RESUMO
A theoretical drawback to alpha-particle therapy with 213Bi is the short range of the particle track coupled with the short half-life of the radionuclide, thereby potentially limiting effective cytotoxicity to rapidly accessible, disseminated individual tumor cells (e.g., as in leukemia). In this work, a prostate carcinoma spheroid model was used to evaluate the feasibility of targeting micrometastatic clusters of tumor cells using 213Bi-labeled anti-prostate-specific membrane antigen (PSMA) antibody, J591. In prostate cancer, vascular dissemination of tumor cells or tumor cell clusters to the marrow constitutes an important step in the progression of this disease to widespread skeletal involvement, an incurable state. Such prevascularized clusters are ideal targets for radiolabeled antibodies because the barriers to antibody penetration that are associated with the capillary basal lamina have not yet formed. Beta- and gamma-emitting radionuclides such as 131I, which are widely used in radioimmunotherapy, are not expected to be effective when targeting single cells or small cell clusters. This is because the range of the emissions is one to two orders of magnitude greater than the target size, and the energy deposited per traversal is insufficient to produce any significant radiobiological effect. Spheroids of the prostate cancer cell line, LNCaP-LN3, were used as a model of prevascularized micrometastases; their response to an anti-PSMA antibody, J591, radiolabeled with the alpha-particle emitter 213Bi (T(1/2), 45.6 min.) has been measured. The time course of spheroid volume reductions was found to be sensitive to the initial spheroid volume. J591 labeled with 0.9 MBq/ml 213Bi resulted in a 3-log reduction in spheroid volume on day 33, relative to control, for spheroids with an initial diameter of 130 microm; 1.8 MBq/ml were required to achieve a similar response for spheroids with an initial diameter of 180 microm. Equivalent spheroid responses were observed after 12 Gy of acute external beam photon irradiation. Monte Carlo-based microdosimetric analyses of the 213Bi decay distribution in individual spheroids of 130-microm diameter yielded an average alpha-particle dose of 3.7 Gy to the spheroids, resulting in a relative biological effectiveness factor of 3.2 over photon irradiation. The activity concentrations used in the experiments were clinically relevant, and this work supports the possibility of using 213Bi-labeled antibodies not only for disseminated single tumor cells, as found in patients with leukemia, but also for micrometastatic tumor deposits up to 180 microm in diameter (1200 cells).
Assuntos
Antígenos de Superfície , Bismuto/farmacologia , Carboxipeptidases/imunologia , Imunotoxinas/farmacologia , Neoplasias da Próstata/radioterapia , Radioisótopos/farmacologia , Partículas alfa/uso terapêutico , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/farmacologia , Divisão Celular/efeitos da radiação , Glutamato Carboxipeptidase II , Humanos , Imunotoxinas/imunologia , Imunotoxinas/metabolismo , Masculino , Microscopia Confocal , Metástase Neoplásica , Neoplasias da Próstata/imunologia , Radioimunoterapia , Esferoides Celulares/imunologia , Esferoides Celulares/efeitos da radiação , Células Tumorais CultivadasRESUMO
A novel alpha-particle emitting monoclonal antibody construct targeting the external domain of prostate-specific membrane antigen (PSMA) was prepared and evaluated in vitro and in vivo. The chelating agent, N-[2-amino-3-(p-isothiocyanatophen-yl)propyl]-trans-cyclohexane-1, 2-diamine-N,N',N',N'',N''-pentaacetic acid, was appended to J591 monoclonal antibody to stably bind the 213Bi radiometal ion. Bismuth-213 is a short-lived (t 1/2 = 46 min) radionuclide that emits high energy alpha-particles with an effective range of 0.07-0.10 mm that are ideally suited to treating single-celled neoplasms and micrometastatic carcinomas. The LNCaP prostate cancer cell line had an estimated 180,000 molecules of PSMA per cell; J591 bound to PSMA with a 3-nM affinity. After binding, the radiolabeled construct-antigen complex was rapidly internalized into the cell, carrying the radiometal inside. [213Bi]J591 was specifically cytotoxic to LNCaP. The LD50 value of [213Bi]J591 was 220 nCi/ml at a specific activity of 6.4 Ci/g. The potency and specificity of [213Bi]J591 directed against LNCaP spheroids, an in vitro model for micrometastatic cancer, also was investigated. [213Bi]J591 effectively stopped growth of LNCaP spheroids relative to an equivalent dose of the irrelevant control [213Bi]HuM195 or unlabeled J591. Cytotoxicity experiments in vivo were carried out in an athymic nude mouse model with an i.m. xenograft of LNCaP cells. [213Bi]J591 was able to significantly improve (P < 0.0031) median tumor-free survival (54 days) in these experiments relative to treatment with irrelevant control [213Bi]HuM195 (33 days), or no treatment (31 days). Prostate-specific antigen (PSA) was also specifically reduced in treated animals. At day 51, mean PSA values were 104 ng/ml +/- 54 ng/ml (n = 4, untreated animals), 66 ng/ml +/- 16 ng/ml (n = 6, animals treated with [213Bi]HuM195), and 28 ng/ml +/- 22 ng/ml (n = 6, animals treated with [213Bi]J591). The reduction of PSA levels in mice treated with [213Bi]J591 relative to mice treated with [213Bi]HuM195 and untreated control animals was significant with P < 0.007 and P < 0.0136, respectively. In conclusion, a novel [213Bi]-radiolabeled J591 has been constructed that selectively delivers alpha-particles to prostate cancer cells for potent and specific killing in vitro and in vivo.
