Search

PDF file - 185K

PDF file - 181K

PDF file - 241K

PDF file - 312K

PDF file - 151K

PDF file- 366K

PDF file - 483K

Supplementary Methods, Figures 1-3 from Selective Killing of Tumor Neovasculature Paradoxically Improves Chemotherapy Delivery to Tumors

Deep-seated bacterial infections caused by pathogens such as Staphylococcus aureus are difficult to diagnose and treat and are thus a major threat to human health. In previous work we demonstrated that positron emission tomography (PET) imaging with 2-[18F]F-p-aminobenzoic acid (2-[18F]F-PABA) could noninvasively identify, localize, and monitor S. aureus infection with excellent sensitivity and specificity in a rodent soft tissue infection model. However, 2-[18F]F-PABA is rapidly N-acetylated and eliminated, and in an attempt to improve radiotracer accumulation in bacteria we adopted a prodrug strategy in which the acid was protected by an ester and the amine was replaced with a nitro group. Metabolite analysis indicated that the nitro group of ethyl 2-[18F]fluoro-4-nitrobenzoate (2-[18F]F-ENB) is converted to the corresponding amine by bacteria-specific nitroreductases while the ester is hydrolyzed in vivo into the acid. PET/CT imaging of 2-[18F]F-ENB and the corresponding acid 2-[18F]F-NB in a rat soft tissue infection model demonstrated colocalization of the radiotracer with the bioluminescent signal arising from S. aureus Xen29, and demonstrated that the tracer could differentiate S. aureus infection from sterile inflammation. Significantly, the accumulation of both 2-[18F]F-ENB and 2-[18F]F-NB at the site of infection was 17-fold higher than at the site of sterile inflammation compared to 8-fold difference observed for 2-[18F]F-PABA, supporting the proposal that the active radiotracer in vivo is 2-[18F]F-NB. Collectively, these data suggest that 2-[18F]F-ENB and 2-[18F]F-NB have the potential for translation to humans as a rapid, noninvasive diagnostic tool to identify and localize S. aureus infections.

Hepatocellular carcinoma (HCC) is difficult to detect, carries a poor prognosis, and is one of few cancers with an increasing yearly incidence. Molecular defects in complement factor H (CFH), a critical regulatory protein of the complement alternative pathway (AP), are typically associated with inflammatory diseases of the eye and kidney. Little is known regarding the role of CFH in controlling complement activation within the liver. While studying aging CFH-deficient (fH(–/–)) mice, we observed spontaneous hepatic tumor formation in more than 50% of aged fH(–/–) males. Examination of fH(–/–) livers (3–24 months) for evidence of complement-mediated inflammation revealed widespread deposition of complement-activation fragments throughout the sinusoids, elevated transaminase levels, increased hepatic CD8(+) and F4/80(+) cells, overexpression of hepatic mRNA associated with inflammatory signaling pathways, steatosis, and increased collagen deposition. Immunostaining of human HCC biopsies revealed extensive deposition of complement fragments within the tumors. Investigating the Cancer Genome Atlas also revealed that increased CFH mRNA expression is associated with improved survival in patients with HCC, whereas mutations are associated with worse survival. These results indicate that CFH is critical for controlling complement activation in the liver, and in its absence, AP activation leads to chronic inflammation and promotes hepatic carcinogenesis.

Staphylococcus aureus is the leading cause of life-threatening infections, frequently originating from unknown or deep-seated foci. Source control and institution of appropriate antibiotics remain challenges, especially with infections due to methicillin-resistant S. aureus (MRSA). In this study, we developed a radiofluorinated analog of para-aminobenzoic acid (2-[18F]F-PABA) and demonstrate that it is an efficient alternative substrate for the S. aureus dihydropteroate synthase (DHPS). 2-[18F]F-PABA rapidly accumulated in vitro within laboratory and clinical (including MRSA) strains of S. aureus but not in mammalian cells. Biodistribution in murine and rat models demonstrated localization at infection sites and rapid renal elimination. In a rat model, 2-[18F]F-PABA positron emission tomography (PET) rapidly differentiated S. aureus infection from sterile inflammation and could also detect therapeutic failures associated with MRSA. These data suggest that 2-[18F]F-PABA has the potential for translation to...

Background I-124 codrituzumab (aka GC33), an antibody directed at Glypican 3, was evaluated in patients with hepatocellular carcinoma (HCC). Fourteen patients with HCC underwent baseline imaging with I-124 codrituzumab (~ 185 MBq, 10 mg). Seven of these patients undergoing sorafenib/immunotherapy with 2.5 or 5 mg/kg of cold codrituzumab had repeat imaging, with co-infusion of I-124 codrituzumab, as part of their immunotherapy treatment. Three patients who progressed while on sorafenib/immunotherapy were re-imaged after a 4-week washout period to assess for the presence of antigen. Serial positron emission tomography (PET) imaging and pharmacokinetics were performed following I-124 codrituzumab. An ELISA assay was used to determine “cold” codrituzumab serum pharmacokinetics and compare it to that of I-124 codrituzumab. Correlation of imaging results was performed with IHC. Short-term safety assessment was also evaluated. Results Thirteen patients had tumor localization on baseline I-124 codrituzumab; heterogeneity in tumor uptake was noted. In three patients undergoing repeat imaging while on immunotherapy/sorafenib, evidence of decreased I-124 codrituzumab uptake was noted. All three patients who underwent imaging after progression while on immunotherapy continued to have I-124 codrituzumab tumor uptake. Pharmacokinetics of I-124 codrituzumab was similar to that of other intact IgG. No significant adverse events were observed related to the I-124 codrituzumab. Conclusions I-124 codrituzumab detected tumor localization in most patients with HCC. Pharmacokinetics was similar to that of other intact iodinated humanized IgG. No visible cross-reactivity with normal organs was observed. Electronic supplementary material The online version of this article (10.1186/s13550-018-0374-8) contains supplementary material, which is available to authorized users.

GPA33 is a colorectal cancer (CRC) antigen with unique retention properties after huA33-mediated tumor targeting. We tested a pretargeted radioimmunotherapy (PRIT) approach for CRC using a tetravalent bispecific antibody with dual specificity for GPA33 tumor antigen and DOTA-Bn–(radiolanthanide metal) complex. PRIT was optimized in vivo by titrating sequential intravenous doses of huA33-C825, the dextran-based clearing agent, and the C825 haptens 177Lu-or 86Y-DOTA-Bn in mice bearing the SW1222 subcutaneous (s.c.) CRC xenograft model. Using optimized PRIT, therapeutic indices (TIs) for tumor radiation-absorbed dose of 73 (tumor/blood) and 12 (tumor/kidney) were achieved. Estimated absorbed doses (cGy/MBq) to tumor, blood, liver, spleen, and kidney for single-cycle PRIT were 65.8, 0.9 (TI 73), 6.3 (TI 10), 6.6 (TI 10), and 5.3 (TI 12), respectively. Two cycles of PRIT (66.6 or 111 MBq 177Lu-DOTA-Bn) were safe and effective, with a complete response of established s.c. tumors (100 – 700 mm3) in nine of nine mice, with two mice alive without recurrence at >140 days. Tumor log kill in this model was estimated to be 2.1 – 3.0 based on time to 500-mm3 tumor recurrence. In addition, PRIT dosimetry/diagnosis was performed by PET imaging of the positron-emitting DOTA hapten 86Y-DOTA-Bn. We have developed anti-GPA33 PRIT as a triple-step theranostic strategy for preclinical detection, dosimetry, and safe targeted radiotherapy of established human colorectal mouse xenografts.

