RESUMO
BACKGROUND/AIM: The standard treatment for localized prostate cancer involves surgical removal of the prostate with curative intent. However, when tumor cells persist in the operation site, there is high risk of local recurrence and tumor spread, leading to stressful follow-up treatments, impaired quality of life, and reduced overall survival. This study examined photoimmunotherapy (PIT) as a new treatment option for prostate cancer cells. MATERIALS AND METHODS: We generated conjugates consisting of either a humanized antibody or Fab fragments thereof targeting the prostate specific membrane antigen (PSMA), along with our silicon phthalocyanine photosensitizer dye WB692-CB1. PSMA-expressing prostate cancer cells were incubated with the antibody dye or Fab dye conjugates and cell binding was measured using flow cytometry. Cells were irradiated with varying doses of red light for dye activation, and cytotoxicity was determined by erythrosin B staining and subsequent analysis using a Neubauer counting chamber. RESULTS: Specific cytotoxicity was induced with the antibody dye conjugate in the prostate cancer cells in a light dose-dependent manner. Treatment of the cells with the Fab dye conjugate resulted in lower cytotoxicity, which could be attributed to a reduced binding affinity and a reduced dye uptake of the Fab fragment. CONCLUSION: Our new antibody dye and Fab dye conjugates offer potential for future intraoperative PIT in patients with localized prostate cancer, with the aim to ensure complete removal of tumor cells from the surgical area, to avoid local recurrence, and to improve clinical outcome.
Assuntos
Antígenos de Superfície , Fragmentos Fab das Imunoglobulinas , Imunoterapia , Neoplasias da Próstata , Humanos , Masculino , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/farmacologia , Neoplasias da Próstata/terapia , Neoplasias da Próstata/radioterapia , Neoplasias da Próstata/patologia , Imunoterapia/métodos , Linhagem Celular Tumoral , Antígenos de Superfície/imunologia , Antígenos de Superfície/metabolismo , Glutamato Carboxipeptidase II/imunologia , Glutamato Carboxipeptidase II/metabolismo , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Fototerapia/métodos , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêuticoRESUMO
INTRODUCTION: Single domain antibody fragments (sdAbs) are an appealing scaffold for radiopharmaceutical development due to their small size (~15 kDa), high solubility, high stability, and excellent tumor penetration. Previously, we developed NB7 sdAb, which has very high affinity for an epitope on PSMA that is different from those targeted by small molecule PSMA inhibitors. Herein, we evaluated NB7 after radioiodination using [*I]SGMIB (1,3,4-isomer) and iso-[*I]SGMIB (1,3,5-isomer), as well as their 211At-labeled analogues. METHODS: [*I]SGMIB, iso-[*I]SGMIB, [211At]SAGMB, and iso-[211At]SAGMB conjugates of NB7 sdAb were synthesized and their binding affinity, cell uptake and internalization were assessed in PSMA+ PC3 PIP and PSMA- PC3 flu cells. Biodistribution studies were performed in mice bearing PSMA+ PC3 PIP xenografts. First, a single-label experiment evaluated the tissue distribution of a NB7 bearing a His6-tag (NB7H6) and labeled with iso-[125I]SGMIB. Three paired-label experiments then were performed to compare: a) NB7 labeled using [*I]SGMIB and iso-[*I]SGMIB, b) 131I- vs 211At-labeled NB7 conjugates and c) [125I]SGMIB-NB7H6 to the small molecule PSMA inhibitor [131I]YF2. RESULTS: All NB7 radioconjugates bound specifically to PSMA with dissociation constants, Kd, in the low nM range (1.4-6.4 nM). An initial biodistribution study demonstrated good tumor uptake for iso-[125I]SGMIB-NB7H6 (7.2 ± 1.5 % ID/g at 1 h) and no deleterious effect of the His6-tag on renal activity levels, which declined to 3.1 ± 1.1 % ID/g by 4 h. Paired-label biodistribution found no distinction between the two SGMIB isomer NB7 conjugates with the [131I]SGMIB-NB7-to-iso-[125I]SGMIB-NB7 tumor uptake ratios not significantly different from unity: 1.06 ± 0.08 at 1 h, 1.04 ± 0.12 at 4 h, and 1.07 ± 0.09 at 24 h. Both isomer conjugates cleared rapidly from normal tissues and exhibited very low uptake in thyroid, lacrimal and salivary glands. Paired-label biodistribution of [131I]SGMIB-NB7H6 and [211At]SAGMB-NB7H6 demonstrated similar tumor uptake and kidney clearance for the two radioconjugates. However, levels of 211At in thyroid, stomach, salivary and lacrimal glands were significantly higher (P < 0.05) that those for 131I suggesting greater dehalogenation for [211At]SAGMB-NB7H6. Finally, co-administration of [125I]SGMIB-NB7H6 and [131I]YF2 demonstrated good tumor uptake for both with considerably more rapid renal clearance for the NB7 radioconjugate. CONCLUSION: NB7 radioconjugates exhibited good accumulation in PSMA-positive xenografts with rapid clearance from kidney and other normal tissues. We conclude that NB7 is a potentially useful scaffold for developing PSMA-targeted theranostics with different characteristics than current small molecule and antibody-based approaches.
