RESUMO
Copper (Cu) is a critical element for cancer cell proliferation and considerably accumulates in the nucleus. 64Cu2+ is an anticancer radiopharmaceutical that targets the copper requirement of cancer cells. However, intravenously injected 64Cu2+ ions primarily accumulate in the liver. Ligand complexation of 64Cu2+ may be a promising method for increasing tumor delivery by reducing liver uptake. In this study, we used three tripodal amine ligands [tris(2-aminoethyl)amine (Tren), diethylenetriamine (Dien), and tris(2-pyridylmethyl)amine (TPMA)] to enclose 64Cu2+ ions and compared their in vivo tumor and liver uptakes using a tumor-bearing xenograft mouse model of the extrahepatic bile duct carcinoma cell line TFK-1. We examined intracellular Cu distribution using microparticle-induced X-ray emission (micro-PIXE) analysis of these compounds. 64Cu2+-Tren and 64Cu2+-Dien showed higher tumor uptake than 64Cu2+-TPMA and 64Cu2+ ions in TFK-1 tumors. Among the three 64Cu2+ complexes and 64Cu2+ ions, liver uptake was inversely correlated with tumor uptake. Micro-PIXE analysis showed that in vitro cellular uptake was similar to in vivo tumor uptake, and nuclear delivery was the highest for 64Cu2+-Tren. Conclusively, an inverse correlation between tumor and liver uptake was observed using three 64Cu2+ complexes of tripodal amine ligands and 64Cu2+ ions. These results provide useful information for the future development of anticancer 64Cu radiopharmaceuticals.
RESUMO
Managing metastasis at the early stage and detecting and treating submillimeter tumors at early metastasis are crucial for improving cancer prognosis. Angiogenesis is a critical target for developing drugs to detect and inhibit submillimeter tumor growth; however, drug development remains challenging because there are no suitable models for observing the submillimeter tumor mass and the surrounding blood vessels in vivo. We have established a xenograft subcutaneous submillimeter tumor mouse model with HT-29-RFP by transplanting a single spheroid grown on radiation-crosslinked gelatin hydrogel microwells. Here, we developed an in vivo dual-observation method to observe the submillimeter tumor mass and tumor-surface blood vessels using this model. RFP was detected to observe the tumor mass, and a fluorescent angiography agent FITC-dextran was administered to observe blood vessels via stereoscopic fluorescence microscopy. The anti-angiogenesis agent regorafenib was used to confirm the usefulness of this method. This method effectively detected the submillimeter tumor mass and tumor-surface blood vessels in vivo. Regorafenib treatment revealed tumor growth inhibition and angiogenesis downregulation with reduced vascular extremities, segments, and meshes. Further, we confirmed that tumor-surface blood vessel areas monitored using in vivo dual-observation correlated with intratumoral blood vessel areas observed via fluorescence microscopy with frozen sections. In conclusion, this method would be useful in developing anti-angiogenesis agents against submillimeter tumors.
Assuntos
Inibidores da Angiogênese , Neoplasias , Humanos , Camundongos , Animais , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/diagnóstico , Proteínas de Fluorescência Verde , Neoplasias/diagnóstico por imagem , Neoplasias/tratamento farmacológico , Neoplasias/patologiaRESUMO
Understanding the physicochemical properties of antibody-drug conjugates is critical to assess their quality at manufacturing and monitor them during subsequent storage. For radiometal-antibody complexes, it is important to control the properties of the antibody-chelator conjugate to maintain the quality of the final product. We have been developing 64Cu-labeled anti-epidermal growth factor receptor antibody NCAB001 (64Cu-NCAB001) for the early diagnosis and therapy of pancreatic cancer with positron-emission tomography. Here, we characterized the larger size variants contained in the antibody-chelator conjugate PCTA-NCAB001 by multi-angle light scattering coupled with size-exclusion chromatography. Secondly, we developed a chromatographic method to remove these size variants. Lastly, we demonstrated the stability of PCTA-NCAB001 after the removal of size variants. Dimer and oligomers were identified in PCTA-NCAB001. These larger size variants, together with some smaller size variants, could be removed by hydrophobic interaction chromatography. The PCTA-NCAB001 product, after the removal of these size variants, could be stored at 4 °C for six months. The methods developed here can be applied to assure the quality of PCTA-NCAB001 and other antibody-drug conjugates to facilitate the development of antibody-radiometal conjugates for positron-emission tomography and radioimmunotherapy of malignant cancers.
