Chameleonicity – or not?

by

Oliv Eidam

Alternative title: Permeability rules for beyond Ro5 molecules

In this post I want to highlight two stories with permeability rules for molecules in the beyond Rule of 5 (Ro5) space. The first story comes from Chugai and consists of at least two JACS papers (Tanada, Ohta) published in 2023 about the discovery of a clinical product called LUNA-18 targeting KRAS. They propose criteria for membrane permeability and metabolic stability specifically for cyclic peptides. The second story comes from Henrik Möbitz at Novartis who proposes a general permeability rule applying to most beyond Ro5 molecules. Both stories have one topic in common: molecular chameleonicity. Chameleonicity is a hypothesis for molecules in beyond Ro5 space and proposes that molecules adopt different conformations in polar vs apolar environments. That way molecules get membrane permeability. The big question: is chameleonicity required to get permeability for beyond Ro5 mols? Let’s dig into that question.
Back to the Chugai molecule LUNA-18. Let me start with this: huge congrats to the Chugai team. The development of the technology culminating in the discovery of LUNA-18 was a huge effort and required the collaboration of many scientists. Kudos!

LUNA-18 is heavily inspired by cyclosporin, the well known oral drug with a molecular weight of 1203 Da which is the prototype of a molecular chameleon. While the chemical composition is very different, they share things in common as the table below highlights.

A table comparing descriptors of LUNA-18 and cyclosporin highlights their similarity:

  • MW > 1200 Da
  • Cyclic peptides with 11 amino acids
  • high number of N-alkylated atoms
  • high Daylight clogP and high TPSA

In the Ohta et al. paper they come up with two rules for permeability of cyclic peptides:

  1. Clog P/(number of residues) of 1.17 or more
  2. N-alkylation ratio of 0.5 or more

They also propose that 11 amino acids appear to be the ideal length (between 10-12) to balance conformational flexibility. I think their first rule regarding clogP is useless because clogP values >14 are a) totally unrealistic, and b) owed to the usage of a specific program. Other programs yield clogP values between 3 to 5 for cyclosporin, which is closer to the measured clogP (4.12, REF?). The second rule regarding N-alkylation sounds more relevant to me, and for sure both cyclosporin and LUNA-18 have a high ratio.
In contrast to the Chugai papers, Henrik Möbitz analyzed a diverse list of permeable bRo5 drugs, and finds that most molecules do not exhibit vastly different conformations in different solvents. He comes up the with the Rule of ~1/5 which is actually two conditions:

  1. minimum 3D PSA < 100 (or 140) Å2
  2. 0.1 < TPSA / MW < 0.3 Å2 / Da

What is the minimum 3D PSA? The calculation of this value is not trivial and requires a program called COSMO-RS. Basically the polar surface area (PSA) is calculated for many conformations, and the 3D PSA of the conformation with the lowest PSA is designated as the minimum 3D PSA.

Scatter plot with 134 beyond Ro5 drugs: TPSA values on the X-axis, the minimum 3D PSA in decadiene on the Y-axis, both in [Å2]; each drug is colored by binned MDCK LE Papp assay values [10-6 cm/s]. Data from Möbitz paper, supp-si_2. Plotted with Spotfire.

Notes to Möbitz study:

  • Showing only one plot does not do justice to this extensive analysis and tour de force with many angles. I am showing the above because I was missing it in the paper. If you want more figures, check the original publication.
  • Most min 3D PSA values in this set of bRo5 mols are below 100/140 Å2 which are classical permeability limits when using TPSA.
  • Cyclosporin has an overestimated min 3D PSA of 131, because the conformation generation tools do not identify a conformation close to the ccdc structure DEKSAN with a PSA of ~100.

What do we learn regarding chameleonicity? Henrik’s study shows that permeable bRo5 mols do not adopt substantially different conformations and have min 3D PSA values below 100 Å2 regardless of the environment. In other words: most bRo5 molecules are no molecular chameleons.
Foremost, both stories are good news: first, Chugai demonstrates that cyclosporin is not the only drug-like exception peptide in the 10-12 amino acid space. LUNA-18 is probably also a chameleon like cyclosporin. At least the authors speculate about it (Figure S10), and this was also my first thought. While I applaud the design success leading to LUNA-18, the lessons learned are for a narrow chemical space. Second, I trust Henrik’s calculations and the conclusion that cyclosporin is an outlier in the permeable bRo5 space. This is good news because following physchem rules is “easier” compared to the design of chameleonicity. The downside of the 3D PSA is that it requires computationally expensive QM calculations which are not available to everyone.

Takeaways: If you make peptides in the cyclosporin space, then by all means learn all you can from that natural product. More generally speaking, if you are in the beyond Ro5 space, then you are well advised to consider a minimum 3D PSA < 100 (or 140) Å2 for your molecules to get permeability.

