RESUMO
PURPOSE: PSMA (prostate-specific membrane antigen) is highly expressed on prostate cancer (PrCa) cells and extensively used as a homing target for PrCa treatment. Most prominently, PSMA-targeting conjugate PSMA-617, carrying a DOTA chelator and labeled with therapeutic radionuclides like beta-emitting lutetium-177 or alpha-emitting actinium-225, has shown clinical activity in PrCa patients. We sought to develop PSMA-targeting small molecule (SMOL) conjugates that show high uptake in PSMA-expressing tumors and fast clearance, and can easily be labeled with the alpha emitter thorium-227 (half-life 18.7 days). METHODS: A novel linker motif with improved competition against 3H-PSMA-617 on PSMA-expressing LNCaP cells was identified. A 2,3-hydroxypyridinone chelator modified with carboxyl groups (carboxy-HOPO) with increased hydrophilicity and robust labeling with thorium-227 was developed and allowed the synthesis of mono-, di-, tri-, and tetrameric conjugates. The resulting monomeric and multimeric PSMA SMOL-TTCs (targeted thorium conjugate) were evaluated for cellular binding, internalization, and antiproliferative activity. The in vivo antitumor efficacy of the PSMA SMOL-TTCs was determined in ST1273 and KUCaP-1 PrCa models in mice, and their biodistribution was assessed in cynomolgus monkeys, minipigs, and mice. RESULTS: The monomeric and multimeric PSMA SMOL conjugates were readily labeled with thorium-227 at room temperature and possessed high stability and good binding, internalization, and antiproliferative activity in vitro. In vivo, the monomeric, dimeric, and trimeric PSMA SMOL-TTCs showed fast clearance, potent antitumor efficacy, and high uptake and retention in prostate tumors in mice. No major uptake or retention in other organs was observed beyond kidneys. Low uptake of free thorium-227 into bone confirmed high complex stability in vivo. Salivary gland uptake remained inconclusive as mini pigs were devalidated as a relevant model and imaging controls failed in cynomolgus monkeys. CONCLUSION: Monomeric and multimeric PSMA SMOL-TTCs show high tumor uptake and fast clearance in preclinical models and warrant further therapeutic exploration.
Assuntos
Neoplasias da Próstata , Tório , Masculino , Humanos , Animais , Camundongos , Suínos , Distribuição Tecidual , Macaca fascicularis/metabolismo , Porco Miniatura/metabolismo , Glutamato Carboxipeptidase II/metabolismo , Antígenos de Superfície/metabolismo , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/metabolismo , Compostos Radiofarmacêuticos , Quelantes/química , Linhagem Celular TumoralRESUMO
BACKGROUND: Thymic epithelial tumours (TETs) are rare tumours comprised of thymomas and thymic carcinoma. Novel therapies are needed, especially in thymic carcinoma where the 5-year survival rate hovers at 30%. Mesothelin (MSLN), a surface glycoprotein that is cleaved to produce mature MSLN (mMSLN) and megakaryocyte potentiating factor (MPF), is expressed in limited tissues. However, its expression is present in various cancers, including thymic carcinoma, where it is expressed in 79% of cases. METHODS: We utilised flow cytometry, in vitro cytotoxicity assays, and an in vivo xenograft model in order to demonstrate the ability of the MSLN targeting antibody-drug conjugate (ADC) anetumab ravtansine (ARav) in inhibiting the growth of thymic carcinoma. RESULTS: Thymoma and thymic carcinoma cell lines express MSLN, and anetumab, the antibody moiety of ARav, was capable of binding MSLN expressing thymic carcinoma cells and internalising. ARav was effective at inhibiting the growth of thymic carcinoma cells stably transfected with mMSLN in vitro. In vivo, 15 mg/kg ARav inhibited T1889 xenograft tumour growth, while combining 7.5 mg/kg ARav with 4 mg/kg cisplatin yielded an additive effect on inhibiting tumour growth. CONCLUSIONS: These data demonstrate that anetumab ravtansine inhibits the growth of MSLN positive thymic carcinoma cells in vitro and in vivo.
