Angiotensin II-derived constrained peptides with antiplasmodial activity and suppressed vasoconstriction

Angiotensin II (Ang II) is a natural mammalian hormone that has been described to exhibit antiplasmodial activity therefore constituting a promising alternative for the treatment of malaria. Despite its promise, the development of Ang II as an antimalarial is limited by its potent induction of vasoconstriction and its rapid degradation within minutes. Here, we used peptide design to perform targeted chemical modifications to Ang II to generate conformationally restricted (disulfide-crosslinked) peptide derivatives with suppressed vasoconstrictor activity and increased stability. Designed constrained peptides were synthesized chemically and then tested for antiplasmodial activity. Two lead constrained peptides were identified (i.e., peptides 1 and 2), each composed of 10 amino acid residues. These peptides exhibited very promising activity in both our Plasmodium gallinaceum ( > 80%) and Plasmodium falciparum ( > 40%) models, an activity that was equivalent to that of Ang II, and led to complete suppression of vasoconstriction. In addition, peptide 5 exhibited selective activity towards the pre-erythrocytic stage (98% of activity against P. gallinaceum), thus suggesting that it may be possible to design peptides that target specific stages of the malaria life cycle. The Ang II derived stable scaffolds presented here may provide the basis for development of a new generation of peptide-based drugs for the treatment of malaria

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Sintetic peptides derivates of human leptin

A leptina, um hormonio proteico secretado pelo tecido adiposo, e responsavel pela sinalizacao do estado nutricional do individuo para o sistema nervoso central e, em parte , pela regulacao do metabolismo energetico, alem de estar envolvida em varios processos fisiologicos. O receptor da leptina (Ob-R) e um membro do tipo I da superfamilia das citoquinas, largamente expresso em diferentes tecidos, porem mais abundantemente no hipotalamo. Em uma tentativa de identificar as regioes da molecula de leptina responsaveis pela sua atividade, e com base na estrutura tridimensional da mesma, sintetizamos e testamos seis fragmentos peptidicos: Ac-hLEP2-26-NH2 (I), Ac-hLEP27-50-NH2(II),Ac-hLEP51_67-NH2(III),Ac-hLEP71_94-NH2(IV), Ac-[Ser96]-hLEP95_119-NH2 (V), AchLEP120-143-NH2 (VI), e seus respectivos dimeros (excecao feita ao peptideo II) obtidos a partir de pontes dissulfeto. As sinteses foram realizadas pela metodologia de fase solida e as purificacoes, por HPLC de fase reversa. A caracterizacao foi feita por LC/ESI-MS, HPLC, CE e AAA. Os fragmentos foram testados, comparativamente com a leptina integra, quanto: a habilidade de induzirem a expressao de Fos nos nucleos hipotalamicos de rato, apos administracao intravenosa; ao efeito causado no peso, na inGestão alimentar e na glicemia de camundongos ob/ob, apos administracoes (ip) diarias; e a atividade funcional (in vitro) em celulas que expressam o receptor da leptina, utilizando-se um microfisiometro cytosensor. O peptideo [D-Leu4]-OB3 e seu analogo da sequencia humana tambem foram testados. Os resultados obtidos com esses ensaios indicaram que: 1- as diferencas entre as sequencias primarias das leptina humana e de camundongo nao sao suficientes para alterar a atividade dessa proteina; 2- os fragmentos de leptina nao foram capazes de induzir a expressao de Fos nos nucleos hipotalamicos (provavelmente, devido a uma possivel degradacao dos mesmos), nem de transpor a a s 0 e oi a r u o i barreira hemoencefalica; 3- os fragmentos IV e V sao reconhecidos pelo receptor leptina (presentes nas celulas HP-75) e que dois residuos de aminoacidos sao important para a interacao com o receptor da leptina, Ser e Leu. Esses aminoacidos estao presentes, respectivamente, nas posicoes 77 e 80 do fragmento IV e nas posicoes 98 do fragmento V; 4- a presenca das pontes dissulfeto nos peptideos dimerizados nao suficiente para induzir a conformacao nativa desses fragmentos, nem para aumentar s atividade biologica. Embora os fragmentos peptidicos necessitem de um ma refinamento, podem vir a ser uteis no desenvolvimento de drogas utilizadas no controle no tratamento da obesidade humana e/ou veterinaria, por mimetizar a acao da leptina sistema nervoso centralBV UNIFESP: Teses e dissertaçõe

