combination with chemotherapy and at multiple patient stages (newly diagnosed neuroblastoma, relapsed or refractory neuroblastoma), with response rates varying from 40% to 60% [20]. Clinical outcomes in relapsed or refractory neuroblastoma are typically poor, so endoradiotherapy with 131I-MIBG can help reduce or stabilize disease burden. Although SPECT imaging with 123I-MIBG is widely used in the staging of neuroblastoma, SPECT imaging gives inferior image resolution compared to PET imaging, and PET imaging also has superior quantitative accuracy [22]. Furthermore, imaging with 123I-MIBG requires thyroid protection and a 2-day imaging schedule which is inconvenient for the patient, so alternative approaches have been explored to evaluate neuroblastoma disease extent. 18 F-metafluorobenzylguanidine (18F-MFBG) has recently shown preliminary success as a PET imaging agent for neuroblastoma. 18F-MFBG has similar radiation exposure as 123IMIBG but faster blood and whole-body elimination that allows for imaging at 1
2 hours post administration rather than next day imaging required by 123I-MIBG (Fig. 10.2).

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10.3.3 Small Molecule Imaging to Determine Intertumor Heterogeneity and Predict Therapy Response—18F-fluoroestradiol (18F-FES) An important biomarker in breast cancer is the estrogen receptor (ER), which is expressed in approximately 75% of breast cancers and correlated with more favorable outcomes [23]. The current method of determining ER status in breast cancer is immunohistochemical staining from biopsied tumor tissue. It is important to note that ER expression at the primary tumor does not predict ER expression at metastatic sites [24]. Because of the intra- and intertumor heterogeneity of ER expression, information gained from primary tumor biopsy is limited in the context of metastatic breast cancer, and attempting to biopsy every metastatic lesion would be both challenging and impractical. An alternative method for determining ER status in breast cancer is molecular imaging using the small molecule 18F-fluoroestradiol (18F-FES) with PET. 18F-FES is a radioactive analog of the female sex hormone estradiol, which is an endogenous agonist of the ER, and multiple clinical trials have validated that high uptake of 18F-FES correlates with ER expression determined by traditional assays from biopsied tissue [23]. 18F-FES PET imaging alone is particularly useful for determining initial ER expression on primary tumors and metastatic lesions, while 18F-FES PET imaging has been combined with 18F-FDG PET or CT imaging to identify ER heterogeneity [23]. Furthermore, low uptake of 18F-FES predicts ER-negative tumor status in biopsied tissue and therefore could act as an alternative to biopsy of metastatic lesions [24]. A variety of breast cancer therapies target the ER pathway by inhibiting the production of estrogen (aromatase inhibitors), blocking estrogen from binding the ER (also known as endocrine therapy, e.g., tamoxifen) or inhibiting the ER signaling cascade (CDK4/6 inhibitors and mTOR inhibitor everolimus), so assessing ER expression and heterogeneity at primary and metastatic lesions is critical for determining whether a patient could benefit from ER-targeted therapies [23]. Multiple clinical trials have evaluated the use of 18F-FES as a companion diagnostic to ER-targeted therapies, as low uptake of 18F-FES in malignant lesions predicts a lack of response in patients undergoing endocrine therapy or aromatase inhibitor therapy (Fig. 10.3). 18 F-FES imaging of breast cancer can be used as a companion diagnostic to preclude patients from ER-targeted therapy, helping the patient avoid potential side effects from ineffective treatment, along with saving the patient time and money. Furthermore, 18F-FES PET imaging allows for dynamic monitoring of ER expression in response to ER-targeted therapies without the need for further tumor biopsy, which can help inform physicians about changes in ER expression and how to proceed with treatment [25].

10.3.4 Small Molecule Imaging to Assess Pharmacodynamics of Novel Therapeutics: 18F-fluorodihydrotestosterone (18F-FDHT) and Enzalutamide Similar to the ER’s role in some breast cancers, the transcription factor androgen receptor (AR) is thought to be an important driver in prostate cancer through overexpression, gain-of-function mutations, and ligand-independent activation [30]. Testosterone and dihydrotestosterone are endogenous agonists of AR, so the PET imaging agent 18Ffluorodihydrotestosterone (18F-FDHT) was developed as an analog to noninvasively image

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F-FES and 18F-FDG PET scans before ER-targeted treatment. (A) The patient showed ERpositive lesions in the vertebrae which had high uptake of 18F-FDG. The 18F-FDG uptake decreased after 3 months of ER-targeted treatment, indicating a therapeutic response. (B) The patient showed ER-negative lesions which had high uptake of 18F-FDG. The 18F-FDG uptake increased after 3 months of ER-targeted treatment, indicating no therapeutic response. Dashed arrows show normal 18F-FES uptake in the liver. PET, Positron emission tomography; 18F-FDG, 18F-fluorodeoxyglucose; F-FES, F-fluoroestradiol; ER, estrogen receptor. Source: Reprinted with permission. r 2006 American Society of Clinical Oncology. All rights reserved. Linden HM, Stekhova, SA, Link JM, et al. Quantitative fluoroestradiol positron emission tomography imaging predicts response to endocrine treatment in breast cancer. J Clin Oncol 2006;24(18):2793
9. Figure 1 [24
29].

