**3. Conclusion**

*Vector-Borne Diseases - Recent Developments in Epidemiology and Control*

reduction in macrophage infection [64].

increased kemptide phosphorylation [64]. These results suggest that cAMP has some direct role in the activation of PKA during transformation in *Leishmania*. Treatment of promastigotes with PKA activators also resulted in growth arrest in the parasite [64]. Parasite survival in the peritoneal macrophages of Balb/c mice was examined using PKA-inhibitor treated parasites and there was a significant

In spite of the discovery of the role played by adenylate cyclases and phosphodiesterases in cAMP homeostasis of *Leishmania*, existence of no specific cAMPbinding effector molecule was known. Bhattacharya et al. [65], in their studies, have identified a regulatory subunit of cAMP-dependent protein kinase (Ldpkar1) in *L. donovani* which was found to be homologous to class I cAMP-dependent protein kinase regulatory subunit of mammals. Studies proved beyond doubt that this regulatory subunit interact with both the catalytic subunits of PKA, thus inhibiting PKA activity. When co-immunoprecipitation assay was performed for both normal and Sp-8-Br-cAMP-pretreated cells, much weaker signal was detected for treated cells as compared to normal cells suggesting Sp-8-Br-cAMP-mediated activation of PKA. Moreover, when activity was analyzed in LdPKAR1-LdPKAC1 and LdPKAR1-LdPKAC2 immunoprecipitated complexes in the presence or absence

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**Figure 6.**

*signaling cascade.*

*cAMP-dependent PKA activity in* Leishmania*: cAMP level increases on phagolysosome condition exposure and cAMP binds with the regulatory subunit of PKA enabling its dissociation from the catalytic subunit rendering the catalytic subunit active. cAMP associated regulatory subunit promotes metacyclogenesis and induces autophagy whereas activated catalytic subunit phosphorylates other proteins downstream to this* 

To conclude we can say that the leading researches in the recent past has enriched our knowledge on the importance of cAMP signaling in kinetoplastid parasites like *Leishmania* and their association with parasite infectivity. These findings provide insight on the functioning of different enzymes associated with cAMP metabolism (**Figure 7**). These studies point toward the fact that modulation of cAMP level in the parasite might be one of the mechanisms to control leishmaniasis and the molecules associated with the same might be tested as potent drug targets against the disease.

Presently, PDE inhibitors are potent drug targets against various human diseases. Study of human PDEs in cAMP signaling pathway has revealed their druggability in various human pathologies leading to various marketed drugs [67]. Moreover, there is a similarity between human and protozoan enzymes and in addition, the availability of human PDE inhibitors as therapeutics has thrown some light on the discovery of some specific protozoan PDE inhibitors as potential drug targets [68]. In kinetoplastid parasites like *Trypanosoma*, PDE inhibitors are being screened as potential drug targets

#### **Figure 7.**

*An overview of cAMP signaling in* Leishmania *during stress condition where receptor adenylate cyclase, PDEA, PDED, PKA the effector molecule of cAMP and acidocalcisomal pyrophosphatases play the major roles in the maintenance of cAMP homeostasis.*

and pharmacological validation of using PDEs as novel drug targets for diseases caused by the kinetoplastid parasites. This study for the first time explores the possibility of using human PDE inhibitors as the starting framework for the design of *Leishmania*selective inhibitors. This proposal is supported by the inhibitory effects of some human PDE inhibitors observed on *T. cruzi* PDEC, e.g., etazolate inhibits human PDE4 and TcrPDEC with the IC50 values of 2 and 0.7 nM [69]. Among the PDE inhibitors used in this study etazolate along with rolipram showed maximum anti-proliferative activity against *Leishmania* parasite with least cytotoxic effect on macrophage cells cultured *in vitro*. Further it was observed that both of them significantly affected the G1 cell cycle arrest and mitochondrial membrane potential of the parasite, therefore we assessed *in vitro* for their ability to clear parasite load within the macrophage cells. Etazolate was found to be more effective in clearing the parasite load when the macrophage cells were pretreated with it compared to rolipram. Etazolate belongs to pyrozopolidine class compound which shows PDE4 enzyme inhibitory activity [70]. Preclinical studies as well as pharmacokinetic and safety profiles in Phase I and Phase IIa of clinical studies revealed that etazolate is a well-established drug of choice with no major side effects reported [70]. Etazolate produced antidepressant like effects in animal models of depression and at the same time it could be used in the treatment of Alzheimer's disease [71]. If through experimentation a minimal dose of etazolate could be determined then etazolate could itself be used as an anti-leishmanial drug. However, if it is not possible to determine the dose concentration which would specifically inhibit parasite PDE, then one can make use of the significant advances made in medicinal chemistry to design compounds which could specifically inhibit parasite PDE but not that of the host. This compound if developed, could act as a potential anti-leishmanial agent in future.

On the other hand, several known PDE inhibitors were tested against *Plasmodium* PDEα, and zaprinast, a known selective inhibitor of human PDE5 which is specific

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provided the original work is properly cited.

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

1 Department of Zoology, Cell and Molecular Biology Laboratory, University of

2 Infectious Diseases and Immunology Division, CSIR-Indian Institute of Chemical

and Pijush K. Das2

\*

*Role of cAMP Homeostasis in Intra-Macrophage Survival and Infectivity of Unicellular Parasites…*

for both cAMP and cGMP, turned out to be the most potent, with an IC50 value of 3.8 μM [72]. High concentration of PDE inhibitor like dipyridamole resulted in the inhibition of promastigote proliferation and macrophage infection in *L. major* [64].

We thank DST-INSPIRE Project grant (IFA-12 LSBM-22), PRG-University of Kalyani and NASI Senior Scientist Platinum Jubilee Fellowship for this work.

*DOI: http://dx.doi.org/10.5772/intechopen.86360*

The authors declare no conflict of interest.

cAMP cyclic adenosine monophosphate

, Anindita Bhattacharjee1

\*Address all correspondence to: pijush52@gmail.com

**Acknowledgements**

**Conflict of interest**

**Author details**

Arunima Biswas1

Kalyani, Kalyani, India

Biology, Kolkata, India

**Acronyms and abbreviations**

AC acidocalcisomes

PDE phosphodiesterase PKA protein kinase A

PPi inorganic pyrophosphate RAC receptor adenylate cyclase *Role of cAMP Homeostasis in Intra-Macrophage Survival and Infectivity of Unicellular Parasites… DOI: http://dx.doi.org/10.5772/intechopen.86360*

for both cAMP and cGMP, turned out to be the most potent, with an IC50 value of 3.8 μM [72]. High concentration of PDE inhibitor like dipyridamole resulted in the inhibition of promastigote proliferation and macrophage infection in *L. major* [64].
