Banca de DEFESA: LUANA CARNEIRO PALMA GONCALVES

Uma banca de DEFESA de DOUTORADO foi cadastrada pelo programa.
DISCENTE : LUANA CARNEIRO PALMA GONCALVES
DATA : 30/06/2021
HORA: 13:30
LOCAL: IGM sala virtual 5
TÍTULO:

Investigation of mechanisms involved in the leishmanicidal action of Hsp90 inhibitors

PALAVRAS-CHAVES:

Leishmania, Hsp90, 17-DMAG

PÁGINAS: 124
GRANDE ÁREA: Ciências Biológicas
ÁREA: Biologia Geral
RESUMO:

Leishmaniasis is an infectious disease caused by protozoan parasites of genus
Leishmania. Regardless of the clinical manifestation of the disease, conventional
treatment is mainly based on the use of pentavalent antimonials and amphotericin B.
However, these drugs exhibit high toxicity, severe side effects and prolonged
administration. Thus, the search for more effective and less toxic compounds for the
treatment of leishmaniasis is urgent. In this context, Hsp90 inhibitors have been
studied for the treatment of leishmaniasis, including inhibitors of the geldanamycin
(GA) family. Studies have shown that 17-N-allyl amino 17-demethoxy geldanamycin
(17-AAG) was able to reduce by 90% the percentage of macrophages infected by L.
amazonensis, as well as the number of intracellular parasites. This effect occurred at
doses that were not toxic to the macrophage, host cell for Leishmania spp. Despite
evidence of the leishmanicidal effect of these inhibitors, little is known about how they
act during Leishmania infection. Thus, to develop an ideal therapeutic scheme for
leishmaniasis based on Hsp90 inhibitors, understanding the mechanisms involved in
the leishmanicidal action of these compounds is essential. Thus, the aim of the
present work was to identify possible mechanisms of action of Hsp90 inhibitors,
including GA, 17-AAG and 17-dimethylaminoethylamino-17-demethoxy
geldanamycin (17-DMAG), on the Leishmania amazonensis death. In the present
thesis we focused on the water soluble compound 17-DMAG. Our hypothesis is that
Hsp90 inhibitors would have a double effect, a direct effect on Leishmania as on the
host cell, modulating the innate immune response, facilitating the intracellular death
of L. amazonensis. Thus, the great efficacy of these compounds in killing Leishmania
parasites in non-toxic concentrations to macrophages could be related to a greater
affinity of the inhibitors of the GA family inhibitors for the Hsp90 of the parasite
compared to their affinity for the Hsp90 of the host cell. This higher affinity would be
the result of possible structural and functional differences between the Hsp90
orthologues, Hsp83 of Leishmania and Hsp90 of the host cell. Furthermore, we
hypothesized that inhibitors of the GA family could contribute to parasite death
through inflammasome activation. To evaluate the direct effect of the inhibitor, we
initially evaluated the parasite and host cell toxicity to inhibitors, GA ,17-AAG and 17-
DMAG, and we observed that L. amazonensis was more sensitive to these Hsp90
inhibitors than the host cell. Then, using an in silico approach, we evaluated the
interactions present between GA, 17-AAG and 17-DMAG and L. amazonensis Hsp83
or human Hsp90. Data showed a predominance of hydrogen bonds mediated by
water molecules in the interaction between the inhibitors and human Hsp90, while
direct hydrogen bonds were observed between the inhibitors and Hsp83 of L.
amazonenis. Then, from docking analysis, it was observed that GA, 17-AAG and 17-
DMAG tend to bind more strongly to L. amazonensis Hsp83 than to human Hsp90,
which may be related to direct hydrogen bonds present between inhibitors and L.
amazonensis Hsp83. Subsequently, the effect of the Hsp90 inhibitor, 17-DMAG, on
the modulation of the host cell's immune response, specifically on the activation of
the inflammasome, was evaluated. We found that infected cells treated with 17-
DMAG produced greater levels of IL-1 when compared to untreated infected cells,
suggesting that inflammasome has been activated in response to Hsp90 inhibitor
treatment. However, 17-DMAG-induced parasite death was not altered in cells
treated with caspase 1 inhibitor in mouse macrophages and in C57BL6 NLRP3-/-
mouse macrophages, suggesting that the NLRP3 inflammasome has no influence on
the leishmanicidal effect of this Hsp90 inhibitor. In conclusion, we presented
evidence that 17-DMAG has a direct effect on the death of Leishmania, which can be
explained by the greater tendency of binding between the inhibitor and Hsp83 of the
parasite than human Hsp90. Studies need to be conducted in the future to clarify
whether IL-1 production in 17-DMAG-treated macrophages influences the death of
L. amazonensis in infected and 17-DMAG-treated macrophages.

MEMBROS DA BANCA:
Interna - 024.952.207-11 - CAMILA INDIANI DE OLIVEIRA - USP
Interno - 249.563.328-58 - LEONARDO PAIVA FARIAS - USP
Presidente - 466.269.055-20 - PATRICIA SAMPAIO TAVARES VERAS - Fiocruz - RJ
Externo à Instituição - VINICIUS PINTO COSTA ROCHA
Externo à Instituição - WANDER ROGÉRIO PAVANELLI - UEL