Protide

Last updated
General representation of a ProTide Protide-general.png
General representation of a ProTide

The ProTide technology is a prodrug approach used in molecular biology and drug design. It is designed to deliver nucleotide analogues (as monophosphate) into the cell (ProTide: PROdrug + nucleoTIDE). This technology was invented by Professor Chris McGuigan from the School of Pharmacy and Pharmaceutical Sciences at Cardiff University in the early 1990s. ProTides form a critical part of antiviral drugs such as sofosbuvir, tenofovir alafenamide, and remdesivir. [1]

Contents

Development

A generic diaryl phosphate pro-drug of AZT Azt-diarylphopsphate.png
A generic diaryl phosphate pro-drug of AZT

The first demonstration of the ProTide approach was made in 1992, when the efficiency of aryloxy phosphates and phosphoramidates was noted. [2] In particular, diaryl phosphates were prepared from zidovudine (AZT) using simple phosphorochloridate chemistry. For the first time, the anti-HIV activity of these phosphate derivatives of AZT exceeded that of the parent nucleoside in some cases. Moreover, while AZT was almost inactive (EC50 100μM) in the JM cell line, the substituted diaryl phosphate was 10 times more active (EC50 10μM). At the time, JM was considered AZT-insensitive due to poor phosphorylation. It later emerged that an AZT-efflux pump was the source of this poor AZT sensitivity. However, the conclusion remains valid that the diaryl phosphate was more able to retain activity in the JM cell line and that this may imply a (small) degree of intracellular phosphate delivery. The electron-withdrawing power of the p-nitro groups and putative enhancements in aryl leaving group ability were suggested as the major driving force of this SAR.

AZT protide AZT-ProTide.png
AZT protide

Subsequently, a series of aryloxy phosphoramidates of AZT were prepared with various p-aryl substituents and several amino acids. [3] These compounds were studied exclusively in the AZT-resistant JM cell line to explore potential (implied) AZT-monophosphate release, with the alanine phosphoramidate proving to be exceptionally effective. In all HIV-1 infected JM cultures, AZT was inhibited at a concentration of 100μM, while the phenyl methoxy alaninyl phosphoramidate was active at 0.8μM. This was taken as the first evidence of a successful nucleotide delivery. It was also noted that in other series, there was a marked preference for alanine over leucine (10-fold) and glycine (>100-fold). Furthermore, although electron-withdrawing aryl substitution had proven highly effective in diaryl systems, it was detrimental in this context. Para fluoro substitution had a slight adventitious effect, but not significantly so, while para-nitro substitution led to a 100-fold loss of activity. In a subsequent study, the range of aryl substituents was expanded, and compounds were tested in both TK+ (thymidine kinase competent) and TK- (thymidine kinase deficient) cell lines. None of the phosphoramidates retained the high (2–4 nM) potency of AZT in TK-competent cell lines (CEM and MT-4) against either HIV-1 or HIV-2. [4] However, while AZT lost all of its activity in the TK- deficient cell line CEM/TK-, most of the phosphoramidates retained antiviral activity, thus being ca >10–35-fold more active than AZT in this assay. Again, alanine emerged as an important component, with the glycine analogue being inactive in HIV-infected CEM/TK- all cultures. In this assay, leucine and phenylalanine were as effective as alanine, although they were less so in CEM/TK+ assays. Thus, the parent phenyl methoxy alanyl phosphoramidate emerged as an important lead compound.

Phenyl isoproyl Ala Abacavir monophosphate Abacavir-protide.png
Phenyl isoproyl Ala Abacavir monophosphate

Stavudine (d4T) was an early application of the ProTide approach. [5] This was a rational choice based on the known kinetics of phosphorylation of d4T. Thus, while the second phosphorylation (AZT-monophosphate to AZT-diphosphate) but not the first phosphorylation (AZT to AZT-monophosphate) is regarded as rate limiting for AZT activation to the triphosphate, the first step (d4T to d4T monophosphate) is thought in general to be the slow step for d4T. Thus, an intracellular (mono)nucleotide delivery should have a maximal impact for d4T and similar nucleosides. In the first instance (halo)alkyloxy phosphoramidates of d4T were prepared and found to retain activity in d4T-resistant JM cells. The activity was dependent on the haloalkyl group; the parent propyl system was poorly active. Subsequent studies in HIV-infected CEM/TK- cell cultures revealed the aryloxy phosphoramidates of d4T to be highly effective and, notably, to retain their full activity in CEM/TK- cells. In this study the benzyl ester emerged as slightly more potent than the parent methyl compound, being almost 10-times more active than d4T in CEM/TK+ assays and thus ca 300-500 fold more active than d4T, in CEM/TK- assays.

