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1.
Microorganisms ; 12(3)2024 Feb 27.
Article in English | MEDLINE | ID: mdl-38543531

ABSTRACT

Membranolytic molecules constitute the first line of innate immune defense against pathogenic microorganisms. Plasmodium sporozoites are potentially exposed to these cytotoxic molecules in the hemolymph and salivary glands of mosquitoes, as well as in the skin, blood, and liver of the mammalian host. Here, we show that sporozoites are resistant to bacteriolytic concentration of cecropin B, a cationic amphipathic antimicrobial insect peptide. Intriguingly, anti-tumoral cell-penetrating peptides derived from the anti-apoptotic protein AAC11 killed P. berghei and P. falciparum sporozoites. Using dynamic imaging, we demonstrated that the most cytotoxic peptide, called RT39, did not significantly inhibit the sporozoite motility until the occurrence of a fast permeabilization of the parasite membrane by the peptide. Concomitantly, the cytosolic fluorescent protein constitutively expressed by sporozoites leaked from the treated parasite body while To-Pro 3 and FITC-labeled RT39 internalized, respectively, binding to the nucleic acids and membranes of sporozoites. This led to an increase in the parasite granularity as assessed by flow cytometry. Most permeabilization events started at the parasite's posterior end, resulting in the appearance of a fluorescent dot in the anterior part of sporozoites. Understanding and exploiting the susceptibility of sporozoites and other plasmodial stages to membranolytic molecules might foster strategies to eliminate the parasite and block its transmission.

2.
Biomolecules ; 14(1)2024 Jan 22.
Article in English | MEDLINE | ID: mdl-38275765

ABSTRACT

Apoptosis, or programmed cell death, is a fundamental process that maintains tissue homeostasis, eliminates damaged or infected cells, and plays a crucial role in various biological phenomena. The deregulation of apoptosis is involved in many human diseases, including cancer. One of the emerging players in the intricate regulatory network of apoptosis is apoptosis inhibitor 5 (API5), also called AAC-11 (anti-apoptosis clone 11) or FIF (fibroblast growth factor-2 interacting factor). While it may not have yet the same level of notoriety as some other cancer-associated proteins, API5 has garnered increasing attention in the cancer field in recent years, as elevated API5 levels are often associated with aggressive tumor behavior, resistance to therapy, and poor patient prognosis. This review aims to shed light on the multifaceted functions and regulatory mechanisms of API5 in cell fate decisions as well as its interest as therapeutic target in cancer.


Subject(s)
Apoptosis Regulatory Proteins , Neoplasms , Humans , Apoptosis Regulatory Proteins/metabolism , Nuclear Proteins/metabolism , Apoptosis , Neoplasms/genetics , Cell Differentiation
3.
Cancers (Basel) ; 15(22)2023 Nov 09.
Article in English | MEDLINE | ID: mdl-38001606

ABSTRACT

Breast cancer is the most common cancer in women, the so-called "Triple-Negative Breast Cancer" (TNBC) subtype remaining the most challenging to treat, with low tumor-free survival and poor clinical evolution. Therefore, there is a clear medical need for innovative and more efficient treatment options for TNBC. The aim of the present study was to evaluate the potential therapeutic interest of the association of the tumor-penetrating BR2 peptide with monophosphoester 2-aminoethyl dihydrogen phosphate (2-AEH2P), a monophosphoester involved in cell membrane turnover, in TNBC. For that purpose, viability, migration, proliferative capacity, and gene expression analysis of proteins involved in the control of proliferation and apoptosis were evaluated upon treatment of an array of TNBC cells with the BR2 peptide and 2-AEH2P, either separately or combined. Our data showed that, while possessing limited single-agent activity, the 2-AEH2P+BR2 association promoted significant cytotoxicity in TNBC cells but not in normal cells, with reduced proliferative potential and inhibition of cell migration. Mechanically, the 2-AEH2P+BR2 combination promoted an increase in cells expressing p53 caspase 3 and caspase 8, a reduction in cells expressing tumor progression and metastasis markers such as VEGF and PCNA, as well as a reduction in mitochondrial electrical potential. Our results indicate that the combination of the BR2 peptide with 2-AEH2P+BR2 may represent a promising therapeutic strategy in TNBC with potential use in clinical settings.