FIGURE 10.3

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AR. Quantitative tumor uptake of 18F-FDHT by PET imaging correlates to AR expression in primary and metastatic lesions, confirming its utility as a noninvasive assay for AR expression [31]. The binding of testosterone, dihydrotestosterone, and 18F-FDHT on AR can be blocked by the use of AR antagonists (which are used therapeutically in androgen deprivation therapy), so molecular imaging offers a method of evaluating the pharmacodynamics of novel AR antagonists in early clinical trials. An example of using 18F-FDHT to evaluate AR antagonists for therapy is enzalutamide [32]. A 18F-FDHT PET/CT scan was first performed in order to establish a baseline of AR expression in primary and metastatic sites. After establishing the baseline of AR expression, 30
600 mg/day of enzalutamide was administered orally to patients, and 18F-FDHT PET/CT scans were repeated after 4 weeks of the treatment (Fig. 10.4).

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FIGURE 10.4 Sagittal and coronal 18F-FDHT PET scans in a patient with prostate cancer (A) before and (B) after 4 weeks of enzalutamide therapy, showing decreased uptake in the vertebrae metastases [32]. PET, Positron emission tomography; F-FDHT, F-fluorodihydrotestosterone. Source: Scher HI, Beer TM, Higano CS, Anand A, Taplin ME, Efstathiou E, et al. Prostate Cancer Foundation/ Department of Defense Prostate Cancer Clinical Trials C. Antitumour activity of MDV3100 in castrationresistant prostate cancer: a phase 1-2 study. Lancet 2010;375 (9724):1437
46. Figure 1B.

The standard uptake value (SUV) of the 18F-FDHT decreased for all who received 4 weeks of enzalutamide therapy, confirming that enzalutamide was binding the AR. The extent of SUV reduction from pre- to posttreatment scans helped determine the optimal dose of enzalutamide, as patients receiving 60 mg/day showed a lower change in SUV, while increasing the dose to 150 mg/day caused an increase in SUV change. Notably, doses higher than 150 mg/day did not show significant differences in SUV change [32]. Because of this and further studies, enzalutamide was approved by the FDA in 2012 with a recommended dosing schedule of 160 mg daily [33].

10.3.5 Small Molecule Imaging to Assess EGFR Mutations for Targeted TKI Therapeutics EGFR is a receptor tyrosine kinase expressed in more than 60% of non
small cell lung carcinomas (NSCLCs) [34]. Within NSCLC, 10%
15% of the patients bear an activating EGFR mutation, such as a deletion in exon 19 or the point mutation T790M [10]. Because

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EGFR promotes growth in NSCLC, enormous effort has been put in to develop small molecules with inhibitory therapeutics against EGFR (herein referred to as tyrosine kinase inhibitors, or TKIs). NSCLCs have differing sensitivities to various TKIs based on the mutational status of the EGFRs [35]. Mutational status is also heterogeneous across metastases (e.g., brain lesions), so understanding mutations in all lesions is necessary to better inform treatment [10]. Molecular imaging can be used as a companion diagnostic to differentiate EGFR mutations across all patient lesions. One example of this is the exon 19 deletion in EGFRs. Patients with an exon 19 deletion are more sensitive to gefitinib, a first-generation TKI, so the small molecule imaging agent [18F]F-IRS was developed as a companion diagnostic to specifically image EGFRs with the exon 19 deletion [35,36]. Another example of a small molecule imaging agent for mutated EGFR is the third-generation TKI osimertinib. [11C]C-osimertinib was used preclinically to study brain penetration in order to validate that osimertinib could be used to treat brain metastases in patients with the T790M mutation [37]. Other EGFR targeting TKIs have also been radiolabeled to study their pharmacokinetic and pharmacodynamics profiles; for a more extensive review on small molecules for EGFR mutational imaging, please refer to Waaijer et al. [10]. EGFR is one example of how small molecules can be used as companion diagnostics to assess mutational status for mutation-targeted therapies.

10.4 PEPTIDES FOR MOLECULAR IMAGING Using peptides as companion diagnostics for molecular imaging has several benefits. Like small molecules, their small size makes them fast diffusing resulting in maximal uptake at the target as soon as 30 minutes postinjection, which allows for patients to be injected and imaged on the same day [38]. Peptides also show rapid clearance through a combination of renal and hepatobiliary excretion which can substantially increase the tumor to background ratios leading to better images. However, natural peptide ligands are quickly degraded in vivo, necessitating chemical modifications to increase their stability and biological half-life. The chemical modifications that improve the stability of peptides can also dramatically affect the ability of peptides to bind to their target, so confirming binding affinity postmodification is necessary before in vivo studies can be conducted. Peptide companion diagnostics have become increasingly relevant with the expanding use of peptide receptor radionuclide therapy (PRRT) in Europe and to a limited extent in the United States. One of the most common radionuclides used for PRRT is lutetium-177, which emits a medium energy beta particle that can damage and kill cancer cells as well as a low energy photon emission which can be used for SPECT imaging [39,40]. Despite the ability of lutetium-177 to be used in SPECT imaging of peptides, corresponding PET imaging peptides are also developed in parallel to therapeutic peptides in order to improve image resolution and quantification. The peptides used in molecular imaging can be divided into agonists (first generation) and antagonists (second generation) [41]. First-generation peptides for radiotherapy and imaging were designed as agonists of their targets, because it was thought that cellular internalization was necessary for sufficient accumulation at the target site. However, the