Side notes:

  • Beyond Rule of five (bRo5) drugs are often required for challenging targets. A recent example is zosurabalpin (Pahil KS et al., 2024; Zampaloni C et al., 2024), a new antibiotic with MW of 791 Da. Zosurabalpin targets Lpt transporters in the outer membrane and was published by my former colleagues from Roche.
  • It will be interesting to watch how new biotechs such as Orbis Medicines tackle permeability for their nCycles. They also work with cyclic peptides in the bRo5 space but they are smaller compared to cyclosporin and LUNA-18.
  • I would like to mention a recent paper by Dahlia Weiss et al. from BMS. Not limited to bRo5 drugs, and geared towards CNS availability, it proposes the Balanced Permeability Index (BPI) which aggregates three important parameters into one:
    BPI = 1000(LogD/PSA·HAC)
    HAC: stands for Heavy Atom Count in the above formula.
    PSA: they do not rely on TPSA and use a EPSA instead.
    The BPI is something to be checked and I love this statement: “The BPI is easily understood and can guide compound design toward smaller, less polar, and more lipophilic compounds.”

References

Development of Orally Bioavailable Peptides Targeting an Intracellular Protein: From a Hit to a Clinical KRAS Inhibitor.
Tanada M, Tamiya M, Matsuo A, Chiyoda A, Takano K, Ito T, Irie M, Kotake T, Takeyama R, Kawada H, Hayashi R, Ishikawa S, Nomura K, Furuichi N, Morita Y, Kage M, Hashimoto S, Nii K, Sase H, Ohara K, Ohta A, Kuramoto S, Nishimura Y, Iikura H, Shiraishi T.
J Am Chem Soc. 2023 Aug 2;145(30):16610-16620. doi: 10.1021/jacs.3c03886. Epub 2023 Jul 18.
PMID: 37463267

Validation of a New Methodology to Create Oral Drugs beyond the Rule of 5 for Intracellular Tough Targets.
Ohta A, Tanada M, Shinohara S, Morita Y, Nakano K, Yamagishi Y, Takano R, Kariyuki S, Iida T, Matsuo A, Ozeki K, Emura T, Sakurai Y, Takano K, Higashida A, Kojima M, Muraoka T, Takeyama R, Kato T, Kimura K, Ogawa K, Ohara K, Tanaka S, Kikuchi Y, Hisada N, Hayashi R, Nishimura Y, Nomura K, Tachibana T, Irie M, Kawada H, Torizawa T, Murao N, Kotake T, Tanaka M, Ishikawa S, Miyake T, Tamiya M, Arai M, Chiyoda A, Akai S, Sase H, Kuramoto S, Ito T, Shiraishi T, Kojima T, Iikura H.
J Am Chem Soc. 2023 Nov 8;145(44):24035-24051. doi: 10.1021/jacs.3c07145. Epub 2023 Oct 24.
PMID: 37874670

Design Principles for Balancing Lipophilicity and Permeability in beyond Rule of 5 Space.
Möbitz H.
ChemMedChem. 2024 Mar 1;19(5):e202300395. doi: 10.1002/cmdc.202300395. Epub 2023 Nov 30.
PMID: 37986275

A new antibiotic traps lipopolysaccharide in its intermembrane transporter.
Pahil KS, Gilman MSA, Baidin V, Clairfeuille T, Mattei P, Bieniossek C, Dey F, Muri D, Baettig R, Lobritz M, Bradley K, Kruse AC, Kahne D.
Nature. 2024 Jan;625(7995):572-577. doi: 10.1038/s41586-023-06799-7. Epub 2024 Jan 3.
PMID: 38172635 Free PMC article.

A novel antibiotic class targeting the lipopolysaccharide transporter.
Zampaloni C, Mattei P, Bleicher K, Winther L, Thäte C, Bucher C, Adam JM, Alanine A, Amrein KE, Baidin V, Bieniossek C, Bissantz C, Boess F, Cantrill C, Clairfeuille T, Dey F, Di Giorgio P, du Castel P, Dylus D, Dzygiel P, Felici A, García-Alcalde F, Haldimann A, Leipner M, Leyn S, Louvel S, Misson P, Osterman A, Pahil K, Rigo S, Schäublin A, Scharf S, Schmitz P, Stoll T, Trauner A, Zoffmann S, Kahne D, Young JAT, Lobritz MA, Bradley KA.
Nature. 2024 Jan;625(7995):566-571. doi: 10.1038/s41586-023-06873-0. Epub 2024 Jan 3.
PMID: 38172634 Free PMC article.

Orbis Medicines Launches with €26 Million Seed Financing to Transform Macrocycle Drug Development through Next-Generation Orally Dosable ‘nCycles’. Press release from Feb 29, 2024

Balanced Permeability Index: A Multiparameter Index for Improved In Vitro Permeability.
Weiss DR, Baylon JL, Evans ED, Paiva A, Everlof G, Cutrone J, Broccatelli F.
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