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1.
Front Immunol ; 13: 936129, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36059502

RESUMO

With the clinical approval of T-cell-dependent immune checkpoint inhibitors for many cancers, therapeutic cancer vaccines have re-emerged as a promising immunotherapy. Cancer vaccines require the addition of immunostimulatory adjuvants to increase vaccine immunogenicity, and increasingly multiple adjuvants are used in combination to bolster further and shape cellular immunity to tumor antigens. However, rigorous quantification of adjuvants' synergistic interactions is challenging due to partial redundancy in costimulatory molecules and cytokine production, leading to the common assumption that combining both adjuvants at the maximum tolerated dose results in optimal efficacy. Herein, we examine this maximum dose assumption and find combinations of these doses are suboptimal. Instead, we optimized dendritic cell activation by extending the Multidimensional Synergy of Combinations (MuSyC) framework that measures the synergy of efficacy and potency between two vaccine adjuvants. Initially, we performed a preliminary in vitro screening of clinically translatable adjuvant receptor targets (TLR, STING, NLL, and RIG-I). We determined that STING agonist (CDN) plus TLR4 agonist (MPL-A) or TLR7/8 agonist (R848) as the best pairwise combinations for dendritic cell activation. In addition, we found that the combination of R848 and CDN is synergistically efficacious and potent in activating both murine and human antigen-presenting cells (APCs) in vitro. These two selected adjuvants were then used to estimate a MuSyC-dose optimized for in vivo T-cell priming using ovalbumin-based peptide vaccines. Finally, using B16 melanoma and MOC1 head and neck cancer models, MuSyC-dose-based adjuvating of cancer vaccines improved the antitumor response, increased tumor-infiltrating lymphocytes, and induced novel myeloid tumor infiltration changes. Further, the MuSyC-dose-based adjuvants approach did not cause additional weight changes or increased plasma cytokine levels compared to CDN alone. Collectively, our findings offer a proof of principle that our MuSyC-extended approach can be used to optimize cancer vaccine formulations for immunotherapy.


Assuntos
Vacinas Anticâncer , Neoplasias , Adjuvantes Imunológicos/farmacologia , Adjuvantes Farmacêuticos/farmacologia , Animais , Vacinas Anticâncer/uso terapêutico , Citocinas , Humanos , Imunoterapia/métodos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/terapia , Eficácia de Vacinas
2.
Proc Natl Acad Sci U S A ; 118(37)2021 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-34508005

RESUMO

Macromolecules such as monoclonal antibodies (mAbs) are likely to experience poor tumor penetration because of their large size, and thus low drug exposure of target cells within a tumor could contribute to suboptimal responses. Given the challenge of inadequate quantitative tools to assess mAb activity within tumors, we hypothesized that measurement of accessible target levels in tumors could elucidate the pharmacologic activity of a mAb and could be used to compare the activity of different mAbs. Using positron emission tomography (PET), we measured the pharmacodynamics of immune checkpoint protein programmed-death ligand 1 (PD-L1) to evaluate pharmacologic effects of mAbs targeting PD-L1 and its receptor programmed cell death protein 1 (PD-1). For PD-L1 quantification, we first developed a small peptide-based fluorine-18-labeled PET imaging agent, [18F]DK222, which provided high-contrast images in preclinical models. We then quantified accessible PD-L1 levels in the tumor bed during treatment with anti-PD-1 and anti-PD-L1 mAbs. Applying mixed-effects models to these data, we found subtle differences in the pharmacodynamic effects of two anti-PD-1 mAbs (nivolumab and pembrolizumab). In contrast, we observed starkly divergent target engagement with anti-PD-L1 mAbs (atezolizumab, avelumab, and durvalumab) that were administered at equivalent doses, correlating with differential effects on tumor growth. Thus, we show that measuring PD-L1 pharmacodynamics informs mechanistic understanding of therapeutic mAbs targeting PD-L1 and PD-1. These findings demonstrate the value of quantifying target pharmacodynamics to elucidate the pharmacologic activity of mAbs, independent of mAb biophysical properties and inclusive of all physiological variables, which are highly heterogeneous within and across tumors and patients.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Antígeno B7-H1/antagonistas & inibidores , Neoplasias da Mama/tratamento farmacológico , Radioisótopos de Flúor/farmacocinética , Fragmentos de Peptídeos/farmacocinética , Tomografia por Emissão de Pósitrons/métodos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Animais , Apoptose , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Compostos Radiofarmacêuticos/farmacocinética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Cancer Immunol Res ; 7(2): 244-256, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30659055