Assuntos
Antígenos de Superfície , Glutamato Carboxipeptidase II , Neoplasias da Próstata , Anticorpos de Domínio Único , Masculino , Humanos , Animais , Camundongos , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Glutamato Carboxipeptidase II/imunologia , Glutamato Carboxipeptidase II/antagonistas & inibidores , Glutamato Carboxipeptidase II/metabolismo , Anticorpos de Domínio Único/química , Anticorpos de Domínio Único/imunologia , Antígenos de Superfície/metabolismo , Antígenos de Superfície/imunologia , Linhagem Celular Tumoral , Distribuição Tecidual , Transformação Celular NeoplásicaRESUMO
INTRODUCTION: Whilst prostate cancer is the fourth most common cancer globally, effective therapies for patients with advanced disease are lacking. In recent years, interest in using theranostic agents to treat castrate-resistant prostate cancer (CRPC) and metastatic prostate cancer has emerged. Lu-TLX591 monoclonal antibody is a potential agent of significance; however, to date, reports on its toxicity and efficacy have been limited to small clinical trials in heavily pretreated patients. This retrospective study describes the real-world toxicity and efficacy profile of Lu-TLX591. METHODS: Eighteen patients received Lu-TLX591 at two private oncology centres in Australia. Patients were eligible if they had CRPC or metastatic prostate cancer and prostate-specific membrane antigen (PSMA)-avid disease confirmed by PSMA-positron emission tomography (PET). Patients received two cycles of Lu-TLX591 monoclonal antibody (177 Lu-DOTA-rosopatamab) each dosed from 1.01-2.85 GBq, 14 days apart. Patient side effects, blood test results and radiology reports were recorded on the patient's electronic medical record (eMR). RESULTS: Prominent side effects included fatigue (55.6%), anorexia (16.7%), nausea (11.1%), and transfusion reactions (11.1%). All-grade haematological toxicities included lymphopenia (61.1%), anaemia (22.2%), leukopenia (27.8%), neutropenia (27.8%), and thrombocytopenia (27.8%). Grade 4 toxicity included lymphopenia (6.7%) and thrombocytopenia (6.7%). Patients' prostate-specific antigen (PSA) responses were as follows; ≥ 30% PSA decline (27.8%), ≥ 50% PSA decline (11.4%) and any PSA decline (38.9%). Follow-up radiology revealed 54.5% stable disease, 45.4% disease progression and 9.1% disease regression. CONCLUSION: Lu-TLX591 was safely administered at acceptable toxicity and its efficacy reflects previous clinical trials. Larger studies are required and are underway (NCT04786847; NCT05146973; NCT04876651) to determine Lu-TLX591 effectiveness amongst different prostate cancer populations and compare its efficacy against peptide-based radiopharmaceutical agents.
Assuntos
Anticorpos Monoclonais , Lutécio , Radioisótopos , Humanos , Masculino , Idoso , Lutécio/uso terapêutico , Lutécio/efeitos adversos , Pessoa de Meia-Idade , Radioisótopos/efeitos adversos , Radioisótopos/uso terapêutico , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais/efeitos adversos , Estudos Retrospectivos , Glutamato Carboxipeptidase II/imunologia , Glutamato Carboxipeptidase II/antagonistas & inibidores , Resultado do Tratamento , Idoso de 80 Anos ou mais , Metástase Neoplásica , Neoplasias da Próstata/patologia , Neoplasias da Próstata/tratamento farmacológico , Antígenos de Superfície/imunologia , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Compostos Radiofarmacêuticos/efeitos adversos , Compostos Radiofarmacêuticos/uso terapêutico , Antígeno Prostático Específico/sangueAssuntos
Antígenos de Superfície , Neoplasias da Bexiga Urinária , Humanos , Neoplasias da Bexiga Urinária/patologia , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/imunologia , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismo , Imunoconjugados/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma de Células de Transição/tratamento farmacológico , Carcinoma de Células de Transição/patologia , Carcinoma de Células de Transição/imunologia , Carcinoma de Células de Transição/genética , Neoplasias Urológicas/imunologia , Neoplasias Urológicas/patologia , Neoplasias Urológicas/tratamento farmacológico , Neoplasias Urológicas/genética , Glutamato Carboxipeptidase II/metabolismo , Glutamato Carboxipeptidase II/imunologia , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/genéticaRESUMO
BACKGROUND: Accurate diagnosis of localized prostate cancer (PCa) is limited by inadequacy of multiparametric (mp) MRI to fully identify and differentiate localized malignant tissue from benign pathologies. Prostate-specific membrane antigen (PSMA) represents an excellent target for molecular imaging. IAB2M, an 85-kD minibody derived from a de-immunized monoclonal antibody directed at the extracellular domain of human PSMA (huJ591), and PSMA-11, a small molecule ligand have been previously tested as probes for visualization of recurrent/metastatic PCa with PET/CT. This pilot, non-randomized trial studied their diagnostic utility in patients (pts) with localized PCa. METHODS: Pts planned for radical prostatectomy (RP) were enrolled and underwent mpMRI and PET/CT imaging with 89 Zr-df-IAB2M and/or 68 Ga-PSMA-PET/CT. Image results were read by a radiologist blinded to clinical information and pathology results, mapped and compared to corresponding histopathology findings from all lesions, both clinically significant and nonsignificant. The detection rates of all three imaging modalities were measured and correlated. RESULTS: 20 pts with median age of 64.5 (46-79) years and PSA level of 7.5 (1.6-36.56) ng/ml were enrolled. 19 pts underwent RP and were imaged pre-operatively with 89 Zr-Df-IAB2M PET/CT and mpMRI. Nine of those were imaged using 68 Ga-PSMA-11 as well. Out of 48 intraprostatic lesions verified on surgical pathology, IAB2M PET/CT was able to detect 36 (75%). A similar proportion of pathologically confirmed, clinically significant lesions (22/29, 76%) was detected. IAB2M PET/CT was also able to identify 14/19 (74%) extraprostatic lesions. The performance of mpMRI was inferior, with 24/48 detectable lesions (50%) and 18/29 clinically significant intraprostatic lesions (62%). Compared to the current standard (mpMRI), IAB2M PET/CT had a sensitivity of 88%, specificity 38%, positive predictive value 58%, and accuracy 63%. In 9 pts who underwent Ga-PSMA-11 as well, the latter yielded a detection rate of 70% (14/20), which was also seen in clinically significant lesions (10/14, 71%). Ga-PSMA-11 PET/CT also detected 4/6 (67%) extraprostatic lesions. CONCLUSIONS: In this pilot study, the performance of 89 Zr-df-IAB2M was superior to mpMRI and similar to 68 Ga-PSMA-11 PET/CT. The higher detection rate of PSMA-PET supports its use as a diagnostic tool with consequent management change implications in men with localized PCa.