4.
Article in English | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: bud-5166

ABSTRACT

Breast cancer is the most common cancer in women, the so-called “Triple-Negative Breast Cancer” (TNBC) subtype remaining the most challenging to treat, with low tumor-free survival and poor clinical evolution. Therefore, there is a clear medical need for innovative and more efficient treatment options for TNBC. The aim of the present study was to evaluate the potential therapeutic interest of the association of the tumor-penetrating BR2 peptide with monophosphoester 2-aminoethyl dihydrogen phosphate (2-AEH2P), a monophosphoester involved in cell membrane turnover, in TNBC. For that purpose, viability, migration, proliferative capacity, and gene expression analysis of proteins involved in the control of proliferation and apoptosis were evaluated upon treatment of an array of TNBC cells with the BR2 peptide and 2-AEH2P, either separately or combined. Our data showed that, while possessing limited single-agent activity, the 2-AEH2P+BR2 association promoted significant cytotoxicity in TNBC cells but not in normal cells, with reduced proliferative potential and inhibition of cell migration. Mechanically, the 2-AEH2P+BR2 combination promoted an increase in cells expressing p53 caspase 3 and caspase 8, a reduction in cells expressing tumor progression and metastasis markers such as VEGF and PCNA, as well as a reduction in mitochondrial electrical potential. Our results indicate that the combination of the BR2 peptide with 2-AEH2P+BR2 may represent a promising therapeutic strategy in TNBC with potential use in clinical settings.

5.
Cells ; 11(19)2022 09 20.
Article in English | MEDLINE | ID: mdl-36230895

ABSTRACT

Sézary syndrome (SS) is an aggressive cutaneous T cell lymphoma with poor prognosis mainly characterized by the expansion of a tumor CD4+ T cell clone in both skin and blood. So far, the development of new therapeutic strategies has been hindered by a lack of reproducible in vivo models closely reflecting patients' clinical features. We developed an SS murine model consisting of the intravenous injection of Sézary patients' PBMC, together with a mixture of interleukins, in NOD-SCID-gamma mice. Thirty-four to fifty days after injection, mice showed skin disorders similar to that observed in patients, with the detection of epidermis thickening and dermal tumor T cell infiltrates. Although experimental variability was observed, Sézary cells could be tracked in the blood stream, confirming that our model could efficiently exhibit both skin and blood involvement. Using this model, we evaluated the therapeutic potential of RT39, a cell-penetrating peptide derived from the survival protein anti-apoptosis clone 11 (AAC-11), that we previously characterized as specifically inducing apoptosis of Sézary patients' malignant clone ex vivo. Systemic administration of RT39 led to cutaneous tumor T cells depletion, demonstrating efficient malignant cells' targeting and a favorable safety profile. These preclinical data confirmed that RT39 might be an innovative therapeutic tool for Sézary syndrome.


Subject(s)
Cell-Penetrating Peptides , Sezary Syndrome , Skin Neoplasms , Animals , Disease Models, Animal , Heterografts , Humans , Interleukins , Leukocytes, Mononuclear/metabolism , Mice , Mice, Inbred NOD , Mice, SCID , Sezary Syndrome/drug therapy , Skin Neoplasms/metabolism
6.
Biomed Pharmacother ; 153: 113398, 2022 Sep.
Article in English | MEDLINE | ID: mdl-36076530

ABSTRACT

Triple-negative breast cancer is the most aggressive subtype of breast cancer, with worse clinical evolution and tumor-free survival, leading to the need to develop new effective therapies for its control. The present study evaluated the action of tumor-penetrating peptide BR2 associated with 2-aminoethyl dihydrogen phosphate (2-AEH2P) on triple-negative breast tumor cells. Cell viability was evaluated by the MTT colorimetric method, mitochondrial electrical potential, and proteins involved in cell proliferation and death control were evaluated by flow cytometry and structural and morphological analysis by confocal microscopy. The results obtained showed that the peptide BR2 and the association 2-AEH2P + BR2 promoted significant cytotoxicity in tumor lines, compared to 2-AEH2P alone. In addition, the association 2-AEH2P + BR2 promoted tumor cells arrest in the G0/G1 phases. Interestingly, both treatments modulated the expression of markers CD44, CD34, CD24, cyclin D1, and Bcl-2, increased p21, Bax, and released cytochrome c. The association proved to be more effective, providing modulation of proteins involved in cell death and senescence, more pronounced cytotoxicity for tumor cells compared to normal cells, and the reduction of markers related to aggressiveness profile, progression, and tumor metastasis.