RESUMO

A limitation to antitumor immunity is the dysfunction of T cells in the tumor microenvironment, in part due to upregulation of coinhibitory receptors such as PD-1. Here, we describe that poliovirus receptor-related immunoglobulin domain protein (PVRIG) acts as a coinhibitory receptor in mice. Murine PVRIG interacted weakly with poliovirus receptor (PVR) but bound poliovirus receptor-like 2 (PVRL2) strongly, making the latter its principal ligand. As in humans, murine NK and NKT cells constitutively expressed PVRIG. However, when compared with humans, less PVRIG transcript and surface protein was detected in murine CD8+ T cells ex vivo However, activated CD8+ T cells upregulated PVRIG expression. In the mouse tumor microenvironment, infiltrating CD8+ T cells expressed PVRIG whereas its ligand, PVRL2, was detected predominantly on myeloid cells and tumor cells, mirroring the expression pattern in human tumors. PVRIG-deficient mouse CD8+ T cells mounted a stronger antigen-specific effector response compared with wild-type CD8+ T cells during acute Listeria monocytogenes infection. Furthermore, enhanced CD8+ T-cell effector function inhibited tumor growth in PVRIG-/- mice compared with wild-type mice and PD-L1 blockade conferred a synergistic antitumor response in PVRIG-/- mice. Therapeutic intervention with antagonistic anti-PVRIG in combination with anti-PD-L1 reduced tumor growth. Taken together, our results suggest PVRIG is an inducible checkpoint receptor and that targeting PVRIG-PVRL2 interactions results in increased CD8+ T-cell function and reduced tumor growth.See related article on p. 257.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Neoplasias/imunologia , Neoplasias/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Antígeno B7-H1/antagonistas & inibidores , Biomarcadores , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Humanos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Camundongos , Camundongos Knockout , Neoplasias/patologia , Interferência de RNA , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Carga Tumoral , Microambiente Tumoral/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
5.
PLoS One ; 13(6): e0199130, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29927979

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is expected to be the second leading cause of cancer mortality by 2030. PDAC remains resistant to the majority of systemic chemotherapies. In this paper, we explore if epigenetic sensitization can improve chemotherapy response in PDAC. Multiple PDAC cell lines were tested with serial concentrations of the epigenetic modulators 5-azacitidine (Aza) and guadecitabine (SGI-110). Guadecitabine was effective at inhibiting the expression of DNA Methyltransferase 1 (DNMT1) and in decreasing cell viability at nanomolar concentrations. We also report that guadecitabine has increased efficacy following a delay period or as we reference, a 'rest period'. Sensitization with guadecitabine improved response to the chemotherapeutic agent-Irinotecan- as measured by decreased cell viability and accompanied by an increase in caspase activity. Additional studies are needed to understand the mechanism of action.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/patologia , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Epigênese Genética/efeitos dos fármacos , Irinotecano/farmacologia , Neoplasias Pancreáticas/patologia , Inibidores da Topoisomerase I/farmacologia , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Carcinoma Ductal Pancreático/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Pâncreas/efeitos dos fármacos , Pâncreas/metabolismo , Pâncreas/patologia , Neoplasias Pancreáticas/metabolismo
6.
Oncotarget ; 9(27): 19379-19395, 2018 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-29721210

RESUMO

Leiomyosarcomas are rare mesenchymal neoplasms characterized by a smooth muscle differentiation pattern. Due to the extremely poor prognosis in patients, the development of novel chemotherapeutic regimens remains critically important. In this study, multiple leiomyosarcoma cell lines, SK-UT1, SK-LMS1, and MES-SA were treated with varying doses of the DNA Methyltransferase Inhibitors (DNMTi) 5-azacitidine (Aza), 5-aza-2-deoxycytidine (DAC), and guadecitabine (SGI-110). The effect of these epigenetic modulators was measured using both in-vitro and in-vivo models. Of the three epigenetic modulators, Guadecitabine was the most effective at decreasing cell survival in LMS cell lines. SK-UT1 was found to be the more sensitive to all three epigenetic modulators, while SK-LMS1 and MES-SA were more resistant. The contrast in sensitivity seen was also represented by the increase in apoptosis in Aza and guadecitabine. In parallel with Aza, guadecitabine was observed to also arrest the cell cycle. Treatment with guadecitabine led to a decrease in growth across the spectrum of sensitivity in LMS cell lines, both in a delayed in vitro and in vivo model; in parallel experiments, apoptotic pathways were activated in sensitive and less sensitive lines. Additional studies are required to explore potential therapeutic applications and mechanisms for leiomyosarcoma treatment.

