Macrolide Inspired Macrocycles as a Promising Templates for Unmet Medical Needs
1. Introduction

            A novel macrolide inspired macrocyclic library was designed and prepared by FideltaMacro™ technology,1 using our long-term experience and in-house knowledge on the chemistry and pharmacology of macrolides and natural products. Macrocycles possessing different ring sizes are designed to diversify and enrich chemical space, incorporating a variety of 3D shapes and potential pharmacophoric features. The FideltaMacro™ library consist of 1500 unique compounds with over 50 novel macrocyclic rings. Compounds populate favourable lipophilic space, exhibit good solubility and low affinity for plasma proteins. Metabolic stability varies from low to high, mostly depending on the ring decorations while permeability in the standard high-throughput assays remains low. Different approaches to characterize the permeability for molecules beyond the rule-of-five have been investigated. Target based as well as phenotypic screening in the anti-inflammatory area, has been performed and several promising chemical scaffolds have been identified.

2. Library design

The FideltaMacro™ library was designed using fingerprint mapping to; enrich amino-acid side-chains most prevalent in protein-protein interactions; to introduce different spatial arrangement of pharmacophoric groups; combine side-chains with different physico–chemical properties; introduce stereochemical diversity and flexibility constrains, as schematically shown in Figure 1.

Figure 1. Schematic presentation of fingerprint mapping in FideltaMacro™ library.

3. Library synthesis

The general synthetic strategy (Scheme 1) comprises several steps as described previously.1 Step 1 is ring opening of macrolide scaffold by oxidative cleavage of the 11,12-diol moiety to afford the seco compound B, which is suitable for further modification of secondary amino group and insertion of desired motifs, fragments, potential pharmacophores and hot spots from known protein-protein interactions (Step 2). Seco compound B still contains the 11C-13C fragment of the macrolide which serves as a protecting group for the carboxylic acid group on 1C. After deprotection of the amino group, a variety of chemical reactions can be used for insertion and further modifications of desired fragments followed by macrocyclisation (Step 3). Once the macrocycle is formed, the side chain modification (Step 4) can be performed as a final step towards specific target molecules or for preparation of small focused libraries around each macrocyclic scaffold.

Scheme 1. Synthetic strategy for the preparation of FideltaMacroTM compounds based on azithromycin scaffold.

4. Physico-chemical properties and 3D shape

Structural characteristics of FideltaMacro™ compounds are shown in Figure 2. Compounds mostly fit within the boundaries for oral macrocycles2 regarding molecular weight, lipophilicity (cLogP), polar surface area (PSA), number of hydrogen bond donors (HBD, with slightly higher number of hydrogen bond acceptors (HBA) and rotational bonds (NRB).

Figure 2. Distribution of structural parameters of the FideltaMacro™ library3 together with boundaries for oral macrocycles defined by Villar et. al.2

Due to structural complexity and number of stereochemical centers, compounds are rich in the 3D character as outlined of the PMI plot shown in Figure 3a.

Figure 3. 3D character described with: a) normalized PMI (principal moments of inertia) ratios (pmi1/pmi3; pmi2/pm3) (structures are from the DrugBank: light grey—200 centroids; blue—approved macrocyclic drugs; red— representative scaffolds from FideltaMacro™ library); b) variations in size and shape due to different diversification vectors.

In general, the macrocycles exhibit; good solubility; low plasma protein binding; lipophilicity that fits into the range reported for oral macrocycles; no effect on hERG and CYP enzymes; low-to-high in vitro microsomal stability ranges depending on ring size and appended functionality; low permeability in the MDCK in vitro assay. It has been shown previously that in vitro permeability does not correlate with observed oral absorption and that cellular accumulation is a better in vitro predictor for the macrolide class of compounds4; whether this observation is maintained for FideltaMacro™ scaffolds still needs to be explored. Measured kinetic solubility, ChromLogD and microsomal stability values for a subset of the FideltaMacro™ library are shown in Figure 4.

Figure 4. PhysChem and in vitro ADME profile of the FideltaMacro™ library.

In order to address the conformational complexity of prepared macrocycles NMR constrained conformational analysis is usually undertaken, due to inefficiency of computational techniques to reliably predict macrocyclic conformations. An ensemble of conformations that satisfy NMR constrains is generated followed by MD simulations in solvents with different dielectric constants mimicking physiological conditions and membrane environment. Potential for chameleonic behavior, e.g. significant solvent induced conformational changes, that would increase membrane permeability and therefore oral exposure is also explored. Conformational differences for two epimers of a 14-membered Br-intermediate are shown in Figure 5. Although the macrocyclic ring is quite small, molecular dynamic simulations reveal different H-bond potential resulting in different shape, PSA and consequently lipophilicity.

Figure 5. Stereochemistry influence on the 3D conformations of macrocycles: a) NMR constrained 3D conformations for two epimers of the Br-intermediate; b) summary of MD simulation in chloroform

5. Biological profiling

A representative set of compounds from the FideltaMacro™ library has been profiled in a battery of phenotypic screens. The workflow and the outcome of the phenotypic screening are summarized in Figure 6.

Figure 6. Phenotypic screening workflow.

Screening the FideltaMacro™ library has resulted in some notable discoveries, including; 1) a compound with significant in vivo anti-fibrotic effects,5 which was subsequently outsourced to Galapagos and is currently in pre-clinical development; 2) a novel scaffold with anti-infective profile comparable to azithromycin and active in an in vivo gram negative model; 3) several diverse scaffolds possessing in vivo anti-inflammatory activity, see Figure 7. In vivo active scaffolds identified from phenotypic screening cascades are promising starting points for anti-inflammatory and immuno-modulatory diseases.

Figure 7. Results of phenotypic profiling and in vivo activity of three diverse macrocyclic scaffolds.

In addition to the aforementioned discoveries, the FideltaMacro™ library has been developed to include target oriented scaffolds for MDM2/p53 and IL-17A/IL17-RA disruptors; see our recently published paper in collaboration with LEO Pharma6  for more information on nM inhibitors of IL-17A.


  1. A. Fajdetić et. al. Int. Pat. Appl. WO2017194452A1, 2017.
  2. E.A.Villar et al. Nature Chemical Biology10 (2014) 723.
  3. ACD Percepta, version 2017.2.1, Advanced Chemistry Development, Inc., Toronto, Canada.
  4. V. Stepanić et. al. J. Med. Chem. 53 (2011) 719.
  5. B. Hrvačić et. al. poster available at
  6. S. Koštrun et. al. J. Med. Chem. 64(2021) 8354.