Subject(s)
Triple Negative Breast Neoplasms , Apoptosis , Cell Line, Tumor , Cell Proliferation , Humans , Phosphates , Triple Negative Breast Neoplasms/pathology
7.
Biomolecules ; 12(7)2022 06 28.
Article in English | MEDLINE | ID: mdl-35883457

ABSTRACT

Since the Nobel Prize award more than twenty years ago for discovering the core apoptotic pathway in C. elegans, apoptosis and various other forms of regulated cell death have been thoroughly characterized by researchers around the world. Although many aspects of regulated cell death still remain to be elucidated in specific cell subtypes and disease conditions, many predicted that research into cell death was inexorably reaching a plateau. However, this was not the case since the last decade saw a multitude of cell death modalities being described, while harnessing their therapeutic potential reached clinical use in certain cases. In line with keeping research into cell death alive, francophone researchers from several institutions in France and Belgium established the French Cell Death Research Network (FCDRN). The research conducted by FCDRN is at the leading edge of emerging topics such as non-apoptotic functions of apoptotic effectors, paracrine effects of cell death, novel canonical and non-canonical mechanisms to induce apoptosis in cell death-resistant cancer cells or regulated forms of necrosis and the associated immunogenic response. Collectively, these various lines of research all emerged from the study of apoptosis and in the next few years will increase the mechanistic knowledge into regulated cell death and how to harness it for therapy.


Subject(s)
Caenorhabditis elegans , Neoplasms , Animals , Apoptosis , Cell Death , Humans , Necrosis
8.
Biomed Pharmacother, v. 153, 113398, set. 2022
Article in English | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: bud-4472

ABSTRACT

Triple-negative breast cancer is the most aggressive subtype of breast cancer, with worse clinical evolution and tumor-free survival, leading to the need to develop new effective therapies for its control. The present study evaluated the action of tumor-penetrating peptide BR2 associated with 2-aminoethyl dihydrogen phosphate (2-AEH2P) on triple-negative breast tumor cells. Cell viability was evaluated by the MTT colorimetric method, mitochondrial electrical potential, and proteins involved in cell proliferation and death control were evaluated by flow cytometry and structural and morphological analysis by confocal microscopy. The results obtained showed that the peptide BR2 and the association 2-AEH2P + BR2 promoted significant cytotoxicity in tumor lines, compared to 2-AEH2P alone. In addition, the association 2-AEH2P + BR2 promoted tumor cells arrest in the G0/G1 phases. Interestingly, both treatments modulated the expression of markers CD44, CD34, CD24, cyclin D1, and Bcl-2, increased p21, Bax, and released cytochrome c. The association proved to be more effective, providing modulation of proteins involved in cell death and senescence, more pronounced cytotoxicity for tumor cells compared to normal cells, and the reduction of markers related to aggressiveness profile, progression, and tumor metastasis.

9.
J Invest Dermatol ; 141(9): 2261-2271.e5, 2021 09.
Article in English | MEDLINE | ID: mdl-33745910

ABSTRACT

Sézary syndrome is an aggressive form of cutaneous T-cell lymphoma characterized by the presence of a malignant CD4+ T-cell clone in both blood and skin. Its pathophysiology is still poorly understood, and the development of targeted therapies is hampered by the absence of specific target proteins. AAC-11 plays important roles in cancer cell progression and survival and thus has been considered as an anticancer therapeutic target. In this study, we show that a peptide called RT39, comprising a portion of AAC-11‒binding site to its protein partners coupled to the penetratin sequence, induces the specific elimination of the malignant T-cell clone both ex vivo on the circulating cells of patients with Sézary syndrome and in vivo in a subcutaneous xenograft mouse model. RT39 acts by direct binding to PAK1 that is overexpressed, located in the plasma membrane, and constitutively activated in Sézary cells, resulting in their selective depletion by membranolysis. Along with the absence of toxicity, our preclinical efficacy evidence suggests that RT39 might represent a promising alternative therapeutic tool for Sézary syndrome because it spares the nonmalignant immune cells and, contrary to antibody-based immunotherapies, does not require the mobilization of the cellular immunity that shows heavy deficiencies at advanced stages of the disease.