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activation of the target receptor by agonists led to negative side effects when evaluated in the clinic. Therefore second-generation peptides for molecular imaging are in the form of antagonists, which are not internalized significantly but do not cause activation of their target receptor. Peptide theranostic pairs for molecular imaging can be used in several contexts including disease diagnosis and staging, interrogating the heterogeneity of metastatic lesions in a single patient, choosing patients who are more likely to respond to PRRT, and monitoring the response of a variety of treatments.

10.4.1 Somatostatin Receptor Targeting Most of the clinical data on PRRT has come from the treatment of NETs. Neuroendocrine cells overexpress somatostatin receptors (SSTRs) on their surface allowing them to be used as targets for imaging and therapeutic agents [42]. Peptide receptor agonists in the form of somatostatin analogs, such as octreotide, have been used since the early 1990s for imaging of NETs. OctreoScan, [111In]In-pentetreotide (diethylenetriaminepentaacetic acid (DTPA)-octreotide), shows high affinity toward the SSTR2 receptors and was the first FDA-approved peptide-imaging agent in 1994. OctreoScan comes in a kit formulation and continues to be used routinely in the clinic (Fig. 10.5). By using dual mode SPECT/CT imaging, it is possible to identify the location of the NET, define tumor stage, and to assess disease burden at follow-up. Preclinically, two somatostatin analogs, DOTA-octreotide (DOTA-TOC) and DOTAoctreotate (DOTA-TATE), were studied in vivo to determine which compound had higher tumor uptake and better tumor volume reduction [44]. [177Lu]Lu-DOTA-TATE had approximately double the tumor uptake and caused superior tumor reduction compared

FIGURE 10.5 [111In]In-DTPA-octreotide anterior abdominal images in a patient with liver metastases of a neuroendocrine tumor, 24 hours postinjection. Source: This research was originally published in Kwekkeboom DJ, Kooij PP, Bakker WH, Macke HR, Krenning EP. Comparison of 111In-DOTA-Tyr3-octreotide and 111In-DTPA-octreotide in the same patients: biodistribution, kinetics, organ, and tumor uptake. J Nucl Med 1999;40(5):762
7. Figure 4. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [43].

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to [177Lu]Lu-DOTA-TOC. This study and others paved the way for [177Lu]Lu-DOTA-TATE clinical trials and clinical use outside the United States. In early 2017, results from a phase III clinical trial (NETTER-1) were released demonstrating increased progression-free survival and response rate as compared to high-dose octreotide long-acting release [45]. Under the trade name Lutathera, [177Lu]Lu-DOTA-TATE was approved for use in the European Union in September 2017 and in the United States in January 2018. Clinical response rates for Lutathera in clinical trials were less than 20%. Due to its expanding use, a companion diagnostic to determine which patients are most likely to respond to PRRT was needed. More recently, somatostatin analogs have been labeled with positron emitters for PET/ CT imaging, as PET instruments can achieve higher spatial resolution than traditional gamma cameras used for SPECT imaging. These agents utilize DOTA as the chelator for the incorporation of gallium-68 into the peptide (Fig. 10.6). Using gallium-68 instead of indium-111 reduces the total radiation dose to the patient, and the scan can be completed in less time. To determine if [68Ga]Ga-DOTA-TATE could serve as a companion diagnostic for Lutathera, the biodistribution of both the [68Ga]Ga- and [177Lu]Lu-labeled peptide was compared in a preclinical setting [47]. The biodistribution was not significantly affected by the change in radionuclide, demonstrating the potential of the gallium labeled peptide as a companion diagnostic. Under the trade name NETSPOT, [68Ga]Ga-DOTA-TATE was approved for use in the United States in June 2016. It is also approved in the European Union where it is used as a companion diagnostic for Lutathera. [68Ga]Ga-DOTA-TATE is also used after Lutathera treatment to assess therapeutic efficacy and patient restaging (Fig. 10.7).

