Polycyclic Guanidine Alkaloids from Poecilosclerida Marine Sponges

Sessile marine sponges provide an abundance of unique and diversified scaffolds. In particular, marine guanidine alkaloids display a very wide range of biological applications. A large number of cyclic guanidine alkaloids, including crambines, crambescins, crambescidins, batzelladines or netamins have been isolated from Poecilosclerida marine sponges. In this review, we will explore the chemodiversity of tri- and pentacyclic guanidine alkaloids. NMR and MS data tools will also be provided, and an overview of the wide range of bioactivities of crambescidins and batzelladines derivatives will be given.


Introduction
The most primitive benthic marine organisms are also among the most chemodiversified producers of secondary metabolites (SM). These compounds are used for predation and competition for space, as well as for communication, and protection against potential surrounding aggressors. Due to the high water dilution, the produced metabolites exhibit several potent biological activities [1]. Several reviews dealing with bioactive marine natural products, in particular, alkaloids have been published in the last few decades [2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19]. Of the many different marine species that have been researched, sponges are likely to be the most studied sources of marine natural products due to the large amount of structurally diverse SM scaffolds they produce. Even so, the last few years have seen a slight decline in the number of newly described metabolites [17,18].
In this review, we focused on the chemodiversity and biological activities of batzelladine-and crambescidin-like guanidine alkaloids isolated from Poecilosclerida marine sponges. These polycyclic guanidine alkaloids are extremely versatile SM [20]. Since their original isolation in 1989, several revisions have been made and many aspects remain to be studied. However, what is clear is that they seem to be specific to Poecilosclerida marine sponges. Over 53 derivatives have been isolated and their original structures, based on chemical degradation studies and extensive NMR and MS studies, have been revised since the development of their total synthesis. The structural and stereochemical complexity of this class of natural products have inspired the development of a number of new synthetic methodologies [21][22][23][24][25][26][27][28][29][30][31][32][33][34], which in turn has led to several total syntheses [31,33,[35][36][37][38][39][40][41][42][43][44][45][46][47][48][49][50] beyond the scope of this review. Finally, these unique and fascinating structures are coupled with a wide range of biological activities due to the typical shape of their tricyclic skeleton. However, little is known about their exact mechanism of action.  [72]. Used with permission.  [72]. Used with permission. Since 1989, the number of new TGA discovered each year is considered as almost constant, although it should be noted that in some years no new TGA were described ( Figure 2).

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Since 1989, the number of new TGA discovered each year is considered as almost constant, although it should be noted that in some years no new TGA were described ( Figure 2).
Cumulative number of new TGA discovered Figure 2. Number of new TGA discovered since 1989. The number of TGA discovered per year (grey bars) and the cumulative data representing the total number of TGA discovered (black curve) are presented from 1989 to 2015.

TGA Structures
Each TGA is structurally closely related to the others. Crambescidins-like GA differ from one another, by the C-8 spiro ring, the presence or absence of a hydroxyl group at C-13 or the nature of the side chain terminus at C-14.
The structures of 22 related crambescidin-like GA are summarized in Figure 3.

TGA Structures
Each TGA is structurally closely related to the others. Crambescidins-like GA differ from one another, by the C-8 spiro ring, the presence or absence of a hydroxyl group at C-13 or the nature of the side chain terminus at C-14.
The structures of 22 related crambescidin-like GA are summarized in Figure 3.

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Since 1989, the number of new TGA discovered each year is considered as almost constant, although it should be noted that in some years no new TGA were described ( Figure 2).
Cumulative number of new TGA discovered Figure 2. Number of new TGA discovered since 1989. The number of TGA discovered per year (grey bars) and the cumulative data representing the total number of TGA discovered (black curve) are presented from 1989 to 2015.