Subject(s)
Apoptosis Regulatory Proteins/metabolism , CD4-Positive T-Lymphocytes/immunology , Nuclear Proteins/metabolism , Peptides/metabolism , Sezary Syndrome/therapy , Skin Neoplasms/therapy , p21-Activated Kinases/metabolism , Animals , Apoptosis Regulatory Proteins/genetics , Carcinogenesis , Cell-Penetrating Peptides/metabolism , Clone Cells , Gene Expression Regulation, Neoplastic , Heterografts , Humans , Mice , Nuclear Proteins/genetics , Peptides/genetics , Protein Binding , p21-Activated Kinases/genetics
10.
Cancers (Basel) ; 12(7)2020 Jul 10.
Article in English | MEDLINE | ID: mdl-32664285

ABSTRACT

In this study, we have identified a novel cell-penetrating sequence, termed hAP10, from the C-terminus of the human protein Acinus. hAP10 was able to efficiently enter various normal and cancerous cells, likely through an endocytosis pathway, and to deliver an EGFP cargo to the cell interior. Cell penetration of a peptide, hAP10DR, derived from hAP10 by mutation of an aspartic acid residue to an arginine was dramatically increased. Interestingly, a peptide containing a portion of the heptad leucine repeat region domain of the survival protein AAC-11 (residues 377-399) fused to either hAP10 or hAP10DR was able to induce tumor cells, but not normal cells, death both ex vivo on Sézary patients' circulating cells and to inhibit tumor growth in vivo in a sub-cutaneous xenograft mouse model for the Sézary syndrome. Combined, our results indicate that hAP10 and hAP10DR may represent promising vehicles for the in vitro or in vivo delivery of bioactive cargos, with potential use in clinical settings.

11.
J Virol ; 94(14)2020 07 01.
Article in English | MEDLINE | ID: mdl-32350074

ABSTRACT

HIV-1 successfully establishes long-term infection in its target cells despite viral cytotoxic effects. We have recently shown that cell metabolism is an important factor driving CD4+ T cell susceptibility to HIV-1 and the survival of infected cells. We show here that expression of antiapoptotic clone 11 (AAC-11), an antiapoptotic factor upregulated in many cancers, increased with progressive CD4+ T cell memory differentiation in association with the expression of cell cycle, activation, and metabolism genes and was correlated with susceptibility to HIV-1 infection. Synthetic peptides based on the LZ domain sequence of AAC-11, responsible for its interaction with molecular partners, were previously shown to be cytotoxic to cancer cells. Here, we observed that these peptides also blocked HIV-1 infection by inducing the death of HIV-1-susceptible primary CD4+ T cells across all T cell subsets. The peptides targeted metabolically active cells and had the greatest effect on effector and transitional CD4+ T cell memory subsets. Our results suggest that the AAC-11 survival pathway is potentially involved in the survival of HIV-1-infectible cells and provide proof of principle that some cellular characteristics can be targeted to eliminate the cells offering the best conditions to sustain HIV-1 replication.IMPORTANCE Although antiretroviral treatment efficiently blocks HIV multiplication, it cannot eliminate cells already carrying integrated proviruses. In the search for an HIV cure, the identification of new potential targets to selectively eliminate infected cells is of the outmost importance. We show here that peptides derived from antiapoptotic clone 11 (AAC-11), whose expression levels correlated with susceptibility to HIV-1 infection of CD4+ T cells, induced cytotoxicity in CD4+ T cells showing the highest levels of activation and metabolic activity, conditions known to favor HIV-1 infection. Accordingly, CD4+ T cells that survived the cytotoxic action of the AAC-11 peptides were resistant to HIV-1 replication. Our results identify a new potential molecular pathway to target HIV-1 infection.


Subject(s)
Apoptosis Regulatory Proteins/pharmacology , CD4-Positive T-Lymphocytes/immunology , HIV Infections/immunology , HIV-1/physiology , Immunologic Memory/drug effects , Nuclear Proteins/pharmacology , Peptides/pharmacology , Virus Replication/drug effects , Apoptosis Regulatory Proteins/chemistry , CD4-Positive T-Lymphocytes/pathology , CD4-Positive T-Lymphocytes/virology , Disease Susceptibility , HIV Infections/drug therapy , HIV Infections/pathology , Humans , Nuclear Proteins/chemistry , Peptides/chemistry , Protein Domains , Virus Replication/immunology
12.
Oncoimmunology ; 9(1): 1728871, 2020.
Article in English | MEDLINE | ID: mdl-32158621