Assuntos
Imunoconjugados/administração & dosagem , Maitansina/análogos & derivados , Mesotelina/genética , Mesotelina/metabolismo , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Timoma/tratamento farmacológico , Neoplasias do Timo/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cisplatino/administração & dosagem , Cisplatino/farmacologia , Sinergismo Farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HT29 , Humanos , Imunoconjugados/farmacologia , Maitansina/administração & dosagem , Maitansina/farmacologia , Camundongos , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Epiteliais e Glandulares/metabolismo , Timoma/genética , Timoma/metabolismo , Neoplasias do Timo/genética , Neoplasias do Timo/metabolismo , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Initially, prostate cancer responds to hormone therapy, but eventually resistance develops. Beta emitter-based prostate-specific membrane antigen (PSMA)-targeted radionuclide therapy is approved for the treatment of metastatic castration-resistant prostate cancer. Here we introduce a targeted alpha therapy (TAT) consisting of the PSMA antibody pelgifatamab covalently linked to a macropa chelator and labeled with actinium-225 and compare its efficacy and tolerability with other TATs. EXPERIMENTAL DESIGN: The in vitro characteristics and in vivo biodistribution, antitumor efficacy, and tolerability of 225Ac-macropa-pelgifatamab (225Ac-pelgi) and other TATs were investigated in cell line- and patient-derived prostate cancer xenograft models. The antitumor efficacy of 225Ac-pelgi was also investigated in combination with the androgen receptor inhibitor darolutamide. RESULTS: Actinium-225-labeling of 225Ac-pelgi was efficient already at room temperature. Potent in vitro cytotoxicity was seen in PSMA-expressing (LNCaP, MDA-PCa-2b, and C4-2) but not in PSMA-negative (PC-3 and DU-145) cell lines. High tumor accumulation was seen for both 225Ac-pelgi and 225Ac-DOTA-pelgi in the MDA-PCa-2b xenograft model. In the C4-2 xenograft model, 225Ac-pelgi showed enhanced antitumor efficacy with a T/Cvolume (treatment/control) ratio of 0.10 compared with 225Ac-DOTA-pelgi, 225Ac-DOTA-J591, and 227Th-HOPO-pelgifatamab (227Th-pelgi; all at 300 kBq/kg) with T/Cvolume ratios of 0.37, 0.39, and 0.33, respectively. 225Ac-pelgi was less myelosuppressive than 227Th-pelgi. 225Ac-pelgi showed dose-dependent treatment efficacy in the patient-derived KuCaP-1 model and strong combination potential with darolutamide in both cell line- (22Rv1) and patient-derived (ST1273) xenograft models. CONCLUSIONS: These results provide a strong rationale to investigate 225Ac-pelgi in patients with prostate cancer. A clinical phase I study has been initiated (NCT06052306).
Assuntos
Actínio , Partículas alfa , Antígenos de Superfície , Glutamato Carboxipeptidase II , Ensaios Antitumorais Modelo de Xenoenxerto , Masculino , Humanos , Animais , Camundongos , Linhagem Celular Tumoral , Glutamato Carboxipeptidase II/antagonistas & inibidores , Glutamato Carboxipeptidase II/metabolismo , Antígenos de Superfície/metabolismo , Partículas alfa/uso terapêutico , Distribuição Tecidual , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/patologia , Compostos Radiofarmacêuticos/administração & dosagemRESUMO
The [(18)F]fluorocyclobutyl group has the potential to be a metabolically stable prosthetic group for PET tracers. The synthesis of the radiolabeling precursor cis-cyclobutane-1,3-diyl bis(toluene-4-sulfonate) 8 was obtained from epibromohydrin in 7 steps (2% overall yield). The radiolabeling of this precursor 8 and its conjugation to L-tyrosine as a model system was successfully achieved to give the new non-natural amino acid 3-[(18)F]fluorocyclobutyl-L-tyrosine (L-3-[(18)F]FCBT) [(18)F]17 in 8% decay-corrected yield from the non-carrier-added [(18)F]fluoride. L-3-[(18)F]FCBT was investigated in vitro in different cancer cell lines to determine the uptake and stability. The tracer [(18)F]17 showed a time dependent uptake into different tumor cell lines (A549, NCI-H460, DU145) with the best uptake of 5.8% injected dose per 5×10(5) cells after 30min in human lung carcinoma cells A549. The stability of L-3-[(18)F]FCBT in human and rat plasma and the stability of the non-radioactive L-3-FCBT in rat hepatocytes were both found to be excellent. These results show that the non-natural amino acid L-3-[(18)F]FCBT is a promising metabolically stable radiotracer for positron emission tomography.
Assuntos
Ciclobutanos/farmacocinética , Radioisótopos de Flúor/farmacocinética , Compostos Radiofarmacêuticos/farmacocinética , Tirosina/farmacocinética , Animais , Linhagem Celular Tumoral , Ciclobutanos/síntese química , Ciclobutanos/química , Radioisótopos de Flúor/química , Hepatócitos/química , Humanos , Modelos Moleculares , Estrutura Molecular , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/química , Ratos , Ratos Wistar , Tirosina/químicaRESUMO
Targeted alpha therapies (TAT) are an innovative class of therapies for cancer treatment. The unique mode-of-action of TATs is the induction of deleterious DNA double-strand breaks. Difficult-to-treat cancers, such as gynecologic cancers upregulating the chemoresistance P-glycoprotein (p-gp) and overexpressing the membrane protein mesothelin (MSLN), are promising targets for TATs. Here, based on the previous encouraging findings with monotherapy, we investigated the efficacy of the mesothelin-targeted thorium-227 conjugate (MSLN-TTC) both as monotherapy and in combination with chemotherapies and antiangiogenic compounds in ovarian and cervical cancer models expressing p-gp. MSLN-TTC monotherapy showed equal cytotoxicity in vitro in p-gp-positive and -negative cancer cells, while chemotherapeutics dramatically lost activity on p-gp-positive cancer cells. In vivo, MSLN-TTC exhibited dose-dependent tumor growth inhibition with treatment/control ratios of 0.03-0.44 in various xenograft models irrespective of p-gp expression status. Furthermore, MSLN-TTC was more efficacious in p-gp-expressing tumors than chemotherapeutics. In the MSLN-expressing ST206B ovarian cancer patient-derived xenograft model, MSLN-TTC accumulated specifically in the tumor, which combined with pegylated liposomal doxorubicin (Doxil), docetaxel, bevacizumab, or regorafenib treatment induced additive-to-synergistic antitumor efficacy and substantially increased response rates compared with respective monotherapies. The combination treatments were well tolerated and only transient decreases in white and red blood cells were observed. In summary, we demonstrate that MSLN-TTC treatment shows efficacy in p-gp-expressing models of chemoresistance and has combination potential with chemo- and antiangiogenic therapies.
