The therapeutic potential of modulators of the Hedgehog-Gli signaling pathway
© BioMed Central Ltd 2002
Published: 6 November 2002
The discovery of small molecules that act as agonists and antagonists of the Hedgehog-Gli signaling pathway, which plays important roles in the embryo and adult, opens a new avenue for the treatment of diseases caused by aberrant suppression or activation of this complex pathway.
The Hedgehog-Gli signaling pathway regulates numerous events during the normal development of many cell types and organs, including the brain, bone, skin, gonads, lung, prostate, gastrointestinal tract and blood. The hedgehog (hh) gene - like many of the components of the signaling pathway triggered by Hedgehog (Hh) protein - was first identified in Drosophila, where it affects pattern formation very early in embryonic development. The binding of Hh to cell membranes triggers a signaling cascade that results in the regulation of transcription by zinc-finger transcription factors of the Gli family.
Of the three hh-family genes in mammals - Sonic hedgehog (Shh), Indian hedgehog (Ihh) and Desert hedgehog (Dhh) - Shh has been the most studied, mainly because it is expressed in various tissues but also because experiments with Shh protein are generally also applicable to other members of the family. The correct regulation of the Hh-Gli signaling pathway is essential not only for normal development but also to prevent a number of human diseases associated with abnormally increased or decreased signaling. Here, we discuss the potential use of small-molecule modulators of the Hh-signaling system, including those reported by Frank-Kamenetsky et al. in this issue , as therapeutic agents.
Mutations in components of the HH-GLI pathway in humans (human gene and protein names are given in capitals) lead to several diseases that result from either loss of function or ectopic activation of the pathway (reviewed in ). For example, haploinsufficiency of SHH or mutation in the human PTCH1 gene are associated with holoprosencephaly, a common syndrome affecting development of the forebrain and mid-face [6–8]. Moreover, ectopic expression of Shh, Gli1 or Gli2 in model systems leads to the formation of tumors that resemble basal cell carcinomas (BCCs) ([9–12]; reviewed in ), and sporadic human BCCs consistently express GLI, suggesting that all sporadic BCCs have this pathway active . Similarly, human mutations in the Suppressor of Fused - SU(FU)- gene predispose the carrier to medulloblastoma ; sporadic medulloblastomas can carry PTCH1 mutations and express GLI1 - again suggesting that they harbor an active pathway - and Ptc+/- mice can develop medulloblastomas ([15–19]; reviewed in ).
From an examination of the different mutations that cause aberrant suppression or activation of the HH-GLI pathway in humans, it seems clear that the development of small molecules that could act as agonists or antagonists of the function of proteins such as PTCH1, SMO or GLI might provide an effective therapeutic approach. One such drug could be SHH protein itself, a natural agonist. For example, it has been reported that injection of Shh into the striatum reduces behavioral deficits in a rat model of Parkinson's disease , that Shh can induce dopaminergic neuronal differentiation [21, 22] and that Shh is a neuroprotective agent . But Shh has a relatively short half-life in serum  and its therapeutic effects have been difficult to evaluate in vivo. The use of synthetic Hh agonists could therefore provide a viable alternative to Shh protein. Frank-Kamenetsky et al. have now identified a synthetic non-peptidyl small molecule that faithfully activates the Hh-Gli pathway, triggering the known biological effects of Hh signaling. They have shown that this agonist promotes proliferation and differentiation in a cell-type-specific manner in vitro, while in vivo it rescues developmental defects of Shh-null mouse embryos. But this agonist, unlike Shh protein, appears to bypass the Ptc1-regulatory step, by interacting directly with Smo (see Figure 1). Similar results with a near-identical agonist have now been obtained by another group . From a therapeutic point of view, the fact that the molecule retains its activity after oral administration is a great advantage and, if its ability to cross the blood-brain and placental barriers occurs in humans, it could be a very valuable therapeutic agent. Nevertheless, systemic side effects are to be expected, as there are many HH-responsive cell populations in the body.
Examples of diseases caused by loss of or ectopic function of the HH-GLI signaling pathway, and the possible agents that could, in principle, be used as therapeutics
Basal cell carcinoma
A number of studies suggest that cyclopamine specifically inhibits Smo activity [27–29] and that it can affect disease states caused by activation of the HH-GLI pathway. For example, the proliferation of a number of human brain-tumor cell lines and primary tumor cultures, including those from medulloblastomas and some gliomas  as well as medulloblastoma allografts , are inhibited by treatment with cyclopamine. This suggests that pathway activation is required for tumor maintenance. Other experiments suggest that the activity of Gli proteins, the terminal elements of the pathway, is sufficient to induce tumor development ([10–12]; reviewed in ). Thus, HH-pathway activity may be involved in the initiation as well as the maintenance of different tumors. This provides an additional opportunity to inhibit the growth of a number of tumors in different organs and tissues, such as basal cell carcinoma in the skin and medulloblastoma in the brain, with the same agent. Cyclopamine could be such an agent if the diseases to be treated arise from activation of the HH-signaling pathway at the level of SMOH or above. In addition, Frank-Kamenetsky et al. report the use of a new, synthetic, small-molecule inhibitor, Cur61414, which has inhibitory properties similar to those of cyclopamine and also acts at the level of Smo . Whether Cur61414, or four additional small-molecule antagonists (SANT1-4) that also act on Smo and were recently identified , will prove to be better and easier to use than cyclopamine remains to be determined, but testing them against skin  and brain tumors is warranted from a biological point of view.
Finally, given that carboxy-terminally truncated repressor forms of GLI3 are potent inhibitors of the activating output of the HH-signaling pathway [31, 34, 35], these could be used as antagonists for the treatment of tumors. The difficulty of delivering them into cells might require the development of in vivo transducing strategies, taking advantage, for example, of the ability of the Penetratin peptide to cross cell membranes while loaded with cargo . It also suggests that it would be useful to search for and design small molecules that inhibit GLI's transcription-activating function, perhaps by promoting endogenous GLI-repressor formation. This may be very difficult, but such drugs would be very specific and would be usable in cases where the cancer is due to mutation in the pathway at any level, from the extracellular ligand, the HH proteins, to the final mediators, the GLI proteins.
Agents that inhibit HH signaling may induce the regression of tumors that are dependent on a deregulated HH-GLI pathway, but these agents are likely also to affect the behavior of other normal pathway-dependent cells in the patient. This may, however, be a small price to pay in order to combat cancer, and the agents may have fewer side effects than current non-specific cytotoxic anti-cancer chemotherapies.
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