Does native Trypanosoma cruzi calreticulin mediate growth inhibition of a mammary tumor during infection?

Abstract

BackgroundFor several decades now an antagonism between Trypanosoma cruzi infection and tumor development has been detected. The molecular basis of this phenomenon remained basically unknown until our proposal that T. cruzi Calreticulin (TcCRT), an endoplasmic reticulum-resident chaperone, translocated-externalized by the parasite, may mediate at least an important part of this effect. Thus, recombinant TcCRT (rTcCRT) has important in vivo antiangiogenic and antitumor activities. However, the relevant question whether the in vivo antitumor effect of T. cruzi infection is indeed mediated by the native chaperone (nTcCRT), remains open. Herein, by using specific modified anti-rTcCRT antibodies (Abs), we have neutralized the antitumor activity of T. cruzi infection and extracts thereof, thus identifying nTcCRT as a valid mediator of this effect.MethodsPolyclonal anti-rTcCRT F(ab’)2 Ab fragments were used to reverse the capacity of rTcCRT to inhibit EAhy926 endothelial cell (EC) proliferation, as detected by BrdU uptake. Using these F(ab’)2 fragments, we also challenged the capacity of nTcCRT, during T. cruzi infection, to inhibit the growth of an aggressive mammary adenocarcinoma cell line (TA3-MTXR) in mice. Moreover, we determined the capacity of anti-rTcCRT Abs to reverse the antitumor effect of an epimastigote extract (EE). Finally, the effects of these treatments on tumor histology were evaluated.ResultsThe rTcCRT capacity to inhibit ECs proliferation was reversed by anti-rTcCRT F(ab’)2 Ab fragments, thus defining them as valid probes to interfere in vivo with this important TcCRT function. Consequently, during infection, these Ab fragments also reversed the in vivo experimental mammary tumor growth. Moreover, anti-rTcCRT Abs also neutralized the antitumor effect of an EE, again identifying the chaperone protein as an important mediator of this anti mammary tumor effect. Finally, as determined by conventional histological parameters, in infected animals and in those treated with EE, less invasive tumors were observed while, as expected, treatment with F(ab’)2 Ab fragments increased malignancy.ConclusionWe have identified translocated/externalized nTcCRT as responsible for at least an important part of the anti mammary tumor effect of the chaperone observed during experimental infections with T. cruzi.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2764-5) contains supplementary material, which is available to authorized users.

Trypanosoma cruzi Evades the Complement System as an Efficient Strategy to Survive in the Mammalian Host: The Specific Roles of Host/Parasite Molecules and Trypanosoma cruzi Calreticulin

“…In parallel, these animals were treated with anti-TcCRT antibodies. As expected, these antibodies, but not their pre-immune counterparts, neutralized the anti-tumor effect of the infection, indicating that native endogenous TcCRT, in the context of the parasite, is responsible for at least an important part of the anti-tumor effect of T. cruzi infection ( Abello-Caceres et al, 2016 ). Thus, our model proposes that, native TcCRT (secreted or on the parasite) mediates fundamental alterations in the tumor cell microenvironment leading to an adaptive immune response, with significant anti-tumor consequences.…”

“…Additionally, this TcCRT/C1q-mediated infectivity correlates with a significant increase in TcCRT mRNA levels in the early infection steps ( Ramirez et al, 2011b ). Unlike the infective forms, epimastigotes are highly sensitive to the complement activation, most likely due, at least in part, to the marginal levels of TcCRT expressed on their surfaces ( Ferreira et al, 2004 ; Abello-Caceres et al, 2016 ). However, epimastigotes bind exogenously added TcCRT and are internalized by fibroblasts in a C1q-dependent manner.…”

“…Our laboratory has contributed to elucidate the role of TcCRT as a mediator of at least an important part of this effect. TcCRT is anti-angiogenic in vitro ( Lopez et al, 2010 ), ex vivo ( Toledo et al, 2010 ) and in vivo ( Lopez et al, 2010 ; Aguilar-Guzman et al, 2014 ; Abello-Caceres et al, 2016 ), inhibiting the growth of mammary adenocarcinoma and melanoma in murine models ( Molina et al, 2005 ; Lopez et al, 2010 ; Toledo et al, 2010 ; Aguilar-Guzman et al, 2014 ). A recombinant TcCRT and its specific N-terminal domain (aa 120-180) inhibit capillary growth ex vivo in Rattus rattus aortic rings, morphogenesis, proliferation and chemotaxis in human umbilical cord ECs (HUVEC) ( Lopez et al, 2010 ) and an in vivo angiogenesis model in Gallus gallus chorioallantoic membrane assay ( Molina et al, 2005 ).…”

“…However, this internalization is susceptible to be inhibited by Fucoidan, a ligand for SR ( Lopez et al, 2010 ), indicating SR as an additional possible receptor to TcCRT on ECs. In addition, the interaction of TcCRT and ECs can be inhibited by anti-TcCRT antibodies or Fucoidan, thus strengthening the participation of SR in this interaction ( Abello-Caceres et al, 2016 ). These antibodies also revert the TcCRT effect on EC proliferation ( Abello-Caceres et al, 2016 ).…”

“…In addition, the interaction of TcCRT and ECs can be inhibited by anti-TcCRT antibodies or Fucoidan, thus strengthening the participation of SR in this interaction ( Abello-Caceres et al, 2016 ). These antibodies also revert the TcCRT effect on EC proliferation ( Abello-Caceres et al, 2016 ).…”