TGA Structures
Each TGA is structurally closely related to the others. Crambescidins-like GA differ from one another, by the C-8 spiro ring, the presence or absence of a hydroxyl group at C-13 or the nature of the side chain terminus at C-14.
The structures of 22 related crambescidin-like GA are summarized in Figure 3.
The structures of several members of the batzelladines have been revised since their original isolation. The originally proposed structures were based on chemical degradation studies, NMR spectroscopy analysis (1D and 2D), and comparison of the data to previously reported polycyclic guanidine such as ptilomycalin A (1). Since the original isolation work, the structures of batzelladines A (26), D (29), E (30), and F (31) have all been revised to their current structures after their partial or total synthesis [27][28][29][30]35,39]. As a consequence of these reassignments, the relative stereochemistry of batzelladines G (32), H (33), and I (34) has been reexamined [30].
The current structures of 22 related batzelladine-like GA are summarized in Figure 6.
The structures of several members of the batzelladines have been revised since their original isolation. The originally proposed structures were based on chemical degradation studies, NMR spectroscopy analysis (1D and 2D), and comparison of the data to previously reported polycyclic guanidine such as ptilomycalin A (1). Since the original isolation work, the structures of batzelladines A (26), D (29), E (30), and F (31) have all been revised to their current structures after their partial or total synthesis [27][28][29][30]35,39]. As a consequence of these reassignments, the relative stereochemistry of batzelladines G (32), H (33), and I (34) has been reexamined [30].
The current structures of 22 related batzelladine-like GA are summarized in Figure 6.
The structures of several members of the batzelladines have been revised since their original isolation. The originally proposed structures were based on chemical degradation studies, NMR spectroscopy analysis (1D and 2D), and comparison of the data to previously reported polycyclic guanidine such as ptilomycalin A (1). Since the original isolation work, the structures of batzelladines A (26), D (29), E (30), and F (31) have all been revised to their current structures after their partial or total synthesis [27][28][29][30]35,39]. As a consequence of these reassignments, the relative stereochemistry of batzelladines G (32), H (33), and I (34) has been reexamined [30].
The current structures of 22 related batzelladine-like GA are summarized in Figure 6.
Other batzelladine-like GA have been isolated such as batzelladine C (28), K (37), and E (30); dehydrobatzelladine C (35); clathriadic acid (41); and batzellamide A (50) (Figure 7).  In summary, the structurally unique tricyclic guanidinium ring system (hydro-5,6,6atriazaacenapthalene) that defines this class of natural products can be found in over 53 different alkaloids. While each of these natural products share this common structural motif, the substituents around the tricyclic core of these molecules leads to a significant structural diversity, which relate to a wide-range of biological properties. For instance, a large number of these molecules including batzelladine F (31) and ptilomycalin A (1) feature esters that tether the tricyclic core to a diverse array of different functional groups, including other tricyclic guanidine subunits. The other area of structural diversity within this family of alkaloids is both the C1 and the C8 alkyl chains, which vary in terms of length, units of unsaturation, and oxidation. Remote oxidation of the alkyl branches is characteristic of the crambescidin alkaloids, including ptilomycalin A (1) and crambescidin 359 (11), which feature two spirocyclic hemiaminals as well as the tricyclic guanidine framework. In addition to structural diversity, batzelladines-like GA and crambescidins-like GA alkaloids feature different stereochemical configurations of the tricyclic core. Both the trans-and cis-configurations of the pyrrolidine subunit have been reported in this class of natural products. Batzelladine F (31) highlights this stereochemical diversity as it contains two distinct tricyclic guanidine subunits, with each featuring one of the pyrrolidine configurations. The stereochemical diversity is generally limited to the configuration of the pyrrolidine unit, as all of the alkaloids in this class feature a trans relationship between the C4 (and/or C6) proton and the C1 (and/or C8) alkyl chain, with the exception of merobatzelladines A (42) and B (43). Both natural products feature a cis relationship between the C6 proton and the C8 alkyl chain.