7.
Nanomedicine ; 14(2): 237-246, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29127039

RESUMO

Therapeutic cancer vaccines require adjuvants leading to robust type I interferon and proinflammatory cytokine responses in the tumor microenvironment to induce an anti-tumor response. Cyclic dinucleotides (CDNs), a potent Stimulator of Interferon Receptor (STING) agonist, are currently in phase I trials. However, their efficacy may be limited to micromolar concentrations due to the cytosolic residence of STING in the ER membrane. Here we utilized biodegradable, poly(beta-amino ester) (PBAE) nanoparticles to deliver CDNs to the cytosol leading to robust immune response at >100-fold lower extracellular CDN concentrations in vitro. The leading CDN PBAE nanoparticle formulation induced a log-fold improvement in potency in treating established B16 melanoma tumors in vivo when combined with PD-1 blocking antibody in comparison to free CDN without nanoparticles. This nanoparticle-mediated cytosolic delivery method for STING agonists synergizes with checkpoint inhibitors and has strong potential for enhanced cancer immunotherapy.


Assuntos
Antineoplásicos/administração & dosagem , Imunoterapia , Melanoma Experimental/terapia , Proteínas de Membrana/agonistas , Nanopartículas/administração & dosagem , Nucleotídeos Cíclicos/administração & dosagem , Animais , Antineoplásicos/química , Feminino , Fator Regulador 3 de Interferon/metabolismo , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/química , Nucleotídeos Cíclicos/química , Polímeros/química , Células Tumorais Cultivadas
8.
Head Neck ; 39(6): 1086-1094, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28323387

RESUMO

BACKGROUND: Cyclic dinucleotides (CDNs) are bacterial intracellular messengers that have demonstrated antitumor activity in melanoma and breast tumors, although their role in immunotherapy of head and neck squamous cell cancers (HNSCCs) has not been well investigated. METHODS: We measured primary tumor growth rates, mechanism of antitumor activity, and efficacy of programmed death-L1 blockade combinatorial therapy in SCCFVII tumor-bearing C3H/HeOUJ mice undergoing intratumoral injections with RR-cyclic-di-guanine (synthetic CDG), CDG (natural cyclic-di-guanine), R848 (TLR 7/8 agonist), or phosphate buffered saline (PBS, control). RESULTS: Intratumoral CDN treatment groups showed decreased tumor size and enhanced splenocyte Th1 response when compared to the PBS treatment control group (p < .05). The RR-CDG tumor microenvironment showed upregulated interferon (IFN)-γ+CD8+ and programmed death-L1. Combining programmed death-L1 blocking antibody with RR-CDG induced regression of established tumors. CONCLUSION: This study demonstrates the antitumor effects of CDNs in a HNSCC cell line. These preclinical data strongly support the future clinical development of intratumoral CDN in patients with HNSCC. © 2017 Wiley Periodicals, Inc. Head Neck 39: 1086-1094, 2017.


Assuntos
Anticorpos Antineoplásicos/imunologia , Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Imunoterapia/métodos , Proteínas de Membrana/efeitos dos fármacos , Receptor de Morte Celular Programada 1/administração & dosagem , Animais , Anticorpos Antineoplásicos/efeitos dos fármacos , Carcinoma de Células Escamosas/patologia , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Neoplasias de Cabeça e Pescoço/patologia , Injeções Intralesionais , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C3H , Receptor de Morte Celular Programada 1/imunologia , Distribuição Aleatória , Sensibilidade e Especificidade , Carcinoma de Células Escamosas de Cabeça e Pescoço
9.
Oncotarget ; 8(2): 2053-2068, 2017 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-28008146