ABSTRACT

Despite considerable progress, the treatment of acute leukemia continues to be a challenge for a significant majority of patients. Using a well-characterized preclinical mouse model of acute promyelocytic leukemia (APL), we evaluated here the antileukemic efficacy of RT53, an anticancer peptide with potential immunological properties. Our results indicate that RT53 possesses a direct antileukemic effect, even at a late stage. We also demonstrate that a single injection of a vaccine consisting of leukemic blasts exposed to RT53, which induces the hallmarks of immunogenic cell death, was highly effective in preventing leukemia development in both prophylactic and therapeutic settings. The vaccine comprising RT53-treated APL cells generated long-term antileukemic protection and depletion experiments indicated that CD4 + T cells were of crucial importance for vaccine efficacy. Combined, our results provide the rationale for the exploration of RT53-based therapies for the treatment of acute leukemia.


Subject(s)
Leukemia, Myeloid, Acute , Leukemia, Promyelocytic, Acute , Animals , Humans , Mice , Peptides , T-Lymphocytes
13.
Molecules ; 24(5)2019 Mar 07.
Article in English | MEDLINE | ID: mdl-30866424

ABSTRACT

Cell-penetrating-peptides (CPPs) are small amino-acid sequences characterized by their ability to cross cellular membranes. They can transport various bioactive cargos inside cells including nucleic acids, large proteins, and other chemical compounds. Since 1988, natural and synthetic CPPs have been developed for applications ranging from fundamental to applied biology (cell imaging, gene editing, therapeutics delivery). In recent years, a great number of studies reported the potential of CPPs as carriers for the treatment of various diseases. Apart from a good efficacy due to a rapid and potent delivery, a crucial advantage of CPP-based therapies is the peptides low toxicity compared to most drug carriers. On the other hand, they are quite unstable and lack specificity. Higher specificity can be obtained using a cell-specific CPP to transport the therapeutic agent or using a non-specific CPP to transport a cargo with a targeted activity. CPP-cargo complexes can also be conjugated to another moiety that brings cell- or tissue-specificity. Studies based on all these approaches are showing promising results. Here, we focus on recent advances in the potential usage of CPPs in the context of cancer therapy, with a particular interest in CPP-mediated delivery of anti-tumoral proteins.


Subject(s)
Cell-Penetrating Peptides/chemistry , Drug Carriers/chemistry , Neoplasms/drug therapy , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/therapeutic use , Cell-Penetrating Peptides/adverse effects , Drug Carriers/adverse effects , Humans , Organ Specificity
14.
PLoS One ; 13(8): e0201220, 2018.
Article in English | MEDLINE | ID: mdl-30080874

ABSTRACT

In recent years, immunogenic cell death (ICD) has emerged as a revolutionary concept in the development of novel anticancer therapies. This particular form of cell death is able, through the spatiotemporally defined emission of danger signals by the dying cell, to induce an effective antitumor immune response, allowing the immune system to recognize and eradicate malignant cells. To date, only a restricted number of chemotherapeutics can trigger ICD of cancer cells. We previously reported that a peptide, called RT53, spanning the heptad leucine repeat region of the survival protein AAC-11 fused to a penetrating sequence, selectively induces cancer cell death in vitro and in vivo. Interestingly, B16F10 melanoma cells treated by RT53 were able to mediate anticancer effects in a tumor vaccination model. Stimulated by this observation, we investigated whether RT53 might mediate ICD of cancer cells. Here, we report that RT53 treatment induces all the hallmarks of immunogenic cell death, as defined by the plasma membrane exposure of calreticulin, release of ATP and the exodus of high-mobility group box 1 protein (HMGB1) from dying cancer cells, through a non-regulated, membranolytic mode of action. In a prophylactic mouse model, vaccination with RT53-treated fibrosarcomas prevented tumor growth at the challenge site. Finally, local intratumoral injection of RT53 into established cancers led to tumor regression together with T-cell infiltration and the mounting of an inflammatory response in the treated animals. Collectively, our results strongly suggest that RT53 can induce bona fide ICD of cancer cells and illustrate its potential use as a novel antitumor and immunotherapeutic strategy.


Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis Regulatory Proteins/pharmacology , Cell Death/drug effects , Cell Death/immunology , Cell-Penetrating Peptides/pharmacology , Nuclear Proteins/pharmacology , Peptides/pharmacology , Amino Acid Sequence , Animals , Antineoplastic Agents/chemistry , Apoptosis Regulatory Proteins/chemistry , Cell Line, Tumor , Cell-Penetrating Peptides/chemistry , HL-60 Cells , Humans , Jurkat Cells , Mice , Mice, Inbred C57BL , Nuclear Proteins/chemistry , Peptide Fragments/chemistry , Peptide Fragments/pharmacology , Peptides/chemistry , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/pharmacology , Sarcoma, Experimental/drug therapy , Sarcoma, Experimental/immunology , Sarcoma, Experimental/pathology , Xenograft Model Antitumor Assays
15.
Am J Nucl Med Mol Imaging ; 8(6): 397-406, 2018.
Article in English | MEDLINE | ID: mdl-30697459

ABSTRACT

Bioluminescence imaging (BLI) is widely used for in-vivo monitoring of anti-cancer therapy in mice. [18F]MEL050 is a Positron Emission Tomography (PET) radiotracer which specifically targets melanin. We evaluated planar BLI and [18F]MEL050-PET/CT for therapy (pro-apoptotic peptide LZDP) monitoring in a mouse model of metastatic pigmented melanoma. Twelve B6-albino mice were intravenously injected with B16-F10-luc2 cells on day 0 (D0). The mice received daily from D2 to D17 either an inactive peptide (G1, n=6), or LZDP (G2, n=6). They underwent both BLI and [18F]MEL050-PET/CT imaging on D2, D8 and D17. The number of visible tumors was determined on BLI and PET/CT. [18F]MEL050 uptake in tumor sites was quantified on PET/CT. After sacrifice (D17), the number of black tumors was counted ex-vivo. On D2, BLI and PET/CT images were visually negative. On D8, BLI detected 8 tumor sites in 4/6 mice of G1 vs 5 in 3/6 mice of G2 (NS); PET/CT was visually negative. On D17, BLI detected 17 tumor sites in 5/6 mice of G1 vs 10 in 4/6 mice of G2 (NS). PET/CT detected 18 tumor sites in 4/4 mice of G1 vs 14 in 3/4 mice of G2 (NS). Mean %ID/g of [18F]MEL050 in tumor sites was lower in G2 than in G1 on D17 (P<0.001), whereas bioluminescence intensity was not different between the 2 groups. Ex-vivo examination confirmed lower number of tumors in G2 (P<0.03). In the small number of animals tested in this study, [18F]MEL050-PET/CT and ex-vivo examination could affirm anti-tumoral effect of LZDP, but not BLI.

16.
Nucl Med Biol ; 43(12): 773-780, 2016 Dec.
Article in English | MEDLINE | ID: mdl-27693672

ABSTRACT

INTRODUCTION: Melanoma is a highly malignant cutaneous tumor of melanin-producing cells. MEL050 is a synthetic benzamide-derived molecule that specifically binds to melanin with high affinity. Our aim was to implement a fully automated radiosynthesis of [18F]MEL050, using for the first time, the AllInOne™ synthesis module (Trasis), and to evaluate the potential of [18F]MEL050 for the detection of pigmented melanoma in mice primary subcutaneous tumors and pulmonary metastases, and to compare it with that of [18F]FDG. METHODS: Automated radiosynthesis of [18F]MEL050, including HPLC purification and formulation, were performed on an AllInOne™ synthesis module. [18F]MEL050 was synthesized using a one-step bromine-for-fluorine nucleophilic heteroaromatic substitution. Melanoma models were induced by subcutaneous (primary tumor) or intravenous (pulmonary metastases) injection of B16-F10-luc2 cells in NMRI mice. The maximum percentage of [18F]MEL050 Injected Dose per g of lung tissue (%ID/g Max) was determined on PET images, compared to [18F]FDG and correlated to in vivo bioluminescence imaging. RESULTS: The automated radiosynthesis of [18F]MEL050 required an overall radiosynthesis time of 48min, with a yield of 13-18% (not-decay corrected) and radiochemical purity higher than 99%. [18F]MEL050 PET/CT images were concordant with bioluminescence imaging, showing increased radiotracer uptake in all primary subcutaneous tumors and pulmonary metastases of mice. PET quantification of radiotracers uptake in tumors and muscles demonstrated similar tumor-to-background ratio (TBR) with [18F]MEL050 and [18F]FDG in subcutaneous tumors and higher TBR with [18F]MEL050 than with [18F]FDG in pulmonary metastases. CONCLUSION: We successfully implemented the radiosynthesis of [18F]MEL050 using the AllInOne™ module, including HPLC purification and formulation. In vivo PET/CT validation of [18F]MEL050 was obtained in mouse models of pigmented melanoma, where higher [18F]MEL050 uptake was observed in sub-millimetric pulmonary metastases, comparatively to [18F]FDG.