Assuntos
Antígenos de Superfície , Radioisótopos de Gálio , Glutamato Carboxipeptidase II , Imageamento por Ressonância Magnética Multiparamétrica , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Neoplasias da Próstata/diagnóstico por imagem , Radioisótopos , Zircônio , Idoso , Anticorpos Monoclonais , Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Prostatectomia , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Prostate-specific membrane antigen (PSMA)-targeted radioligand therapy (RLT) may be associated with renal toxicity. We aimed to identify predictive parameters for the development of chronic kidney disease (CKD) in patients with metastatic castration resistant prostate cancer (mCRPC) undergoing RLT. METHODS: In 46 mCRPC patients scheduled for Lu-177-PSMA-RLT, pretherapeutic estimated glomerular filtration rate (eGFR [ml/min/1.73 m2 ]), Tc-99m-mercaptoacetyltriglycine (Tc-99m-MAG3) clearance and baseline Ga-68-PSMA-ligand positron emission tomography (PET)-derived renal cortical uptake and PSMA-tumor volume (TV) were determined. We tested the predictive capability of these parameters and clinical risk factors for the occurrence of CKD (defined as CTCAE vers. 5.0 grade 2 or higher) during follow-up. RESULTS: After 4 ± 3 cycles of RLT average eGFR declined from 76 ± 17 to 72 ± 20 ml/min/1.73 m2 (p = 0.003). Increased estimated renal radiation dose (eRRD) was significantly associated with renal functional decline (p = 0.008). During follow-up, 16/46 (30.4%) developed CKD grade 2 (no grade 3 or higher). In receiver operating characteristic (ROC) analysis, pretherapeutic eGFR was highly accurate in identifying the occurrence of CKD vs no CKD with an area under the curve (AUC) of 0.945 (p < 0.001; best threshold, 77 ml/min/1.73 m2 ), followed by Tc-99m-MAG3-derived tubular extraction rate (TER; AUC, 0.831, p < 0.001; best threshold, 200 ml/min/1.73 m2 ). Renal PET signal (p = 0.751) and PSMA-TV (p = 0.942), however, were not predictive. Kaplan-Meier analyses revealed adverse renal outcome for patients with lower eGFR (p = 0.001) and lower scintigraphy-derived TER (p = 0.009), with pretherapeutic eGFR emerging as the sole predictive parameter in multivariate analysis (p = 0.007). CONCLUSION: Serious adverse renal events are not a frequent phenomenon after PSMA-targeted RLT. However, in patients developing moderate CKD after RLT, pretherapeutic eGFR is an independent predictor for renal impairment during follow-up.