RESUMO

Correlative studies from checkpoint inhibitor trials have indicated that better understanding of human leukocytic trafficking into the human tumor microenvironment can expedite the translation of future immune-oncologic agents. In order to directly characterize signaling pathways that can regulate human leukocytic trafficking into the tumor, we have developed a completely autologous xenotransplantation method to reconstitute the human tumor immune microenvironment in vivo. We were able to genetically mark the engrafted CD34+ bone marrow cells as well as the tumor cells, and follow the endogenous leukocytic infiltration into the autologous tumor. To investigate human tumor intrinsic factors that can potentially regulate the immune cells in our system, we silenced STAT3 signaling in the tumor compartment. As expected, STAT3 signaling suppression in the tumor compartment in these autologously reconstituted humanized mice showed increased tumor infiltrating lymphocytes and reduction of arginase-1 in the stroma, which were associated with slower tumor growth rate. We also used this novel system to characterize human myeloid suppressor cells as well as to screen novel agents that can alter endogenous leukocytic infiltration into the tumor. Taken together, we present a valuable method to study individualized human tumor microenvironments in vivo without confounding allogeneic responses.


Assuntos
Linfócitos do Interstício Tumoral/patologia , Transplante de Neoplasias/imunologia , Transplante de Neoplasias/patologia , Neoplasias/imunologia , Neoplasias/patologia , Microambiente Tumoral/imunologia , Animais , Carcinoma de Células Escamosas/imunologia , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Antígeno HLA-A2/genética , Neoplasias de Cabeça e Pescoço/imunologia , Neoplasias de Cabeça e Pescoço/patologia , Xenoenxertos , Humanos , Linfócitos do Interstício Tumoral/fisiologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Carcinoma de Células Escamosas de Cabeça e Pescoço , Transgenes , Transplante Autólogo
10.
PLoS One ; 10(10): e0139657, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26436418

RESUMO

BACKGROUND: Current chemotherapeutic agents based on apoptosis induction are lacking in desired efficacy. Therefore, there is continuous effort to bring about new dimension in control and gradual eradication of cancer by means of ever evolving therapeutic strategies. Various forms of PCD are being increasingly implicated in anti-cancer therapy and the complex interplay among them is vital for the ultimate fate of proliferating cells. We elaborated and illustrated the underlying mechanism of the most potent Andrographolide analogue (AG-4) mediated action that involved the induction of dual modes of cell death-apoptosis and autophagy in human leukemic U937 cells. PRINCIPAL FINDINGS: AG-4 induced cytotoxicity was associated with redox imbalance and apoptosis which involved mitochondrial depolarisation, altered apoptotic protein expressions, activation of the caspase cascade leading to cell cycle arrest. Incubation with caspase inhibitor Z-VAD-fmk or Bax siRNA decreased cytotoxic efficacy of AG-4 emphasising critical roles of caspase and Bax. In addition, AG-4 induced autophagy as evident from LC3-II accumulation, increased Atg protein expressions and autophagosome formation. Pre-treatment with 3-MA or Atg 5 siRNA suppressed the cytotoxic effect of AG-4 implying the pro-death role of autophagy. Furthermore, incubation with Z-VAD-fmk or Bax siRNA subdued AG-4 induced autophagy and pre-treatment with 3-MA or Atg 5 siRNA curbed AG-4 induced apoptosis-implying that apoptosis and autophagy acted as partners in the context of AG-4 mediated action. AG-4 also inhibited PI3K/Akt/mTOR pathway. Inhibition of mTOR or Akt augmented AG-4 induced apoptosis and autophagy signifying its crucial role in its mechanism of action. CONCLUSIONS: Thus, these findings prove the dual ability of AG-4 to induce apoptosis and autophagy which provide a new perspective to it as a potential molecule targeting PCD for future cancer therapeutics.