Subject(s)
Lung Neoplasms/diagnostic imaging , Melanins/metabolism , Melanoma/diagnostic imaging , Niacinamide/analogs & derivatives , Positron Emission Tomography Computed Tomography/methods , Radiochemistry/methods , Animals , Automation , Cell Line, Tumor , Fluorodeoxyglucose F18 , Lung Neoplasms/secondary , Melanoma/metabolism , Melanoma/pathology , Mice , Niacinamide/chemical synthesis , Niacinamide/chemistry , Niacinamide/metabolism , Pigmentation
17.
Cancer Res ; 76(18): 5479-90, 2016 09 15.
Article in English | MEDLINE | ID: mdl-27406828

ABSTRACT

AAC-11 is an antiapoptotic protein that is upregulated in most cancer cells. Increased expression of AAC-11 confers a survival advantage when cancer cells are challenged with various stresses and contributes to tumor invasion and metastases, whereas its deregulation reduces resistance to chemotherapeutic drugs. The antiapoptotic effect of AAC-11 may be clinically relevant as its expression correlates with poor prognosis in several human cancers. Thus, inactivation of AAC-11 might constitute an attractive approach for developing cancer therapeutics. We have developed an AAC-11-derived cell-penetrating peptide, herein named RT53, mimicking in part the heptad leucine repeat region of AAC-11, which functions as a protein-protein interaction module, and that can prevent AAC-11 antiapoptotic properties. In this study, we investigated the anticancer effects of RT53. Our results indicate that RT53 selectively kills cancer cells while sparing normal cells. RT53 selectively inserts into the membranes of cancer cells, where it adopts a punctate distribution and induces membranolysis and release of danger-associated molecular pattern molecules. Systemic administration of RT53 inhibited the growth of preexisting BRAF wild-type and V600E mutant melanoma xenograft tumors through induction of apoptosis and necrosis. Toxicological studies revealed that repetitive injections of RT53 did not produce significant toxicity. Finally, RT53-killed B16F10 cells induced tumor growth inhibition in immunocompetent mice following a rechallenge with live cancer cells of the same type. Collectively, our data demonstrate that RT53 possesses tumor-inhibitory activity with no toxicity in mice, suggesting its potential as a therapeutic agent for the treatment of melanoma and probably other cancers. Cancer Res; 76(18); 5479-90. ©2016 AACR.


Subject(s)
Apoptosis Regulatory Proteins/antagonists & inhibitors , Apoptosis/drug effects , Cell-Penetrating Peptides/pharmacology , Melanoma, Experimental/drug therapy , Nuclear Proteins/antagonists & inhibitors , Animals , Cell Line, Tumor , Humans , Immunoprecipitation , In Situ Nick-End Labeling , Melanoma, Experimental/pathology , Mice , Mice, Nude , Xenograft Model Antitumor Assays
18.
Oncotarget ; 6(34): 36269-77, 2015 Nov 03.
Article in English | MEDLINE | ID: mdl-26474455

ABSTRACT

Little is known about inherited factors associated with the risk of developing chronic myelogenous leukemia (CML). We used a dedicated DNA chip containing 16 561 single nucleotide polymorphisms (SNPs) covering 1 916 candidate genes to analyze 437 CML patients and 1 144 healthy control individuals. Single SNP association analysis identified 139 SNPs that passed multiple comparisons (1% false discovery rate). The HDAC9, AVEN, SEMA3C, IKBKB, GSTA3, RIPK1 and FGF2 genes were each represented by three SNPs, the PSM family by four SNPs and the SLC15A1 gene by six. Haplotype analysis showed that certain combinations of rare alleles of these genes increased the risk of developing CML by more than two or three-fold. A classification tree model identified five SNPs belonging to the genes PSMB10, TNFRSF10D, PSMB2, PPARD and CYP26B1, which were associated with CML predisposition. A CML-risk-allele score was created using these five SNPs. This score was accurate for discriminating CML status (AUC: 0.61, 95%CI: 0.58-0.64). Interestingly, the score was associated with age at diagnosis and the average number of risk alleles was significantly higher in younger patients. The risk-allele score showed the same distribution in the general population (HapMap CEU samples) as in our control individuals and was associated with differential gene expression patterns of two genes (VAPA and TDRKH). In conclusion, we describe haplotypes and a genetic score that are significantly associated with a predisposition to develop CML. The SNPs identified will also serve to drive fundamental research on the putative role of these genes in CML development.