RESUMO
[64Cu]Cu-diacetyl-bis(N4-methylthiosemicarbazone) ([64Cu]Cu-ATSM) is a radioactive hypoxia-targeting therapeutic agent being investigated in clinical trials for malignant brain tumors. For the quality management of [64Cu]Cu-ATSM, understanding trace metal impurities' effects on the chelate formation of 64Cu and ATSM is important. In this study, we conducted coordination chemistry studies on metal-ATSM complexes. First, the effects of nonradioactive metal ions (Cu2+, Ni2+, Zn2+, and Fe2+) on the formation of [64Cu]Cu-ATSM were evaluated. When the amount of Cu2+ or Ni2+ added was 1.2 mol or 288 mol, equivalent to ATSM, the labeling yield of [64Cu]Cu-ATSM fell below 90%. Little effect was observed even when excess amounts of Zn2+ or Fe2+ were added to the ATSM. Second, these metals were reacted with ATSM, and chelate formation was measured using ultraviolet-visible (UV-Vis) absorption spectra. UV-Vis spectra showed a rapid formation of Cu2+ and the ATSM complex upon mixing. The rate of chelate formation by Ni2+ and ATSM was lower than that by Cu-ATSM. Zn2+ and Fe2+ showed much slower reactions with the ATSM than Ni2+. Trace amounts of Ni2+, Zn2+, and Fe2+ showed little effect on [64Cu]Cu-ATSM' quality, while the concentration of impurity Cu2+ must be controlled. These results can provide process management tools for radiopharmaceuticals.
RESUMO
Detecting tumor lesions <1 cm in size using current imaging methods remains a clinical challenge, especially in pancreatic cancer. Previously, we developed a method to identify pancreatic tumor lesions ≥3 mm using positron emission tomography (PET) with an intraperitoneally administered 64Cu-labeled anti-epidermal growth factor receptor (EGFR) antibody (64Cu-NCAB001 ipPET). Here, we conducted an extended single-dose toxicity study of 64Cu-NCAB001 ipPET in mice based on approach 1 of the current ICH M3 [R2] guideline, as our new drug formulation contains 45 µg of the antibody. We used NCAB001 labeled with stable copper isotope instead of 64Cu. The total content of size variants was approximately 6.0% throughout the study. The relative binding potency of Cu-NCAB001 to recombinant human EGFR was comparable to that of cetuximab. The general and neurological toxicities of Cu-NCAB001 ipPET at 62.5 or 625 µg/kg were assessed in mice. The no-observed-adverse-effect level of Cu-NCAB001 was 625 µg/kg, a dose approximately 1000-fold higher at the µg/kg level than the dose of 64Cu-NCAB001 in our formulation (45 µg). The size variants did not affect the safety of the formulation. Therefore, clinical studies on the efficacy of 64Cu-NCAB001 ipPET for early detection of pancreatic cancer using PET imaging can be safely conducted.
RESUMO
Background: Gastric cancer is a common cause of cancer-related death worldwide, and peritoneal dissemination is the most frequent metastatic pattern of gastric cancer. However, the treatment of this disease condition remains difficult. It has been demonstrated that intraperitoneal radioimmunotherapy (ipRIT) with 64Cu-labeled cetuximab (anti-epidermal growth factor receptor antibody; 64Cu-cetuximab) is a potential treatment for peritoneal dissemination of gastrointestinal cancer in vivo. Recent preclinical and clinical studies have also shown that a histone deacetylase inhibitor, vorinostat, effectively sensitized gastrointestinal cancer to external radiation. Aim: In the present study, we examined the efficacy of the combined use of vorinostat, as a radiosensitizer during ipRIT with 64Cu-cetuximab in a peritoneal dissemination mouse model with human gastric cancer NUGC4 cells stably expressing red fluorescent protein. Methods: The mouse model was treated by ipRIT with 64Cu-cetuximab plus vorinostat, each single treatment, or saline (control). Side effects, including hematological and biochemical parameters, were evaluated in similarly treated, tumor-free mice. Results: Coadministration of ipRIT with 64Cu-cetuximab + vorinostat significantly prolonged survival compared to control and each single treatment. No significant toxicity signals were observed in all treatment groups. Conclusions: Our data suggest that vorinostat is a potentially effective radiosensitizer for use during the treatment of peritoneal dissemination of gastric cancer by ipRIT with 64Cu-cetuximab.