Assuntos
Antígenos de Superfície , Glutamato Carboxipeptidase II , Lutécio , Neoplasias de Próstata Resistentes à Castração , Radioimunoterapia , Radioisótopos , Insuficiência Renal Crônica , Antígenos de Superfície/imunologia , Antígenos de Superfície/metabolismo , Taxa de Filtração Glomerular , Glutamato Carboxipeptidase II/imunologia , Glutamato Carboxipeptidase II/metabolismo , Humanos , Estimativa de Kaplan-Meier , Lutécio/administração & dosagem , Lutécio/efeitos adversos , Masculino , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons/métodos , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/terapia , Radioimunoterapia/efeitos adversos , Radioimunoterapia/métodos , Radioisótopos/administração & dosagem , Radioisótopos/efeitos adversos , Eliminação Renal , Insuficiência Renal Crônica/induzido quimicamente , Insuficiência Renal Crônica/diagnóstico , Insuficiência Renal Crônica/prevenção & controle , Risco Ajustado/métodos , Fatores de Risco , Tecnécio/farmacologiaRESUMO
BACKGROUND: Therapeutic options currently available for metastatic castration-resistant prostate cancer (mCRPC) do not extend median overall survival >6 months. Therefore, the development of novel and effective therapies for mCRPC represents an urgent medical need. T cell engagers (TCEs) have emerged as a promising approach for the treatment of mCRPC due to their targeted mechanism of action. However, challenges remain in the clinic due to the limited efficacy of TCEs observed thus far in solid tumors as well as the toxicities associated with cytokine release syndrome (CRS) due to the usage of high-affinity anti-CD3 moieties such as OKT3. METHODS: Using genetically engineered transgenic rats (UniRat and OmniFlic) that express fully human IgG antibodies together with an NGS-based antibody discovery pipeline, we developed TNB-585, an anti-CD3xPSMA TCE for the treatment of mCRPC. TNB-585 pairs a tumor-targeting anti-PSMA arm together with a unique, low-affinity anti-CD3 arm in bispecific format. We tested TNB-585 in T cell-redirected cytotoxicity assays against PSMA+ tumor cells in both two-dimensional (2D) cultures and three-dimensional (3D) spheroids as well as against patient-derived prostate tumor cells. Cytokines were measured in culture supernatants to assess the ability of TNB-585 to induce tumor killing with low cytokine release. TNB-585-mediated T cell activation, proliferation, and cytotoxic granule formation were measured to investigate the mechanism of action. Additionally, TNB-585 efficacy was evaluated in vivo against C4-2 tumor-bearing NCG mice. RESULTS: In vitro, TNB-585 induced activation and proliferation of human T cells resulting in the killing of PSMA+ prostate tumor cells in both 2D cultures and 3D spheroids with minimal cytokine release and reduced regulatory T cell activation compared with a positive control antibody that contains the same anti-PSMA arm but a higher affinity anti-CD3 arm (comparable with OKT3). In addition, TNB-585 demonstrated potent efficacy against patient-derived prostate tumors ex vivo and induced immune cell infiltration and dose-dependent tumor regression in vivo. CONCLUSIONS: Our data suggest that TNB-585, with its low-affinity anti-CD3, may be efficacious while inducing a lower incidence and severity of CRS in patients with prostate cancer compared with TCEs that incorporate high-affinity anti-CD3 domains.
Assuntos
Anticorpos Biespecíficos/administração & dosagem , Antígenos de Superfície/imunologia , Complexo CD3/imunologia , Glutamato Carboxipeptidase II/imunologia , Imunoglobulina G/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Animais , Anticorpos Biespecíficos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Macaca fascicularis , Masculino , Camundongos , Células PC-3 , Neoplasias de Próstata Resistentes à Castração/imunologia , Ratos , Ratos Transgênicos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Prostate cancer (PCa) is the second most common cancer in men, causing more than 300,000 deaths every year worldwide. Due to their superior cell-killing ability and the relative simplicity of their preparation, immunotoxin molecules have great potential in the clinical treatment of cancer, and several such molecules have been approved for clinical application. In this study, we adopted a relatively simple strategy based on a single-domain antibody (sdAb) and an improved Pseudomonas exotoxin A (PE) toxin (PE24X7) to prepare a safer immunotoxin against prostate-specific membrane antigen (PSMA) for PCa treatment. The designed anti-PSMA immunotoxin, JVM-PE24X7, was conveniently prepared in its soluble form in an Escherichia coli (E. coli) system, avoiding the complex renaturation process needed for immunotoxin preparation by the conventional strategy. The product was very stable and showed a very strong ability to bind the PSMA receptor. Cytotoxicity assays showed that this molecule at a very low concentration could kill PSMA-positive PCa cells, with an EC50 value (concentration at which the cell viability decreased by 50%) of 15.3 pM against PSMA-positive LNCaP cells. Moreover, this molecule showed very good killing selectivity between PSMA-positive and PSMA-negative cells, with a selection ratio of more than 300-fold. Animal studies showed that this molecule at a very low dosage (5 × 0.5 mg/kg once every three days) completely inhibited the growth of PCa tumors, and the maximum tolerable dose (MTD) was more than 15 mg/kg, indicating its very potent tumor-treatment ability and a wide therapeutic window. Use of the new PE toxin, PE24X7, as the effector moiety significantly reduced off-target toxicity and improved the therapeutic window of the immunotoxin. The above results demonstrate that the designed anti-PSMA immunotoxin, JVM-PE24X7, has good application value for the treatment of PCa.