Assuntos
Mesotelina , Humanos , Feminino , Proteínas Ligadas por GPI , Linhagem Celular Tumoral , Resistência a MedicamentosRESUMO
In vivo cell-death imaging is still a challenging issue. Until now, only (99m)Tc-labeled HYNIC-rh-annexin A5 has been extensively studied in clinical trials. In the ongoing search for an alternative imaging agent, we synthesized a series of fluorescent zinc-cyclen complexes as annexin A5 mimics and studied structural variations on the uptake behavior of cells undergoing apoptosis/necrosis. The number of cyclen chelators was varied and the spacer separating cyclen from the central scaffold was modified. Five zinc-cyclen complexes were labeled with fluorescein for flow cytometric studies and one was labeled with (18)F for in vivo applications. Jurkat cells were treated with staurosporine to induce apoptosis/necrosis, incubated with the fluorescein-labeled zinc complexes and analyzed them by flow cytometry. Fluorescent annexin A5 and propidium iodide were applied as reference dyes. Flow cytometry revealed greater accumulation of zinc-cyclen complexes in staurosporine treated cells. The uptake was contingent on the presence of zinc and the fluorescence intensity was dependent on the number of zinc-cyclen groups. Confocal laser scanning microscopy showed the {bis[Zn(cyclen)]}(4+) complex distributed throughout the cytosol different to annexin A5. Owing to the structural similarity of the bis-cyclen ligands with CXCR4 binding bis-cyclam derivatives the zinc-cyclen complex uptake was challenged with the meta derivative of AMD3100. Lack of uptake depletion in staurosporine treated cells ruled out measurable CXCR4 interaction. PET imaging using the (18)F labeled zinc-cyclen complex revealed significantly higher uptake in an irradiated Dunning R3327-AT1 prostate tumor as compared to the contralateral control tumor. PET imaging of a HelaMatu tumor model additionally showed an increased uptake after taxol treatment. It could be demonstrated that the fluorescent zinc-cyclen complexes offer potential as new agents for flow cytometry and microscopic imaging of cell death. In addition, the (18)F labeled analogue holds promise for in vivo applications providing informations about cell death after radiation therapy and cytostatic drug treatment.
Assuntos
Apoptose , Complexos de Coordenação , Corantes Fluorescentes , Compostos Heterocíclicos , Necrose/diagnóstico , Zinco , Animais , Complexos de Coordenação/química , Ciclamos , Citometria de Fluxo , Corantes Fluorescentes/química , Compostos Heterocíclicos/química , Humanos , Células Jurkat , Masculino , Microscopia Confocal , Neoplasias/diagnóstico , Tomografia por Emissão de Pósitrons , Ratos , Zinco/químicaRESUMO
The transfer of peptides identified through the phage display technology to clinical applications is difficult. Major drawbacks are the metabolic degradation and label instability. The aim of our work is the optimization of DUP-1, a peptide which was identified by phage display to specifically target human prostate carcinoma. To investigate the influence of chelate conjugation, DOTA was coupled to DUP-1 and labeling was performed with ¹¹¹In. To improve serum stability cyclization of DUP-1 and targeted D-amino acid substitution were carried out. Alanine scanning was performed for identification of the binding site and based on the results peptide fragments were chemically synthesized. The properties of modified ligands were investigated in in vitro binding and competition assays. In vivo biodistribution studies were carried out in mice, carrying human prostate tumors subcutaneously. DOTA conjugation resulted in different cellular binding kinetics, rapid in vivo renal clearance and increased tumor-to-organ ratios. Cyclization and D-amino acid substitution increased the metabolic stability but led to binding affinity decrease. Fragment investigation indicated that the sequence NRAQDY might be significant for target-binding. Our results demonstrate challenges in optimizing peptides, identified through phage display libraries, and show that careful investigation of modified derivatives is necessary in order to improve their characteristics.
Assuntos
Radioisótopos de Índio/química , Peptídeos/química , Neoplasias da Próstata/química , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Quelantes/química , Compostos Heterocíclicos com 1 Anel/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Dados de Sequência Molecular , Biblioteca de Peptídeos , Peptídeos/genética , Distribuição TecidualRESUMO
The aim of this study was to investigate effects of high LET α-radiation in combination with inhibitors of DDR (DNA-PK and ATM) and to compare the effect with the radiosensitizing effect of low LET X-ray radiation. The various cell lines were irradiated with α-radiation and with X-ray. Clonogenic survival, the formation of micronuclei and cell cycle distribution were studied after combining of radiation with DDR inhibitors. The inhibitors sensitized different cancer cell lines to radiation. DNA-PKi affected survival rates in combination with α-radiation in selected cell lines. The sensitization enhancement ratios were in the range of 1.6-1.85 in cancer cells. ATMi sensitized H460 cells and significantly increased the micronucleus frequency for both radiation qualities. ATMi in combination with α-radiation reduced survival of HEK293. A significantly elicited cell cycle arrest in G2/M phase after co-treatment of ATMi with α-radiation and X-ray. The most prominent treatment effect was observed in the HEK293 by combining α-radiation and inhibitions. ATMi preferentially sensitized cancer cells and normal HEK293 cells to α-radiation. DNA-PKi and ATMi can sensitize cancer cells to X-ray, but the effectiveness was dependent on cancer cells itself. α-radiation reduced proliferation in primary fibroblast without G2/M arrest.
Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/farmacologia , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Reparo do DNA/efeitos da radiação , Proteína Quinase Ativada por DNA/farmacologia , Tolerância a Radiação/efeitos dos fármacos , Radiossensibilizantes/farmacologia , Partículas alfa , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/efeitos da radiação , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Células HEK293 , Histonas/metabolismo , Humanos , Transferência Linear de Energia , Testes para Micronúcleos , Radiação Ionizante , Radiometria , Raios XRESUMO
PURPOSE: Androgen receptor (AR) inhibitors are well established in the treatment of castration-resistant prostate cancer and have recently shown efficacy also in castration-sensitive prostate cancer. Although most patients respond well to initial therapy, resistance eventually develops, and thus, more effective therapeutic approaches are needed. Prostate-specific membrane antigen (PSMA) is highly expressed in prostate cancer and presents an attractive target for radionuclide therapy. Here, we evaluated the efficacy and explored the mode of action of the PSMA-targeted thorium-227 conjugate (PSMA-TTC) BAY 2315497, an antibody-based targeted alpha-therapy, in combination with the AR inhibitor darolutamide. EXPERIMENTAL DESIGN: The in vitro and in vivo antitumor efficacy and mode of action of the combination treatment were investigated in preclinical cell line-derived and patient-derived prostate cancer xenograft models with different levels of PSMA expression. RESULTS: Darolutamide induced the expression of PSMA in androgen-sensitive VCaP and LNCaP cells in vitro, and the efficacy of darolutamide in combination with PSMA-TTC was synergistic in these cells. In vivo, the combination treatment showed synergistic antitumor efficacy in the low PSMA-expressing VCaP and in the high PSMA-expressing ST1273 prostate cancer models, and enhanced efficacy in the enzalutamide-resistant KUCaP-1 model. The treatments were well tolerated. Mode-of-action studies revealed that darolutamide induced PSMA expression, resulting in higher tumor uptake of PSMA-TTC, and consequently, higher antitumor efficacy, and impaired PSMA-TTC-mediated induction of DNA damage repair genes, potentially contributing to increased DNA damage. CONCLUSIONS: These results provide a strong rationale to investigate PSMA-TTC in combination with AR inhibitors in patients with prostate cancer.
Assuntos
Antagonistas de Receptores de Andrógenos , Antígenos de Superfície , Glutamato Carboxipeptidase II , Neoplasias da Próstata , Pirazóis , Tório , Animais , Humanos , Masculino , Camundongos , Antagonistas de Receptores de Andrógenos/uso terapêutico , Antígenos de Superfície/efeitos dos fármacos , Combinação de Medicamentos , Glutamato Carboxipeptidase II/efeitos dos fármacos , Modelos Biológicos , Neoplasias da Próstata/tratamento farmacológico , Pirazóis/uso terapêutico , Tório/uso terapêuticoRESUMO
Eukaryotes have evolved two major pathways to repair potentially lethal DNA double-strand breaks. Homologous recombination represents a precise, DNA-template-based mechanism available during the S and G2 cell cycle phase, whereas non-homologous end joining, which requires DNA-dependent protein kinase (DNA-PK), allows for fast, cell cycle-independent but less accurate DNA repair. Here, we report the discovery of BAY-8400, a novel selective inhibitor of DNA-PK. Starting from a triazoloquinoxaline, which had been identified as a hit from a screen for ataxia telangiectasia and Rad3-related protein (ATR) inhibitors with inhibitory activity against ATR, ATM, and DNA-PK, lead optimization efforts focusing on potency and selectivity led to the discovery of BAY-8400. In in vitro studies, BAY-8400 showed synergistic activity of DNA-PK inhibition with DNA damage-inducing targeted alpha therapy. Combination of PSMA-targeted thorium-227 conjugate BAY 2315497 treatment of human prostate tumor-bearing mice with BAY-8400 oral treatment increased antitumor efficacy, as compared to PSMA-targeted thorium-227 conjugate monotherapy.