Assuntos
Anticorpos Monoclonais/farmacologia , Bismuto/farmacologia , Imunotoxinas/farmacologia , Neoplasias da Próstata/radioterapia , Radioimunoterapia , Radioisótopos/farmacologia , Partículas alfa/uso terapêutico , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/metabolismo , Complexo Antígeno-Anticorpo/metabolismo , Sítios de Ligação , Morte Celular/efeitos da radiação , Humanos , Imunotoxinas/imunologia , Imunotoxinas/metabolismo , Cinética , Masculino , Camundongos , Camundongos Nus , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Esferoides Celulares/efeitos da radiação , Especificidade por Substrato , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cells from the prostate tumor cell line LNCaP have been grown as spheroids. The growth kinetics of the spheroids have been characterized by fitting a Gompertz equation to spheroid growth curves. The proliferation state of cells within spheroids of different diameters was assessed by bromodeoxyuridine staining. Scanning and electron transmission microscopy were performed to determine the ultrastructure of the spheroids. Prostate-specific antigen (PSA) secretion was monitored throughout spheroid growth. Consistent with Gompertzian kinetics, the volume of LNCaP spheroids initially increased exponentially and then reached a plateau. The doubling time during the exponential phase was 29 +/- 4 h. A core of nonproliferating cells was seen in spheroids with a diameter of 400 microm; at a diameter of 600 microm, a necrotic core had formed. In smaller, 200-microm diameter spheroids, a core of nonproliferating cells was not seen, but proliferating cells were concentrated at the spheroid periphery. Electron microscopy showed that the spheroids were enveloped by an extracellular matrix and that cell adhesion within the spheroids was due in part to desmosomes. PSA secretion by the spheroids could be modeled as originating from a spherical shell whose thickness was independent of overall spheroid diameter. The shell thickness obtained by fitting an appropriate equation to the data was consistent with that determined from the bromodeoxyuridine studies. LNCaP cells exhibit several important features of prostate cancer cells; in vivo, they are androgen responsive, and they express prostatic acid phosphatase, PSA, and prostate-specific membrane antigen. LNCaP spheroids provide a simple but relevant model for the study of drug delivery and response in prostate cancer.
Assuntos
Neoplasias da Próstata/patologia , Esferoides Celulares/patologia , Bromodesoxiuridina/metabolismo , Divisão Celular , Humanos , Masculino , Antígeno Prostático Específico/sangue , Células Tumorais CultivadasRESUMO
UNLABELLED: Data from nine patients with leukemia participating in a phase I activity-escalation study of HuM195, labeled with the alpha-particle emitter 213Bi (half-life = 45.6 min), were used to estimate pharmacokinetics and dosimetry. This is the first trial using an alpha-particle emitter in humans. The linear energy transfer of alpha particles is several hundredfold greater than that of beta emissions. The range in tissue is approximately 60-90 microm. METHODS: The activity administered to patients ranged from 0.6 to 1.6 GBq. Patient imaging was initiated at the start of each injection. Thirty 1-min images followed by ten 3-min images were collected in dynamic mode; a 20% photopeak window centered at 440 keV was used. Blood samples were collected until 3 h postinjection and counted in a gamma counter. Contours around the liver and spleen were drawn on the anterior and posterior views and around a portion of the spine on the posterior views. No other organs were visualized. RESULTS: The percentage injected dose in the liver and spleen volumes increased rapidly over the first 10-15 min to a constant value for the remaining hour of imaging, yielding a very rapid uptake followed by a plateau in the antibody uptake curves. The kinetic curves were integrated to yield cumulated activity. The mean energy emitted per nuclear transition for 213Bi and its daughters, adjusted by a relative biologic effectiveness of 5 for alpha emissions, was multiplied by the cumulated activity to yield the absorbed dose equivalent. Photon dose to the total body was determined by calculating a photon-absorbed fraction. The absorbed dose equivalent to liver and spleen volumes ranged from 2.4 to 11.2 and 2.9 to 21.9 Sv, respectively. Marrow (or leukemia) mean dose ranged from 6.6 to 12.2 Sv. The total-body dose (photons only) ranged from 2.2 x 10(-4) to 5.8 x 10(-4) Gy. CONCLUSION: This study shows that patient imaging of 213Bi, an alpha-particle emitter, labeled to HuM195 is possible and may be used to derive pharmacokinetics and dosimetry. The absorbed dose ratio between marrow, liver and spleen volumes and the whole body for 213Bi-HuM195 is 1000-fold greater than that commonly observed with beta-emitting radionuclides used for radioimmunotherapy.