TGA Classification
Different guanidine alkaloids classifications can be made, and, as such, Santos et al. (2015) have described four GA chemotypes [70]. The first class is constituted of a monocyclic pyrimidinamine skeleton, for example, crambescin C1 (55) or the bicyclic cyclopentapyrimidinamine skeleton, such as crambine A (56) (Figure 8). The second one is a tricyclic triazaacenaphthylene skeleton, which only contains one guanidine moiety like crambescidins, and the third one possess the same skeleton as for class 2, but contains, at least, one more guanidine moiety, as can be seen in the case of most batzelladines. The fourth one is a tricyclic cyclopentaquinazolinamine skeleton like netamine M (57) or ptilocaulin (58).    In summary, the structurally unique tricyclic guanidinium ring system (hydro-5,6,6atriazaacenapthalene) that defines this class of natural products can be found in over 53 different alkaloids. While each of these natural products share this common structural motif, the substituents around the tricyclic core of these molecules leads to a significant structural diversity, which relate to a wide-range of biological properties. For instance, a large number of these molecules including batzelladine F (31) and ptilomycalin A (1) feature esters that tether the tricyclic core to a diverse array of different functional groups, including other tricyclic guanidine subunits. The other area of structural diversity within this family of alkaloids is both the C1 and the C8 alkyl chains, which vary in terms of length, units of unsaturation, and oxidation. Remote oxidation of the alkyl branches is characteristic of the crambescidin alkaloids, including ptilomycalin A (1) and crambescidin 359 (11), which feature two spirocyclic hemiaminals as well as the tricyclic guanidine framework. In addition to structural diversity, batzelladines-like GA and crambescidins-like GA alkaloids feature different stereochemical configurations of the tricyclic core. Both the transand cis-configurations of the pyrrolidine subunit have been reported in this class of natural products. Batzelladine F (31) highlights this stereochemical diversity as it contains two distinct tricyclic guanidine subunits, with each featuring one of the pyrrolidine configurations. The stereochemical diversity is generally limited to the configuration of the pyrrolidine unit, as all of the alkaloids in this class feature a trans relationship between the C4 (and/or C6) proton and the C1 (and/or C8) alkyl chain, with the exception of merobatzelladines A (42) and B (43). Both natural products feature a cis relationship between the C6 proton and the C8 alkyl chain.

TGA Classification
Different guanidine alkaloids classifications can be made, and, as such, Santos et al. (2015) have described four GA chemotypes [70]. The first class is constituted of a monocyclic pyrimidinamine skeleton, for example, crambescin C1 (55) or the bicyclic cyclopentapyrimidinamine skeleton, such as crambine A (56) (Figure 8). The second one is a tricyclic triazaacenaphthylene skeleton, which only contains one guanidine moiety like crambescidins, and the third one possess the same skeleton as for class 2, but contains, at least, one more guanidine moiety, as can be seen in the case of most batzelladines. The fourth one is a tricyclic cyclopentaquinazolinamine skeleton like netamine M (57) or ptilocaulin (58). In summary, the structurally unique tricyclic guanidinium ring system (hydro-5,6,6atriazaacenapthalene) that defines this class of natural products can be found in over 53 different alkaloids. While each of these natural products share this common structural motif, the substituents around the tricyclic core of these molecules leads to a significant structural diversity, which relate to a wide-range of biological properties. For instance, a large number of these molecules including batzelladine F (31) and ptilomycalin A (1) feature esters that tether the tricyclic core to a diverse array of different functional groups, including other tricyclic guanidine subunits. The other area of structural diversity within this family of alkaloids is both the C1 and the C8 alkyl chains, which vary in terms of length, units of unsaturation, and oxidation. Remote oxidation of the alkyl branches is characteristic of the crambescidin alkaloids, including ptilomycalin A (1) and crambescidin 359 (11), which feature two spirocyclic hemiaminals as well as the tricyclic guanidine framework. In addition to structural diversity, batzelladines-like GA and crambescidins-like GA alkaloids feature different stereochemical configurations of the tricyclic core. Both the trans-and cis-configurations of the pyrrolidine subunit have been reported in this class of natural products. Batzelladine F (31) highlights this stereochemical diversity as it contains two distinct tricyclic guanidine subunits, with each featuring one of the pyrrolidine configurations. The stereochemical diversity is generally limited to the configuration of the pyrrolidine unit, as all of the alkaloids in this class feature a trans relationship between the C4 (and/or C6) proton and the C1 (and/or C8) alkyl chain, with the exception of merobatzelladines A (42) and B (43). Both natural products feature a cis relationship between the C6 proton and the C8 alkyl chain.