The use of exosomes as a drug delivery vehicle has gained considerable interest. To establish if exosomes could be utilized effectively for drug delivery, a better understanding of their in vivo fate must be established. Through comparisons to liposomal formulations, which have been studied extensively for the last thirty years, we were able to make some comprehensive conclusions about the fate of unmodified tumor-derived exosomes in vivo. We observed a comparable rapid clearance and minimal tumor accumulation of intravenously-injected exosomes, PC:Chol liposomes, and liposomes formulated with the lipid extract of exosomes, suggesting that the unique protein and lipid composition of exosomes does not appreciably impact exosomes' rate of clearance and biodistribution. This rapid clearance along with minimal tumor accumulation of unmodified exosomes limits their use as an anti-cancer drug delivery vehicle; however, when delivered intratumorally, exosomes remained associated with tumor tissue to a significantly greater extent than PC:Chol liposomes. Furthermore, experiments utilizing mice with impaired adaptive or innate immune systems, revealed the significance of the innate immune system along with the complement protein C5 on exosomes' rate of clearance.

Codrituzumab, a humanized antibody against glypican-3, is highly expressed in HCC. A phase I study evaluated the combination with sorafenib in HCC. In a 3 + 3 design, codrituzumab was given intravenously in various doses with sorafenib 400 mg twice daily to patients with advanced HCC, age ≥18, ECOG 0–1, Child-Pugh A and B7, adequate organ functions, and no prior systemic therapy, with tumor assessment by RECIST 1.0 and safety by CTCAE 3.0. PK and pre, during, and post-therapy 124I radiolabeled codrituzumab PET scan imaging were performed. 41 patients were enrolled: 2.5 mg/kg weekly (qw) (12), 5 mg/kg qw (12), 10 mg/kg qw (3), 1600 mg every 2 weeks (q2w) (6), and 1600 mg qw (7). Two drug limiting toxicities occurred: grade 3 hyponatremia at 5 mg/kg and grade 3 hyponatremia and hyperglycemia at 1600 mg q2w. Adverse events occurred in 80% of patients, including at least one ≥grade 3: ten (25%) increased AST, three (7.5%) increased ALT, and ten (25%) increased lipase. There were no responses and nine (25.7%) had stable disease. PK C max and AUCt of codrituzumab and sorafenib were comparable to single-agent data. Thirteen out of 14 patients showed 124I radiolabeled codrituzumab uptake in tumor. In all three patients who underwent a post-progression PET, glypican-3 remained expressed. Codrituzumab plus sorafenib were tolerated at 1600 mg q2w and 400 mg bid, respectively, with no responses. Codrituzumab exerts selective distribution to HCC cells, and GPC3 does not show any down-regulation post-progression (NCT00976170).

5-[18F]F-pyrazinamide (5-[18F]F-PZA), a radiotracer analog of the first-line tuberculosis drug pyrazinamide (PZA), was employed to determine the biodistribution of PZA using PET imaging and ex vivo analysis. 5-[18F]F-PZA was synthesized in 60 min using a halide exchange reaction. The overall decay-corrected yield of the reaction was 25% and average specific activity was 2.6 × 106 kBq (70 mCi)/μmol. The biodistribution of 5-[18F]F-PZA was examined in a pulmonary Mycobacterium tuberculosis mouse model, where rapid distribution of the tracer to the lung, heart, liver, kidney, muscle, and brain was observed. The concentration of 5-[18F]F-PZA was not significantly different between infected and uninfected lung tissue. Biochemical and microbiological studies revealed substantial differences between 5-F-PZA and PZA. 5-F-PZA was not a substrate for pyrazinamidase, the bacterial enzyme that activates PZA, and the minimum inhibitory concentration for 5-F-PZA against M. tuberculosis was more than 100-fold higher than that for PZA.

Given the bone tropism of prostate cancer, conventional imaging modalities poorly identify or quantify metastatic disease. (89)Zr-huJ591 positron emission tomography (PET) imaging was performed in patients with metastatic prostate cancer to analyze and validate this as an imaging biomarker for metastatic disease. The purpose of this initial study was to assess safety, biodistribution, normal organ dosimetry, and optimal imaging time post-injection for lesion detection.Ten patients with metastatic prostate cancer received 5 mCi of (89)Zr-huJ591. Four whole-body scans with multiple whole-body count rate measurements and serum activity concentration measurements were obtained in all patients. Biodistribution, clearance, and lesion uptake by (89)Zr-huJ591 immuno-PET imaging was analyzed and dosimetry was estimated using MIRD techniques. Initial assessment of lesion targeting of (89)Zr-huJ591 was done. Optimal time for imaging post-injection was determined.The dose was well tolerated with mild chills and rigors seen in two patients. The clearance of (89)Zr-huJ591 from serum was bi-exponential with biological half-lives of 7 ± 4.5 h (range 1.1-14 h) and 62 ± 13 h (range 51-89 h) for initial rapid and later slow phase. Whole-body biological clearance was 219 ± 48 h (range 153-317 h). The mean whole-body and liver residence time was 78.7 and 25.6 h, respectively. Dosimetric estimates to critical organs included liver 7.7 ± 1.5 cGy/mCi, renal cortex 3.5 ± 0.4 cGy/mCi, and bone marrow 1.2 ± 0.2 cGy/mCi. Optimal time for patient imaging after injection was 7 ± 1 days. Lesion targeting of bone or soft tissue was seen in all patients. Biopsies were performed in 8 patients for a total 12 lesions, all of which were histologically confirmed as metastatic prostate cancer. One biopsy-proven lesion was not positive on (89)Zr-huJ591, while the remaining 11 lesions were (89)Zr-huJ591 positive. Two biopsy-positive nodal lesions were noted only on (89)Zr-huJ591 study, while the conventional imaging modality was negative.(89)Zr-huJ591 PET imaging of prostate-specific membrane antigen expression is safe and shows good localization of disease in prostate cancer patients. Liver is the critical organ for dosimetry, and 7 ± 1 days is the optimal imaging time. A larger study is underway to determine lesion detection in an expanded cohort of patients with metastatic prostate cancer.

Targeted α-particle radiation using the radioisotope 225Ac is a promising form of therapy for various types of cancer. Historic obstacles to the use of 225Ac have been the difficulty in finding suitable chelators to stably attach it to targeting vehicles such as peptides and monoclonal antibodies, the low specific activities of the products, and the lack of cost-effective radiolabeling procedures. We initially solved the first problem with a procedure involving 2 chemical steps that has been used as a standard in preclinical and clinical studies. However, this procedure involves the loss of 90% of the input 225Ac. A more efficient, economical process is needed to facilitate the more widespread use of 225Ac. Methods: We conjugated representative antibodies with 2 forms of DOTA as well as other chelators as controls. We developed conditions to radiolabel these constructs in 1 chemical step and characterized their stability, immunoreactivity, biodistribution, and therapeutic efficacy in healthy and tumor-bearing mice. Results: DOTA–antibody constructs were labeled to a wide range of specific activities in 1 chemical step at 37°C. Radiochemical yields were approximately 10-fold higher, and specific activities were up to 30-fold higher than with the previous approach. The products retained immunoreactivity and were stable to serum challenge in vitro and in mice. Labeling kinetics of DOTA–antibody constructs linked through a benzyl isothiocyanate linkage were more favorable than those linked through an N-hydroxysuccinimide linkage. Tissue distribution was similar but not identical between the constructs. The constructs produced specific therapeutic responses in a mouse model of acute myeloid leukemia. Conclusion: We have characterized an efficient, 1-step radiolabeling method that produces stable, therapeutically active conjugates of antibodies with 225Ac at high specific activity. We propose that this technology greatly expands the possible clinical applications of 225Ac monoclonal antibodies.