Assuntos
Radiossensibilizantes , Neoplasias Gástricas , Animais , Linhagem Celular Tumoral , Cetuximab/uso terapêutico , Modelos Animais de Doenças , Inibidores de Histona Desacetilases/farmacologia , Humanos , Camundongos , Radiossensibilizantes/farmacologia , Radioimunoterapia , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/radioterapia , VorinostatRESUMO
Peritoneal dissemination of pancreatic cancer has a poor prognosis. We have reported that intraperitoneal radioimmunotherapy using a 64Cu-labeled antibody (64Cu-ipRIT) is a promising adjuvant therapy option to prevent this complication. To achieve personalized 64Cu-ipRIT, we developed a new in vitro tumor cell-binding assay (64Cu-TuBA) system with a panel containing nine candidate 64Cu-labeled antibodies targeting seven antigens (EGFR, HER2, HER3, TfR, EpCAM, LAT1, and CD98), which are reportedly overexpressed in patients with pancreatic cancer. We investigated the feasibility of 64Cu-TuBA to select the highest-binding antibody for individual cancer cell lines and predict the treatment response in vivo for 64Cu-ipRIT. 64Cu-TuBA was performed using six human pancreatic cancer cell lines. For three cell lines, an in vivo treatment study was performed with 64Cu-ipRIT using high-, middle-, or low-binding antibodies in each peritoneal dissemination mouse model. The high-binding antibodies significantly prolonged survival in each mouse model, while low-and middle-binding antibodies were ineffective. There was a correlation between in vitro cell binding and in vivo therapeutic efficacy. Our findings suggest that 64Cu-TuBA can be used for patient selection to enable personalized 64Cu-ipRIT. Tumor cells isolated from surgically resected tumor tissues would be suitable for analysis with the 64Cu-TuBA system in future clinical studies.
Assuntos
Neoplasias Pancreáticas , Radioimunoterapia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Estudos de Viabilidade , Humanos , Camundongos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/radioterapia , Neoplasias PancreáticasRESUMO
BACKGROUND: [64Cu]Cu-diacethyl-bis(N4-methylthiosemicarbazone) ([64Cu]Cu-ATSM) is a radioactive hypoxia-targeting therapeutic agent, and the efficacy and safety of [64Cu]Cu-ATSM in the treatment of malignant brain tumors are evaluated in clinical trials. For the clinical application of [64Cu]Cu-ATSM, we determined a drug formulation incorporating a stabilizer against radiolysis and confirmed its radiochemical stability. This study aimed to identify trace chemical impurities derived from the degradation of ATSM contained in the [64Cu]Cu-ATSM investigational drug formulation and assess their potential hazards by quantitative structure-activity relationship (QSAR) assessment. METHODS: We hypothesized that the chemical impurities contained in the [64Cu]Cu-ATSM formulation were derived from the degradation of ATSM. Therefore, we first identified the degradants of ATSM using LC-MS/MS. ATSM was dissolved with the drug formulation of [64Cu]Cu-ATSM, except for 64Cu, and analyzed by LC-MS/MS at 0 and 48 h after sample preparation. Subsequently, the chemical impurities contained in the [64Cu]Cu-ATSM formulation were measured at 0, 5, and 24 h after preparation by HPLC, and the results were compared to the degradants of ATSM. The potential hazards of the chemical impurities contained in the [64Cu]Cu-ATSM formulation were assessed using the QSAR Toolbox (ver. 4.3). RESULTS: Six ATSM degradants were detected and identified by LC-MS/MS analysis, indicating that the functional groups around the nitrogen and sulfur atoms of ATSM were affected. The same peaks were detected as trace chemical impurities in the [64Cu]Cu-ATSM formulation at 24 h, while no apparent peaks were detected at 0 and 5 h. The estimated LD50 values of these chemical impurities showed 4.31 mg/kg or more by QSAR assessment. In contrast, the estimated amount of each chemical impurity exposed to patients was 31.8 ng/kg or less per dose. The smallest margin between the amount of chemical impurities and smallest estimated LD50 value of the corresponding impurity was a ratio of approximately 1:700,000. CONCLUSIONS: We identified trace chemical impurities derived from ATSM in the [64Cu]Cu-ATSM formulation. This suggests that the potential risk of the systemic exposure of patients to these chemical impurities is substantially low.
