The human filarial parasites represent a major impediment to global public health. The lymphatic filaria (Wuchereria bancrofti
, Brugia malayi
and Brugia timori
) together infect roughly 4.4 million people worldwide and are estimated to result in the loss of 5.7 million disability adjusted life years [1
]. Onchocerciasis, or river blindness, is caused by the filarial parasite Onchocerca volvulus
, and historically has represented the second most important cause of infectious blindness worldwide [2
]. The significant socio-economic impact of these diseases has attracted the attention of the international community, which is currently supporting several control and elimination programs whose overall goal is to either eliminate the diseases as public health problems or to regionally eliminate the parasites altogether [3
]. All of these programs currently rely primarily upon mass distribution of a very limited number of drugs which must be given repeatedly over a prolonged period of time to control transmission. This is logistically difficult and leaves the programs vulnerable to failure if drug resistance develops. Furthermore, some of the drugs used by these programs may induce adverse reactions, complicating their use in mass drug administration programs. For example diethylcarbamazine, while effective against the lymphatic filaria, causes severe reactions in individuals infected with O. volvulus
], precluding its use in much of Africa where W. bancrofti
and O. volvulus
are co-endemic. Similarly, ivermectin can cause severe neurological complications in individuals infected with Loa loa
, complicating ivermectin distribution for onchocerciasis in much of central Africa, where L. loa
is endemic [5
]. Thus, there is a need to develop new drugs that can supplement the currently available compounds used to treat these infections.
The filarial parasites are ecdysozoans, a group of organisms whose developmental profile is characterized by a series of molts. The developmental program of the ecdysozoans is not shared with vertebrates, making this process an attractive potential chemotherapeutic target. In insects, molting is controlled through the release of the molting steroid hormone 20-OH ecdysone. 20-OH ecdysone mediates its effect through the ecdysone receptor, a heterodimer consisting of two different members of the zinc finger-containing nuclear hormone receptor family of transcription factors, the ecdysone receptor protein (EcR) and the retinoic acid X receptor (RXR) [6
]. Binding of 20-OH ecdysone activates the ecdysone receptor, which then binds to a specific motif (the ecdysone response element [EcRE]) in a number of ecdysone responsive genes, thereby activating transcription of these genes. These in turn control the specific expression of a number of other genes, resulting in the ecdysone cascade, which eventually leads to molting. Mutation of the ecdysone receptor in insects leads to a wide variety of developmental defects, including embryonic and larval lethality, aberrant neuronal remodeling and defects in vitellogenesis [7
]. These findings suggest that the EcR is a central developmental regulator in insects, playing a role in embryogenesis and other developmental processes in addition to controlling molting.
The mechanisms controlling the developmental processes of the filarial parasites remain poorly understood. However, several lines of evidence suggest that ecdysone may play important roles in these processes. For example, ecdysone and related compounds (hereafter referred to for simplicity as ecdysteroids) have been found in many parasitic nematodes [9
]. Ecdysteroids also have been shown to have developmental effects on parasitic nematodes. For example, ecdysone was shown to stimulate microfilarial release in Brugia pahangi
, and to promote embryogenesis in ovaries of Dirofilaria immitis
adult females [10
]. Finally, two genes encoding orthologues of both partners of the insect ecdysone receptor heterodimer (EcR and RXR) have been recently characterized in B. malayi
]. The protein encoded by the EcR gene is capable of dimerizing with RXR homologues from B. malayi
and other organisms, a property identical to that of the insect EcRs [6
]. Finally, synthetic promoter constructs containing a consensus EcRE inserted into a B. malayi
ribosomal protein promoter were capable of being induced by 20-OH ecdysone when transfected into B. malayi
]. Together, these data suggest that B. malayi
contains a functional ecdysone regulated developmental pathway.
As a first step in deciphering the physiological role of this ecdysone regulatory pathway in B. malayi, we have studied the effect of ecdysteroid exposure on protein expression in cultured adult female B. malayi and have conducted a bioinformatic analysis of the B. malayi genome to identify endogenous genes containing consensus EcREs. We further demonstrate that a native B. malayi promoter containing an endogenous EcRE is up-regulated by 20-OH-ecdysone, and that the endogenous EcRE is necessary for this ecdysone mediated up-regulation of the promoter.