OBJECTIVE 68Ga-1,4,7,10-Tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA)-F(ab')2-trastuzumab [68Ga-DOTA-F(ab')2-trastuzumab] has been developed at our institution as a positron imaging reagent for assessing human epidermal growth factor receptor 2 (HER2) expression status by in-vivo imaging. Initial studies on animals demonstrated promising results in the monitoring of treatment response to heat shock protein 90-targeted drugs that inhibit the client protein HER2. We report here our initial clinical experience in the assessment of the toxicity, pharmacokinetics, biodistribution, and dosimetry profile of 68Ga-DOTA-F(ab')2-trastuzumab with PET/computed tomography using a mean of 236 MBq/5 mg administered intravenously. MATERIALS AND METHODS A group of 16 women with breast cancer were enrolled in this study. The one patient who did not receive 68Ga-DOTA-F(ab')2-trastuzumab was excluded from analysis. Both HER2-negative (n=7) and HER2-positive (n=8) cases were studied. Among the latter, seven had undergone trastuzumab treatment previously and one had not. RESULTS It was determined that 68Ga-DOTA-F(ab')2-trastuzumab was well tolerated, with a T½ of ≈ 3.6 ± 0.9 h; the critical organ was the kidney, with a mean dose of 0.383 cGy/37 MBq; and tumor targeting was seen in 4/8 patients with HER2-positive disease. CONCLUSION The reagent is safe, and assessments through additional studies in a better-defined group of patients, using larger administered masses of antibodies, with a better immunoreactive fraction are needed.

Background The current study aims to assess the safety, pharmacokinetics, feasibility, and reproducibility of immunoPET imaging with copper-64 (64Cu) trastuzumab. Methods An IV injection of 296-370 MBq/5 mg 64Cu-trastuzumab was administered between 1 to 4 hours after routine trastuzumab treatment. Whole-body PET scans were performed immediately post-injection and at 24 hours post-injection. Serial pharmacokinetics were performed. Of 11 patients (median age of 52; range of 31-61), 8 underwent a repeat study with 64Cu-trastuzumab to assess image and pharmacokinetic reproducibility. Patients were monitored for toxicity. Results Patients experienced no allergic reactions or significant adverse effects from 64Cu-trastuzumab. Eight patients successfully completed a repeat 64Cu-trastuzumab study, with acceptable reproducibility of both the biodistribution and pharmacokinetic clearance. Study 1 versus study 2 showed similar serum concentration post-injection (mean 42.4±7.8 %ID/L vs. 44.7±12.6 %ID/L) and similar T1/2 (single exponential 46.1 vs. 44.2 hours), P>0.5. The volume of distribution (median 2.50 L) was in the range reported for trastuzumab and close to the estimated plasma volume of 2.60 L. Of 11 patients, two had 64Cu-trastuzumab localization corresponding to known tumor sites - one in liver and one in breast. Conclusions Preliminary results suggest that scanning with 64Cu-trastuzumab is feasible, safe, and reproducible. Tumor uptake of 64Cu-trastuzumab was observed, but tumor detection exhibited low sensitivity in this study in which imaging was performed in the presence of trastuzumab therapy.

Heat shock protein 90 (Hsp90) is an ATP dependent molecular chaperone protein whose function is critical for maintaining several key proteins involved in survival and proliferation of cancer cells. PU-H71 (1), is a potent purine-scaffold based ATP pocket binding Hsp90 inhibitor which has been shown to have potent activity in a broad range of in vivo cancer models and is currently in Phase I clinical trials in patients with advanced solid malignancies, lymphomas, and myeloproliferative neoplasms. In this report, we describe the radiosynthesis of [(124)I]-PU-H71(5); this was synthesized from the corresponding Boc-protected stannane precursor 3 by iododestannylation with [(124)I]-NaI using chloramine-T as an oxidant for 2 min, followed by Boc deprotection with 6 N HCl at 50 °C for 30 min to yield the final compound. The final product 5 was purified using HPLC and was isolated with an overall yield of 55 ± 6% (n = 6, isolated) from 3, and >98% purity and an average specific activity of 980 mCi/µmol. Our report sets the stage for the introduction of [(124)I]-PU-H71 as a potential non-invasive probe for understanding biodistribution and pharmacokinetics of PU-H71 in living subjects using positron emission tomography imaging.

Only about 1 in 5,000 investigational agents in a preclinical stage acquires Food and Drug Administration approval. Among many reasons for this includes an inefficient transition from preclinical to clinical phases, which exponentially increase the cost and the delays the process of drug development. Positron emission tomography (PET) is a nuclear imaging technique that has been used for the diagnosis, risk stratification, and guidance of therapy. However, lately with the advance of radiochemistry and of molecular imaging technology, it became evident that PET could help novel drug development process. By using a PET radioligand to report on receptor occupancy during novel agent therapy, it may help assess the effectiveness, efficacy, and safety of such a new medication in an early preclinical stage and help design successful clinical trials even at a later phase. In this article, we explore the potential implications of PET in the development of new heart failure therapies and review PET's application in the respective pathophysiologic pathways such as myocardial perfusion, metabolism, innervation, inflammation, apoptosis, and cardiac remodeling.

Continued reliance on the androgen receptor (AR) is now understood as a core mechanism in castration-resistant prostate cancer (CRPC), the most advanced form of this disease. While established and novel AR pathway–targeting agents display clinical efficacy in metastatic CRPC, dose-limiting side effects remain problematic for all current agents. In this study, we report the discovery and development of ARN-509, a competitive AR inhibitor that is fully antagonistic to AR overexpression, a common and important feature of CRPC. ARN-509 was optimized for inhibition of AR transcriptional activity and prostate cancer cell proliferation, pharmacokinetics, and in vivo efficacy. In contrast to bicalutamide, ARN-509 lacked significant agonist activity in preclinical models of CRPC. Moreover, ARN-509 lacked inducing activity for AR nuclear localization or DNA binding. In a clinically valid murine xenograft model of human CRPC, ARN-509 showed greater efficacy than MDV3100. Maximal therapeutic response in this model was achieved at 30 mg/kg/d of ARN-509, whereas the same response required 100 mg/kg/d of MDV3100 and higher steady-state plasma concentrations. Thus, ARN-509 exhibits characteristics predicting a higher therapeutic index with a greater potential to reach maximally efficacious doses in man than current AR antagonists. Our findings offer preclinical proof of principle for ARN-509 as a promising therapeutic in both castration-sensitive and castration-resistant forms of prostate cancer. Cancer Res; 72(6); 1494–503. ©2012 AACR.

Most cancers are characterized by multiple molecular alterations, but identification of the key proteins involved in these signaling pathways is currently beyond reach. We show that the inhibitor PU-H71 preferentially targets tumor-enriched Hsp90 complexes and affinity captures Hsp90-dependent oncogenic client proteins. We have used PU-H71 affinity capture to design a proteomic approach that, when combined with bioinformatic pathway analysis, identifies dysregulated signaling networks and key oncoproteins in chronic myeloid leukemia. The identified interactome overlaps with the well-characterized altered proteome in this cancer, indicating that this method can provide global insights into the biology of individual tumors, including primary patient specimens. In addition, we show that this approach can be used to identify previously uncharacterized oncoproteins and mechanisms, potentially leading to new targeted therapies. We further show that the abundance of the PU-H71-enriched Hsp90 species, which is not dictated by Hsp90 expression alone, is predictive of the cell’s sensitivity to Hsp90 inhibition.

