Blanc SIMI 7 - Sciences de l'information, de la matière et de l'ingénierie : Chimie moléculaire, organique, de coordination, catalyse et chimie biologique

Thapsigargin and novel analogues: Synthesis and hemisynthesis of SERCA inhibitors with specific therapeutic profile – THASER

Treatment of androgen-independent metastatic prostate cancer: thapsigargin analogues

Design and synthesis of thapsigargin analogues for treatment of androgen-independent metastatic prostate cancer

Treatment of androgen-independent metastatic prostate cancer by thapsigargin analogues

Thapsigargin (Tg) is a natural product that selectively inhibits (nM scale) SERCA calcium pumps in the endoplasmic reticulum, initiating a molecular cascade resulting in cell apoptosis. Therefore, unlike most cytotoxic cancer drugs, Tg is not cell cycle-dependent and can kill non dividing cancer cells. Preclinical results achieved by means of a Tg prodrug disclosed broad prostate cancer regression associated with minimal toxic effects. To date, only succinct structure activity relationship studies and a single total synthesis of this molecule were conducted. Significant progress is needed to effectively develop Tg analogues, especially with modified backbone. We propose a «multifaceted« research program to develop miscellaneous Tg analogues. These molecules should contribute to a better understanding of the mechanism of action of Tg and thus ultimately help to define pharmacologically more targeted compounds.

A multifaceted research program has been initiated, which relied on original accesses to novel Tg-like molecules, through total and/or partial synthesis. The final objective is to provide new, potent and if possible, simplified analogues of Tg, prostate cancer being specifically focused.
The program is composed of three closely interactive parts:
- The first effort concerns the elaboration of a viable synthetic route to access to Tg.
- The aim of the second part of the program is to elaborate original analogues through two complementary strategic approaches: hemisynthesis and total synthesis.
In the third part of the project, these molecules will be assayed on the basis of their SERCA-bound interactions through cocrystallization experiments and their ATPase activity. The cytotoxicity (particularly on prostate cancer cell lines) and the apoptosis effect of the synthesized analogues will be investigated. Finally, if necessary, in vivo trials will be realized.

It was previously demonstrated the relevance of the structure of Tg analogues regarding their SERCA binding. Thapsigargin was extracted on a large scale, from roots of Thapsia garganica with an extractor funded by ANR. High yields allowed the isolation of pure thapsigargin in large quantities. First hemisynthetic trials were made. Anti- SERCA and cytotoxic activities of these analogues were evaluated and very interesting results were observed. The most active analogues will be co-crystallized with SERCA for a better understanding of the interactions with the enzyme. These results allow us to develop other similar structures. The key steps of the total synthesis of Tg have been validated with elaboration of advanced intermediates. It remains to establish some chemical functions to complete the total synthesis of Tg. Based on this work, new analogues are being developed.

The objective of this work is to obtain original and active Tg analogues with simplified structure. These analogues are prepared either by partial synthesis from Tg or by total synthesis. All analogues will be tested for their anti SERCA activity and cytotoxic activity. Most active compounds will be co-crystallized with the SERCA for a better understanding of structure-activity relationships and the design of new analogues. These analogues will be evaluated in the prostate cancer.

Until now, the project has resulted in three publications (international journals):
Two publications concerned the total synthesis of Tg :
A. Tap, M. Jouanneau, G. Galvani, G. Sorin, M.-I. Lannou, J.-P. Férézou, J. Ardisson. Asymmetric synthe

Thapsigargin (Tg) is a highly oxygenated sesquiterpene belonging to the 6,12-guaianolide family, isolated from the Mediterranean plant species Thapsia. Tg presents an interesting and challenging complex chemical structure, with a polyoxygenated [5,7,5] tricyclic core and eight stereogenic centres. Tg is a selective inhibitor of the sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA)-dependent pumps at subnanomolar concentrations, leading to apoptotic cell death. The availability of crystal structures of Tg-bound enzyme facilitates understanding the structure-activity relationship (SAR) for SERCA inhibitors and enables molecular modelling techniques. Tg, which kills the cancer cells in all phases of the cell cycle, is equally effective in high and low growth, in contrast with classic antimitotics. Tg induces apoptosis in human androgen-independent prostatic cancer lines with nM IC50 values. Lack of selectivity, however, prevents the use of Tg as a systemic drug. Recently, very promising results for the treatment of non-hormonal dependent prostate cancer were described (current clinical investigation) based on the selective targeting of prostate cancer cells with Tg prodrugs.
Tg has been targeted as a lead, but its use is limited by dependence on natural sources and the diversity in the preparation of new analogues from the natural product itself is currently restricted to peripheral modifications of the core structure. Moreover, so far, only one total synthesis of Tg has been reported. Therefore, the availability of hemisynthetic or synthetic thapsigargin-related compounds would be welcomed.
Hereby, we propose a multifaceted research program which relies on original accesses to novel Tg-like molecules, through total and/or partial synthesis. The final objective is to provide a better understanding of the mechanism of action of this family of molecules as well as to lead to powerful and efficient biologically active molecules with promising therapeutic profile, especially with selective prostate cancer properties.
The proposed THASER program will be composed of three closely interactive parts:
- The first and most important efforts will concern the elaboration of a viable synthetic route to access to Tg, which stills represents a significant challenge for the chemist. Tight partnership between two renowned research groups in the field will help to design the more efficient route.
- The aim of the second part of the program is to elaborate original analogues through two complementary strategic approaches. A reasonable first objective to get pertinent molecules to be tested in a reasonable delay will involve hemisynthetic transformations from natural Tg itself. On the other hand, the modular character of the envisioned pathway for the total synthesis of Tg will allow a potentially good diversity in the structural definition of new active compounds.
- In a more advanced part of the project, these molecules will be assayed on the basis of their SERCA-bound interactions through co-crystallization experiments. The ATPase activity of the different Tg analogues will be tested biologically. The cytotoxicity (particularly on prostate cancer cell lines) and the apoptosis effect of the synthesized analogues will be investigated. Finally, if necessary, in vivo trials will be realized.
Furthermore, in prevision of still unknown potential alternative pharmaceutical applications, particularly against some human parasitoses, a collaboration with a Brazilian team specialized in SERCAs involved in tropical diseases will be undertaken.
In the international context, the importance of the challenge represented by the research of efficient multi-steps syntheses of new pharmacologically active molecules including anticancer agents requires to be highly competitive. Our original mult- partnership, rarely encountered in France in the academic area, should meet with these requirements.

Project coordinator

Madame janick ardisson (UNIVERSITE DE PARIS V - RENE DESCARTES) – janick.ardisson@parisdescartes.fr

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

Université Paris Descartes/LCRB UNIVERSITE DE PARIS V - RENE DESCARTES
UPS/ICMMO UNIVERSITE DE PARIS XI [PARIS- SUD]
UNIVERSITE DE PARIS V - RENE DESCARTES
UNIVERSITE DE PARIS V - RENE DESCARTES

Help of the ANR 569,998 euros
Beginning and duration of the scientific project: - 48 Months

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