Tài liệu Báo cáo khoa học:Symmetric fluoro-substituted diol-based HIV protease inhibitors

Symmetric fluoro-substituted diol-based HIV protease inhibitors

Ortho-fluorinated and meta-fluorinated P1/P1¢-benzyloxy side groups significantly

improve the antiviral activity and preserve binding efficacy

Jimmy Lindberg1

, David Pyring2

, Seved Lo¨ wgren1

, A˚ sa Rosenquist2

, Guido Zuccarello2

,

Ingemar Kvarnstro¨ m2

, Hong Zhang3

, Lotta Vrang3

, Bjo¨ rn Classon3,4, Anders Hallberg5

,

Bertil Samuelsson3,4 and Torsten Unge1

1

Department of Cell and Molecular Biology, BMC, Uppsala University, Sweden; 2

Department of Chemistry, Linko¨ping University,

Sweden; 3

Medivir AB, Huddinge, Sweden; 4

Department of Organic Chemistry, Arrhenius Laboratory, Stockholm University, Sweden; 5

Department of Organic Pharmaceutical Chemistry, Uppsala University, BMC, Sweden

HIV-1 protease is a pivotal enzyme in the later stages of the

viral life cycle which is responsible for the processing and

maturation of the virus particle into an infectious virion. As

such, HIV-1 protease has become an important target for the

treatment of AIDS, and efficient drugs have been developed.

However, negative side effects and fast emerging resistance

to the current drugs have necessitated the development of

novel chemical entities in order to exploit different phar￾macokinetic properties as well as new interaction patterns.

We have used X-ray crystallography to decipher the struc￾ture–activity relationship of fluoro-substitution as a strategy

to improve the antiviral activity and the protease inhibition

of C2-symmetric diol-based inhibitors. In total we present six

protease–inhibitor complexes at 1.8–2.3 A˚ resolution, which

have been structurally characterized with respect to their

antiviral and inhibitory activities, in order to evaluate the

effects of different fluoro-substitutions. These C2-symmetric

inhibitors comprise mono- and difluoro-substituted benzyl￾oxy side groups in P1/P1¢ and indanoleamine side groups in

P2/P2¢. The ortho- and meta-fluorinated P1/P1¢-benzyloxy

side groups proved to have the most cytopathogenic effects

compared with the nonsubstituted analog and related

C2-symmetric diol-based inhibitors. The different fluoro￾substitutions are well accommodated in the protease S1/S1¢

subsites, as observed by an increase in favorable Van der

Waals contacts and surface area buried by the inhibitors.

These data will be used in the development of potent

inhibitors with different pharmacokinetic profiles towards

resistant protease mutants.

Keywords: AIDS; aspartic protease; crystal structure; fluor￾ine; HIV.

Human immunodeficiency virus 1 (HIV) is the causative

agent of AIDS [1–3]. The single-stranded RNA genome of

HIV encodes a dimeric aspartyl protease (protease) which

processes the viral gag and gag-pol precursor polyproteins

into structural and functional proteins. The HIV protease

has been shown to be essential in the production of mature

and infectious virions [4,5], hence inhibition of this enzyme

has become an attractive target for effective antiviral agents;

several protease inhibitors are currently in clinical trials.

Despite the initial success of the FDA approved protease

inhibitors (saquinavir [6], ritonavir [7], indinavir [8], nelfin￾avir [9], amprenavir [10], lopinavir [11] and atazanavir [12]),

there is an urgent need for improved drugs against HIV

protease because of increasing viral resistance and unfavor￾able pharmacokinetic profiles [13–16].

Our research group has utilized carbohydrates as building

blocks in the design and synthesis of C2-symmetric protease

inhibitors. The applied method of synthesis produces a

symmetry core unit with the C2-symmetry axis in the center

of an asymmetric inhibitor using L-mannaric acid as the

building block [17–20]. Subsequent benzylation and coup￾ling with amino acid or amines gave a series of symmetric or

asymmetric diol-based inhibitors which were further opti￾mized on the P1/P1¢ and P2/P2¢ side groups, providing a

variety of inhibitors with efficient antiviral profiles [21–26].

This class of protease inhibitors has previously been

associated with poor absorption profiles in cell assays and

unsatisfactory pharmacokinetics in rats, which led us to

investigate the effect of fluoro-substituted inhibitors on cell

absorption. The substitution of fluorine for hydrogen

introduces a minor increase in molecular mass and minimal

steric changes accompanied by increased lipophilicity

(Table 1) [27–29]. Previously, these properties of fluorine

have been utilized successfully in the development of

receptor-subtype-selective cholinergic and adrenergic drugs

[30–32]. To study these effects of fluorine on symmetric diol￾based protease inhibitors, we synthesized a series of fluoro

inhibitors, with either mono- or di-substituted P1/P1¢-

benzyloxy side groups [33].

Correspondence to T. Unge, Department of Cell and Molecular

Biology, BMC, Box 596, Uppsala University, SE-751 24, Uppsala,

Sweden. Fax: +46 18 530396, Tel.: +46 18 4714985,

E-mail: [email protected]

Enzyme: HIV-1 protease, POL_HV1B1 (P03366) (EC 3.4.23.16).

Note: The refined coordinates and associated structure factors of

HIV-1 protease in complex with inhibitors 1–6 have been deposited

in the RCSB Protein Data Bank with accession codes: 1EBY, 1EC0,

1W5V, 1W5W, 1W5X, and 1W5Y.

(Received 20 August 2004, revised 28 September 2004,

accepted 12 October 2004)

Eur. J. Biochem. 271, 4594–4602 (2004)
FEBS 2004 doi:10.1111/j.1432-1033.2004.04431.x