Subject(s)
Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Female , Genetic Predisposition to Disease , Humans , Male , Middle Aged , Polymorphism, Single Nucleotide
19.
Cell Cycle ; 14(8): 1242-51, 2015.
Article in English | MEDLINE | ID: mdl-25695197

ABSTRACT

Beside its central role in the mitochondria-dependent cell death pathway, the apoptotic protease activating factor 1 (Apaf-1) is involved in the DNA damage response through cell-cycle arrest induced by genotoxic stress. This non-apoptotic function requires a nuclear translocation of Apaf-1 during the G1-to-S transition. However, the mechanisms that trigger the nuclear accumulation of Apaf-1 upon DNA damage remain to be investigated. Here we show that the main 4 isoforms of Apaf-1 can undergo nuclear translocation and restore Apaf-1 deficient MEFs cell cycle arrest in the S phase following genotoxic stress through activation of Chk-1. Interestingly, DNA damage-dependent nuclear accumulation of Apaf-1 occurs independently of p53 and the retinoblastoma (pRb) pathway. We demonstrated that Apaf-1 associates with the nucleoporin Nup107 and this association is necessary for Apaf-1 nuclear import. The CED-4 domain of Apaf-1 directly binds to the central domain of Nup107 in an ATR-regulated, phosphorylation-dependent manner. Interestingly, expression of the Apaf-1-interacting domain of Nup107 interfered with Apaf-1 nuclear translocation upon genotoxic stress, resulting in a marked reduction of Chk-1 activation and cell cycle arrest. Thus, our results confirm the crucial role of Apaf-1 nuclear relocalization in mediating cell-cycle arrest induced by genotoxic stress and implicate Nup107 as a critical regulator of the DNA damage-induced intra-S phase checkpoint response.


Subject(s)
Apoptotic Protease-Activating Factor 1/metabolism , Cell Nucleus/metabolism , DNA Damage , Nuclear Pore Complex Proteins/metabolism , Animals , Apoptotic Protease-Activating Factor 1/deficiency , Apoptotic Protease-Activating Factor 1/genetics , Ataxia Telangiectasia Mutated Proteins/antagonists & inhibitors , Ataxia Telangiectasia Mutated Proteins/genetics , Ataxia Telangiectasia Mutated Proteins/metabolism , Cell Line , Checkpoint Kinase 1 , Cisplatin/toxicity , DNA Damage/drug effects , Humans , Mice , Nuclear Pore Complex Proteins/antagonists & inhibitors , Nuclear Pore Complex Proteins/genetics , Phosphorylation/drug effects , Protein Binding , Protein Isoforms/metabolism , Protein Kinases/metabolism , RNA Interference , RNA, Small Interfering/metabolism , Retinoblastoma Protein/metabolism , S Phase Cell Cycle Checkpoints/drug effects , Tumor Suppressor Protein p53/deficiency , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism
20.
Mol Inform ; 33(6-7): 414-437, 2014 Jun.
Article in English | MEDLINE | ID: mdl-25254076

ABSTRACT

[Formula: see text] Fundamental processes in living cells are largely controlled by macromolecular interactions and among them, protein-protein interactions (PPIs) have a critical role while their dysregulations can contribute to the pathogenesis of numerous diseases. Although PPIs were considered as attractive pharmaceutical targets already some years ago, they have been thus far largely unexploited for therapeutic interventions with low molecular weight compounds. Several limiting factors, from technological hurdles to conceptual barriers, are known, which, taken together, explain why research in this area has been relatively slow. However, this last decade, the scientific community has challenged the dogma and became more enthusiastic about the modulation of PPIs with small drug-like molecules. In fact, several success stories were reported both, at the preclinical and clinical stages. In this review article, written for the 2014 International Summer School in Chemoinformatics (Strasbourg, France), we discuss in silico tools (essentially post 2012) and databases that can assist the design of low molecular weight PPI modulators (these tools can be found at www.vls3d.com). We first introduce the field of protein-protein interaction research, discuss key challenges and comment recently reported in silico packages, protocols and databases dedicated to PPIs. Then, we illustrate how in silico methods can be used and combined with experimental work to identify PPI modulators.

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