Assuntos
Adenocarcinoma/tratamento farmacológico , Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Glutamato Carboxipeptidase II/antagonistas & inibidores , Imunotoxinas/uso terapêutico , Terapia de Alvo Molecular , Neoplasias da Próstata/tratamento farmacológico , Anticorpos de Domínio Único/uso terapêutico , Animais , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Antígenos de Superfície/imunologia , Antineoplásicos Imunológicos/toxicidade , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Glutamato Carboxipeptidase II/imunologia , Humanos , Imunotoxinas/toxicidade , Masculino , Dose Máxima Tolerável , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Modelos Moleculares , Conformação Proteica , Proteínas Recombinantes de Fusão/uso terapêutico , Proteínas Recombinantes de Fusão/toxicidade , Anticorpos de Domínio Único/toxicidade , Ensaios Antitumorais Modelo de XenoenxertoAssuntos
Indústria Farmacêutica/organização & administração , Compostos Radiofarmacêuticos/farmacologia , Antígenos de Superfície/imunologia , Sistemas de Liberação de Medicamentos , Glutamato Carboxipeptidase II/imunologia , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/radioterapia , Compostos Radiofarmacêuticos/síntese química , Radioterapia/métodosRESUMO
We illustrate the development of NaDyF4-NaGdF4 core-shell nanoparticles (NPs) for targeting prostate cancer cells using a preclinical 9.4 T magnetic resonance imaging (MRI) of live animals. The NPs composed of paramagnetic Dy3+ and Gd3+ (T2- and T1-contrast agents, respectively) demonstrate proton relaxivities of r1 = 20.2 mM-1 s-1 and r2 = 32.3 mM-1 s-1 at clinical 3 T and r1 = 9.4 mM-1 s-1 and r2 = 144.7 mM-1 s-1 at preclinical 9.4 T. The corresponding relaxivity values per NP are r1 = 19.4 × 105 mMNP-1 s-1 and r2 = 33.0 × 105 mMNP-1 s-1 at 3 T and r1 = 9.0 × 105 mMNP-1 s-1 and r2 = 147.0 × 105 mMNP-1 s-1 at 9.4 T. In vivo active targeting of human prostate tumors grown in nude mice revealed docking of anti-prostate-specific membrane antigen (PSMA) antibody-tagged NPs at tumor sites post-24 h of their intravenous injection. On the other hand, in vivo passive targeting showed preferential accumulation of NPs at tumor sites only within 2 h of their injection, ascribed to the enhanced permeation and retention effect of the tumor. A biodistribution study employing the harvested organs of mice, post-24 h injection of NPs, quantified active targeting as nearly twice as efficient as passive targeting. These outcomes provide potential opportunities for noninvasive diagnosis using NaDyF4-NaGdF4 core-shell NPs for target-specific MRI.
Assuntos
Adenocarcinoma/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Nanopartículas/química , Neoplasias da Próstata/diagnóstico por imagem , Animais , Glutamato Carboxipeptidase II/imunologia , Masculino , Glicoproteínas de Membrana/imunologia , Camundongos , Camundongos NusRESUMO
PURPOSE: The gold nanoparticle (GNP) as a promising theranostic probe has been increasingly studied. The tumor-targeting efficiency of GNPs is crucial to increase the therapeutic ratio. In this study, we developed PSMA-targeted GNPs to enhance GNP uptake in prostate cancer and developed an x-ray fluorescence imaging system to noninvasively monitor and assess GNP delivery. METHODS AND MATERIALS: For targeted therapy of prostate cancer, anti-prostate-specific membrane antigen (PSMA) antibodies were conjugated onto PEGylated GNPs through 1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) (EDC/NHS) chemistry. In vivo imaging was implemented using an in-house-developed dual-modality computed tomography (CT) and x-ray fluorescence CT (XFCT) system on mice bearing subcutaneous LNCaP prostate tumors. After intravenous administration of GNPs (15 mg/mL, 200 µL), the x-ray fluorescence signals from the tumor were collected at various time points (5 minutes to approximately 30 hours) for GNP pharmacokinetics analysis. At 24 hours after administration, x-ray fluorescence projection (XRFproj) and XFCT imaging were conducted to evaluate the prostate tumor uptake of active- and passive-targeting GNPs. Inductively coupled plasma mass spectrometry analysis was adopted as a benchmark to verify the quantification accuracy of XRFproj/XFCT imaging. RESULTS: Fluorescence microscopic imaging confirmed the enhanced (approximately 4 times) targeting efficiency of PSMA-targeted GNPs in vitro. The pharmacokinetics analysis showed enhanced tumor uptake/retention of PSMA-targeted GNPs and revealed that the peak tumor accumulation appeared at approximately 24 hours after intravenous administration. Both XRFproj and XFCT imaging presented their accuracy in quantifying GNPs within tumors noninvasively. Moreover, XFCT imaging verified its unique capabilities to simultaneously determine the heterogeneous spatial distribution and the concentration of GNPs within tumors in vivo. CONCLUSIONS: In conjunction with PSMA-targeted GNPs, XRFproj/XFCT would be a highly sensitive tool for targeted imaging of prostate cancer, benefiting the elucidation of mechanisms of GNP-assisted prostate-cancer therapy.
Assuntos
Antígenos de Superfície/análise , Glutamato Carboxipeptidase II/análise , Ouro/farmacocinética , Nanopartículas Metálicas , Imagem Óptica/métodos , Neoplasias da Próstata/diagnóstico por imagem , Tomografia Computadorizada por Raios X/métodos , Animais , Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Humanos , Masculino , Camundongos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/terapiaRESUMO
BACKGROUND: Chimeric antigen receptor (CAR) T cells are effective in B-cell malignancies. However, heterogeneous antigen expression and antigen loss remain important limitations of targeted immunotherapy in solid tumors. Therefore, targeting multiple tumor-associated antigens simultaneously is expected to improve the outcome of CAR-T cell therapies. Due to the instability of single-chain variable fragments, it remains challenging to develop the simultaneous targeting of multiple antigens using traditional single-chain fragment variable (scFv)-based CARs. METHODS: We used Humabody VH domains derived from a transgenic mouse to obtain fully human prostate-specific membrane antigen (PSMA) VH and mesothelin (MSLN) VH sequences and redirect T cell with VH based-CAR. The antitumor activity and mode of action of PSMA VH and MSLN VH were evaluated in vitro and in vivo compared with the traditional scFv-based CARs. RESULTS: Human VH domain-based CAR targeting PSMA and MSLN are stable and functional both in vitro and in vivo. VH modules in the bispecific format are capable of binding their specific target with similar affinity as their monovalent counterparts. Bispecific CARs generated by joining two human antibody VH domains can prevent tumor escape in tumor with heterogeneous antigen expression. CONCLUSIONS: Fully human antibody VH domains can be used to generate functional CAR molecules, and redirected T cells elicit antitumoral responses in solid tumors at least as well as conventional scFv-based CARs. In addition, VH domains can be used to generate bispecific CAR-T cells to simultaneously target two different antigens expressed by tumor cells, and therefore, achieve better tumor control in solid tumors.