Imatinib mesylate is a tyrosine kinase inhibitor that was approved by the U.S. Food and Drug Administration in 2001 for treatment of many different stages of chronic myeloid leukemia and in 2002 for treatment of gastrointestinal stromal tumors. Imatinib is known to inhibit the dysregulated proliferation of chronic myeloid leukemia, which is associated with the Bcr-Abl kinase; in gastrointestinal stromal tumors, imatinib is known to act via c-Kit kinase inhibition. The objective of this study was to synthesize an 18F-labeled analog of imatinib not as a primary imaging agent but rather as a tracer for in vivo drug distribution and tracer concentration that can be used as a PET imaging surrogate for imatinib. Methods: Molecular modeling studies based on the crystal structure of imatinib bound to the active site of Abl were performed for designing the fluorinated analog. A 2-fluoroethyl analog of imatinib (SKI696) was synthesized using well-established procedures. The selectivity and binding affinity of SKI696 were compared with those of imatinib in vitro. Mice bearing K562 tumor xenografts, which are known to overexpress Bcr-Abl, were imaged with 18F-SKI696 PET. A biodistribution study was also performed on K562 tumor–bearing mice to which our radiolabeled tracer was administered. Results: The kinase assay verified that imatinib and SKI696 bind to the same targets with similar affinities. The feasibility of using 18F-SKI696 as a PET agent was examined in vivo, and 18F-SKI696 PET was shown to visualize K562 tumor xenografts in nude mice. The tumor was visible on PET 1 h after injection, with uptake of 1% of the injected dose. Biodistribution studies in K562-bearing mice were also performed, and the uptake of 18F-SKI696 (percentage injected dose per gram) for each organ was calculated. Conclusion: The results of the binding assay showed that SKI696 has selectivity and binding affinity comparable to imatinib. Small-animal PET of K562 tumor–bearing mice using 18F-SKI696 as a PET agent displayed promising tumor uptake and tumor-to-nontarget contrast. Because 18F-SKI696 has been taken up in vivo by tumors that overexpress Bcr-Abl, we are exploring a possible role for identifying tumors that will respond to imatinib before therapy.

Humanized A33 (huA33) is a promising monoclonal antibody that recognizes A33 antigen, which is present in more than 95% of colorectal cancers and in normal bowel. In this study, we took advantage of quantitative PET to evaluate 124I huA33 targeting, biodistribution, and safety in patients with colorectal cancer. We also determined the biodistribution of 124I-huA33 when a large dose of human intravenous IgG (IVIG) was administered to manipulate the Fc receptor or when 124I-huA33 was given via hepatic arterial infusion (HAI). Methods: We studied 25 patients with primary or metastatic colorectal cancer; 19 patients had surgical exploration or resection. Patients received a median of 343 MBq (44.4–396 MBq) and 10 mg of 124I-huA33. Nineteen patients received the antibody intravenously and 6 patients via HAI, and 5 patients also received IVIG. Results: Ten of 12 primary tumors were visualized in 11 patients. The median concentration in primary colon tumors was 0.016% injected dose per gram, compared with 0.004% in normal colon. The PET-based median ratio of hepatic tumor uptake to normal-liver uptake was 3.9 (range, 1.8–22.2). Quantitation using PET, compared with well counting of serum and tissue, showed little difference. Prominent uptake in bowel hindered tumor identification in some patients. Pharmacokinetics showed that patients receiving IVIG had a significantly shorter serum half-time (41.6 ± 14.0 h) than those without (65.2 ± 9.8 h). There were no differences in clearance rates among the intravenous group, IVIG group, and HAI group, nor was there any difference in serum area under the curve, maximum serum concentration, or volume of distribution. Weak titers of human–antihuman antibodies were observed in 6 of 25 patients. No acute side effects or significant toxicities were associated with huA33. Conclusion: Good localization of 124I-huA33 in colorectal cancer with no significant toxicity has been observed. PET-derived 124I concentrations agreed well with those obtained by well counting of surgically resected tissue and blood, confirming the quantitative accuracy of 124I-huA33 PET. The HAI route had no advantage over the intravenous route. No clinically significant changes in blood clearance were induced by IVIG.

Despite encouraging clinical results with next generation drugs (MDV3100 and abiraterone) that inhibit androgen receptor (AR) signaling in patients with castration-resistant prostate cancer (CRPC), responses are variable and short-lived. There is an urgent need to understand the basis of resistance to optimize their future use. We reasoned that a radiopharmaceutical that measures intratumoral changes in AR signaling could substantially improve our understanding of AR pathway directed therapies. Expanding on previous observations, we first show that prostate-specific membrane antigen (PSMA) is repressed by androgen treatment in multiple models of AR-positive prostate cancer in an AR-dependent manner. Conversely, antiandrogens up-regulate PSMA expression. These expression changes, including increased PSMA expression in response to treatment with the antiandrogen MDV3100, can be quantitatively measured in vivo in human prostate cancer xenograft models through PET imaging with a fully humanized, radiolabeled antibody to PSMA, 64 Cu-J591. Collectively, these results establish that relative changes in PSMA expression levels can be quantitatively measured using a human-ready imaging reagent and could serve as a biomarker of AR signaling to noninvasively evaluate AR activity in patients with CRPC.

Positron emission tomography (PET) of epidermal growth factor receptor (EGFR) kinase-specific radiolabeled tracers could provide a means for non-invasively characterizing EGFR expression and signaling activity in patients' tumors before, during, and after therapy with EGFR inhibitors. Towards this goal, our group has developed PET tracers which irreversibly bind to EGFR. However, tumor uptake is relatively low because of both the lipophilicity of such tracers (e.g. the morpholino-[124I]-IPQA [SKI 212243]), with octanol-to-water partition coefficients of up to 4, and a short dwell time in the blood and significant hepatobiliary clearance and intestinal reuptake. Liposomal nanoparticle delivery systems may favorably alter the pharmacokinetic profile and improve tumor targeting of highly lipophilic but otherwise promising cancer imaging tracers, such as the EGFR inhibitor SKI 243. SKI 243 is therefore an interesting model molecule for incorporation into lipid-based nanoparticles, as it would not only improve their solubility but also increase the circulation time, availability and, potentially, targeting of tumors. In the current study, we compared the pharmacokinetics and tumor targeting of the bare EGFR kinase-targeting radiotracer SKI 212243 (SKI 243) with that of the same tracer embedded in liposomes. SKI 243 and liposomal SKI 243 are both taken up by tumor xenografts but liposomal SKI 243 remained in the blood longer and consequently exhibited a 3- to 6-fold increase in uptake in the tumor among several other organs.

The human somatostatin receptor subtype 2 (hSSTr2)–68Ga-DOTATOC reporter system has several attractive features for potential translation to human studies. These include a low expression of hSSTr2 in most organs, a rapid internalized accumulation of 68Ga-DOTATOC in the SSTr2-expressing cells, and a rapid excretion of unbound radioligand by the renal system. We performed a series of in vitro and in vivo validation studies of this reporter system. Methods: A retroviral vector containing a dual reporter, pQCXhSSTr2-IRES-GFP (IRES: internal ribosome entry site; GFP: green fluorescent protein), was constructed and transduced into Jurkat, C6, and U87 cells. Stably transduced reporter cells were characterized in vitro using optical and radiometric methods. Multiple tumor-bearing mice were evaluated with 68Ga-DOTATOC PET studies. Results: The dual-reporter genes were incorporated into all tumor cell lines, and their expression levels were confirmed by fluorescence-activated cell sorting (FACS), GFP visualization, and reverse-transcriptase polymerase chain reaction (RT-PCR) analysis for hSSTr2. In vitro, hSSTr2 cell membrane expression was 36,000, 280,000, and 1,250,000 copies per cell for the SSTR2-transfected Jurkat, U87, and C6 cell lines. Small-animal PET of 68Ga-DOTATOC in tumor-bearing mice demonstrated that the in vivo uptake of this radioligand was directly proportional to the in vitro expression of hSSTr2. The in vivo uptake of 68Ga-DOTATOC, at 2 h after injection, was low in all organs except the kidneys (7.8 percentage of injected dose per gram [%ID/g]) and as high as 15.2 %ID/g in transduced C6 tumors. The corresponding transduced–to–nontransduced tumor uptake ratio was 64, and the tumor-to-muscle uptake ratio was around 500. Conclusion:68Ga-DOTATOC is an excellent specific ligand for this hSSTr2 reporter system and for hSSTr2 reporter gene PET. Because DOTATOC has undergone extensive clinical testing, this human reporter system has the potential for translation to human studies.