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Proteína Quinase Ativada por DNA/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células , Proteína Quinase Ativada por DNA/genética , Sinergismo Farmacológico , Quimioterapia Combinada , Hepatócitos/efeitos dos fármacos , Humanos , Camundongos , Estrutura Molecular , Fosfatidilinositol 3-Quinases/genética , Ratos , Relação Estrutura-Atividade , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Targeted thorium-227 conjugates (TTCs) are an emerging class of targeted alpha therapies (TATs). Their unique mode of action (MoA) is the induction of difficult-to-repair clustered DNA double-strand breaks. However, thus far, their effects on the immune system are largely unknown. Here, we investigated the immunostimulatory effects of the mesothelin-targeted thorium-227 conjugate (MSLN-TTC) in vitro and in vivo in monotherapy and in combination with an inhibitor of the immune checkpoint programmed death receptor ligand 1 (PD-L1) in immunocompetent mice. METHODS: The murine cell line MC38 was transfected with the human gene encoding for MSLN (hMSLN) to enable binding of the non-cross-reactive MSLN-TTC. The immunostimulatory effects of MSLN-TTC were studied in vitro on human cancer cell lines and MC38-hMSLN cells. The efficacy and MoA of MSLN-TTC were studied in vivo as monotherapy or in combination with anti-PD-L1 in MC38-hMSLN tumor-bearing immunocompetent C57BL/6 mice. Experiments were supported by RNA sequencing, flow cytometry, immunohistochemistry, mesoscale, and TaqMan PCR analyses to study the underlying immunostimulatory effects. In vivo depletion of CD8+ T cells and studies with Rag2/Il2Rg double knockout C57BL/6 mice were conducted to investigate the importance of immune cells to the efficacy of MSLN-TTC. RESULTS: MSLN-TTC treatment induced upregulation of DNA sensing pathway transcripts (IL-6, CCL20, CXCL10, and stimulator of interferon genes (STING)-related genes) in vitro as determined by RNASeq analysis. The results, including phospho-STING activation, were confirmed on the protein level. Danger-associated molecular pattern molecules were upregulated in parallel, leading to dendritic cell (DC) activation in vitro. MSLN-TTC showed strong antitumor activity (T:C 0.38, p<0.05) as a single agent in human MSLN-expressing MC38 tumor-bearing immunocompetent mice. Combining MSLN-TTC with anti-PD-L1 further enhanced the efficacy (T:C 0.08, p<0.001) as evidenced by the increased number of tumor-free surviving animals. MSLN-TTC monotherapy caused migration of CD103+ cDC1 DCs and infiltration of CD8+ T cells into tumors, which was enhanced on combination with anti-PD-L1. Intriguingly, CD8+ T-cell depletion decreased antitumor efficacy. CONCLUSIONS: These in vitro and in vivo data on MSLN-TTC demonstrate that the MoA of TTCs involves activation of the immune system. The findings are of relevance for other targeted radiotherapies and may guide clinical combination strategies.
Assuntos
Antígeno B7-H1/antagonistas & inibidores , Imunoconjugados/uso terapêutico , Tório/uso terapêutico , Animais , Perfilação da Expressão Gênica , Imunoconjugados/farmacologia , Imunoterapia , Camundongos , Tório/farmacologia , Transfecção , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Targeted α therapy (TAT) offers the potential for the targeted delivery of potent α-particle-emitting radionuclides that emit high linear energy transfer radiation. This leads to a densely ionizing radiation track over a short path. Localized radiation induces cytotoxic, difficult-to-repair, clustered DNA double-strand breaks (DSBs). To date, radium-223 (223Ra) is the only TAT approved for the treatment of patients with metastatic castration-resistant prostate cancer. Thorium-227 (227Th), the progenitor nuclide of 223Ra, offers promise as a wider-ranging alternative due to the availability of efficient chelators, such as octadentate 3,2-hydroxypyridinone (3,2-HOPO). The 3,2-HOPO chelator can be readily conjugated to a range of targeting moieties, enabling the generation of new targeted thorium-227 conjugates (TTCs). This review provides a comprehensive overview of the advances in the preclinical development of TTCs for hematological cancers, including CD22-positive B cell cancers and CD33-positive leukemia, as well as for solid tumors overexpressing renal cell cancer antigen CD70, membrane-anchored glycoprotein mesothelin in mesothelioma, prostate-specific membrane antigen in prostate cancer, and fibroblast growth factor receptor 2. As the mechanism of action for TTCs is linked to the formation of DSBs, the authors also report data supporting combinations of TTCs with inhibitors of the DNA damage response pathways, including those of the ataxia telangiectasia and Rad3-related protein, and poly-ADP ribose polymerase. Finally, emerging evidence suggests that TTCs induce immunogenic cell death through the release of danger-associated molecular patterns. Based on encouraging preclinical data, clinical studies have been initiated to investigate the safety and tolerability of TTCs in patients with various cancers.