TGA Classification
Different guanidine alkaloids classifications can be made, and, as such, Santos et al. (2015) have described four GA chemotypes [70]. The first class is constituted of a monocyclic pyrimidinamine skeleton, for example, crambescin C1 (55) or the bicyclic cyclopentapyrimidinamine skeleton, such as crambine A (56) (Figure 8). The second one is a tricyclic triazaacenaphthylene skeleton, which only contains one guanidine moiety like crambescidins, and the third one possess the same skeleton as for class 2, but contains, at least, one more guanidine moiety, as can be seen in the case of most batzelladines. The fourth one is a tricyclic cyclopentaquinazolinamine skeleton like netamine M (57) or ptilocaulin (58).   Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins   (Table 3).
Interestingly, a chemotaxonomic study suggested that the biogenetically related guanidine alkaloids isolated from Crambe crambe, Monanchora arbuscula, Ptilocaulis spiculifer, and Hemimycale sp., should eventually be united in a single genus, preferentially Crambe [53]. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and 2D) and MS analyses, and chemical degradation.
The following section is a guideline for the structure determination of crambescidin-or batzelladine-like GA derivatives by their characteristic fragments via the use of NMR and MS analyses. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and 2D) and MS analyses, and chemical degradation.
The following section is a guideline for the structure determination of crambescidin-or batzelladine-like GA derivatives by their characteristic fragments via the use of NMR and MS analyses. Another classification can be made with the same first, second and fourth classes, an modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and and MS analyses, and chemical degradation. The following section is a guideline for the structure determination of crambescidinbatzelladine-like GA derivatives by their characteristic fragments via the use of NMR and analyses. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and 2D) and MS analyses, and chemical degradation.
The following section is a guideline for the structure determination of crambescidin-or batzelladine-like GA derivatives by their characteristic fragments via the use of NMR and MS analyses. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).
Interestingly, a chemotaxonomic study suggested that the biogenetically related guanidine alkaloids isolated from Crambe crambe, Monanchora arbuscula, Ptilocaulis spiculifer, and Hemimycale sp., should eventually be united in a single genus, preferentially Crambe [53]. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).
Interestingly, a chemotaxonomic study suggested that the biogenetically related guanidine alkaloids isolated from Crambe crambe, Monanchora arbuscula, Ptilocaulis spiculifer, and Hemimycale sp., should eventually be united in a single genus, preferentially Crambe [53]. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and 2D) and MS analyses, and chemical degradation.
The following section is a guideline for the structure determination of crambescidin-or batzelladine-like GA derivatives by their characteristic fragments via the use of NMR and MS analyses. Another classification can be made with the same first, second and fourth classes, and a modified third class describing pentacyclic triazaacenaphthylene skeletons like crambescidins (1-25) ( Table 3).

Analytical Tools for TGA Structural Analysis
TGA structural determinations have been accomplished based on extensive NMR (1D and 2D) and MS analyses, and chemical degradation.
The following section is a guideline for the structure determination of crambescidin-or batzelladine-like GA derivatives by their characteristic fragments via the use of NMR and MS analyses.

NMR Spectroscopy
TGA NMR spectra have a high content of information. Due to the relative signal richness, the 1 H-1 H COSY experiment is appropriate in helping to find whether one or another TGA class is present by analyzing the NMR spectra between 0.8 and 5 ppm. Nonetheless, several characteristic signals from different atoms listed below (Figure 9) should be found in the 1 H NMR spectra (Table 4).

NMR Spectroscopy
TGA NMR spectra have a high content of information. Due to the relative signal richness, the 1 H-1 H COSY experiment is appropriate in helping to find whether one or another TGA class is present by analyzing the NMR spectra between 0.8 and 5 ppm. Nonetheless, several characteristic signals from different atoms listed below (Figure 9) should be found in the 1 H NMR spectra (Table 4)  Crambescidin Case: Ptilomycalin A (1) As an example, we choose ptilomycalin A (1), which is part of the pentacyclic TGA class. First of all, the H1 signal (triplet) and the H4 and H5 signals of the double bond are very characteristic. There are also clear correlations from H1 to H5, and finally, the NMR chemical shifts of Ha, Hb, Hc, and Hd are also very characteristic within the crambescidin family ( Figure 10) (Table 4).  Crambescidin Case: Ptilomycalin A (1) As an example, we choose ptilomycalin A (1), which is part of the pentacyclic TGA class. First of all, the H 1 signal (triplet) and the H 4 and H 5 signals of the double bond are very characteristic. There are also clear correlations from H 1 to H 5 , and finally, the NMR chemical shifts of H a , H b, H c, and H d are also very characteristic within the crambescidin family ( Figure 10) (Table 4). TGA NMR spectra have a high content of information. Due to the relative signal richness, the 1 H-1 H COSY experiment is appropriate in helping to find whether one or another TGA class is present by analyzing the NMR spectra between 0.8 and 5 ppm. Nonetheless, several characteristic signals from different atoms listed below (Figure 9) should be found in the 1 H NMR spectra (Table 4)  Crambescidin Case: Ptilomycalin A (1) As an example, we choose ptilomycalin A (1), which is part of the pentacyclic TGA class. First of all, the H1 signal (triplet) and the H4 and H5 signals of the double bond are very characteristic. There are also clear correlations from H1 to H5, and finally, the NMR chemical shifts of Ha, Hb, Hc, and Hd are also very characteristic within the crambescidin family ( Figure 10) (Table 4). In general, all the NMR signals reported in Table 4 are more shielded in batzelladines than in crambescidins. Moreover, the NMR spectra are more complex and are often different since the guanidinium core can be once or more dehydrogenated.
Within the tricyclic TGA class 2, we used the example of batzelladine F (31). The major difference between batzelladine F and ptilomycalin A spectra is the absence of the signals for the protons H4 and H5, and the corresponding correlation. On the other hand, two correlations are very characteristic within batzelladines: Ha with Hb, and Hc with Hd ( Figure 11 In general, all the NMR signals reported in Table 4 are more shielded in batzelladines than in crambescidins. Moreover, the NMR spectra are more complex and are often different since the guanidinium core can be once or more dehydrogenated. Within the tricyclic TGA class 2, we used the example of batzelladine F (31). The major difference between batzelladine F and ptilomycalin A spectra is the absence of the signals for the protons H 4 and H 5 , and the corresponding correlation. On the other hand, two correlations are very characteristic within batzelladines: H a with H b , and H c with H d (Figure 11). In general, all the NMR signals reported in Table 4 are more shielded in batzelladines than in crambescidins. Moreover, the NMR spectra are more complex and are often different since the guanidinium core can be once or more dehydrogenated.
Within the tricyclic TGA class 2, we used the example of batzelladine F (31). The major difference between batzelladine F and ptilomycalin A spectra is the absence of the signals for the protons H4 and H5, and the corresponding correlation. On the other hand, two correlations are very characteristic within batzelladines: Ha with Hb, and Hc with Hd ( Figure 11