Assuntos
Complexos de Coordenação , Compostos Organometálicos , Tiossemicarbazonas , Cromatografia Líquida , Radioisótopos de Cobre , Humanos , Relação Quantitativa Estrutura-Atividade , Espectrometria de Massas em TandemRESUMO
Actinium-225 (225Ac) is a promising radionuclide used in targeted alpha therapy (TAT). Although 225Ac labeling of bifunctional chelating ligands is effective, previous in vivo studies reported that free 225Ac can be released from the drugs and that such free 225Ac is predominantly accumulated in the liver and could cause unexpected toxicity. To accelerate the clinical development of 225Ac TAT with a variety of drugs, preparing methods to deal with any unexpected toxicity would be valuable. The aim of this study was to evaluate the feasibility of various chelators for reducing and excreting free 225Ac and compare their chemical structures. Nine candidate chelators (D-penicillamine, dimercaprol, Ca-DTPA, Ca-EDTA, CyDTA, GEDTA TTHA, Ca-TTHA, and DO3A) were evaluated in vitro and in vivo. The biodistribution and dosimetry of free 225Ac were examined in mice before an in vivo chelating study. The liver exhibited pronounced 225Ac uptake, with an estimated human absorbed dose of 4.76 SvRBE5/MBq. Aminopolycarboxylate chelators with five and six carboxylic groups, Ca-DTPA and Ca-TTHA, significantly reduced 225Ac retention in the liver (22% and 30%, respectively). Significant 225Ac reductions were observed in the heart and remainder of the body with both Ca-DTPA and Ca-TTHA, and in the lung, kidney, and spleen with Ca-TTHA. In vitro interaction analysis supported the in vivo reduction ability of Ca-DTPA and Ca-TTHA. In conclusion, aminopolycarboxylate chelators with five and six carboxylic groups, Ca-DTPA and Ca-TTHA, were effective for whole-body clearance of free 225Ac. This feasibility study provides useful information for reducing undesirable radiation exposure from free 225Ac.
RESUMO
OBJECTIVES: To improve the prognosis of pancreatic cancer, new imaging methods to identify tumor lesions at a size of <1 cm are urgently needed. To approach this clinical issue, we developed a new method to detect small tumor lesions in the pancreas (≥3 mm) by positron emission tomography (PET) using an intraperitoneally (ip)-administered 64Cu-labeled new anti-epidermal growth factor receptor (EGFR) antibody (encoded as NCAB001), called 64Cu-NCAB001 ipPET. METHODS: NCAB001 was manufactured under cGMP conditions and labeled with 64Cu. The radiochemical and biological properties of 64Cu-NCAB001 were evaluated. Tumor uptake of an ip-administered 64Cu-NCAB001 in mice with orthotopic pancreatic tumor xPA1-DC xenografts was also evaluated. Pharmacokinetics and radiation dosimetry were examined using PET images acquired after the ip administration of 64Cu-NCAB001 into cynomolgus monkeys with pharmacologic safety monitoring. RESULTS: Radio-chromatography, cell-binding assays, and biodistribution of 64Cu-NCAB001 in mice were identical to those of our previous data with clinically available cetuximab. Small tumor lesions in the pancreas (≥3 mm) of mice could be identified by 64Cu-NCAB001 ipPET. The ip administration of 64Cu-NCAB001 into monkeys was safely conducted using ultrasound imaging. PET images in monkeys showed that ip-administered 64Cu-NCAB001 was distributed throughout the intraperitoneal cavity for up to 6 h and cleared thereafter. Most of the radioactivity was distributed in the liver and the large intestine. The radioactivity around the pancreas became negligible 24 h after administration. The estimated human effective dose was 0.0174 mSv/MBq. CONCLUSION: Our data support the initiation of clinical trials of 64Cu-NCAB001 ipPET to transfer this promising tool for the early diagnosis of pancreatic cancers.
RESUMO
BACKGROUND: In pancreatic cancer surgery, accurate identification and resection of intrapancreatic residual tumors are quite difficult. We have developed a novel open-typed PET system (called 'OpenPET'), which enables high-resolution PET-guided surgery in real time, and demonstrated that OpenPET-guided surgery with intraperitoneally administered 64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab is useful to detect and resect primary pancreatic cancer. Here, we investigated applicability of OpenPET-guided surgery for unexpected residual intrapancreatic tumors and examined its survival benefit over conventional surgery. METHODS: A mouse model with large (>1 cm) resectable pancreatic cancer of xPA-1-DC cells expressing red fluorescent protein was used. OpenPET-guided surgery was conducted 24 h after intraperitoneal administration of 64Cu-labeled cetuximab (7.4 MBq/mouse). For comparison, similar surgical procedures were conducted, and conventional tumor resection was attempted using only the naked eye (control). Survival rate after OpenPET-guided surgery was compared to that after control operations. RESULTS: Intraoperative OpenPET guidance enabled detection and resection of small residual tumors. Ten residual tumor specimens (3-10 mm in diameter) were intraoperatively isolated with OpenPET guidance (n = 7 mice). All isolated specimens showed tumor RFP signals. No resection of tumor tissue was performed in control group because the tumor could not be clearly detected with the naked eye alone. Mice after OpenPET-guided surgery showed significantly longer survival rates than those in control group. CONCLUSIONS: OpenPET-guided surgery with 64Cu-labeled-cetuximab enabled intraoperative identification and resection of intrapancreatic small residual tumors. This technology could be useful to prevent tumor residuals during surgery and improve pancreatic cancer survival.
