**5. Conclusions**

*4.1.3. Marine toxins*

There is a plethora of marine toxins with distinctive neurological effects. These toxins are mainly known for the poisoning risk they carry. Nonetheless, a few of them have valuable biological activities as research chemicals and one has intriguing effects relative to AD. Using cell-based assays, the toxin 13-desmethyl spirolide C from the dinoflagellate *Alexandrium ostenfeldii* was shown to reduce Aβ accumulation in cells by just over 40% [117]. The antibody used (6E10, Covance) detects Aβ without distinguishing between monomeric or oligomeric or fibril forms. It would therefore be valuable to repeat these analyses using fold-specific anti-Aβ antibodies or another fold-sensitive method to see if any effect there could be detected.

A variety of chemical chaperones, which are loosely defined as small molecules that promote folding of many proteins, have gained interest in terms of wide-spectrum protection of proteins from misfolding (reviewed in [118]). In marine species faced with protein misfolding risk, the accumulation of one or more chemical chaperones is a common adaptation because these small chaperoning molecules promote general proteostasis [111, 119]. Therefore, at first glance, these molecules would appear to be ideal in terms of proteins such as Aβ and αS. They could be produced naturally or synthetically, depending upon their molecular features, and they may stabilize many different proteins. However, some chemical chaperones may have problematic effects with respect to amyloidotic protein misfolding. Glycerol at supraphysiological concen‐ trations (molar range) and trimethylamine oxide (TMAO) at moderate concentrations were both shown to favour the transition of Aβ from its unfolded conformation to the β-sheet form requisite for fiber formation [120]. Protofibril to fibril conversion was also enhanced [120]. The situation appears more complex for αS, with elevated concentrations (molar range) of TMAO favouring a partially folded form with high propensity for fibril formation and even higher concentrations of TMAO favouring an oligomeric α-helical conformation [121], which may be consistent with a native α-helical form of the protein that resists misfolding [29]. Although these chaperone concentrations used were far in excess of those that would be reached *in vivo*, the effects suggest a possibility to be aware of if a novel folding modulator appears to be non-specific. Stabilization by chemical chaperones may bring an increased risk of aggregation for some proteins [119]. Furthermore, stabilization of a wider range of cell proteins or protein complexes may cause unanticipated problems. Therefore, examination of unusual and off-

Otherwise, it may act on Aβ in other beneficial ways that would also be of interest.

target effects would be prudent in the evaluation of chemical chaperones.

A challenge presented by many rare or remote marine species that may produce pharmaceut‐ ical or nutraceutical molecules is the difficulty in obtaining sufficient product in a sustainable manner, from both the environmental and economic standpoints [122]. Options for these products include the development of culture methods for the species or synthesis of the molecule, although in many cases the molecules are too complex to be efficiently synthesized [122, 123]. In contrast, an advantage for nutraceutical or drug development from many

**4.2. Sources for marine products**

*4.1.4. Marine-sourced chemical chaperones and conflicting results*

108 Using Old Solutions to New Problems - Natural Drug Discovery in the 21st Century

With protein misfolding identified as a causative event in multiple neurodegenerative diseases and with no definitive means of prevention or cure in place, the progress being made in the discovery and development of natural protein misfolding modulators is of great interest. Although several promising leads from terrestrial sources have been identified, none have been shown to provide a clear result in disease outcome to date. Therefore, while current products are being developed and refined, it would be logical to investigate a wider range of sources for protein folding modulators. In this context, marine species deserve a second (or, in some cases, a first) look.

There has already been extensive work on marine molecules from some sources, but crucial to the proposed endeavour is screening for the most relevant activities. For example, testing marine molecules or extracts for effects in model Aβ or αS cell survival assays may not indicate protein folding modulation. However, the reverse is likely to be true in most cases. In other words, appropriate protein folding modulation activity should lead to enhanced cell survival. It is also important to be mindful of assay subtleties. Testing for anti-fibrillization activity may uncover molecules that cause toxic oligomers to accumulate instead of amyloid fibrils, leading to increased pathology instead of amelioration. Similarly, searching for molecules that bring about a reduction in total Aβ (or αS) may not be prudent, as we have little understanding of the normal roles of these proteins in the brain and therefore drastic reduction in levels may have undesirable consequences [18]. For these reasons, an assay suite that allows determina‐ tion of toxic oligomers as well as other (non-toxic) oligmers, monomers, amyloid fibrils and other aggregates would be ideal. In order to identify a marine extract or molecule that provides protective protein folding modulation, the spectrum of its effects on protein misfolding and association protein would need to be investigated. Furthermore, because a number of natural molecules appear to affect the misfolding trajectories of both Aβ and αS, these would be of particular interest because of the potential for wider-ranging protection from toxic misfolding.

In closing, examination of marine biota for protein folding modulators using a suitable suite of assays may offer a promising opportunity to identify a safe, effective and sustainable protein folding modulator that addresses the cause of a neurodegenerative disease. The untapped diversity of marine species combined with a rational selection of assays for folding modulator identification based upon examples used in terrestrial investigations and coupled with appropriate downstream *in vivo* analysis would be ideal. For proteins such as Aβ and αS, which form similar amyloids and can even cross-seed in spite of their structural differences [129], a further tantalizing prospect would be the identification of a molecule that would be effective for both AD and PD. Given the paucity of treatments currently available, discovery of an active molecule from a marine resource could bring hope at this time.