Assuntos
4-Butirolactona/análogos & derivados , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Diterpenos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , 4-Butirolactona/farmacologia , Caspases/metabolismo , Ativação Enzimática/efeitos dos fármacos , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteínas de Neoplasias/antagonistas & inibidores , Oxirredução , Estresse Oxidativo , RNA Interferente Pequeno/farmacologia , Células U937/efeitos dos fármacos
11.
Planta Med ; 81(12-13): 1029-37, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26085047

RESUMO

Diverse solvent extracts of Artemisia indica leaves originating from the West Bengal region (India) were assessed for the content of artemisinin and characteristic Artemisia polymethoxyflavonoids, namely eupatin (1), casticin (2), chrysoplenetin (3), cirsilineol (4), chrysophenol-D (5), and artemetin (6). HPLC-DAD and HPLC-MS were used to investigate the extracts macerated by solvents of increasing polarity, i.e., petroleum ether, n-hexane, dichloromethane, acetone, MeOH, or EtOH (either 96, 80, or 60 % v/v), and hot water. Artemisinin was absent in all extracts. The acetone and EtOH extracts comprised the highest levels of polymethoxyflavonoids, whereas no flavonoid could be detected in the infusion. None of the remaining extracts contained chryosphenol-D (5) or artemetin (6), while chrysoplenetin (3) was found in all extracts. The essential oil of the plant was also obtained by hydrodistillation and analysed by gas chromatography and gas chromatography-mass spectrometry simultaneously. Of the 92 compounds detected in the oil, camphor (13.0 %) and caryophyllene oxide (10.87 %) were the major components. All solvent extracts and the volatile oil showed in vitro antimalarial activity, plus a potential malaria prophylactic effect by inhibiting at least two recombinant plasmodial fatty acid biosynthesis (PfFAS-II) enzymes. Except for the infusion, all extracts were also active against other parasitic protozoa and displayed low cytotoxicity against mammalian cells. This is the first detailed study investigating both artemisinin and polymethoxyflavonoid content as well as in vitro malaria prophylactic and detailed antiprotozoal potential of A. indica extracts against a panel of protozoan parasites. This is also the first report of antiparasitic activity of the essential oil of the plant.


Assuntos
Antimaláricos/farmacologia , Antiprotozoários/farmacologia , Artemisia/química , Óleos Voláteis/farmacologia , Óleos de Plantas/farmacologia , Animais , Antimaláricos/química , Antimaláricos/isolamento & purificação , Antiprotozoários/química , Antiprotozoários/isolamento & purificação , Artemisininas/química , Artemisininas/isolamento & purificação , Artemisininas/farmacologia , Linhagem Celular , Flavonoides/química , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Índia , Estrutura Molecular , Óleos Voláteis/química , Óleos Voláteis/isolamento & purificação , Folhas de Planta/química , Óleos de Plantas/química , Óleos de Plantas/isolamento & purificação , Ratos
12.
Mol Cancer Ther ; 11(9): 2033-44, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22778153

RESUMO

Ponatinib is a novel tyrosine kinase inhibitor with potent activity against BCR-ABL with mutations, including T315I, and also against fms-like tyrosine kinase 3. We tested interactions between ponatinib at pharmacologically relevant concentrations of 50 to 200 nmol/L and the MDR-associated ATP-binding cassette (ABC) proteins ABCB1, ABCC1, and ABCG2. Ponatinib enhanced uptake of substrates of ABCG2 and ABCB1, but not ABCC1, in cells overexpressing these proteins, with a greater effect on ABCG2 than on ABCB1. Ponatinib potently inhibited [(125)I]-IAAP binding to ABCG2 and ABCB1, indicating binding to their drug substrate sites, with IC(50) values of 0.04 and 0.63 µmol/L, respectively. Ponatinib stimulated ABCG2 ATPase activity in a concentration-dependent manner and stimulated ABCB1 ATPase activity at low concentrations, consistent with it being a substrate of both proteins at pharmacologically relevant concentrations. The ponatinib IC(50) values of BCR-ABL-expressing K562 cells transfected with ABCB1 and ABCG2 were approximately the same as and 2-fold higher than that of K562, respectively, consistent with ponatinib being a substrate of both proteins, but inhibiting its own transport, and resistance was also attenuated to a small degree by ponatinib-induced downregulation of ABCB1 and ABCG2 cell-surface expression on resistant K562 cells. Ponatinib at pharmacologically relevant concentrations produced synergistic cytotoxicity with ABCB1 and ABCG2 substrate chemotherapy drugs and enhanced apoptosis induced by these drugs, including daunorubicin, mitoxantrone, topotecan, and flavopiridol, in cells overexpressing these transport proteins. Combinations of ponatinib and chemotherapy drugs warrant further testing.