Assuntos
Radioisótopos de CobreRESUMO
Early diagnosis of pancreatic cancer using current imaging modalities remains challenging. We have developed a new approach to identify tumor lesions ≥ 3 mm in the pancreas by positron emission tomography (PET) with a new intraperitoneally administered 64Cu-labeled anti-epidermal growth factor receptor (EGFR) antibody (encoded as NCAB001), called 64Cu-NCAB001 ipPET. Generally, in clinical research, a radiometal-antibody complex must be prepared immediately before use at the imaging site. To make 64Cu-NCAB001 ipPET available to daily clinical practices in a sustainable way, the NCAB001-chelator conjugate and 64Cu-NCAB001 must be characterized and stabilized. NCAB001 was manufactured under cGMP conditions. NCAB001 was conjugated with a bifunctional chelator (p-SCN-Bn-PCTA), and the antibody-chelator conjugate (PCTA-NCAB001) was characterized by LC/MS and ELISA. Thereafter, to effectively manufacture 64Cu-NCAB001, we developed a new formulation to stabilize PCTA-NCAB001 and 64Cu-NCAB001. An average of three PCTA chelators were conjugated per molecule of NCAB001. The relative binding potency of PCTA-NCAB001 was comparable to cetuximab. The formulation consisting of acetate buffer, glycine, and polysorbate-80 stabilized PCTA-NCAB001 for a year-long storage. Additionally, this formulation enabled the stabilization of 64Cu-NCAB001 for up to 24 h after radiolabeling with a sufficient radioactivity concentration for clinical use. These results may accelerate the future use of 64Cu-NCAB001 ipPET in clinical settings for the early diagnosis and treatment of pancreatic cancer.
RESUMO
Primary central nervous system lymphoma (PCNSL) is an isolated type of lymphoma of the central nervous system and has a dismal prognosis despite intensive chemotherapy. Recent genomic analyses have identified highly recurrent mutations of MYD88 and CD79B in immunocompetent PCNSL, whereas LMP1 activation is commonly observed in Epstein-Barr virus (EBV)-positive PCNSL. However, a lack of clinically representative preclinical models has hampered our understanding of the pathogenic mechanisms by which genetic aberrations drive PCNSL disease phenotypes. Here, we establish a panel of 12 orthotopic, patient-derived xenograft (PDX) models from both immunocompetent and EBV-positive PCNSL and secondary CNSL biopsy specimens. PDXs faithfully retained their phenotypic, metabolic, and genetic features, with 100% concordance of MYD88 and CD79B mutations present in PCNSL in immunocompetent patients. These models revealed a convergent functional dependency upon a deregulated RelA/p65-hexokinase 2 signaling axis, codriven by either mutated MYD88/CD79B or LMP1 with Pin1 overactivation in immunocompetent PCNSL and EBV-positive PCNSL, respectively. Notably, distinct molecular alterations used by immunocompetent and EBV-positive PCNSL converged to deregulate RelA/p65 expression and to drive glycolysis, which is critical for intracerebral tumor progression and FDG-PET imaging characteristics. Genetic and pharmacologic inhibition of this key signaling axis potently suppressed PCNSL growth in vitro and in vivo. These patient-derived models offer a platform for predicting clinical chemotherapeutics efficacy and provide critical insights into PCNSL pathogenic mechanisms, accelerating therapeutic discovery for this aggressive disease. SIGNIFICANCE: A set of clinically relevant CNSL xenografts identifies a hyperactive RelA/p65-hexokinase 2 signaling axis as a driver of progression and potential therapeutic target for treatment and provides a foundational preclinical platform. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/23/5330/F1.large.jpg.