10.4.2 Gastrin-Releasing Peptide Receptor Targeting Along with somatostatin, analogs of the bombesin peptide are being explored as theranostic agents targeting gastrin-releasing peptide receptors (GRPRs), which are overexpressed in prostate, breast, and gastrointestinal cancers [49]. While bombesin is a natural ligand to GRPR, it suffers from low in vivo stability which precludes its use for imaging or PRRT. Two bombesin analogs have been studied extensively preclinically and beyond, the agonist PESIN and the antagonist NeoBomb1. Both of these peptides were developed as theranostic pairs so that the imaging peptide could be given as a diagnostic to determine whether the therapeutic peptide would be beneficial. PESIN, a GRPR agonist, was coupled to either a DOTA or DTPA chelate for incorporation of a range of radionuclides into the peptide (e.g., indium-111 for SPECT and gallium-68 for PET) [50]. When evaluated with a panel of other bombesin analogs, PESIN showed significant tumor uptake enabling imaging of PC-3 prostate xenografts in athymic mice. The tumor uptake remained constant over 24 hours, which allowed for imaging at later time points when the tumor-to-blood ratio had increased. In another study focusing on PESIN alone, receptor-mediated uptake was seen in several normal organs including the pancreas, pituitary, adrenals, spleen, bowel, and stomach [51]. High uptake in the pancreas was seen in the biodistribution of PESIN, and was observed in all mouse studies of GRPR-targeting peptides. In patients, this is not expected since the human pancreas does

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FIGURE 10.6 Comparison of (A) [111In] In-DTPA-octreotide SPECT/CT and (B) [68Ga]Ga-DOTA-TATE PET/CT in a patient with suspected recurrence of small bowel NET in the liver. Nine liver metastases were found with PET/CT resulting in a change in the surgical plan. CT, Computed tomography; PET, positron emission tomography; SPECT, single-photon emission CT; NET, neuroendocrine tumor. Source: This research was originally published in Deppen SA, Blume J, Bobbey AJ, et al. 68Ga-DOTATATE compared with 111In-DTPA-octreotide and conventional imaging for pulmonary and gastroenteropancreatic neuroendocrine tumors: a systematic review and meta-analysis. J Nucl Med 2016;57 (6):872
8. Figure 3. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [46].

not express GRPR at measurable levels. PESIN demonstrated suitable pharmacokinetics with high tumor uptake, slow tumor washout, and high tumor-to-liver and tumor-tokidney ratios. Despite promising preclinical data, PESIN was never translated to the clinic. Due to its agonistic effect on GRPRs, PESIN caused gastrointestinal side effects [52,53]. Because they do not activate GRPRs, peptide antagonists were expected to be better for clinical use with fewer adverse side effects. This hypothesis was confirmed when the peptide antagonist RM2 was used in patients to target GRPR, and no adverse side effects were observed [54]. Due to this key finding, second-generation GRPR peptides focused on antagonists. One of

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(A) Pretreatment with [68Ga]Ga-DOTA-TATE PET/CT images showing lesions with avid uptake. (B) Repeated imaging with [68Ga]Ga-DOTA-TATE after the patient underwent three cycles of [177Lu]LuDOTA-TATE showed a partial metabolic response in lesions in the liver and vertebral body with no new lesions detected. CT, Computed tomography; PET, positron emission tomography. Source: This research was originally published in Gains JE, Bomanji JB, Fersht NL, et al. 177Lu-DOTATATE molecular radiotherapy for childhood neuroblastoma. J Nucl Med 2011;52(7):1041
7. Figure 2. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [48].

FIGURE 10.7

these antagonists, NeoBomb1, is another bombesin analog and has demonstrated success in preclinical studies [55]. The NeoBomb1 peptide is conjugated to a DOTA chelator and has been shown to have a very high affinity for GRPR. In vitro studies with the NeoBomb1 peptide labeled with gallium-68, indium-111, or lutetium-177 showed that changing the radionuclide did not significantly affect the cell binding. In binding assays, high specific binding was seen for all labeled peptides, and less than 10% of the peptides were found to be internalized, which is expected for antagonistic peptides. Labeled peptides also showed encouraging in vivo biodistribution in PC-3 xenografted mice (Fig. 10.8). The [68Ga]Ga- and [177Lu]Lu-labeled NeoBomb1 peptides were studied further to assess their potential as a theranostic pair for clinical use [56]. A comprehensive biodistribution study was conducted for both peptides so that mouse dosimetry could be calculated as well as a human dosimetry estimate. PET/CT imaging allowed for the clear visualization of

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FIGURE 10.8 (A) PET/CT imaging 1 hour postinjection of [68Ga]Ga-NeoBomb1. (B) SPECT/MR imaging 4 hours postinjection of [177Lu]Lu-NeoBomb1, tumor located on the right shoulder. CT, Computed tomography; PET, positron emission tomography; SPECT, single-photon emission CT; GRPR, gastrin-releasing peptide receptor. Source: This research was originally published in Dalm SU, Bakker IL, de Blois E, et al. 68Ga/177Lu-NeoBomb1, a novel radiolabeled GRPR antagonist for theranostic use in oncology. J Nucl Med 2017;58(2):293
9. Figures 2 and 3. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [56].