Antiangiogenic therapies are frequently used with concomitantly administered cancer chemotherapy to improve outcomes, but the mechanism for the benefit of the combination is uncertain. We describe a mechanism by which a specific, cytotoxic antivascular agent causes vascular remodeling and improved chemotherapy results. By selectively killing tumor neovasculature using short-ranged α-particles targeted to vascular endothelial (VE)-cadherin on vascular endothelial cells (by use of 225Ac-labeled E4G10 antibody) we were able both to reduce tumor growth and to increase the efficacy of chemotherapy, an effect seen only when the chemotherapy was administered several days after the vascular targeting agent, but not if the order of administration was reversed. Immunohistochemical and immunofluorescence studies showed that the vasculature of 225Ac-E4G10–treated tumors was substantially depleted; the remaining vessels appeared more mature morphologically and displayed increased pericyte density and coverage. Tumor uptake and microdistribution studies with radioactive and fluorescent small molecule drugs showed better accumulation and more homogenous distribution of the drugs within 225Ac-E4G10–treated tumors. These results show that 225Ac-E4G10 treatment leads to ablation and improvement of the tumor vascular architecture, and also show that the resulting vascular remodeling can increase tumor delivery of small molecules, thus providing a process for the improved outcomes observed after combining antivascular therapy and chemotherapy. This study directly shows evidence for what has long been a speculated mechanism for antiangiogenic therapies. Moreover, targeting the vessel for killing provides an alternative mode of improving chemotherapy delivery and efficacy, potentially avoiding some of the drawbacks of targeting a highly redundant angiogenic pathway. Cancer Res; 70(22); 9277–86. ©2010 AACR.

Radioimmunotherapy (RIT) using (131)I-3F8 injected into cerebrospinal fluid (CSF) was a safe modality for the treatment of leptomeningeal metastases (JCO, 25:5465, 2007). A single-compartment pharmacokinetic model described previously (JNM 50:1324, 2009) showed good fitting to the CSF radioactivity data obtained from patients. We now describe a two-compartment model to account for the ventricular reservoir of (131)I-3F8 and to identify limiting factors that may impact therapeutic ratio.Each parameter was examined for its effects on (1) the area under the radioactivity concentration curve of the bound antibody (AUC[C(IAR)]), (2) that of the unbound antibody AUC[C(IA)], and (3) their therapeutic ratio (AUC[C(IAR)]/AUC[C(IA)]).Data fitting showed that CSF kBq/ml data fitted well using the two-compartment model (R = 0.95 ± 0.03). Correlations were substantially better when compared to the one-compartment model (R = 0.92 ± 0.11 versus 0.77 ± 0.21, p = 0.005). In addition, we made the following new predictions: (1) Increasing immunoreactivity of (131)I-3F8 from 10% to 90% increased both (AUC[C(IAR)]) and therapeutic ratio ([AUC[C(IAR)]/AUC[C(IA)]] by 7.4 fold, (2) When extrapolated to the clinical setting, the model predicted that if (131)I-3F8 could be split into 4 doses of 1.4 mg each and given at ≥24 hours apart, an antibody affinity of K(D) of 4 × 10(-9) at 50% immunoreactivity were adequate in order to deliver ≥100 Gy to tumor cells while keeping normal CSF exposure to10 Gy.This model predicted that immunoreactivity, affinity and optimal scheduling of antibody injections were crucial in improving therapeutic index.

The CA125 antigen, recognized by the OC125 antibody, is a tissue-specific, circulating antigen expressed in ovarian cancer. The CA125 antigen is encoded by the MUC16 gene, cloned by Lloyd and Yin. The full-length gene describes a complex tethered mucin protein present primarily in a variety of gynecologic tissues, especially neoplasms. OC125 and other related antibodies react with glycosylation-dependent antigens present exclusively in the cleaved portion of the molecule. These antibodies are not useful as screening tools, nor can they detect the proximal residual MUC16 protein fragment after cleavage. This has limited its diagnostic and therapeutic applications. Using synthetic peptides we raised novel-specific antibodies to the carboxy-terminal portion of MUC16, retained by the cell, proximal to the putative cleavage site. These antibodies were characterized using fluorescence-activated cell-sorting analysis, enzyme-linked immunoassay, Western blot analysis, and immunohistochemistry. Each of the selected monoclonal antibodies was reactive against recombinant GST-ΔMUC16c114 protein and the MUC16 transfected SKOV3 cell line. Three antibodies, 4H11, 9C9, and 4A5 antibodies demonstrated high affinities by Western blot analysis and saturation-binding studies of transfected SKOV3 cells, and displayed antibody internalization. Immunohistochemical positivity with novel antibody 4H11 was similar to OC125, but with important differences, including diffuse positivity in lobular breast cancer and a small percentage of OC125-negative ovarian carcinomas that showed intense and diffuse 4H11. Development of such antibodies may be useful for the characterization of MUC16 biology and allow for future studies in targeted therapy and diagnostics.

89Zr (half-life, 78.41 h) is a positron-emitting radionuclide that displays excellent potential for use in the design and synthesis of radioimmunoconjugates for immunoPET. In the current study, we report the preparation of 89Zr-desferrioxamine B (DFO)-J591, a novel 89Zr-labeled monoclonal antibody (mAb) construct for targeted immunoPET and quantification of prostate-specific membrane antigen (PSMA) expression in vivo. Methods: The in vivo behavior of 89Zr-chloride, 89Zr-oxalate, and 89Zr-DFO was studied using PET. High-level computational studies using density functional theory calculations have been used to investigate the electronic structure of 89Zr-DFO and probe the nature of the complex in aqueous conditions. 89Zr-DFO-J591 was characterized both in vitro and in vivo. ImmunoPET in male athymic nu/nu mice bearing subcutaneous LNCaP (PSMA-positive) or PC-3 (PSMA-negative) tumors was conducted. The change in 89Zr-DFO-J591 tissue uptake in response to high- and low-specific-activity formulations in the 2 tumor models was measured using acute biodistribution studies and immunoPET. Results: The basic characterization of 3 important reagents—89Zr-chloride, 89Zr-oxalate, and the complex 89Zr-DFO—demonstrated that the nature of the 89Zr species dramatically affects the biodistribution and pharmacokinetics. Density functional theory calculations provide a rationale for the observed high in vivo stability of 89Zr-DFO–labeled mAbs and suggest that in aqueous conditions, 89Zr-DFO forms a thermodynamically stable, 8-coordinate complex by coordination of 2 water molecules. 89Zr-DFO-J591 was produced in high radiochemical yield (>77%) and purity (>99%), with a specific activity of 181.7 ± 1.1 MBq/mg (4.91 ± 0.03 mCi/mg). In vitro assays demonstrated that 89Zr-DFO-J591 had an initial immunoreactive fraction of 0.95 ± 0.03 and remained active for up to 7 d. In vivo biodistribution experiments revealed high, target-specific uptake of 89Zr-DFO-J591 in LNCaP tumors after 24, 48, 96, and 144 h (34.4 ± 3.2 percentage injected dose per gram [%ID/g], 38.0 ± 6.2 %ID/g, 40.4 ± 4.8 %ID/g, and 45.8 ± 3.2 %ID/g, respectively). ImmunoPET studies also showed that 89Zr-DFO-J591 provides excellent image contrast, with tumor-to-muscle ratios greater than 20, for the delineation of LNCaP xenografts between 48 and 144 h after administration. Conclusion: These studies demonstrate that 89Zr-DFO–labeled mAbs show exceptional promise as radiotracers for immunoPET of human cancers. 89Zr-DFO-J591 displays high tumor–to–background tissue contrast in immunoPET and can be used to delineate and quantify PSMA-positive prostate tumors in vivo.