Assuntos
Neoplasias do Sistema Nervoso Central/patologia , Hexoquinase/metabolismo , Linfoma/patologia , Fator de Transcrição RelA/metabolismo , Animais , Antígenos CD79/genética , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Neoplasias do Sistema Nervoso Central/metabolismo , Neoplasias do Sistema Nervoso Central/mortalidade , Feminino , Glicólise , Hexoquinase/genética , Humanos , Linfoma/tratamento farmacológico , Linfoma/metabolismo , Linfoma/mortalidade , Camundongos SCID , Mutação , Fator 88 de Diferenciação Mieloide/genética , NF-kappa B/metabolismo , Peptidilprolil Isomerase de Interação com NIMA/metabolismo , Transdução de Sinais , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Pancreatic cancer (PC) has a poor prognosis owing to difficulties in the diagnosis of resectable PC at early stages. Several clinical studies have indicated that the detection and surgery of small resectable PC (<1 cm) can significantly improve survival; however, imaging diagnosis and accurate resection of small PC remain challenging. Here, we report the feasibility of "immuno-OpenPET" as a novel approach enabling not only early diagnosis but also image-guided surgery, using a small (<1 cm) resectable PC orthotopic xenograft mouse model. For immuno-OpenPET, we utilized our original OpenPET system, which enables high-resolution positron emission tomography (PET) imaging with depth-of-interaction detectors, as well as real-time image-guided surgery, by arranging the detectors to create an open space for surgery and accelerating the image reconstruction process by graphics processing units. For immuno-OpenPET, 64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab was intraperitoneally administered into mice. It clearly identified PC tumors ≥3 mm. In contrast, neither OpenPET with intravenous-administered 64Cu-cetuximab nor intraperitoneal/intravenous-administered 18F-FDG (a traditional PET probe) could detect PC in this model. Immuno-OpenPET-guided surgery accurately resected small PC in mice and achieved significantly prolonged survival. This technology could provide a novel diagnostic and therapeutic strategy for small resectable PC to improve patient survival.
Assuntos
Cetuximab/imunologia , Radioisótopos de Cobre/metabolismo , Detecção Precoce de Câncer/métodos , Neoplasias Pancreáticas/patologia , Tomografia por Emissão de Pósitrons/métodos , Cirurgia Assistida por Computador/métodos , Animais , Antineoplásicos Imunológicos/imunologia , Apoptose , Proliferação de Células , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/diagnóstico por imagem , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/cirurgia , Compostos Radiofarmacêuticos/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Hypoxia plays important roles in the prognosis of malignant brain tumors such as glioblastoma because it causes drug delivery deficiencies and the induction of hypoxia-inducible factor-1α in tumor cells. Extensive hypoxic areas are associated with poor prognosis of these fatal diseases. We previously reported that multiple administrations of the hypoxia-targeted internal radiotherapy agent 64Cu-diacetyl-bis(N4-methylthiosemicarbazone) (64Cu-ATSM), four times at intervals of 1 or 2 weeks, show antitumor effects in glioblastoma without treatment-related adverse events. Before initiating clinical trials, preclinical safety studies using Cu-ATSM composed of stable isotopes and its precursor ATSM were required to understand the potential risks of systemic and repeated chemical exposure of our investigational drug. In this study, the concentrations of Cu-ATSM and ATSM in mouse plasma after intravenous administration were determined by liquid chromatography-tandem mass spectrometry, and the half-lives were estimated to be 21.5 and 22.4â¯minutes for Cu-ATSM and ATSM, respectively. Based on this result, approach 2 of the current ICH M3 [R2] guideline was adopted, and a 7-day intravenous toxicity study was conducted in mice. Cu-ATSM and ATSM in a ratio of 2:25 mimicking our current investigational drug was used, and no adverse effects were observed when Cu-ATSM and ATSM were administered at 81⯵g/kg. These results and those of previous studies suggest that our current investigational drug formulation containing Cu-ATSM and ATSM at a dose of 15⯵g can be safely administered to patients once per week for 4 weeks for treatment with 64Cu-ATSM.