Assuntos
Bismuto , Leucemia Mieloide/radioterapia , Radioimunoterapia , Radioisótopos , Doença Aguda , Partículas alfa , Animais , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapêutico , Câmaras gama , Humanos , Leucemia Mieloide/diagnóstico por imagem , Camundongos , Cintilografia , Dosagem RadioterapêuticaRESUMO
Hypocrellins are perylenequinone pigments with substantial absorption in the red spectral region and high singlet oxygen yield. They are available in pure monomeric form and may be derivatized to optimize properties of red light absorption, tissue biodistribution and toxicity. In vitro screening of synthetic derivatives of the naturally occurring compound, hypocrellin B (HB), for optimal properties of cyto-(dark) toxicity and phototoxicity resulted in selection of three compounds for preclinical evaluation: HBEA-R1 (ethanolaminated HB), HBBA-R2 (butylaminated HB) and HBDP-R1 [2-(N,N-dimethylamino)-propylamine-HB]. Extinction coefficients at 630 nm (epsilon 630) are 6230, 6190 and 4800, respectively; and 1O2 quantum yields, phi, 0.60, 0.32 and 0.42. Intracellular uptake is essentially complete within 2 h (HBEA-R1, HBBA-R2) and 20 h (HBDP-R1). Greatest uptake is associated with lysosomes and Golgi. The HBEA-R1 and HBBA-R2 elicit phototoxicity in vitro primarily via the type II mechanism, with some type I activity under stringently hypoxic conditions. Transcutaneous phototherapy with HBEA-R1 permanently ablates EMT6/Ed tumors growing in the flanks of Balb/c mice, with minimal cutaneous effects. The HBBA-R2 does not elicit mutagenic activity in strains TA98 and TA100 of Salmonella typhimurium. Further development of selected hypocrellin derivatives as photosensitizers for photodynamic therapy is warranted.
Assuntos
Neoplasias Experimentais/tratamento farmacológico , Perileno/análogos & derivados , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Quinonas/uso terapêutico , Animais , Concentração de Íons de Hidrogênio , Camundongos , Camundongos Endogâmicos BALB C , Testes de Mutagenicidade , Oxigênio/metabolismo , Perileno/efeitos adversos , Perileno/farmacocinética , Perileno/uso terapêutico , Fármacos Fotossensibilizantes/efeitos adversos , Fármacos Fotossensibilizantes/farmacocinética , Quinonas/efeitos adversos , Quinonas/farmacocinética , Distribuição TecidualRESUMO
Photodynamic therapy (PDT) is a promising technique for destroying tumours. Photosensitizing drugs presently available are not sufficiently tumour specific; hence, light dosimetry is required in order to control light exposure and thereby restrict cell kill to the target tissue to avoid damage to healthy tissue. Current light dosimetry methods rely on tissue optical characterization by fluence measurements at several points. Fluence-based tissue characterization is impractical for tumours in organs such as prostate where access by optical probes is limited and the tumours are highly optically inhomogeneous. This paper explores the potential of radiance-based light dosimetry as an alternative. Correlation is found between Monte Carlo simulation of radiance in a tissue phantom and radiance measurements made using a new radiance probe. Radiance is sensitive to variations in the tissue optical parameters, absorption coefficient mu(a), scattering coefficient mu(s), and anisotropy factor g, and therefore is potentially useful for tissue characterization. Radiance measurements have several advantages over fluence measurements. Radiance measurements provide more information from a single location, better spatial resolution of the tissue optical parameters, and higher sensitivity in discriminating between different media. However, the Monte Carlo method is too slow to be of practical value for tissue characterization by correlation of measured and simulated radiance. An analytical solution to the transport equation for radiance would be desirable as this would facilitate and increase the speed of tissue characterization.