the tumor with uptake also being evident along the gastrointestinal tract and the pancreas. In a clinical proof-of-principle study in patients with prostate cancer, [68Ga]Ga-NeoBomb1 was well tolerated by all subjects with no side effects reported [55]. The tracer was found to accumulate preferentially in primary prostate-confined and metastatic foci. Along with the primary carcinoma in the prostate, metastases could be detected in the lymph nodes, abdomen, next to the esophagus, and the mediastinum (Fig. 10.9). Along with prostate cancer, the NeoBomb1 theranostic pair may have applications in breast cancer as well [57]. GRPR is found to be overexpressed in more than 60% of tumor biopsies of patients with invasive breast carcinoma. It has also been found that all metastases from the primary GRPR expressing tumors retained this overexpression. In vitro assays were done in the breast cancer cell line T-47D with [67Ga]Ga-NeoBomb1, and the peptide retained its high affinity for GRPR despite the change in cancer context. In vivo

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[68Ga]Ga-NeoBomb1 PET/CT imaging of the patient with prostate adenocarcinoma (A) maximum intensity projection, (B) serial PET transverse, (C) corresponding CT transverse, (D) fusion PET/CT. Multiple metastases evident in mediastinum, abdomen, paraesophagus, and pelvic lymph nodes indicated by arrows and cross bars. CT, Computed tomography; PET, positron emission tomography. Source: This research was originally published in Nock BA, Kaloudi A, Lymperis E, et al. Theranostic perspectives in prostate cancer with the gastrinreleasing peptide receptor antagonist NeoBomb1: preclinical and first clinical results. J Nucl Med 2017;58(1):75
80. Figure 6. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [55].

FIGURE 10.9

stability of the [67Ga]Ga-NeoBomb1 was determined by radio-high pressure liquid chromatography (HPLC), which demonstrated that 90% of the peptide remains intact at 30 minutes postinjection. This was encouraging because most peptides suffer from rapid proteolytic degradation in vivo. The biodistribution of [67Ga]Ga-NeoBomb1 was then determined in severe combined immunodeficient (SCID) mice bearing T-47D xenografts. While no significant change in tumor uptake from 1 to 24 hours postinjection was observed, biodistribution showed that uptake in all other organs gradually decreased over this time, increasing tumor to background ratios and opening the possibility of NeoBomb1 to be used in the context of breast cancer. Based on promising preclinical data across multiple cancer types, a phase I/II clinical trial has been posted (NCT02931929) to evaluate [68Ga]Ga-NeoBomb1 PET/CT imaging in patients with advanced gastrointestinal tumors.

10.5 MOLECULAR IMAGING USING ANTIBODIES In addition to small molecules and peptides, several antibodies have been used as molecular imaging agents [58
60]. Most of these immuno-PET agents are based on FDAapproved antibodies. Antibodies have the ability to bind to antigens overexpressed or selectively expressed on cancer cells with exquisite specificity. Antibody-based molecular imaging agents must bind to antigens present at the cell surface of the tumor or on the extracellular compartment due to their inability to cross the cell membrane. The different applications of antibody-based molecular compounds in cancer imaging and therapy have been reviewed in several papers [7,59,61
64]. Antibody-based imaging agents allow tumor

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diagnosis in both adult and childhood malignancies, allow delineation of surgical margins, predict tumor response to therapy, detect tumor heterogeneity, image metastatic lesions occurring in clinically challenging sites (e.g., brain and bone), and detect tumor recurrence [63
81]. Although few studies explore the potential of antibody-based molecular imaging in pediatric tumors, this methodology seems to be promising to enhance the sensitivity and specificity of diagnosis. Molecular imaging with radiolabeled antibodies is also a valuable methodology during initial clinical studies that aim at screening new antibodies [61]. When testing new therapeutic antibodies which have not yet been validated, immuno-PET imaging using suboptimal doses (i.e., microdosing) can provide information about antibody biodistribution, uptake in the tumor versus normal tissues, dosimetry, and safety [77]. The antigens MET (i.e., the receptor of hepatocyte growth factor), cytotoxic T-lymphocyte-associated antigen 4 (CTLA4), EGFR variant III (EGFRvIII), fibroblast activation protein, the Notch ligand delta-like 3 (DLL3), and the cancer antigen 125 (CA125) are examples of new potential targets for immuno-PET applications which have been studied preclinically [82
87]. One of the drawbacks of using fully intact antibodies as molecular imaging agents is related to the period of time needed for the antibody to accumulate in the tumor tissue. Intact antibodies remain in circulation for days or weeks and can take a few days to significantly accumulate in the tumors, which is necessary to acquire images with good contrast [70]. When compared with small molecules and peptides, the slow kinetics of antibodies requires their conjugation with radionuclides of long physical half-lives, which exposes the patient to a higher radiation dose. After binding to cell surface antigens, the majority of labeled antibodies are internalized and proteolytically degraded. Once degraded, the radioactive label is no longer linked to the targeting moiety resulting in images that no longer reveal whether or not the labeled conjugate is still binding its target. The degradation of the antibody or release of the radionuclide is particularly important when using certain radionuclides, such as zirconium-89, due to its tendency to accumulate in bone and become intracellularly trapped. These internalization and catabolism processes occurring between the administration of the antibody-labeled agent and the acquisition of wholebody imaging can result in false-positive images [76].