Assuntos
Transportadores de Cassetes de Ligação de ATP/antagonistas & inibidores , Antineoplásicos/farmacologia , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Imidazóis/farmacologia , Proteínas de Neoplasias/antagonistas & inibidores , Piridazinas/farmacologia , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/metabolismo , Apoptose/efeitos dos fármacos , Carbocianinas/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Clorofila/análogos & derivados , Clorofila/metabolismo , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Humanos , Concentração Inibidora 50 , Mitoxantrona/farmacologia , Proteínas de Neoplasias/metabolismo , Ligação Proteica , Rodamina 123/metabolismo , Topotecan/farmacologia
13.
Biochimie ; 94(1): 166-83, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22037022

RESUMO

Multidrug resistance (MDR) in cancer, a major obstacle to successful application of cancer chemotherapy, is often characterized by over-expression of multidrug resistance-related proteins such as MRP1, P-gp or elevated glutathione (GSH) level. Efflux of drugs by functional P-gp, MRP1 and elevated GSH level can confer resistance to apoptosis induced by a range of different stimuli. Therefore, it is necessary to develop new cell death inducers with relatively lower toxicity toward non-malignant cells that can overcome MDR by induction of apoptotic or non-apoptotic cell death pathways. Herein we report the synthesis and spectroscopic characterization of a GSH depleting, redox active Schiff's base, viz., potassium-N-(2-hydroxy-3-methoxy-benzaldehyde)-alaninate (PHMBA). Cytotoxic potential of PHMBA has been studied in doxorubicin-resistant and -sensitive T lymphoblastic leukemia cells and Ehrlich ascites carcinoma (EAC) cells. PHMBA kills both the cell types irrespective of their drug-resistance phenotype following apoptotic/necrotic pathways. Moreover, PHMBA-induced cell death is associated with oxidative stress mediated mitochondrial pathway as the H(2)O(2) inhibitor PEG-Catalase abrogated PHMBA-induced apoptosis/necrosis. PHMBA induces anti-tumor activity in both doxorubicin-sensitive and -resistant EAC-tumor-bearing Swiss albino mice. The non-toxicity of PHMBA was also confirmed through cytotoxicity studies on normal cell lines like PBMC, NIH3T3 and Chang Liver. To summarise, our data provide compelling rationale for future clinical use of this redox active Schiff's base in treatment of cancer patients irrespective of their drug-resistance status.


Assuntos
Apoptose/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Mitocôndrias/efeitos dos fármacos , Neoplasias/patologia , Espécies Reativas de Oxigênio/metabolismo , Bases de Schiff/farmacologia , Animais , Cálcio/metabolismo , Calpaína/metabolismo , Caspase 3/metabolismo , Linhagem Celular Tumoral , Citometria de Fluxo , Glutationa/metabolismo , Espectroscopia de Ressonância Magnética , Camundongos , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Necrose , Neoplasias/enzimologia , Neoplasias/metabolismo , Espectrofotometria Ultravioleta , Espectroscopia de Infravermelho com Transformada de Fourier
14.
Phytomedicine ; 18(12): 1056-69, 2011 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-21596544

RESUMO

Diseases caused by insect borne trypanosomatid parasites are significant, yet remain a neglected public health problem. Leishmania, a unicellular protozoan parasite is the causative organism of Leishmaniasis and is transmitted by female phlebotamine sandflies affecting millions of people worldwide. In the wake of resistance to pentavalent antimonial drugs, new therapeutic alternatives are desirable. The plant kingdom has in the past provided several affordable compounds and this review aims to provide an overview of the current status of available leishmanicidal plant derived compounds that are effective singly or in combination with conventional anti-leishmanial drugs, yet are non toxic to mammalian host cells. Furthermore, delineation of the contributory biochemical mechanisms involved in mediating their effect would help develop new chemotherapeutic approaches.