Memorial Sloan-Kettering Center The aim of this study was to develop a clinically applicable non-invasive method to quantify changes in androgen receptor (AR) levels based on F-18-16 beta-fluoro-5 alpha-dihydrotestosterone (F-18-FDHT) PET in prostate cancer patients undergoing therapy. Methods: Thirteen patients underwent dynamic F-18-FDHT PET over a selected tumor. Concurrent venous blood samples were acquired for blood metabolite analysis. A second cohort of 25 patients injected with F-18-FDHT underwent dynamic PET of the heart. These data were used to generate a population-based input function, essential for pharmacokinetic modeling. Linear compartmental pharmacokinetic models of increasing complexity were tested on the tumor tissue data. Four suitable models were applied and compared using the Bayesian information criterion (BIC). Model 1 consisted of an instantaneously equilibrating space, followed by a unidirectional trap. Models 2a and 2b contained a reversible space between the instantaneously equilibrating space and the trap, into which metabolites were excluded (2a) or allowed (2b). Model 3 built on model 2b with the addition of a second reversible space preceding the unidirectional trap and from which metabolites were excluded. Results: the half-life of the F-18-FDHT in blood was between 6 and 7 min. As a consequence, the uptake of F-18-FDHT in prostate cancer lesions reached a plateau within 20 min as the blood-borne activity was consumed. Radiolabeled metabolites were shown not to bind to ARs in in vitro studies with CWR22 cells. Model 1 produced reasonable and robust fits for all datasets and was judged best by the BIC for 16 of 26 tumor scans. Models 2a, 2b, and 3 were judged best in 7, 2, and 1 cases, respectively. Conclusion: Our study explores the clinical potential of using F-18-FDHT PET to estimate free AR concentration. This process involved the estimation of a net uptake parameter such as the k(trap) of model 1 that could serve as a surrogate measure of AR expression in met-astatic prostate cancer. Our initial studies suggest that a simple body mass-normalized standardized uptake value correlates reasonably well to model-based k(trap) estimates, which we surmise may be proportional to AR expression. Validation studies to test this hypothesis are underway. Mem Sloan Kettering Canc Ctr, Dept Med Phys, New York, NY 10021 USA Mem Sloan Kettering Canc Ctr, Dept Neurol, New York, NY 10021 USA Mem Sloan Kettering Canc Ctr, Dept Radiol, New York, NY 10021 USA St Lukes Roosevelt Hosp, Dept Radiol, New York, NY 10025 USA Mem Sloan Kettering Canc Ctr, Dept Med, Genitourinary Oncol Serv, New York, NY 10021 USA Universidade Federal de São Paulo, Dept Radiol, São Paulo, Brazil Stanford Univ, Dept Radiol & Bioengn, Bio X Program, Stanford, CA 94305 USA Department of Radiology, Federal University of Sao Paulo, São Paulo, Brazil Memorial Sloan-Kettering Center: P50-CA92629 Memorial Sloan-Kettering Center: P50 CA086438 Memorial Sloan-Kettering Center: K23: CA102544 Web of Science

Innovation in the management of brain metastases is needed. We evaluated the addition of compartmental intrathecal antibody-based radioimmunotherapy (cRIT) in patients with recurrent metastatic central nervous system (CNS) neuroblastoma following surgery, craniospinal irradiation, and chemotherapy. Twenty one patients treated for recurrent neuroblastoma metastatic to the CNS, received a cRIT-containing salvage regimen incorporating intrathecal (131)I-monoclonal antibodies (MoAbs) targeting GD2 or B7H3 following surgery and radiation. Most patients also received outpatient craniospinal irradiation, 3F8/GMCSF immunotherapy, 13-cis-retinoic acid and oral temozolomide for systemic control. Seventeen of 21 cRIT-salvage patients are alive 7-74 months (median 33 months) since CNS relapse, with all 17 remaining free of CNS neuroblastoma. One patient died of infection at 22 months with no evidence of disease at autopsy, and one of lung and bone marrow metastases at 15 months, and one of progressive bone marrow disease at 30 months. The cRIT-salvage regimen was well tolerated, notable for myelosuppression minimized by stem cell support (n = 5), and biochemical hypothyroidism (n = 5). One patient with a 7-year history of metastatic neuroblastoma is in remission from MLL-associated secondary leukemia. This is significantly improved to published results with non-cRIT based where relapsed CNS NB has a median time to death of approximately 6 months. The cRIT-salvage regimen for CNS metastases was well tolerated by young patients, despite their prior history of intensive cytotoxic therapies. It has the potential to increase survival with better than expected quality of life.

Large structural allografts used for reconstruction of bone defects after revision arthroplasty and tumor resection fracture up to 27% of the time from osteolytic resorption around the fixation screw holes and tendon or ligament attachment sites. Treating structural allografts before implantation with bisphosphonates may inhibit local osteoclastic processes and prevent bone resorption and the development of stress risers, thereby reducing the long-term fracture rate. Taking advantage of allografts’ open-pore structure, we asked whether passive soaking or positive-pressure pumping was a more efficient technique for delivering bisphosphonates. We treated matched pairs of ovine tibial allografts with fluids containing Tc-99m pamidronate and toluidine blue stain to facilitate indicator distribution analysis via microSPECT-microCT imaging and light microscopy, respectively. Surfactants octylphenoxy polyethoxy ethanol or beractant were added to the treatment fluids to reduce flow resistance of solutions pumped through the allografts. Indicator distribution after 1 hour of soaking produced a thin ring around periosteal and endosteal surfaces, while pumping for 10 minutes produced a more even distribution throughout the allograft. Flow resistance was reduced with octylphenoxy polyethoxy ethanol but unaffected with beractant. Pumped allografts displayed a more homogeneous indicator distribution in less time than soaking while surfactants enhanced fluid movement.

The in vitro and in vivo behavior of the radiolabeled monoclonal antibody MORAb-003 was investigated as a prelude to a clinical trial.The cellular retention of 111In- and 131I-labeled MORAb-003 was investigated using IGROV1 and SW620 cells. Biodistribution studies in tumor-bearing mice were performed with the more favorable agent.Five 1,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetraacetic acid (DOTA) molecules were conjugated to MORAb-003 with no apparent loss of immunoreactivity. Radiolabeled MORAb-003 had a high affinity for the folate receptor alpha (FRA) expressed by both IGROV1 and SW620 cells and was found to bind to around 8 x 10(5) and 7 x 10(5) sites/cell, respectively. Both cancer cell lines were found to internalize both 131I- and 111In-labeled MORAb-003, but 111In was retained and 131I was released as iodide. In athymic mice, 111In-DOTA-MORAb-003 was cleared from the blood with a single exponential biological clearance rate of 110 h. The uptake in SW620 tumors was 32+/-5%ID/g after 4 days. The clearance rate of activity from normal organs such as liver, kidney and spleen was similar to the blood clearance and was 5.36%ID/g, 4.03%ID/g and 4.36%ID/g at 1 day postinjection and 2.14%ID/g, 1.65%ID/g and 3.74%ID/g after 8 days, respectively. In a pilot clinical study, the biodistribution and tumor targeting of 111In-MORAb-003 was assessed in three patients undergoing treatment with cold MORAb-003.MORAb-003 is an attractive antibody for radioimmunoscintigraphy and possibly radioimmunotherapy of FRA-expressing cancers in addition to its potential direct therapeutic effects.

In severe pulmonary hypertension (SPH), prior studies have shown an increase in right ventricle (RV) uptake of glucose, but it is unclear whether there is a change in the relative utilization of fatty acids. We hypothesized that in the RV in SPH, as in left ventricular (LV) failure, there is altered substrate utilization, with increased glucose uptake and decreased fatty acid uptake. SPH was induced in rats by treatment with the VEGF receptor inhibitor SU5416 and 3 wk of hypoxia (10% FiO2), followed by an additional 4 wk of normoxia (SU-Hx group). Control rats were treated with carboxymethylcellulose vehicle and 7 wk of normoxia (CMC-Nx group). The rodents then underwent positron emission tomography with sequential administration of two radiotracers, 2-deoxy-2-[18F]fluoroglucose (18F-FDG) and 14-(R,S)-[18F]fluoro-6-thia-heptadecanoic acid (18F-FTHA), analogs of glucose and fatty acid, respectively. Five CMC-Nx and 3 SU-Hx rats completed the entire experimental protocol. In the RV, there was a mild increase in 18F-FDG uptake (1.35-fold, P = 0.085) and a significant decrease in 18F-FTHA uptake (−2.1-fold, P < 0.05) in the SU-Hx rats relative to the CMC-Nx rats. In the LV, SU-Hx rats had less uptake of both radiotracers compared with CMC-Nx rats. Less RV fatty acid uptake in SPH was corroborated by decreased fatty acid transporters and enzymes in the RV tissue, and specifically a decrease in lipoprotein lipase. In the RV in rats with SPH, there is a major shift in metabolic substrate preference, largely due to decreased fatty acid uptake.