RESUMO
Pancreatic cancer (PC) has a very poor prognosis. Surgery is the primary treatment for patients with resectable PC; however, local recurrence, hepatic metastasis, and peritoneal dissemination often occur even after extensive surgery. Adjuvant chemotherapy, typically with gemcitabine, has been used clinically but with only a modest survival benefit. To achieve a better outcome, we investigated the efficacy of 64Cu-intraperitoneal radioimmunotherapy (ipRIT) with 64Cu-labeled antiepidermal growth factor receptor antibody cetuximab as an adjuvant treatment after PC surgery using an orthotopic xenografted mouse model. Methods: The efficacy of adjuvant 64Cu-ipRIT was investigated in a human PC mouse model harboring orthotopic xenografts of xPA-1-DC cells. To reproduce the clinical situation, PC xenografts were surgically resected when pancreatic tumors were readily visible but not metastatic tumors. Increasing doses of 64Cu-cetuximab were intraperitoneally injected, and the mice were monitored for toxicity to determine the safe therapeutic dose. For adjuvant 64Cu-ipRIT, the day after tumor resection, the mice were intraperitoneally administered 22.2 MBq of 64Cu-PCTA-cetuximab and the survival was compared with that in surgery-only controls. For comparison, adjuvant chemotherapy with gemcitabine was also examined using the same model. Results: The mouse model not only developed primary tumors in the pancreas but also subsequently reproduced local recurrence, hepatic metastasis, and peritoneal dissemination after surgery, which is similar to the manifestations that occur with human PC. Adjuvant 64Cu-ipRIT with 64Cu-labeled cetuximab after surgery effectively suppressed local recurrence, hepatic metastasis, and peritoneal dissemination in this model. Significant improvement of the survival with minimal toxicity was achieved by adjuvant 64Cu-ipRIT compared with that in control mice that underwent surgery only. Adjuvant chemotherapy with gemcitabine nominally prolonged the survival, but the effect was not statistically significant. Conclusion:64Cu-ipRIT with cetuximab can be an effective adjuvant therapy after PC surgery.
Assuntos
Radioisótopos de Cobre/farmacologia , Injeções Intraperitoneais , Neoplasias Pancreáticas/radioterapia , Radioterapia Adjuvante , Animais , Linhagem Celular Tumoral , Cetuximab/farmacologia , Terapia Combinada , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Modelos Animais de Doenças , Receptores ErbB/química , Feminino , Células HCT116 , Humanos , Neoplasias Hepáticas/radioterapia , Neoplasias Hepáticas/secundário , Camundongos , Camundongos Endogâmicos BALB C , Metástase Neoplásica , Recidiva Local de Neoplasia , Transplante de Neoplasias , Radioimunoterapia , Resultado do Tratamento , Gencitabina , Neoplasias PancreáticasRESUMO
Peritoneal dissemination is a common cause of death from gastrointestinal cancers and is difficult to treat using current therapeutic options, particularly late-phase disease. Here, we investigated the feasibility of integrated therapy using 64Cu-intraperitoneal radioimmunotherapy (ipRIT), alone or in combination with positron emission tomography (PET)-guided surgery using a theranostic agent (64Cu-labeled anti-epidermal growth factor receptor antibody cetuximab) to treat early- and late-phase peritoneal dissemination in mouse models. In this study, we utilized the OpenPET system, which has open space for conducting surgery while monitoring objects at high resolution in real time, as a novel approach to make PET-guided surgery feasible. 64Cu-ipRIT with cetuximab inhibited tumor growth and prolonged survival with little toxicity in mice with early-phase peritoneal dissemination of small lesions. For late-phase peritoneal dissemination, a combination of 64Cu-ipRIT for down-staging and subsequent OpenPET-guided surgery for resecting large tumor masses effectively prolonged survival. OpenPET clearly detected tumors (≥3 mm in size) behind other organs in the peritoneal cavity and was useful for confirming the presence or absence of residual tumors during an operation. These findings suggest that integrated 64Cu therapy can serve as a novel treatment strategy for peritoneal dissemination.