Assuntos
Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Região Variável de Imunoglobulina/imunologia , Imunoterapia Adotiva , Mesotelina/imunologia , Neoplasias da Próstata/terapia , Receptores de Antígenos Quiméricos/imunologia , Anticorpos de Cadeia Única/imunologia , Linfócitos T/transplante , Animais , Linhagem Celular Tumoral , Técnicas de Cocultura , Citocinas/metabolismo , Citotoxicidade Imunológica , Humanos , Região Variável de Imunoglobulina/genética , Ativação Linfocitária , Masculino , Camundongos Endogâmicos NOD , Fenótipo , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Receptores de Antígenos Quiméricos/genética , Anticorpos de Cadeia Única/genética , Linfócitos T/imunologia , Linfócitos T/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoAssuntos
Adenocarcinoma/terapia , Glutamato Carboxipeptidase II/antagonistas & inibidores , Terapia de Alvo Molecular , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias da Próstata/terapia , Adenocarcinoma/sangue , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/radioterapia , Anticorpos Biespecíficos/uso terapêutico , Anticorpos Antineoplásicos/uso terapêutico , Antígenos de Superfície/sangue , Antígenos de Superfície/imunologia , Biomarcadores Tumorais , Ensaios Clínicos como Assunto , Dipeptídeos/uso terapêutico , Docetaxel/administração & dosagem , Docetaxel/uso terapêutico , Glutamato Carboxipeptidase II/sangue , Glutamato Carboxipeptidase II/imunologia , Compostos Heterocíclicos com 1 Anel/uso terapêutico , Humanos , Imunoconjugados/efeitos adversos , Imunoconjugados/uso terapêutico , Imunoterapia Adotiva , Lutécio/administração & dosagem , Lutécio/efeitos adversos , Lutécio/uso terapêutico , Masculino , Nanopartículas , Proteínas de Neoplasias/análise , Antígeno Prostático Específico , Neoplasias da Próstata/sangue , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/radioterapia , Radioisótopos/administração & dosagem , Radioisótopos/efeitos adversos , Radioisótopos/uso terapêutico , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
PURPOSE: Metastatic castration-resistant prostate cancer (mCRPC) remains a disease with high unmet medical need, as most patients do not achieve durable response with available treatments. Prostate-specific membrane antigen (PSMA) is a compelling target for mCRPC. It is highly expressed by primary and metastatic prostate cancer cells, with increased expression after progression on androgen deprivation therapy. EXPERIMENTAL DESIGN: We developed AMG 160, a half-life extended, bispecific T-cell engager immuno-oncology therapy that binds PSMA on prostate cancer cells and cluster of differentiation 3 on T cells for treatment of mCRPC. AMG 160 was evaluated in vitro and in mCRPC xenograft models. AMG 160 tolerability was assessed in nonhuman primates (NHP). AMG 160 activity as monotherapy and in combination with a PSMA-imaging agent, novel hormonal therapy, and immune checkpoint blockade was evaluated. RESULTS: AMG 160 induces potent, specific killing of PSMA-expressing prostate cancer cell lines in vitro, with half-maximal lysis of 6-42 pmol/L. In vivo, AMG 160 administered weekly at 0.2 mg/kg engages T cells administered systemically and promotes regression of established 22Rv-1 mCRPC xenograft tumors. AMG 160 is compatible with the imaging agent gallium 68-labeled PSMA-11, and shows enhanced cytotoxic activity when combined with enzalutamide or an anti-programmed death-1 antibody. AMG 160 exhibits an extended half-life and has an acceptable safety profile in NHPs. CONCLUSIONS: The preclinical characterization of AMG 160 highlights its potent antitumor activity in vitro and in vivo, and its potential for use with known diagnostic or therapeutic agents in mCRPC. These data support the ongoing clinical evaluation of AMG 160 in patients with mCRPC.See related commentary by Kamat et al., p. 2675.