The molecular specificity of monoclonal antibodies (mAbs) directed against tumor antigens has proven effective for targeted therapy of human cancers, as shown by a growing list of successful antibody-based drug products. We describe a novel, nonlinear compartmental model using PET-derived data to determine the "best-fit" parameters and model-derived quantities for optimizing biodistribution of intravenously injected (124)I-labeled antitumor antibodies.As an example of this paradigm, quantitative image and kinetic analyses of anti-A33 humanized mAb (also known as "A33") were performed in 11 colorectal cancer patients. Serial whole-body PET scans of (124)I-labeled A33 and blood samples were acquired and the resulting tissue time-activity data for each patient were fit to a nonlinear compartmental model using the SAAM II computer code.Excellent agreement was observed between fitted and measured parameters of tumor uptake, "off-target" uptake in bowel mucosa, blood clearance, tumor antigen levels, and percent antigen occupancy.This approach should be generally applicable to antibody-antigen systems in human tumors for which the masses of antigen-expressing tumor and of normal tissues can be estimated and for which antibody kinetics can be measured with PET. Ultimately, based on each patient's resulting "best-fit" nonlinear model, a patient-specific optimum mAb dose (in micromoles, for example) may be derived.

Background: Prostate-specific membrane antigen (PSMA) is a transmembrane glycoprotein primarily expressed on benign and malignant prostatic epithelial cells. J591 is an IgG1 monoclonal antibody that targets the external domain of the PSMA. The relationship among dose, safety, pharmacokinetics, and antibody-dependent cellular cytotoxicity (ADCC) activation for unlabeled J591 has not been explored. Patients and Methods: Patients with progressive metastatic prostate cancer despite androgen deprivation were eligible. Each patient received 10, 25, 50, and 100 mg of J591. Two milligrams of antibody, conjugated with the chelate 1,4,7,10-tetraazacyclododecane-N, N′,N″,N‴-tetraacetic acid, were labeled with 5 mCi indium-111 (111In) as a tracer. One group of patients received unlabeled J591 before the labeled antibody; the other received both together. Toxicities, pharmacokinetic properties, biodistribution, ADCC induction, immunogenicity, and clinical antitumor effects were assessed. Results: Fourteen patients were treated (seven in each group). Treatment was well tolerated. Biodistribution of 111In-labeled J591 was comparable in both groups. The mean T1/2 was .96, 1.9, 2.75, and 3.47 days for the 10, 25, 50, and 100 mg doses, respectively. Selective targeting of 111In-labeled J591 to tumor was seen. Hepatic saturation occurred by the 25-mg dose. ADCC activity was proportional to dose. One patient showed a >50% prostate-specific antigen decline. Conclusions: J591 is well tolerated in repetitive dose-escalating administrations. The rate of serum clearance decreases with increasing antibody mass. ADCC activation is proportional to antibody mass. The optimal dose is 25 mg for radioimmunotherapy and 100 mg for immunotherapy. Phase II studies using J591 as a radioconjugate are under way.

Purpose: To compare two potential positron emission tomography (PET) tracers of tumor hypoxia in an animal model. Methods and Materials: The purported hypoxia imaging agents 18F-fluoromisonidazole (FMISO) and 64Cu(II)-diacetyl-bis(N4-methylthiosemicarbazone) (Cu-ATSM) were compared by serial microPET imaging of Fisher-Copenhagen rats bearing the R3327-AT anaplastic rat prostate tumor. Probe measurements of intratumoral P o 2 were compared with the image data. At the microscopic level, the relationship between the spatial distributions of 64Cu (assessed by digital autoradiography) and tumor hypoxia (assessed by immunofluorescent detection of pimonidazole) was examined. 18F-FMISO and 64Cu-ATSM microPET images were also acquired in nude rats bearing xenografts derived from the human squamous cell carcinoma cell line, FaDu. Results: In R3327-AT tumors, the intratumoral distribution of 18F-FMISO remained relatively constant 1–4 h after injection. However, that of 64Cu-ATSM displayed a significant temporal evolution for 0.5–20 h after injection in most tumors. In general, only when 64Cu-ATSM was imaged at later times (16–20 h after injection) did it correspond to the distribution of 18F-FMISO. Oxygen probe measurements were broadly consistent with 18F-FMISO and late 64Cu-ATSM images but not with early 64Cu-ATSM images. At the microscopic level, a negative correlation was found between tumor hypoxia and 64Cu distribution when assessed at early times and a positive correlation when assessed at later times. For the FaDu tumor model, the early and late 64Cu-ATSM microPET images were similar and were in general concordance with the 18F-FMISO scans. Conclusion: The difference in behavior between the R3327-AT and FaDu tumor models suggests a tumor-specific dependence of Cu-ATSM uptake and retention under hypoxic conditions.

We performed an interim analysis of imaging data collected in 2 phase I radioimmunotherapy trials to determine the ability of monoclonal antibody (mAb) J591 directed to the extracellular domain of prostate specific membrane antigen (PSMA) to target sites of known metastatic prostate cancer accurately.Patients with progressing hormone independent prostate cancer were entered in 2 phase I dose finding trials with radiolabeled mAb J591. J591 is the first mAb targeting the extracellular domain of PSMA as well as the first de-immunized (humanized) mAb to PSMA to be tested in humans. These trials were primarily designed to assess dose limiting toxicity, maximum tolerated dose, pharmacokinetics and organ dosimetry. Planar gamma camera imaging studies obtained on the first 53 patients were reviewed and compared to sites of metastatic prostate cancer visualized on conventional imaging studies including bone scan, computerized tomography and/or magnetic resonance imaging. In 1 trial 29 patients received 111indium-J591 for imaging followed by 90yttrium-J591 for therapy. In the parallel trial 24 patients were treated with 177lutetium-J591, an isotope that can be imaged directly.Of 53 patients reviewed 46 (87%) had evidence of metastatic disease on conventional scans. Overall, of the 43 evaluable patients J591 accurately targeted bone and/or soft tissue lesions in 42 (98%). J591 accurately targeted bone lesions in 32 of 34 (94%) and soft tissue lesions in 13 of 18 (72%) evaluable patients.Radiolabeled J591 accurately targets bone and soft tissue metastatic prostate cancer sites, and may be useful for targeting therapeutic and/or diagnostic imaging agents.

A method for conjugation of ligands to the surface of exosomes was developed using click chemistry. Copper-catalyzed azide alkyne cycloaddition (click chemistry) is ideal for biocojugation of small molecules and macromolecules to the surface of exosomes, due to fast reaction times, high specificity, and compatibility in aqueous buffers. Exosomes cross-linked with alkyne groups using carbodiimide chemistry were conjugated to a model azide, azide-fluor 545. Conjugation had no effect on the size of exosomes, nor was there any change in the extent of exosome adherence/internalization with recipient cells, suggesting the reaction conditions were mild on exosome structure and function. We further investigated the extent of exosomal protein modification with alkyne groups. Using liposomes with surface alkyne groups of a similar size and concentration to exosomes, we estimated that approximately 1.5 alkyne groups were present for every 150 kDa of exosomal protein.