RESUMO
BACKGROUND: The present study proposed a new concept for targeted radionuclide therapy (TRT) to improve the intratumoral distribution of radioactivity using two different radiopharmaceuticals. We examined the efficacy of a combination of a tetrameric cyclic Arg-Gly-Asp (cRGD) peptide-based radiopharmaceutical, 64Cu-cyclam-RAFT-c(-RGDfK-)4 (64Cu-RaftRGD, an αVß3 integrin [αVß3] tracer), and 64Cu-diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM, a supposed tracer for hypoxic metabolism) in a small animal tumor model. RESULTS: Mice with subcutaneous αVß3-positive U87MG glioblastoma xenografts were used. The intratumoral distribution of a near-infrared dye, Cy5.5-labeled RAFT-c(-RGDfK-)4 (Cy5.5-RaftRGD), 64Cu-RaftRGD, and 64Cu-ATSM was visualized by fluorescence imaging and autoradiography of the co-injected Cy5.5-RaftRGD with 64Cu-RaftRGD or 64Cu-ATSM at 3 h postinjection. Mice were treated with a single intravenous dose of the vehicle solution (control), 18.5 or 37 MBq of 64Cu-RaftRGD or 64Cu-ATSM, or a combination (18.5 MBq of each agent). The tumor volume, tumor cell proliferation, body weight, survival, and tumor and organ uptake of radiopharmaceuticals were assessed. It was shown that Cy5.5-RaftRGD colocalized with 64Cu-RaftRGD and could be used as a surrogate for the radioactive agent. The intratumoral distribution of Cy5.5-RaftRGD and 64Cu-ATSM was discordant and nearly complementary, indicating a more uniform distribution of radioactivity achievable with the combined use of 64Cu-RaftRGD and 64Cu-ATSM. Neither 64Cu-RaftRGD nor 64Cu-ATSM showed significant effects on tumor growth at 18.5 MBq. The combination of both (18.5 MBq each) showed sustained inhibitory effects against tumor growth and tumor cell proliferation and prolonged the survival of the mice, compared to that by either single agent at 37 MBq. Interestingly, the uptake of the combination by the tumor was higher than that of 64Cu-RaftRGD alone, but lower than that of 64Cu-ATSM alone. The kidneys showed the highest uptake of 64Cu-RaftRGD, whereas the liver exhibited the highest uptake of 64Cu-ATSM. No obvious adverse effects were observed in all treated mice. CONCLUSIONS: The combination of 64Cu-RaftRGD and 64Cu-ATSM achieved an improved antitumor effect owing to the more uniform intratumoral distribution of radioactivity. Thus, combining different radiopharmaceuticals to improve the intratumoral distribution would be a promising concept for more effective and safer TRT.
RESUMO
Glioblastoma is the most aggressive malignant brain tumor in humans and is difficult to cure using current treatment options. Hypoxic regions are frequently found in glioblastoma, and increased levels of hypoxia are associated with poor clinical outcomes of glioblastoma patients. Hypoxia plays important roles in the progression and recurrence of glioblastoma because of drug delivery deficiencies and induction of hypoxia-inducible factor-1α in tumor cells, which lead to poor prognosis. We focused on a promising hypoxia-targeted internal radiotherapy agent, 64Cu-diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM), to address the need for additional treatment for glioblastoma. This compound can target the overreduced state under hypoxic conditions within tumors. Clinical positron emission tomography studies using radiolabeled Cu-ATSM have shown that Cu-ATSM accumulates in glioblastoma and its uptake is associated with high hypoxia-inducible factor-1α expression. To evaluate the therapeutic potential of this agent for glioblastoma, we examined the efficacy of 64Cu-ATSM in mice bearing U87MG glioblastoma tumors. Administration of single dosage (18.5, 37, 74, 111, and 148 MBq) and multiple dosages (37 MBq × 4) of 64Cu-ATSM was investigated. Single administration of 64Cu-ATSM in high-dose groups dose-dependently inhibited tumor growth and prolonged survival, with slight and reverse signs of adverse events. Multiple dosages of 64Cu-ATSM remarkably inhibited tumor growth and prolonged survival. By splitting the dose of 64Cu-ATSM, no adverse effects were observed. Our findings indicate that multiple administrations of 64Cu-ATSM have effective antitumor effects in glioblastoma without side effects, indicating its potential for treating this fatal disease.
RESUMO
Bevacizumab, an anti-vascular endothelial growth factor (VEGF) antibody, is an antiangiogenic agent clinically used for various cancers. However, repeated use of this agent leads to tumor-decreased vascularity and hypoxia with activation of an HIF-1 signaling pathway, which results in drug delivery deficiency and induction of malignant behaviors in tumors. Here, we developed a novel strategy to treat tumors with bevacizumab-induced vascular decrease and hypoxia using 64Cu-diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM), a potential theranostic agent, which possesses high tissue permeability and can target over-reduced conditions under hypoxia in tumors, with a human colon carcinoma HT-29 tumor-bearing mouse model. The long-term treatment with bevacizumab caused decreased blood vessel density and activation of an HIF-1 signaling pathway; increased uptake of 64Cu-ATSM was also observed despite limited blood vessel density in HT-29 tumors. In vivo high-resolution SPECT/PET/CT imaging confirmed reduced vascularity and increased proportion of 64Cu-ATSM uptake areas within the bevacizumab-treated tumors. 64Cu-ATSM therapy was effective to inhibit tumor growth and prolong survival of the bevacizumab-treated tumor-bearing mice without major adverse effects. In conclusion, 64Cu-ATSM therapy effectively enhanced anti-tumor effects in tumors with bevacizumab-induced vascular decrease and hypoxia. 64Cu-ATSM therapy could represent a novel approach as an add-on to antiangiogenic therapy.