10.5.1 Antibody Fragments The use of antibody fragments and engineered variants—F(ab0 )2, F(ab0 ), Fab, single chain Fv (scFv), and the covalent dimers scFv2, diabodies, and minibodies—has been shown to reduce the period of time between injection and image acquisition from days to hours, due to their faster accumulation in the tumor tissue when compared with fully intact antibodies [59,88]. Because of this, fragments of antibodies can be conjugated with short half-life radionuclides. Recent studies have also suggested that PET imaging with a radiolabeled diabody can be used to determine whether or not a patient is suitable for immunotherapy [89]. For instance, Wu et al. demonstrated in preclinical syngeneic tumor models that a radiolabeled anti-CD8 cys-diabody can be used to determine therapyinduced fluctuations in T-cell population (systemic and tumor-infiltrating CD8) [89]. Although preclinical studies demonstrate that antibody fragments have some advantages

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when compared with intact antibodies, they are not currently used in clinics. Therefore the next subsections discuss mainly the different examples of intact antibody-based molecular imaging agents as companion diagnostics.

10.5.2 HER Targeting Antibodies Although the status of HER2 is used in breast cancer management as a biomarker to select patients for HER2-targeted therapies, HER2 positivity as detected from histologic analysis has not always been correlated with patient benefit from HER2-targeted therapy. As an example, trastuzumab has demonstrated therapeutic efficacy in women with HER2negative breast tumors [90,91]. Tumor heterogeneity between different sites of malignancy in a single patient can account for these findings, suggesting that results based on a single tumor biopsy are insufficient to determine HER2 status. Whole-body molecular imaging allows the detection of heterogeneity in HER2 expression between primary tumors versus metastases (Figs. 10.10 and 10.11). Nevertheless, the use of radiolabeled trastuzumab for imaging of tumor heterogeneity requires optimization of the dose and time of administration [66]. A clinical trial enrolling nine patients with HER2-negative primary breast cancer explored the potential of [89Zr]Zr-DFO-trastuzumab PET/CT to detect tumor heterogeneity [76]. Five of these patients demonstrated suspicious HER2-positive metastases as detected by uptake of [89Zr] Zr-DFO-trastuzumab. HER2 positivity in these lesions was further confirmed by pathological analysis in two of these patients. In addition, the patients with HER2-negative primary breast tumors and HER2-positive metastases saw a benefit from HER2-targeted therapy.

FIGURE 10.10 (A) Immunohistochemistry analysis of HER2 in a 41-year-old with primary breast cancer, showing HER2-negative malignancy. (B) Axial CT and PET/CT images obtained after administration of [89Zr]ZrDFO-trastuzumab demonstrating radiotracer accumulation in metastases. (C) Immunohistochemistry analysis of HER2 showing that those metastases are HER2 positive. (D) Axial CT obtained after 2 months of treatment showing a therapeutic response. CT, Computed tomography; PET, positron emission tomography. Source: This research was originally published in Ulaner GA, Hyman DM, Ross DS, et al. Detection of HER2-positive metastases in patients with HER2-negative primary breast cancer using 89Zr-trastuzumab PET/CT. J Nucl Med 2016;57(10):1523
8. Figure 2. r by the Society of Nuclear Medicine and Molecular Imaging, Inc.

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FIGURE 10.11 Sequential MIP images obtained at different time points (A
D) after administration of [89Zr] Zr-DFO-pertuzumab to a 46-year-old woman with both HER2-positive and HER2-negative primary breast and brain metastases. CT, Computed tomography; PET, positron emission tomography; MIP, maximum-intensity projection. Source: This research was originally published in Ulaner GA, Lyashchenko SK, Riedl C, et al. First-inhuman HER2-targeted imaging using 89Zr-pertuzumab PET/CT: dosimetry and clinical application in patients with breast cancer. J Nucl Med 2018;59(6):900
6. Figure 3. r by the Society of Nuclear Medicine and Molecular Imaging, Inc. [76
78].

The potential of PET/CT to determine HER2 heterogeneity has also been demonstrated with [89Zr]Zr-DFO-pertuzumab (another HER2-targeting humanized monoclonal antibody) [77]. PET/CT imaging of a patient with HER2-negative primary breast cancer demonstrated [89Zr]Zr-DFO-pertuzumab accumulation in chest wall metastases, which were then confirmed to be HER2 positive by pathological analysis. In addition to being used for cancer diagnosis, radiolabeled trastuzumab allows patient selection for therapy [70,78]. The ZEPHIR trial combined [89Zr]Zr-DFO-trastuzumab PET/ CT with standard 18F-FDG to predict therapeutic response with trastuzumab emtansine (a HER2-targeted antibody
drug conjugate) in HER2-positive metastatic breast cancer patients [78]. The combination of [89Zr]Zr-DFO-trastuzumab with 18F-FDG PET scans allowed differentiation of patients with time-to-treatment failure (TTF) of 2.8 months from those with a TTF of 15 months. In other studies, immuno-PET using [89Zr]Zr-DFO-cetuximab [a therapeutic antibody targeting EGFR (also known as HER1)] has been explored as an imaging tool to predict therapeutic response [74]. The tumor uptake of [89Zr]Zr-DFO-cetuximab was positively correlated with overall survival and progression free in patients with K-RAS (Kirsten rat sarcoma viral oncogene homolog) advanced colorectal cancer [74]. Although some studies have suggested a correlation between EGFR expression and response to therapy, others have suggested that imaging mutations of this receptor might be a more accurate strategy to predict therapeutic response to EGFR-targeted therapies [75].