Assuntos
Leishmaniose/tratamento farmacológico , Fitoterapia , Extratos Vegetais/uso terapêutico , Animais , Humanos , Extratos Vegetais/farmacologia
15.
Bioorg Med Chem Lett ; 21(10): 3084-7, 2011 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-21459575

RESUMO

2,2'-Diphenyl-3,3'-diindolylethylene (DPDIE) derivatives 3a-g were regioselectively prepared in one pot from indoles 1a-g in the presence of Lewis acids and were subsequently evaluated for cytotoxic activity against human leukemic cell lines, U937 and K562. The most potent compound 3g exhibited IC(50) of 13.0-17.0 µM.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/toxicidade , Etilenos/síntese química , Etilenos/farmacologia , Indóis/química , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Etilenos/química , Citometria de Fluxo , Humanos , Concentração Inibidora 50 , Estrutura Molecular
16.
J Pharmacol Exp Ther ; 336(1): 206-14, 2011 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-20876229

RESUMO

We have previously demonstrated that resveratrol (Resv)-induced cellular apoptosis occurs after formation of reactive oxygen species (ROS) but the role of GSH has not been well defined. Our experimental data enumerated that Resv treatment (50 µm) induced apoptosis in human leukemic monocyte lymphoma cells, which was preceded by cellular GSH efflux. High concentration of extracellular thiol (GSH, N-acetyl cysteine) and two specific inhibitors of carrier-mediated GSH extrusion, methionine or cystathionine, prevented the process of oxidative burst and cell death. This proved that GSH efflux could be a major molecular switch to modulate Resv-induced ROS generation. Spectrofluorometric data depicted that after 6 h of Resv treatment, ROS generation was evident. Pretreatment of cells with intracellular ROS scavenger (polyethylene glycol-superoxide dismutase and polyethylene glycol-catalase) efficiently reduced ROS generation but failed to prevent intracellular GSH depletion. Thus, it suggested that intracellular GSH depletion was independent of ROS production but dependent on GSH extrusion. Furthermore, to bridge the link between GSH efflux and ROS generation, we carried out confocal microscopy of the localization of Bax protein. Microscopic analysis and small interfering RNA treatment emphasized that cellular GSH efflux triggered Bax translocation to mitochondria, which resulted in the loss of mitochondrial membrane potential, ROS generation, and caspase 3 activation and thus triggered apoptosis.


Assuntos
Apoptose/fisiologia , Glutationa/metabolismo , Líquido Intracelular/metabolismo , Mitocôndrias/metabolismo , Estilbenos/farmacologia , Proteína X Associada a bcl-2/metabolismo , Apoptose/efeitos dos fármacos , Humanos , Líquido Intracelular/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/fisiologia , Mitocôndrias/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Resveratrol , Células U937
17.
Bioorg Med Chem Lett ; 20(23): 6947-50, 2010 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-20974534

RESUMO

A series of analogues of andrographolide, prepared through chemo-selective functionalization at C14 hydroxy, have been evaluated for in vitro cytotoxicities against human leukemic cell lines. Two of the analogues (6a, 9b) exhibited significant potency. Preliminary studies on structure-activity relationship (SAR) revealed that the α-alkylidene-γ-butyrolactone moiety of andrographolide played a major role in the activity profile. The structures of the analogues were established through spectroscopic and analytical data.


Assuntos
Antineoplásicos/química , Diterpenos/síntese química , 4-Butirolactona , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Diterpenos/química , Diterpenos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Leucemia/tratamento farmacológico , Leucemia/patologia , Estrutura Molecular , Análise Espectral , Relação Estrutura-Atividade
18.
Free Radic Res ; 44(11): 1289-95, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20815780

RESUMO

An increasing incidence of unresponsiveness to antimonials in Leishmaniasis has led to identification of plant-derived anti-leishmanial compounds like Artemisinin. Since iron-mediated generation of free radicals sustains the anti-malarial activity of Artemisinin, this study investigated whether similar mechanisms accounted for its activity in Leishmania promastigotes. Artemisinin effectively disrupted the redox potential via an increased generation of free radicals along with a decrease in levels of non-protein thiols. Attenuation of its IC50 by a free radical scavenger N-acetyl L-cysteine and an iron chelator desferoxamine established the pivotal role of free radicals and of the potentiating effect of iron. An enhanced Fluo-4 fluorescence reflected Artemisinin-induced mobilization of intracellular calcium, which triggered apoptosis. However, the absence of any detectable caspase activity indicated that the leishmanicidal activity of Artemisinin is mediated by an iron-dependent generation of reactive intermediates, terminating in a caspase-independent, apoptotic mode of cell death.