Current applications

The ProTide pro-drugs are useful for delivering phosphonate containing drugs to cell types with high expression of CTSA and CES1, such as immune cells. Tenofovir alafenamide is a successful example of this iteration. ProTides are also useful for nucleoside analogues that do not get phosphorylated efficiently by endogenous nucleoside kinases. For the nucleoside GS-334750, the parent of sofosbuvir, phosphorylation by nucleoside kinases is effectively nilled, and the only way to deliver active nucleotide is through ProTide. A major limitation of ProTides is that they require an expression of esterases like CTSA and CES1, which is very high in some cell types like hepatocytes and plays to an advantage for the treatment of Hepatitis C of Sofosbuvir.

Sofosbuvir Sofosbuvir.svg
Sofosbuvir

Extensive studies followed on these promising d4T derivatives and the ProTide technology was successfully applied to a wide range of nucleoside analogues. [6] [7] In particular, the ProTide approach has been used on several clinically evaluated anti-HCV nucleoside analogues, including the 2013 FDA approved compound sofosbuvir, and the 2016 FDA approved compound, Tenofovir alafenamide. Remdesivir also uses the ProTide prodrug technology (self-immolation is the key principle of ProTide nucleotide prodrugs [8] ). Because GS-441524 nucleoside can be phosphorylated and activated, some researchers have argued that the Protide application is an unnecessary complication in Remdesivir's design and that the parent nucleoside would be a cheaper and more effective COVID-19 drug. [9] [10]

ProTides have been tested to deliver key phosphorylated metabolites in inborn errors of metabolism, such as phosphopantothenate for PANK2 deficiency, however these were a clinical failure. [11]

Related Research Articles

<span class="mw-page-title-main">Nucleotide</span> Biological molecules constituting nucleic acids

Nucleotides are organic molecules composed of a nitrogenous base, a pentose sugar and a phosphate. They serve as monomeric units of the nucleic acid polymers – deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), both of which are essential biomolecules within all life-forms on Earth. Nucleotides are obtained in the diet and are also synthesized from common nutrients by the liver.

<span class="mw-page-title-main">Nucleoside</span> Any of several glycosylamines comprising a nucleobase and a sugar molecule

Nucleosides are glycosylamines that can be thought of as nucleotides without a phosphate group. A nucleoside consists simply of a nucleobase and a five-carbon sugar whereas a nucleotide is composed of a nucleobase, a five-carbon sugar, and one or more phosphate groups. In a nucleoside, the anomeric carbon is linked through a glycosidic bond to the N9 of a purine or the N1 of a pyrimidine. Nucleotides are the molecular building blocks of DNA and RNA.

<span class="mw-page-title-main">Thymidine</span> Chemical compound

Thymidine, also known as deoxythymidine, deoxyribosylthymine, or thymine deoxyriboside, is a pyrimidine deoxynucleoside. Deoxythymidine is the DNA nucleoside T, which pairs with deoxyadenosine (A) in double-stranded DNA. In cell biology it is used to synchronize the cells in G1/early S phase. The prefix deoxy- is often left out since there are no precursors of thymine nucleotides involved in RNA synthesis.

<span class="mw-page-title-main">Zidovudine</span> Antiretroviral medication

Zidovudine (ZDV), also known as azidothymidine (AZT), was the first antiretroviral medication used to prevent and treat HIV/AIDS. It is generally recommended for use in combination with other antiretrovirals. It may be used to prevent mother-to-child spread during birth or after a needlestick injury or other potential exposure. It is sold both by itself and together as lamivudine/zidovudine and abacavir/lamivudine/zidovudine. It can be used by mouth or by slow injection into a vein.

Reverse-transcriptase inhibitors (RTIs) are a class of antiretroviral drugs used to treat HIV infection or AIDS, and in some cases hepatitis B. RTIs inhibit activity of reverse transcriptase, a viral DNA polymerase that is required for replication of HIV and other retroviruses.

<span class="mw-page-title-main">Zalcitabine</span> Chemical compound

Zalcitabine, also called dideoxycytidine, is a nucleoside analog reverse-transcriptase inhibitor (NRTI) sold under the trade name Hivid. Zalcitabine was the third antiretroviral to be approved by the Food and Drug Administration (FDA) for the treatment of HIV/AIDS. It is used as part of a combination regimen.