Strutura-activity relation of angiotensin II ciclic analogs

Nos últimos anos, foram feitos muitos esforços na tentativa de se estabelecer a conformação ativa da angiotensina II, utilizando vários métodos físico-químicos, teóricos e espectroscópicos. A utilização de análogos de conformação restrita, especialmente por ciclização, via pontes de dissulfeto ou de lactama tem se mostrado extremamente útil. Optamos pela utilização da restrição conformacional via lactama, onde procuramos explorar: 1) Em que posição da molécula a restrição de conformação deve ser introduzida; 2) Qual deve ser a quiralidade dos aminoácidos que formam os pares para a ciclização; 3) Qual deve ser o tamanho do ciclo e 4) otimizar os efeitos dos análogos descritos na literatura. Para tanto, sintetizamos e testamos uma série limitada de análogos monocíclicos da AII, utilizando como padrão de ciclização, Asp-X-Lys para pontes do tipo i-(i+2) e Asp-X-Y-Lys para pontes do tipo i-(i+3) ao longo de toda a molécula da angiotensina II. Em todos os peptídeos estudados os grupos funcionais principais para atividade da AII (i.e. as cadeia laterais dos resíduos de Tyr, His e Phe e o grupo carboxílico C-terminal), foram preservados. Procuramos também obter informações a respeito do efeito causado pela adição de dois resíduos de aminoácidos na expressão da atividade biológica da AII, através da comparação dos resultados entre os análogos cíclicos e os lineares correspondentes. Os peptídeos foram sintetizados pelo método da fase sólida e ciclizados ainda ligados a resina. A eficiência da reação de ciclização foi estudada com diferentes ativadores (BOP, HATU, TBTU, DIC/HOBt e HBTU), solventes (DMF, NMP, DMSO, 2O por cento DMSO/NMP e 25 por cento DMSO/toluene) e temperaturas (27, 45 e 60ºC). Os peptídeos foram então crivados e desprotegidos em HF, extraídos e liofilizados. Os mesmos foram então purificados em HPLC utilizando-se solventes com os pares iônicos TFA ou TEAP. Os análogos foram então caracterizados por HPLC e CZE, onde apresentaram purezas superiores a 95 por cento. Além disso, ao serem analisados por espectrometria de massa e análise de aminoácidos, apresentaram resultados semelhantes aos teóricos esperado. Os ensaios biológicos foram realizados em três preparações: íleo isolado de cobaia, útero de rata e na pressão sangüínea de rato anestesiados. As atividades agonísticas e antagonísticas dos análogos foram muito baixas ou inexistentes, com exceção dos análagos, ciclo(0-1ª)[Aspº. Endo-...(au).BV UNIFESP: Teses e dissertaçõe

Antiproliferative Modulation and Pro-Apoptotic Effect of BR2 Tumor-Penetrating Peptide Formulation 2-Aminoethyl Dihydrogen Phosphate in Triple-Negative Breast Cancer

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

The Importance of Ring Size and Position for the Antiplasmodial Activity of Angiotensin II Restricted Analogs

Malaria is caused by the protozoa Plasmodium and is responsible for approximately one million deaths annually. the antimalarial effects of angiotensin II and its analogs against Plasmodium gallinaceum and falciparum have recently been reported. Here, 12 angiotensin II restricted analogs that contain i - (i + 2), i - (i + 3) and i - (i + 4) lactam bridges were synthesized to analyze their effect on antiplasmodial activity. To accomplish this, peptides containing two amino acid residues (aspartic or glutamic acids and lysine or ornithine), were synthesized by the t-Boc solid phase method, purified by liquid chromatography, and characterized by mass spectrometry, and conformational studies were performed by circular dichroism. the results indicate that some of the analogs had anti-plasmodium activity similar to angiotensin II (88 % activity). Among those, eight compounds exhibited high activity (> 70 %), measured by fluorescence microscopy. the analogs with smaller lactam rings and an aspartic acid residue as the bridgehead element had lower levels of lytic activity. the results obtained with the new restricted analogs showed that the insertion position (near the N-terminus), the ring size, and the number of residues between the rings are as important as the components of lactam bridge, regardless of their chirality. the circular dichroism studies suggest that the active analogs, and native angiotensin II, adopt a beta-fold conformation in different solutions. in conclusion, this approach provides insight for understanding the effects of restricting the ring size and position on the bioactivity of angiotensin II and provides a new direction for the design of potential chemotherapeutic agents.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Univ Fed ABC, Ctr Ciencias Nat & Humanas, BR-09210170 Santo Andre, SP, BrazilUniversidade Federal de São Paulo, Dept Biofis, São Paulo, BrazilUniversidade Federal de São Paulo, Inst Ciencias Biomed 2, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Biofis, São Paulo, BrazilUniversidade Federal de São Paulo, Inst Ciencias Biomed 2, São Paulo, BrazilFAPESP: 2011/10823-9FAPESP: 2011/15083-3FAPESP: 2011/11448-2Web of Scienc

Anti-Plasmodium Activity of Angiotensin II and Related Synthetic Peptides

Plasmodium species are the causative agents of malaria, the most devastating insect-borne parasite of human populations. Finding and developing new drugs for malaria treatment and prevention is the goal of much research. Angiotensins I and II (ang I and ang II) and six synthetic related peptides designated Vaniceres 1-6 (VC1-VC6) were assayed in vivo and in vitro for their effects on the development of the avian parasite, Plasmodium gallinaceum. Ang II and VC5 injected into the thoraces of the insects reduced mean intensities of infection in the mosquito salivary glands by 88 % and 76%, respectively. Although the mechanism(s) of action is not completely understood, we have demonstrated that these peptides disrupt selectively the P.gallinaceum cell membrane. Additionally, incubation in vitro of sporozoites with VC5 reduced the infectivity of the parasites to their vertebrate host. VC5 has no observable agonist effects on vertebrates, and this makes it a promising dru