Assuntos
Partículas alfa/uso terapêutico , Neoplasias Hematológicas/radioterapia , Imunoconjugados/uso terapêutico , Compostos Radiofarmacêuticos/uso terapêutico , Tório/uso terapêutico , Alarminas/metabolismo , Quelantes/química , Dano ao DNA/efeitos da radiação , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/imunologia , Neoplasias Hematológicas/patologia , Humanos , Imunoconjugados/química , Morte Celular Imunogênica/efeitos da radiação , Medicina de Precisão/métodos , Piridonas/química , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/farmacologia , Tório/química , Tório/farmacologia , Resultado do TratamentoRESUMO
PURPOSE: Prostate-specific membrane antigen (PSMA) is an attractive target for radionuclide therapy of metastatic castration-resistant prostate cancer (mCRPC). PSMA-targeted alpha therapy (TAT) has shown early signs of activity in patients with prostate cancer refractory to beta radiation. We describe a novel, antibody-based TAT, the PSMA-targeted thorium-227 conjugate PSMA-TTC (BAY 2315497) consisting of the alpha-particle emitter thorium-227 complexed by a 3,2-HOPO chelator covalently linked to a fully human PSMA-targeting antibody. EXPERIMENTAL DESIGN: PSMA-TTC was characterized for affinity, mode of action, and cytotoxic activity in vitro. Biodistribution, pharmacokinetics, and antitumor efficacy were investigated in vivo using cell line and patient-derived xenograft (PDX) models of prostate cancer. RESULTS: PSMA-TTC was selectively internalized into PSMA-positive cells and potently induced DNA damage, cell-cycle arrest, and apoptosis in vitro. Decrease in cell viability was observed dependent on the cellular PSMA expression levels. In vivo, PSMA-TTC showed strong antitumor efficacy with T/C values of 0.01 to 0.31 after a single injection at 300 to 500 kBq/kg in subcutaneous cell line and PDX models, including models resistant to standard-of-care drugs such as enzalutamide. Furthermore, inhibition of both cancer and cancer-induced abnormal bone growth was observed in a model mimicking prostate cancer metastasized to bone. Specific tumor uptake and efficacy were demonstrated using various PSMA-TTC doses and dosing schedules. Induction of DNA double-strand breaks was identified as a key mode of action for PSMA-TTC both in vitro and in vivo. CONCLUSIONS: The strong preclinical antitumor activity of PSMA-TTC supports its clinical evaluation, and a phase I trial is ongoing in mCRPC patients (NCT03724747).
Assuntos
Partículas alfa/uso terapêutico , Antígenos de Superfície/metabolismo , Antineoplásicos Imunológicos/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Glutamato Carboxipeptidase II/metabolismo , Imunoconjugados/farmacocinética , Neoplasias da Próstata/radioterapia , Tório/farmacologia , Animais , Apoptose , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos Nus , Camundongos SCID , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Compostos Radiofarmacêuticos/farmacologia , Distribuição Tecidual , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
UNLABELLED: Combination therapy has improved the quality of life for patients with squamous cell carcinomas of the head and neck (HNSCCs) but has not decisively changed prognosis. Targeted therapies, which enhance accumulation of the drug in the tumor, may be realized using tumor-specific binding peptides. This paper identifies and characterizes an HNSCC affine peptide. METHODS: From a phage library comprising 10(9) different displayed peptides, 1 peptide was enriched after 5 in vitro selection rounds on HNO223 tumor cells. Subsequently, the gained peptide sequence H(2)N-SPRGDLAVLGHKY-CONH(2) (HBP-1) was synthesized as an amide and labeled with (125)I. In vitro studies for binding kinetics and competition were performed with 5 different HNSCC cell lines. Furthermore, the stability of the peptide was evaluated in human serum. The in vivo biodistribution of (131)I-labeled peptide was determined in HNSCC tumor-bearing nude mice. The results were further validated in human HNSCC tumor tissue sections using fluorescence-labeled HBP-1. Competition experiments were performed to determine the binding sequence and validate the target. RESULTS: The HBP-1 motif was enriched in 62% of all phages sequenced. Labeled (125)I-HBP-1 showed binding to 5 different HNSCC cell lines and a maximum binding to HNO97 cells, with 11% of the applied dose per 10(6) cells and an inhibitory concentration of 50% of 38.9 nM. Stability experiments in human serum showed a half-life of 55 min. In 2 different HNSCC tumor xenografts, (131)I-HBP-1 accumulated rapidly, with stable uptake until 45 min after intravenous application. Peptide immunohistochemistry of HNSCC tissue sections exhibited tumor staining by HBP-1, whereas normal tissue remained negative. Sequence mutation and competition experiments revealed that the intrinsic RGD motif in combination with the intrinsic LXXL motif is responsible for the binding ability of HBP1. The RGDLXXL sequence within this peptide is known and indicates that binding occurs via the alpha(v)beta(6) rather than the alpha(v)beta(3) integrin. CONCLUSION: Within the sequence of HBP-1 is a RGDLXXL motif, and most likely it is targeting the alpha(v)beta(6) receptor of the integrin family of cell adhesion receptors. HBP-1 represents a promising lead structure for the development of targeted therapies or diagnostic procedures in patients with HNSCC.