Assuntos
Anti-Infecciosos/farmacologia , Apoptose/efeitos dos fármacos , Artemisininas/farmacologia , Ferro/farmacologia , Leishmania donovani/efeitos dos fármacos , Caspases/metabolismo , Radicais Livres/metabolismo , Humanos , Leishmania donovani/metabolismo , Leishmaniose/tratamento farmacológico , Espécies Reativas de Oxigênio/metabolismo , Células U937
19.
J Pharmacol Exp Ther ; 334(2): 381-94, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20484155

RESUMO

Stomach ulceration is a major side effect of most chemopreventive drugs. We have established that although resveratrol is a promising chemopreventive compound, it delays the ulcer healing process. However, its analog hydroxystilbene-1 (HST-1) was devoid of such an ulcerogenic side effect. Consequently, here we tried to explore the chemopreventive efficacy of HST-1 compared with resveratrol in different cancer cell lines and identified the probable signaling pathways responsible for cell death. Our cell viability study established that HST-1, compared with resveratrol, showed better chemopreventive potential in all of the cell lines tested, with U937 and MCF-7 being the cells most affected. Furthermore, in U937 and MCF-7 cell lines, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, cell cycle analysis, and nuclear fragmentation by confocal microscopy established that both HST-1 and resveratrol switched on the apoptotic death cascade to execute cell death. The initiator signal was Fas-independent but synchronized in terms of cytosolic Ca(2+) influx, dissipation of mitochondrial membrane potential, and oxidative burst. It is noteworthy that the executioner signal was cell-specific as in U937 cells; HST-1 and resveratrol treatment induced mitochondrial permealization followed by cardiolipin depletion and cytochrome c release, which eventually activated downstream caspases 9 and 3 to execute the death process. In contrast, in MCF-7 cells the death process was executed in a caspase-independent but calpain-dependent manner as calpain activation induced cleavage of cytosolic alpha-fodrin, stimulated mitochondrial release of apoptotic inducing factor and endonuclease G, and thus harmonized cytosolic and mitochondrial death signals to accomplish apoptosis.


Assuntos
Anticarcinógenos/farmacologia , Apoptose/efeitos dos fármacos , Calpaína/fisiologia , Caspases/fisiologia , Estilbenos/farmacologia , Apoptose/fisiologia , Cálcio/metabolismo , Cardiolipinas/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Citosol/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteínas de Transporte da Membrana Mitocondrial/fisiologia , Poro de Transição de Permeabilidade Mitocondrial , Espécies Reativas de Oxigênio/metabolismo , Explosão Respiratória , Resveratrol , Transdução de Sinais
20.
Int J Antimicrob Agents ; 36(1): 43-9, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20403680

RESUMO

Visceral leishmaniasis (VL), caused by the protozoan Leishmania sp., affects 500000 people annually, with the Indian subcontinent contributing a significant proportion of these cases. Emerging refractoriness to conventional antimony therapy has emphasised the need for safer yet effective antileishmanial drugs. Artemisinin, a widely used antimalarial, demonstrated anti-promastigote activity and the 50% inhibitory concentration (IC(50)) ranged from 100 microM to 120 microM irrespective of Leishmania species studied. Leishmania donovani-infected macrophages demonstrated decreased production of nitrite as well as mRNA expression of inducible nitric oxide synthase, which was normalised by artemisinin, indicating that it exerted both a direct parasiticidal activity as well as inducing a host protective response. Furthermore, in a BALB/c model of VL, orally administered artemisinin (10mg/kg and 25mg/kg body weight) effectively reduced both splenic weight and parasite burden, which was accompanied by a restoration of Th1 cytokines (interferon-gamma and interleukin-2). Taken together, these findings have delineated the therapeutic potential of artemisinin in experimental VL.


Assuntos
Antiprotozoários/administração & dosagem , Artemisininas/administração & dosagem , Leishmaniose Visceral/tratamento farmacológico , Animais , Antiprotozoários/farmacologia , Artemisininas/farmacologia , Modelos Animais de Doenças , Humanos , Concentração Inibidora 50 , Leishmania donovani/efeitos dos fármacos , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Parasitária , Baço/parasitologia , Baço/patologia , Resultado do Tratamento
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