Assuntos
Fotoquimioterapia , Próstata/anatomia & histologia , Neoplasias da Próstata/tratamento farmacológico , Anisotropia , Fenômenos Biofísicos , Biofísica , Humanos , Técnicas In Vitro , Luz , Masculino , Modelos Anatômicos , Modelos Biológicos , Método de Monte Carlo , Fótons , Espalhamento de RadiaçãoRESUMO
BACKGROUND AND OBJECTIVE: Variations in the optical coefficients in tissue and the photosensitizer during photodynamic therapy (PDT) will require adjustment of the light dose during the course of therapy. We have studied the dynamics using light transmission spectra for two different tumor models when tetrasulfonated aluminum phthalocyanine (AlPcS4) was used as photosensitizer. STUDY DESIGN/MATERIALS AND METHODS: Spectra were measured noninvasively in the EMT6/Ed murine tumor model, and with interstitially implanted source and probe fibers in the Dunning R3327-AT rat tumor model. Measurements were performed in the range 600-840 nm, using a tunable dye laser, a diode laser, and a Ti:Sapphire laser. AlPcS4 has absorption in the range 600-700 nm with an absorption peak at 670 nm in saline. RESULTS: The in vivo spectrum of AlPcS4 both in the EMT6/Ed tumor model and the Dunning R3327-AT tumor model differs from the spectrum of AlPcS4 in saline. The absorption at 670 nm was reduced, whereas the absorption at 640 nm increased. Exposure of phototherapeutic levels of light caused reduced light absorption by the photosensitizer and further spectral shift. CONCLUSION: We found that the AIPcS4 absorption spectrum changes in a biological environment, and we also observed increased light transmission at the treatment wavelength during PDT in both tumor models. Instability in the absorption spectrum of the photosensitizer may influence the effectiveness of PDT.
Assuntos
Indóis/uso terapêutico , Neoplasias Experimentais/tratamento farmacológico , Compostos Organometálicos/uso terapêutico , Fotoquimioterapia , Radiossensibilizantes/uso terapêutico , Animais , Antineoplásicos/uso terapêutico , Lasers , Masculino , Camundongos , Transplante de Neoplasias , Imagens de Fantasmas , Neoplasias da Próstata/tratamento farmacológico , Ratos , Espectrometria de Fluorescência , Espectrofotometria , Células Tumorais CultivadasRESUMO
Temporal and illumination-induced variations in the in vivo light transmission spectrum of the photosensitizer will influence light dosimetry for photodynamic therapy (PDT). The present authors have studied the in vivo spectra of four photosensitizers in the EMT6/Ed murine tumour model in Balb/c mice. The following photosensitizers were used: bis(dimethylthexylsiloxy)silicon 2,3-naphthalocyanine (SiNc 8), benzoporphyrin-derivative monoacid ring A (BPD Verteporfin), Photofrin and ethanolamined hypocrellin B (HBEA-R2). Spectra were measured non-invasively in the EMT6/Ed murine tumour model in the spectral range 600-840 nm, using a diode laser, a dye laser and a Ti:sapphire laser. Red-shift and broadening of the SiNc 8 absorption band was observed at 790 nm, and a slight red-shift was observed in the BPD, HBEA-R2 and Photofrin in vivo absorption spectrum. Exposure to 300 J of light at the peak absorption wavelength caused complete photobleaching of BPD at 690 nm, and a reduced absorption by SiNc 8 at 780 nm, Photofrin at 626 nm, and HBEA-R2 at 656 nm.
RESUMO
BACKGROUND AND OBJECTIVE: The goal of this study is to determine if flat cleaved fiber probes are appropriate for interstitial measurements of radiance in tissue. Flat cleaved probes have the advantage of high responsivity, and they are easy to insert into tissue. Owing to the non-isotropic response of flat cleaved probes, a calibration function is required, taking the anisotropy in the radiance in tissue into account. STUDY DESIGN, MATERIALS AND METHODS: The method used to determine this function consists of radiance measurements in tissue, performed with a flat cleaved fiber probe mounted on a stereotactic stage for insertion into the tissue from different directions. Interstitial irradiation at 630 nm was delivered by a spherical source. RESULTS: We found that the degree of anisotropy in the radiance decreases with increasing distance from the interstitially implanted source in two different tissue phantoms and in the Dunning R3327-AT and R3327-H rat tumor models. CONCLUSION: A position-dependent calibration function is required for interstitially implanted flat cleaved fiber probes. An anisotropy function is presented, which modifies the measurements of radiance with a flat cleaved probe, to account for the change in anisotropy in the radiance. The anisotropy functions for the two tumor models differ substantially.