Assuntos
Neoplasias de Cabeça e Pescoço/diagnóstico por imagem , Oligopeptídeos/farmacocinética , Compostos Radiofarmacêuticos/farmacocinética , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Neoplasias de Cabeça e Pescoço/metabolismo , Humanos , Radioisótopos do Iodo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Dados de Sequência Molecular , Transplante de Neoplasias , Biblioteca de Peptídeos , CintilografiaRESUMO
Targeted alpha therapy is an emerging innovative approach for the treatment of advanced cancers, in which targeting agents deliver radionuclides directly to tumors and metastases. The biological effects of α-radiation are still not fully understood - partly due to the lack of sufficiently accurate research methods. The range of α-particles is <100 µm, and therefore, standard in vitro assays may underestimate α-radiation-specific radiation effects. In this report we focus on α-radiation-induced DNA lesions, DNA repair as well as cellular responses to DNA damage. Herein, we used Ra-223 to deliver α-particles to various tumor cells in a Transwell system. We evaluated the time and dose-dependent biological effects of α-radiation on several tumor cell lines by biological endpoints such as clonogenic survival, cell cycle distribution, comet assay, foci analysis for DNA damage, and calculated the absorbed dose by Monte-Carlo simulations. The radiobiological effects of Ra-223 in various tumor cell lines were evaluated using a novel in vitro assay designed to assess α-radiation-mediated effects. The α-radiation induced increasing levels of DNA double-strand breaks (DSBs) as detected by the formation of 53BP1 foci in a time- and dose-dependent manner in tumor cells. Short-term exposure (1-8 h) of different tumor cells to α-radiation was sufficient to double the number of cells in G2/M phase, reduced cell survival to 11-20% and also increased DNA fragmentation measured by tail intensity (from 1.4 to 3.9) dose-dependently. The α-particle component of Ra-223 radiation caused most of the Ra-223 radiation-induced biological effects such as DNA DSBs, cell cycle arrest and micronuclei formation, leading ultimately to cell death. The variable effects of α-radiation onto the different tumor cells demonstrated that tumor cells show diverse sensitivity towards damage caused by α-radiation. If these differences are caused by genetic alterations and if the sensitivity could be modulated by the use of DNA damage repair inhibitors remains a wide field for further investigations.
Assuntos
Morte Celular/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Rádio (Elemento) , Ciclo Celular/efeitos da radiação , Linhagem Celular Tumoral , Relação Dose-Resposta à Radiação , HumanosRESUMO
PURPOSE: The catalytic function of BUB1 is required for chromosome arm resolution and positioning of the chromosomal passenger complex for resolution of spindle attachment errors and plays only a minor role in spindle assembly checkpoint activation. Here, we present the identification and preclinical pharmacologic profile of the first BUB1 kinase inhibitor with good bioavailability. EXPERIMENTAL DESIGN: The Bayer compound library was screened for BUB1 kinase inhibitors and medicinal chemistry efforts to improve target affinity and physicochemical and pharmacokinetic parameters resulting in the identification of BAY 1816032 were performed. BAY 1816032 was characterized for kinase selectivity, inhibition of BUB1 signaling, and inhibition of tumor cell proliferation alone and in combination with taxanes, ATR, and PARP inhibitors. Effects on tumor growth in vivo were evaluated using human triple-negative breast xenograft models. RESULTS: The highly selective compound BAY 1816032 showed long target residence time and induced chromosome mis-segregation upon combination with low concentrations of paclitaxel. It was synergistic or additive in combination with paclitaxel or docetaxel, as well as with ATR or PARP inhibitors in cellular assays. Tumor xenograft studies demonstrated a strong and statistically significant reduction of tumor size and excellent tolerability upon combination of BAY 1816032 with paclitaxel or olaparib as compared with the respective monotherapies. CONCLUSIONS: Our findings suggest clinical proof-of-concept studies evaluating BAY 1816032 in combination with taxanes or PARP inhibitors to enhance their efficacy and potentially overcome resistance.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/genética , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células HeLa , Humanos , Camundongos , Neoplasias/genética , Neoplasias/patologia , Ftalazinas/farmacologia , Piperazinas/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Taxoides/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Despite the recent advances in the treatment of ovarian cancer, it remains an area of high unmet medical need. Epithelial ovarian cancer is associated with high levels of mesothelin expression, and therefore, mesothelin is an attractive candidate target for the treatment of this disease. Herein, we investigated the antitumor efficacy of the mesothelin-targeting antibody-drug conjugate (ADC) anetumab ravtansine as a novel treatment option for ovarian cancer in monotherapy and in combination with the antitumor agents pegylated liposomal doxorubicin (PLD), carboplatin, copanlisib and bevacizumab. Anetumab ravtansine showed potent antitumor activity as a monotherapy in ovarian cancer models with high mesothelin expression. No activity was seen in mesothelin-negative models. The combination of anetumab ravtansine with PLD showed additive anti-proliferative activity in vitro, which translated into improved therapeutic in vivo efficacy in ovarian cancer cell line- and patient-derived xenograft (PDX) models compared to either agents as a monotherapy. The combination of anetumab ravtansine with the PI3Kα/δ inhibitor copanlisib was additive in the OVCAR-3 and OVCAR-8 cell lines in vitro, showing increased apoptosis in response to the combination treatment. In vivo, the combination of anetumab ravtansine with copanlisib resulted in more potent antitumor activity than either of the treatments alone. Likewise, the combination of anetumab ravtansine with carboplatin or bevacizumab showed improved in vivo efficacy in the ST081 and OVCAR-3 models, respectively. All combinations were well-tolerated. Taken together, these data support the development of anetumab ravtansine for ovarian cancer treatment and highlight its suitability for combination therapy with PLD, carboplatin, copanlisib, or bevacizumab.