Assuntos
Transferência Adotiva/métodos , Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Linfócitos T/imunologia , Animais , Complexo CD3/antagonistas & inibidores , Complexo CD3/imunologia , Complexo CD3/metabolismo , Linhagem Celular Tumoral , Citocinas/metabolismo , Citotoxicidade Imunológica , Modelos Animais de Doenças , Relação Dose-Resposta Imunológica , Glutamato Carboxipeptidase II/antagonistas & inibidores , Humanos , Ativação Linfocitária/imunologia , Masculino , Camundongos , Neoplasias de Próstata Resistentes à Castração/patologia , Linfócitos T/metabolismo , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Synthetic small molecules that redirect endogenous antibodies to target cells are promising drug candidates because they overcome the potential shortcomings of therapeutic antibodies, such as immunogenicity and the need for intravenous delivery. Previously, we reported a novel class of bispecific molecules targeting the antibody Fc region and folate receptor, named Fc-binding antibody-recruiting molecules (Fc-ARMs). Fc-ARMs can theoretically recruit most endogenous antibodies, inducing antibody-dependent cell-mediated cytotoxicity (ADCC) to eliminate cancer cells. Herein, we describe new Fc-ARMs that target prostate cancer (Fc-ARM-Ps). Fc-ARM-Ps recruited antibodies to cancer cells expressing prostate-specific membrane antigen but did so with lower efficiency compared with Fc-ARMs targeting the folate receptor. Upon recruitment by Fc-ARM-P, defucosylated antibodies efficiently activated natural killer cells and induced ADCC, whereas antibodies with intact N-glycans did not. The results suggest that the affinity between recruited antibodies and CD16a, a type of Fc receptor expressed on immune cells, could be a key factor controlling immune activation in the Fc-ARM strategy.
Assuntos
Anticorpos Monoclonais/química , Antígenos de Superfície/química , Glutamato Carboxipeptidase II/química , Fragmentos Fc das Imunoglobulinas/química , Anticorpos Monoclonais/imunologia , Reações Antígeno-Anticorpo , Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Estrutura MolecularRESUMO
Aim: We report results of a first-in-human study of pasotuxizumab, a PSMA bispecific T-cell engager (BiTE®) immune therapy mediating T-cell killing of tumor cells in patients with advanced castration-resistant prostate cancer. Patients & methods: We assessed once-daily subcutaneous (SC) pasotuxizumab. All SC patients developed antidrug antibodies; therefore, continuous intravenous (cIV) infusion was assessed. Results: A total of 47 patients received pasotuxizumab (SC: n = 31, 0.5-172 µg/d; cIV: n = 16, 5-80 µg/d). The SC maximum tolerated dose was 172.0 µg/d. A sponsor change stopped the cIV cohort early; maximum tolerated dose was not determined. PSA responders occurred (>50% PSA decline: SC, n = 9; cIV, n = 3), including two long-term responders. Conclusion: Data support pasotuxizumab safety in advanced castration-resistant prostate cancer and represent evidence of BiTE monotherapy efficacy in solid tumors. Clinical trial registration: NCT01723475 (ClinicalTrials.gov).
Assuntos
Anticorpos Biespecíficos , Antineoplásicos Imunológicos , Neoplasias de Próstata Resistentes à Castração , Idoso , Idoso de 80 Anos ou mais , Humanos , Masculino , Pessoa de Meia-Idade , Anticorpos Biespecíficos/imunologia , Anticorpos Biespecíficos/farmacocinética , Anticorpos Biespecíficos/uso terapêutico , Antígenos de Superfície/imunologia , Antineoplásicos Imunológicos/imunologia , Antineoplásicos Imunológicos/farmacocinética , Antineoplásicos Imunológicos/uso terapêutico , Biomarcadores Tumorais/sangue , Complexo CD3/imunologia , Glutamato Carboxipeptidase II/imunologia , Imunoterapia , Infusões Intravenosas , Injeções Subcutâneas , Dose Máxima Tolerável , Neoplasias de Próstata Resistentes à Castração/sangue , Neoplasias de Próstata Resistentes à Castração/imunologia , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/terapia , Resultado do TratamentoRESUMO
Prostate-Specific Membrane Antigen (PSMA) is an established biomarker for the imaging and experimental therapy of prostate cancer (PCa), as it is strongly upregulated in high-grade primary, androgen-independent, and metastatic lesions. Here, we report on the development and functional characterization of recombinant single-chain Fv (scFv) and Fab fragments derived from the 5D3 PSMA-specific monoclonal antibody (mAb). These fragments were engineered, heterologously expressed in insect S2 cells, and purified to homogeneity with yields up to 20 mg/L. In vitro assays including ELISA, immunofluorescence and flow cytometry, revealed that the fragments retain the nanomolar affinity and single target specificity of the parent 5D3 antibody. Importantly, using a murine xenograft model of PCa, we verified the suitability of fluorescently labeled fragments for in vivo imaging of PSMA-positive tumors and compared their pharmacokinetics and tissue distribution to the parent mAb. Collectively, our data provide an experimental basis for the further development of 5D3 recombinant fragments for future clinical use.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Superfície/imunologia , Glutamato Carboxipeptidase II/imunologia , Neoplasias da Próstata/imunologia , Animais , Linhagem Celular , Linhagem Celular Tumoral , Fluorescência , Humanos , Insetos , Masculino , Camundongos , Camundongos Nus , Células PC-3 , Proteínas Recombinantes/imunologia , Anticorpos de Cadeia Única/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Due to the technical innovations in generating bispecific antibodies (BsAbs) in recent years, BsAbs have become important reagents for diagnostic and therapeutic applications. However, the difficulty of producing a heterodimer consisting of two different arms with high yield and purity constituted a major limitation for their application in academic and clinical settings. Here, we describe a novel Fc-containing BsAb format (Fab × sdAb-Fc) composed of a conventional antigen-binding fragment (Fab), and a single domain antibody (sdAb), which avoids heavy-light chain mis-pairing during antibody assembly. In this study, the Fab x sdAb-Fc BsAbs were efficiently produced by three widely used heavy-heavy chain heterodimerization methods: Knobs-into-holes (KIH), Charge-pairs (CP) and controlled Fab-arm exchange (cFAE), respectively. The novel Fab x sdAb-Fc format provided a rapid and efficient strategy to generate BsAb with high purity and a unique possibility to further purify desired BsAbs from undesired antibodies based on molecular weight (MW). Compared to conventional BsAb formats, the advantages of Fab x sdAb-Fc format may thus provide a straightforward opportunity to apply bispecific antibody principles to research and development of novel targets and pathways in diseases such as cancer and autoimmunity.