Long acting somatostatin-14 (SST) analogs are used clinically to inhibit tumor growth and proliferation of various tumor types via binding to specific receptors (R). We have developed a 111In-/90Y-labeled SST analog, DOTA-(d)βNal1-lanreotide (DOTALAN), for tumor diagnosis and therapy. 111In-/90Y-DOTALAN bound with high affinity (dissociation constant, Kd, 1–12 nm) to a number of primary human tumors (n = 31) such as intestinal adenocarcinoma (n = 17; 150-4000 fmol/mg protein) or breast cancer (n = 4; 250-9000 fmol/mg protein). 111In-/90Y-DOTALAN exhibited a similar high binding affinity (Kd, 1–15 nm) for the human breast cancer cell lines T47D and ZR75–1, the prostate cancer cell lines PC3 and DU145, the colonic adenocarcinoma cell line HT29, the pancreatic adenocarcinoma cell line PANC1, and the melanoma cell line 518A2. When expressed in COS7 cells, 111In-DOTALAN bound with high affinity to hsst2 (Kd, 4.3 nm), hsst3 (Kd, 5.1 nm), hsst4 (Kd, 3.8 nm), and hsst5 (Kd, 10 nm) and with lower affinity to hsst1...

Item does not contain fulltext BACKGROUND: Patients with metastatic clear cell renal cell carcinoma (ccRCC) have a dismal prognosis. Therefore, new and less toxic treatments are needed. OBJECTIVE: We determined the maximum tolerated dose (MTD) and potential therapeutic efficacy of multiple infusions of lutetium 177 ((177)Lu)-girentuximab (cG250) on various dose levels in a phase 1 trial in patients with progressive metastasized ccRCC. DESIGN, SETTING, AND PARTICIPANTS: In this uncontrolled case series in 23 patients with progressive ccRCC metastases, cG250 accumulation was verified by diagnostic indium 111-cG250 imaging. Patients then received a high-activity dose of (177)Lu-cG250. INTERVENTION: Groups of three patients received (177)Lu-cG250, starting at a dose level of 1110 MBq/m(2)(177)Lu-cG250, with dose increments of 370 MBq/m(2) per group. In the absence of persistent toxicity, progressive disease, and accelerated blood clearance, patients were eligible for retreatment after 3 mo with 75% of the previous activity dose. Patients could receive a total of three treatment cycles. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Determination of the MTD was the primary and therapeutic efficacy was the secondary outcome measurement of the study. RESULTS AND LIMITATIONS: The MTD was 2405 MBq/m(2) because higher doses resulted in dose-limiting myelotoxicity. Some patients received second (13 of 23 [56%]) and third (4 of 23 [17%]) treatment cycles. Most patients (17 of 23 [74%]) demonstrated stable disease 3 mo after the first treatment, and one patient showed a partial response that lasted for 9 mo. Mean growth of target tumor lesions was reduced from 40.4% (95% confidence interval [CI], +/-17.0) during the last 3 mo before study entry to 5.5% (95% CI, +/-5.3; p

Bombesin receptors are under intense investigation as molecular targets since they are overexpressed in several prevalent solid tumors. We rationally designed and synthesized a series of modified bombesin (BN) peptide analogs to study the influence of charge and spacers at the N-terminus, as well as amino acid substitutions, on both receptor binding affinity and pharmacokinetics. This enabled development of a novel (64/67)Cu-labeled BN peptide for PET imaging and targeted radiotherapy of BN receptor-positive tumors. Our results show that N-terminally positively charged peptide ligands had significantly higher affinity to human gastrin releasing peptide receptor (GRPr) than negatively charged or uncharged ligands (IC(50): 3.2±0.5 vs 26.3±3.5 vs 41.5±2.5 nM). The replacement of Nle(14) by Met, and deletion of D-Tyr(6), further resulted in 8-fold higher affinity. Contrary to significant changes to human GRPr binding, modifications at the N-terminal and at the 6(th), 11(th), and 14(th) position of BN induced only slight influences on affinity to mouse GRPr. [Cu(II)]-CPTA-[βAla(11)] BN(7-14) ([Cu(II)]-BZH7) showed the highest internalization rate into PC-3 cells with relatively slow efflux because of its subnanomolar affinity to GRPr. Interestingly, [(64/67)Cu]-BZH7 also displayed similar affinities to the other 2 human BN receptor subtypes. In vivo studies showed that [(64/67)Cu]-BZH7 had a high accumulation in PC-3 xenografts and allowed for clear-cut visualization of the tumor in PET imaging. In addition, a CPTA-glycine derivative, forming a hippurane-type spacer, enhanced kidney clearance of the radiotracer. These data indicate that the species variation of BN receptor plays an important role in screening radiolabeled BN. As well, the positive charge from the metallated complex at the N-terminal significantly increases affinity to human GRPr. Application of these observations enabled the novel ligand [(64/67)Cu]-BZH7 to clearly visualize PC-3 tumors in vivo. This study provides a strong starting point for optimizing radiopeptides for targeting carcinomas that express any of the BN receptor subtypes.

The primary aim of this analysis was to examine the quantitative features of antibody–antigen interactions in tumors and normal tissue after parenteral administration of antitumor antibodies to human patients. Methods: Humanized anti-A33 antibody (10 mg) labeled with the positron-emitting radionuclide 124I (124I-huA33) was injected intravenously in 15 patients with colorectal cancer. Clinical PET/CT was performed approximately 1 wk later, followed by a detailed assay of surgically removed tissue specimens including radioactivity counting, autoradiography, immunohistochemistry, and antigen density determination. Results: PET/CT showed high levels of antibody targeting in tumors and normal bowel. In tissue specimens, the spatial distribution of 124I-huA33 conformed to that of A33 antigen, and there was a linear relationship between the amount of bound antibody and antigen concentration. Antibody uptake was high in 1- to 2-mm regions of antigen-positive tumor cells (mean, ∼0.05 percentage injected dose per gram) and in antigen-positive normal colonic mucosa (mean, ∼0.03 percentage injected dose per gram). The estimated binding site occupancy for tumor and normal colon was 20%–50%. Conclusion: The in vivo biodistribution of 124I-huA33 in human patients 1 wk after antibody administration was determined by A33 antigen expression. Our data imply that the optimal strategy for A33-based radioimmunotherapy of colon cancer will consist of a multistep treatment using a radionuclide with short-range (α- or β-particle) emissions.

Introduction The present investigation focuses on the chemical and biological fate of 89 Zr in mice. Electrophoreses of 89 Zr solvated or chelated in different conditions are here presented. The biological fate of mice injected with [ 89 Zr]Zr-oxalate, [ 89 Zr]Zr-chloride, [ 89 Zr]Zr-phosphate, [ 89 Zr]Zr-desferrioxamine and [ 89 Zr]Zr-citrate is studied with the biodistribution, the clearances and positron emission tomography images. A special focus is also given regarding the quality of 89 Zr bone accumulation. Methods Electrophoreses were carried out on chromatography paper and read by gamma counting. Then, the solutions were intravenously injected in mice, imaged at different time points and sacrificed. The bones, the epiphysis and the marrow substance were separated and evaluated with gamma counts. Results The clearances of [ 89 Zr]Zr-chloride and [ 89 Zr]Zr-oxalate reached 20% of injected dose (ID) after 6 days whereas [ 89 Zr]Zr-phosphate was only 5% of ID. [ 89 Zr]Zr-citrate and [ 89 Zr]Zr-DFO were noticeably excreted after the first day postinjection (p.i.). [ 89 Zr]Zr-chloride and [ 89 Zr]Zr-oxalate resulted in a respective bone uptake of ∼15% ID/g and∼20% ID/g at 8 h p.i. with minor losses after 6 days. [ 89 Zr]Zr-citrate bone uptake was also observed, but [ 89 Zr]Zr-phosphate was absorbed in high amounts in the liver and the spleen. The marrow cells were insignificantly radioactive in comparison to the calcified tissues. Conclusion Despite the complexity of Zr coordination, the electrophoretic analyses provided detailed evidences of Zr charges either as salts or as complexes. This study also shows that weakly chelated, 89 Zr is a bone seeker and has a strong affinity for phosphate.