168
5 Future Possibilities
activity in vitro against Plasmodium falciparum. Quinn and collaborators have also
used native mass spectrometry in the bacterial domain applied to natural product
fractions in which activity against Mycobacterium tuberculosis had been identified
through high throughput screening. These fractions were then checked by native MS
against a range of purified possible possible targets from Mycobacterium tuberculosis. Further analysis then uncovered a promising new natural product lead, altholactone, which binds, probably via irreversible covalent binding, to the mycobacterial protein Rv1466. Altholactone had moderate activity against Mycobacterium
tuberculosis in vitro (Elnaas et al. 2020). Extensions of the methodology might
embody looking for triple or higher order binding by ligands to one or more bacterial protein targets. Also three possible binding sites could be reasonably proposed
through computer-based modelling and docking and then checked through bioaffinity
mass spectrometry with the appropriate proteins. Alternatively one could increase
the number of proteins and look for any binding ligand. Some bacterial targets are
not proteins though and they would also need to be accessible in sufficient amount
and purity for the mass spectrometric studies.
It has also been shown recently that the strategy named PROSPECT (primary
screening of strains to prioritize expanded chemistry and targets) can be used to identify new hit compounds which in Mycobacterium tuberculosis (Mtb) inhibited a new
target found to be the essential efflux pump EfpA. These hit compounds can then be
developed or optimised into good leads. With PROSPECT, library compounds were
screened against pools of engineered Mtb hypomorph strains, in which vital bacterial
targets were greatly diminished. Specific activity-enriched and unbiased compound
libraries were used to identify new inhibitors targeting DNA gyrase, mycolic acid
biosynthesis, folate and tryptophan biosynthesis, RNA polymerase, as well as the
EfpA efflux pump. Significantly, this target depletion-based strategy identified new
inhibitors against targets that would not have been found using screening based on
wild-type Mtb alone. Also, this approach should be applicable to other pathogenic
bacterial strains, and potentially provide information on compounds with more than
one mode of action which possibly would not be revealed by conventional screening
methodology (Johnson et al. 2019).
Toxins and toxin inhibitors
Nullification by small molecules of bacterial endo- or exotoxins and their actions is
a potentially rich area for further development of multi-active agents. Various steps
could be targeted including their synthesis and release from bacteria, binding of
ligands to key sites on the toxins required for activity, and antagonising the binding
of toxins to host cell receptors. Bacterial toxin inhibitors have been developed based
on multivalent scaffolds with multiple sites for interaction. These inhibitors are based
on glycopolymers, glycodendrimers, or glycoclusters and are quite large molecules
(Branson and Turnbull 2013). Monovalent and multivalent carbohydrate receptorbinding inhibitors of cholera toxin, the entero toxin secreted by Vibrio cholera, a
pathogenic Gram-negative facultative anaerobe and cause of cholera, have also been
described by Kumar and Turnbull (2018), although again these mostly tend to have
5 Future Possibilities
activity in vitro against Plasmodium falciparum. Quinn and collaborators have also
used native mass spectrometry in the bacterial domain applied to natural product
fractions in which activity against Mycobacterium tuberculosis had been identified
through high throughput screening. These fractions were then checked by native MS
against a range of purified possible possible targets from Mycobacterium tuberculosis. Further analysis then uncovered a promising new natural product lead, altholactone, which binds, probably via irreversible covalent binding, to the mycobacterial protein Rv1466. Altholactone had moderate activity against Mycobacterium
tuberculosis in vitro (Elnaas et al. 2020). Extensions of the methodology might
embody looking for triple or higher order binding by ligands to one or more bacterial protein targets. Also three possible binding sites could be reasonably proposed
through computer-based modelling and docking and then checked through bioaffinity
mass spectrometry with the appropriate proteins. Alternatively one could increase
the number of proteins and look for any binding ligand. Some bacterial targets are
not proteins though and they would also need to be accessible in sufficient amount
and purity for the mass spectrometric studies.
It has also been shown recently that the strategy named PROSPECT (primary
screening of strains to prioritize expanded chemistry and targets) can be used to identify new hit compounds which in Mycobacterium tuberculosis (Mtb) inhibited a new
target found to be the essential efflux pump EfpA. These hit compounds can then be
developed or optimised into good leads. With PROSPECT, library compounds were
screened against pools of engineered Mtb hypomorph strains, in which vital bacterial
targets were greatly diminished. Specific activity-enriched and unbiased compound
libraries were used to identify new inhibitors targeting DNA gyrase, mycolic acid
biosynthesis, folate and tryptophan biosynthesis, RNA polymerase, as well as the
EfpA efflux pump. Significantly, this target depletion-based strategy identified new
inhibitors against targets that would not have been found using screening based on
wild-type Mtb alone. Also, this approach should be applicable to other pathogenic
bacterial strains, and potentially provide information on compounds with more than
one mode of action which possibly would not be revealed by conventional screening
methodology (Johnson et al. 2019).
Toxins and toxin inhibitors
Nullification by small molecules of bacterial endo- or exotoxins and their actions is
a potentially rich area for further development of multi-active agents. Various steps
could be targeted including their synthesis and release from bacteria, binding of
ligands to key sites on the toxins required for activity, and antagonising the binding
of toxins to host cell receptors. Bacterial toxin inhibitors have been developed based
on multivalent scaffolds with multiple sites for interaction. These inhibitors are based
on glycopolymers, glycodendrimers, or glycoclusters and are quite large molecules
(Branson and Turnbull 2013). Monovalent and multivalent carbohydrate receptorbinding inhibitors of cholera toxin, the entero toxin secreted by Vibrio cholera, a
pathogenic Gram-negative facultative anaerobe and cause of cholera, have also been
described by Kumar and Turnbull (2018), although again these mostly tend to have