10.5.3 Carbonic Anhydrase-IX Targeting Antibodies Divgi et al. have demonstrated the potential of a radiolabeled chimeric antibody (girentuximab) in detecting clear-cell renal carcinomas (ccRCCs) [67]. Girentuximab targets

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carbonic anhydrase-IX (a cell surface antigen overexpressed in ccRCCs) and when radiolabeled with iodine-124, it showed to be a powerful imaging agent in detecting ccRCCs. Immuno-PET with this conjugate also correlated positively with biopsy analysis [68].

10.5.4 Vascular Endothelial Growth Factor Targeting Antibodies The radiolabeled bevacizumab (a monoclonal antibody targeting the vascular endothelial growth factor-A) is an example of a radiolabeled antibody that has been used to predict therapeutic response of patients with non
small cell lung carcinoma (NSCLC) after chemotherapy with carboplatin, paclitaxel, and bevacizumab (CPB) [73]. Although there was a positive correlation between tracer uptake and patient response to CPB, the trend was not significant due to the small number of patients enrolled in the study. When bevacizumab contains both a radiolabel and a fluorescent label, the same molecule can be used for preoperative PET detection and intraoperative fluorescent detection [70
72]. Therefore the first dual-labeled probe ([111In]In-DOTA-girentuximab-IR-800CW) is being exploited in clinical trials in patients with RCCs (NCT024975599).

10.5.5 Immunopositron Emission Tomography to Detect Brain and Bone Metastases Antibodies were expected to have limited use as imaging agents of brain tumors due to their large size, which interferes with their ability to cross the blood
brain barrier. Nevertheless, recent studies have demonstrated that radiolabeled antibodies are able to specifically detect brain metastases of primary breast cancer tumors and gliomas [66,77,92]. Their ability to reach brain tumors might be related to alterations in the vasculature during the tumor development [60]. As an example, immuno-PET with radiolabeled trastuzumab can be used to determine HER2 status in brain metastases [77]. In another study, de Hollander et al. demonstrated for the first time that PET imaging of patients with high-grade gliomas (brain tumors) is possible using [89Zr]Zr-DFO-fresolimumab (an antibody against isoforms of transforming growth factor-β) [92]. In addition to brain metastases, PET imaging allows detection of lesions in the bone. As an example, [89Zr]Zr-DFO-huJ591 (a humanized monoclonal antibody that targets the extracellular domain of prostate-specific membrane antigen) exhibited a maximum SUV higher for bone lesions when compared with soft tissues in patients with metastatic prostate cancer (Fig. 10.12). Other studies have also demonstrated that immuno-PET with [89Zr]Zr-DFO-5B1 is able to detect metastatic lesions in the bone of patients with CA19-9 positive metastatic malignancies (eight pancreatic and one of unknown origin) [79].

10.5.6 Programmed Death-Ligand 1 Targeting Antibodies The programmed death receptor 1 (PD-1 and CD279) and its ligand PD-L1 (B7-H1 and CD274) are major therapeutic targets in the clinical cancer setting of immune checkpoint blockade. PD-L1 upregulation in tumors results in immune suppression due to

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FIGURE 10.12 Bone scans demonstrating that [89Zr]Zr-DFO-huJ591 allows the detection of bone lesions to a higher extent than 18F-FDG or [99mTc]Tc-MDP. PET, Positron emission tomography; 18F-FDG, 18F-fluorodeoxyglucose; MDP, methyl diphosphonate. Source: Republished with permission of Springer Science and Bus Media B V, from 89 Zr-huJ591 immuno-PET imaging in patients with advanced metastatic prostate cancer, Pandit-Taskar N, et al. 41,11 2014. Figure 2; permission conveyed through Copyright Clearance Center, Inc. [80].

deactivation of PD-1 expressing tumor-infiltrating lymphocytes. Although PD-L1 expression has been associated with therapeutic response to PD-1/PD-L1 inhibition, patients with low expression of PD-L1 (as assayed by immunohistochemistry) have also demonstrated a response to therapy. To interrogate the dynamic and heterogeneous expression of PD-L1, PET imaging with a PD-L1 targeting antibody (MPDL3280A, atezolizumab) has been performed in patients with metastatic NSCLC, bladder cancer, and triple-negative breast cancer [93]. [89Zr]Zr-DFO-atezolizumab demonstrated a heterogeneous uptake in tumors within the same patient and between different patients. The radiotracer also accumulated in tumors that did not express PD-L1 as determined by immunohistochemistry. Recent preclinical studies have demonstrated that immuno-PET with a novel recombinant human IgG1 (termed C4) detects low endogenous levels and acute changes in the expression of PD-L1 [94]. Nevertheless, further studies are warranted to determine the potential clinical application of radiolabeled C4.