Mass Spectrometry
Several TGA were detected by positive electrospray mass spectrometry ionization studies. Usually, authors notify the TGA quasi-molecular ion [M + H] + with the exception of batzelladines M (39) and N (40), which were detected through their dicharged ion [M + 2H] 2+ [73]. On the other hand, both quasi-molecular and discharged ions were observed for batzelladine L (38) and monanchomycalin C (25). Mass spectrometry data for TGA are summarized in Table 5. Tandem mass spectrometry (MS²) experiments were also performed to confirm a hypothesis or provide additional information concerning the TGA side chains, as reported in Table 6. Tandem mass spectrometry (MS²) experiments were also performed to confirm a hypothesis or provide additional information concerning the TGA side chains, as reported in Table 6.

Biological Activities
TGA exhibited a wide range of biological activities with mainly antiviral, antimicrobial (including antifungal, antibacterial, anti-yeast, and antiparasitic), and antitumor properties.

Antiviral Activities
Several TGA derivatives were evaluated for their antiviral activities against different viruses such as Human immunodeficiency virus (HIV-1), Herpes simplex virus (HSV-1), and Human hepatitis B virus (HBV). The reported data for 17 TGA are summarized in Table 7. Tandem mass spectrometry (MS²) experiments were also performed to confirm a hypothesis or provide additional information concerning the TGA side chains, as reported in Table 6.
Surprisingly, neofolitispates (8)(9)(10) are the only crambescidin-like GA derivatives that have been tested on HBV and exhibited an anti-HBV activity [57]. Unfortunately, few details have been reported.
On the other hand, batzelladines A-E (26)(27)(28)(29)(30) were shown to block the interaction between the surface of the HIV envelope glycoprotein gp120 and the extracellular domains of human CD4 receptor protein [77]. This binding is vital to the replication of the virus as it controls its entry into the human cells, since without access to the biochemical environment within the cell the virus is unable to replicate. As a consequence, batzelladines have a therapeutic interest in the treatment of HIV [76].
To date, batzelladine-like GA have not been tested against HSV or HBV viruses.
In some cases, Batzelladine-like GA present similar activities compared to crambescidin-like GA. Batzelladines C (28) and L (38) are often the more potent molecules [73]. Nevertheless, batzelladine M (39) is the less active TGA class 2 and surprisingly, the same authors showed batzelladine N (40) to be nine times more efficient compared to batzelladine M (39) against M. tuberculosis (minimum inhibitory concentration (MIC) 3.18 and 28.5 µg¨mL´1, respectively), despite their chemical structures being closely related [73].