RESUMO
Compared with follicular lymphoma, high PI3Kα expression was more prevalent in diffuse large B cell lymphoma (DLBCL), although both tumor types expressed substantial PI3Kδ. Simultaneous inhibition of PI3Kα and PI3Kδ dramatically enhanced the anti-tumor profile in ABC-DLBCL models compared with selective inhibition of PI3Kδ, PI3Kα, or BTK. The anti-tumor activity was associated with suppression of p-AKT and a mechanism of blocking nuclear factor-κB activation driven by CD79mut, CARD11mut, TNFAIP3mut, or MYD88mut. Inhibition of PI3Kα/δ resulted in tumor regression in an ibrutinib-resistant CD79BWT/MYD88mut patient-derived ABC-DLBCL model. Furthermore, rebound activation of BTK and AKT was identified as a mechanism limiting CD79Bmut-ABC-DLBCL to show a robust response to PI3K and BTK inhibitor monotherapies. A combination of ibrutinib with the PI3Kα/δ inhibitor copanlisib produced a sustained complete response in vivo in CD79Bmut/MYD88mut ABC-DLBCL models.
Assuntos
Classe I de Fosfatidilinositol 3-Quinases/antagonistas & inibidores , Linfoma Difuso de Grandes Células B/tratamento farmacológico , NF-kappa B/fisiologia , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas c-akt/fisiologia , Receptores de Antígenos de Linfócitos B/fisiologia , Adenina/análogos & derivados , Adulto , Tirosina Quinase da Agamaglobulinemia , Idoso , Animais , Linhagem Celular Tumoral , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Linfoma Difuso de Grandes Células B/mortalidade , Linfoma Difuso de Grandes Células B/patologia , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Piperidinas , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirazóis/farmacologia , Pirimidinas/farmacologia , Quinazolinas/farmacologiaRESUMO
UNLABELLED: (D)-(18)F-fluoromethyltyrosine (d-(18)F-FMT), or BAY 86-9596, is a novel (18)F-labeled tyrosine derivative rapidly transported by the l-amino acid transporter (LAT-1), with a faster blood pool clearance than the corresponding l-isomer. The aim of this study was to demonstrate the feasibility of tumor detection in patients with non-small cell lung cancer (NSCLC) or head and neck squamous cell cancer (HNSCC) compared with inflammatory and physiologic tissues in direct comparison to (18)F-FDG. METHODS: 18 patients with biopsy-proven NSCLC (n = 10) or HNSCC (n = 8) were included in this Institutional Review Board-approved, prospective multicenter study. All patients underwent (18)F-FDG PET/CT scans within 21 d before d-(18)F-FMT PET/CT. For all patients, safety and outcome data were assessed. RESULTS: No adverse reactions were observed related to d-(18)F-FMT. Fifty-two lesions were (18)F-FDG-positive, and 42 of those were malignant (34 histologically proven and 8 with clinical reference). Thirty-two of the 42 malignant lesions were also d-(18)F-FMT-positive, and 10 lesions had no tracer uptake above the level of the blood pool. Overall there were 34 true-positive, 8 true-negative, 10 false-negative, and only 2 false-positive lesions for d-(18)F-FMT, whereas (18)F-FDG was true-positive in 42 lesions, with 10 false-positive and only 2 false-negative, resulting in a lesion-based detection rate for d-(18)F-FMT and (18)F-FDG of 77% and 95%, respectively, with an accuracy of 78% for both tracers. A high d-(18)F-FMT tumor-to-blood pool ratio had a negative correlation with overall survival (P = 0.050), whereas the (18)F-FDG tumor-to-blood pool ratio did not correlate with overall survival. CONCLUSION: d-(18)F-FMT imaging in patients with NSCLC and HNSCC is safe and feasible. The presented preliminary results suggest a lower sensitivity but higher specificity for d-(18)F-FMT over (18)F-FDG, since there is no d-(18)F-FMT uptake in inflammation. This increased specificity may be particularly beneficial in areas with endemic granulomatous disease and may improve clinical management. Further clinical investigations are needed to determine its clinical value and relevance for the prediction of survival prognosis.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/diagnóstico por imagem , Carcinoma de Células Escamosas/diagnóstico por imagem , Radioisótopos de Flúor/química , Neoplasias de Cabeça e Pescoço/diagnóstico por imagem , Neoplasias Pulmonares/diagnóstico por imagem , Tirosina/análogos & derivados , Tirosina/química , Adulto , Idoso , Biópsia , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma de Células Escamosas/diagnóstico , Reações Falso-Positivas , Feminino , Fluordesoxiglucose F18/química , Neoplasias de Cabeça e Pescoço/diagnóstico , Humanos , Inflamação , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/diagnóstico , Masculino , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons , Prognóstico , Estudos Prospectivos , Sensibilidade e Especificidade , Tomografia Computadorizada por Raios XRESUMO
F18 2-Fluoro 2-deoxyglucose (FDG) has been the gold standard in positron emission tomography (PET) oncologic imaging since its introduction into the clinics several years ago. Seeking to complement FDG in the diagnosis of breast cancer using radio labeled fructose based analogs, we investigated the expression of the chief fructose transporter-GLUT 5 in breast cancer cells and human tissues. Our results indicate that GLUT 5 is not over-expressed in breast cancer tissues as assessed by an extensive immunohistochemistry study. RT-PCR studies showed that the GLUT 5 mRNA was present at minimal amounts in breast cancer cell lines. Further knocking down the expression of GLUT 5 in breast cancer cells using RNA interference did not affect the fructose uptake in these cell lines. Taken together these results are consistent with GLUT 5 not being essential for fructose uptake in breast cancer cells and tissues.