10.5.7 Shedding Targets and Pretargeting Antigen shedding or secretion by some cancer cells is a challenge in using antibody-based imaging agents. For example, [89Zr]Zr-DFO-trastuzumab demonstrated rapid clearance in trastuzumab-naı¨ve patients probably due to trastuzumab binding to the HER2 extracellular domains, which are shed by the cancer cells and now present in the plasma [66]. The antigen CA19-9 (carbohydrate antigen 19-9) is well known as a serum marker for pancreatic cancer, and it is also overexpressed in pancreatic tumor tissues. Recent studies in patients with pancreatic cancer have demonstrated that CA19-9 is a valuable antigen for PET imaging with the humanized monoclonal antibody 5B1 [79]. In addition, the use of a

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pretargeting PET imaging system has demonstrated to be a promising strategy when the aim is to image a shedding target, such as CA19-9 [95]. This pretargeting methodology involves the administration of a bispecific unlabeled antibody that has a binding site for the antigen and another for binding a radiolabeled small molecule. The small molecule is administered after the antibody has accumulated at the tumor, usually 1
3 days postantibody injection. Since pretargeting is a two-step strategy, its success is dependent on optimization of molar ratios of antibody and small labeled probe and the period of time between the two injections [96]. The first clinical studies using an immuno-PET pretargeting strategy were reported in 2013 in colorectal carcinoma patients using the tri-Fab molecule TF2 (targeting CEACAM5) and the peptide [111In]In-IMP288 (Fig. 10.13).

FIGURE 10.13

(A) SPECT/CT image of a 38-year-old patient with primary colon tumors acquired 24 hours postinjection with [111In]In-IMP288 and pretargeting with tri-Fab molecule TF2 (targeting CEACAM5). The image shows radiotracer accumulation in axillary lymph-node metastases and low accumulation in normal tissues. (B) and (C) contrast-enhanced CT and fused 18F-FDG-PET/CT scans obtained for the same patient. (D) SPECT, (E) CT scan, and (F) 18F-FDG-PET/CT images of the primary colon tumors of the same patient demonstrating radiotracer specificity on those tumors. CT, Computed tomography; PET, positron emission tomography; SPECT, single-photon emission CT; 18F-FDG, 18F-fluorodeoxyglucose. Source: Reprinted by permission from Macmillan Publishers Ltd. on behalf of Cancer Research UK: Schoffelen R, Boerman OC, Goldenberg DM, Sharkey RM, et al. Development of an imaging-guided CEA-pretargeted radionuclide treatment of advanced colorectal cancer: first clinical results. Br J Cancer 2013;109(4):934
42. Figure 4, copyright 2014. [96].

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The study was successful in showing that a pretargeting strategy accelerates tumor uptake, allowing for images to be obtained within a few hours, as well as increased tumor-to-background ratios which may make it possible to use short-lived isotopes.

10.6 CONCLUSION Preclinical and clinical data support the enormous potential of using radiolabeled small molecules, peptides, or antibodies as companion diagnostics for cancer management. Because molecular imaging techniques are noninvasive and can be used to monitor patient status over time, they can offer complementary information to traditional diagnostic techniques, such as tumor biopsy. However, few clinical studies have explored the relationship between molecular imaging and analysis of biopsied tumor specimens/circulating cancer cells, so this relationship should be further examined. When compared with peptides, radiolabeled antibodies and small molecules are the most frequently studied constructs. Small molecule imaging probes tend to mimic endogenous small molecules, while immuno-PET imaging probes are usually antibodies that have already been FDA approved for cancer treatment. Once approved for therapy, antibodies can be easily repurposed for diagnostics. Notably, a construct with excellent therapeutic properties does not necessarily result in a construct with sufficient diagnostic properties but can be an informative approach for companion diagnostic development. Between small molecules, peptides, and antibodies, one class of targeting vectors is not necessarily better than the others. The choice of the targeting vector is highly dependent on the cellular target and the type of information that one wants to obtain. In addition, careful planning in the labeling methodology, choice of chelate, and radionuclide are important to generate stable and selective constructs. While antibody-labeled constructs might be superior for determining extracellular receptor expression, small molecules could better assess intracellular processes, such as metabolism. Although peptide-labeled constructs have demonstrated use as theranostic pairs for PRRT, their clinical use in the United States has been limited by regulatory approval of therapeutic-based peptides. The combination of different imaging agents and/or distinct imaging modalities shows promise in the field of companion diagnostics, and its potential will continue to grow in the era of precision medicine.

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