Antitumoral Activities
Over 20 TGA were tested for their cytotoxicity against several cancer cell lines such as prostate, ovary, breast, melanoma leukemia, pancreas, colon, and cervix (Table 9).   Ptilomycalin A (1) is once again the most active TGA tested as its half maximal growth inhibition values (GI 50 ) are always below 0.1 µg/mL on all the cell lines tested [73]. Crambescidin 800 (2) shows similar activities [73]. Compared to TGA class 3, TGA class 2 is less active, although batzelladine C (28) and L (38) and dehydrobatzelladine C (35), in general, exhibited a GI 50 below 1 µg/mL [73]. Finally, crambescidin 816 (3) was shown to inhibit cell migration by altering the cytoskeleton dynamics and induced cell death by apoptosis [75,78].
In summary, both TGA class 2 and 3 have been tested against viruses, microbes, and several cancer cell lines. The wide-ranging biological activity of this class of natural products can be in part attributed to the cationic nature of the guanidinium functional group that can participate in a large number of non-covalent molecular interactions [77]. In general, TGA from class 3 exhibited better activities compared to TGA from class 2 [73]. Within TGA class 3, ptilomycalin A (1) and crambescidin 800 (2) showed similar activities in all assays, suggesting that the hydroxyl group of the right-handed portion in crambescidin 800 (2) did not affect the bioactivity. Nevertheless, 16β-hydroxycrambescidin 359 (16) did not show any significant antimicrobial activity, suggesting that the hydroxyl group, located on the pentacycle at C-16, diminished the activity [73]. Further studies are needed to confirm this hypothesis. This could be assessed by evaluating the inhibitory activities of crambescidins 816 (3), 830 (4) and 844 (5), as they carry the same hydroxyl moiety as crambescidin 800 (2) and another hydroxyl located in the pentacycle at C-13. In parallel, several batzelladines have been reported to disrupt protein-protein interactions. Elucidation of the mechanism by which protein-protein interactions are modulated by these molecules is of great interest, since protein-protein associations are important in all the aspects of cell biochemistry. Moreover, small molecules that influence protein-protein association would be new biological tools and potential therapeutic agents. In particular, batzelladines or their derivatives may prove to be applicable for AIDS treatment. Batzelladines A-E (26)(27)(28)(29)(30) block interaction between the surface of the HIV envelope glycoprotein gp120 and the extracellular domains of human CD4 receptor protein [66]. A subset of batzelladines exhibited also potential immunosuppressive activity as they induce dissociation of the complex between the protein kinase p56 lck and CD4 [67]. Synthetic derivatives of batzelladines were reported to disrupt Nef-p53, Nef-actin, and Nef-p56 lck interactions [77]. Bewley et al. tested a series of 28 synthetic batzelladine-like GA analogues on HIV-1 cell fusion assay, to find structure-activity relationships [76]. According to this study, the greater the rigidity of the molecule, the less biologically active it is. Moreover, they have shown that the most active compounds tested were compounds which contain two tricyclic guanidine moieties connected by an alkyl ester linkage including eight heavy atoms, such as batzelladines F (31) and G (32). Batzelladines biological properties could be dramatically affected by the ester side chains. For example, batzelladine A (26) inhibits the binding of HIV glycoprotein gp120 to CD4 receptors, whereas batzelladine D (29) has no known biological activity [77].
Finally, several derivatives that do not feature an ester side chain may still exhibit interesting biological properties such as the antibacterial and antimalarial activities of merobatzelladines A (42) and B (43) [69].

Conclusions
Several batzelladine-and crambescidin-like guanidine alkaloids have been isolated from Poecilosclerida marine sponges. Their biosynthesis, or the biosynthesis of their precursors, may involve symbiotic microorganisms [79]. This class of natural products is structurally unique as all the derivatives are constituted by a tricyclic guanidinium ring system, to which are appended different substituents. This significant structural diversity has led to wide-ranging biological properties with mainly antiviral, antimicrobial (including antifungal, antibacterial, anti-yeast, and antiparasitic), and antitumor activities. In addition to the biomimetic strategy, a number of groups completed the total synthesis of several of these alkaloids by developing new synthetic methodologies. Although there are many efficient and stereoselective synthetic routes towards this class of natural products, there is not yet one method that would allow entry into all of the cores of this family. Nonetheless, a number of these syntheses have enabled the establishment of their relative and absolute stereochemical configurations. In conclusion, several of these alkaloids and their synthetic analogs were prepared on a large enough scale to allow further biological testing, including few structure-activity relationship studies. Ongoing studies may provide us with further clues regarding this class of Marine Natural Products.