Assuntos
Anticorpos Biespecíficos/imunologia , Receptores ErbB/imunologia , Glutamato Carboxipeptidase II/imunologia , Fragmentos Fab das Imunoglobulinas/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Imunoglobulina G/imunologia , Glicoproteínas de Membrana/imunologia , Anticorpos de Domínio Único/imunologia , Animais , Anticorpos Biespecíficos/biossíntese , Anticorpos Biespecíficos/genética , Especificidade de Anticorpos , Células CHO , Cricetulus , Receptores ErbB/genética , Receptores ErbB/metabolismo , Glutamato Carboxipeptidase II/genética , Glutamato Carboxipeptidase II/metabolismo , Fragmentos Fab das Imunoglobulinas/biossíntese , Fragmentos Fab das Imunoglobulinas/genética , Fragmentos Fc das Imunoglobulinas/biossíntese , Fragmentos Fc das Imunoglobulinas/genética , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Peso Molecular , Mutação , Estudo de Prova de Conceito , Multimerização Proteica , Anticorpos de Domínio Único/biossíntese , Anticorpos de Domínio Único/genéticaRESUMO
The clinical translation of theranostic 177Lu-radiopharmaceuticals based on inhibitors of the prostate-specific membrane antigen (PSMA) has demonstrated positive clinical responses in patients with advanced prostate cancer (PCa). However, challenges still remain, particularly regarding their pharmacokinetic and dosimetric properties. We developed a potential PSMA-immunotheranostic agent by conjugation of a single-chain variable fragment of the IgGD2B antibody (scFvD2B) to DOTA, to obtain a 177Lu-labelled agent with a better pharmacokinetic profile than those previously reported. The labelled conjugated 177Lu-scFvD2B was obtained in high yield and stability. In vitro, 177Lu-scFvD2B disclosed a higher binding and internalization in LNCaP (PSMA-positive) compared to PC3 (negative control) human PCa cells. In vivo studies in healthy nude mice revealed that 177Lu-scFvD2B present a favorable biokinetic profile, characterized by a rapid clearance from non-target tissues and minimal liver accumulation, but a slow wash-out from kidneys. Micro-SPECT/CT imaging of mice bearing pulmonary microtumors evidenced a slow uptake by LNCaP tumors, which steadily rose up to a maximum value of 3.6 SUV at 192 h. This high and prolonged tumor uptake suggests that 177Lu-scFvD2B has great potential in delivering ablative radiation doses to PSMA-expressing tumors, and warrants further studies to evaluate its preclinical therapeutic efficacy.
Assuntos
Antígenos de Superfície/metabolismo , Antineoplásicos Imunológicos/farmacocinética , Glutamato Carboxipeptidase II/metabolismo , Imunoconjugados/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Compostos Radiofarmacêuticos/farmacocinética , Animais , Antígenos de Superfície/imunologia , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/farmacologia , Linhagem Celular Tumoral , Glutamato Carboxipeptidase II/imunologia , Compostos Heterocíclicos com 1 Anel/química , Humanos , Imunoconjugados/química , Imunoconjugados/farmacocinética , Região Variável de Imunoglobulina/química , Lutécio/farmacocinética , Masculino , Camundongos Nus , Células PC-3 , Medicina de Precisão/métodos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Radioisótopos/farmacocinética , Compostos Radiofarmacêuticos/química , Tomografia Computadorizada com Tomografia Computadorizada de Emissão de Fóton Único , Anticorpos de Cadeia Única/química , Distribuição Tecidual , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The repertoire of methods for the detection and chemotherapeutic treatment of prostate cancer (PCa) is currently limited. Prostate-specific membrane antigen (PSMA) is overexpressed in PCa tumors and can be exploited for both imaging and drug delivery. We developed and characterized four nanobodies that present tight and specific binding and internalization into PSMA+ cells and that accumulate specifically in PSMA+ tumors. We then conjugated one of these nanobodies to the cytotoxic drug doxorubicin, and we show that the conjugate internalizes specifically into PSMA+ cells, where the drug is released and induces cytotoxic activity. In vivo studies show that the extent of tumor growth inhibition is similar when mice are treated with commercial doxorubicin and with a 42-fold lower amount of the nanobody-conjugated doxorubicin, attesting to the efficacy of the conjugated drug. These data highlight nanobodies as promising agents for the imaging of PCa tumors and for the targeted delivery of chemotherapeutic drugs.