A nucleoside triphosphate is a nucleoside containing a nitrogenous base bound to a 5-carbon sugar, with three phosphate groups bound to the sugar. They are the molecular precursors of both DNA and RNA, which are chains of nucleotides made through the processes of DNA replication and transcription. Nucleoside triphosphates also serve as a source of energy for cellular reactions and are involved in signalling pathways.

<span class="mw-page-title-main">Thymidine kinase</span> Enzyme found in most living cells

Thymidine kinase is an enzyme, a phosphotransferase : 2'-deoxythymidine kinase, ATP-thymidine 5'-phosphotransferase, EC 2.7.1.21. It can be found in most living cells. It is present in two forms in mammalian cells, TK1 and TK2. Certain viruses also have genetic information for expression of viral thymidine kinases. Thymidine kinase catalyzes the reaction:

<span class="mw-page-title-main">Nucleoside analogue</span> Biochemical compound

Nucleoside analogues are structural analogues of a nucleoside, which normally contain a nucleobase and a sugar. Nucleotide analogues are analogues of a nucleotide, which normally has one to three phosphates linked to a nucleoside. Both types of compounds can deviate from what they mimick in a number of ways, as changes can be made to any of the constituent parts. They are related to nucleic acid analogues.

<span class="mw-page-title-main">Cyclic nucleotide phosphodiesterase</span> Class of enzymes

3′,5′-cyclic-nucleotide phosphodiesterases (EC 3.1.4.17) are a family of phosphodiesterases. Generally, these enzymes hydrolyze a nucleoside 3′,5′-cyclic phosphate to a nucleoside 5′-phosphate:

<span class="mw-page-title-main">Nucleic acid metabolism</span> Process

Nucleic acid metabolism is a collective term that refers to the variety of chemical reactions by which nucleic acids are either synthesized or degraded. Nucleic acids are polymers made up of a variety of monomers called nucleotides. Nucleotide synthesis is an anabolic mechanism generally involving the chemical reaction of phosphate, pentose sugar, and a nitrogenous base. Degradation of nucleic acids is a catabolic reaction and the resulting parts of the nucleotides or nucleobases can be salvaged to recreate new nucleotides. Both synthesis and degradation reactions require multiple enzymes to facilitate the event. Defects or deficiencies in these enzymes can lead to a variety of diseases.

<span class="mw-page-title-main">Deoxycytidine kinase</span> Protein-coding gene in the species Homo sapiens

Deoxycytidine kinase (dCK) is an enzyme which is encoded by the DCK gene in humans. dCK predominantly phosphorylates deoxycytidine (dC) and converts dC into deoxycytidine monophosphate. dCK catalyzes one of the initial steps in the nucleoside salvage pathway and has the potential to phosphorylate other preformed nucleosides, specifically deoxyadenosine (dA) and deoxyguanosine (dG), and convert them into their monophosphate forms. There has been recent biomedical research interest in investigating dCK's potential as a therapeutic target for different types of cancer.

<span class="mw-page-title-main">Organic anion transporter 1</span> Protein-coding gene in the species Homo sapiens

The organic anion transporter 1 (OAT1) also known as solute carrier family 22 member 6 (SLC22A6) is a protein that in humans is encoded by the SLC22A6 gene. It is a member of the organic anion transporter (OAT) family of proteins. OAT1 is a transmembrane protein that is expressed in the brain, the placenta, the eyes, smooth muscles, and the basolateral membrane of proximal tubular cells of the kidneys. It plays a central role in renal organic anion transport. Along with OAT3, OAT1 mediates the uptake of a wide range of relatively small and hydrophilic organic anions from plasma into the cytoplasm of the proximal tubular cells of the kidneys. From there, these substrates are transported into the lumen of the nephrons of the kidneys for excretion. OAT1 homologs have been identified in rats, mice, rabbits, pigs, flounders, and nematodes.

Discovery and development of nucleoside and nucleotide reverse-transcriptase inhibitors began in the 1980s when the AIDS epidemic hit Western societies. NRTIs inhibit the reverse transcriptase (RT), an enzyme that controls the replication of the genetic material of the human immunodeficiency virus (HIV). The first NRTI was zidovudine, approved by the U.S. Food and Drug Administration (FDA) in 1987, which was the first step towards treatment of HIV. Six NRTI agents and one NtRTI have followed. The NRTIs and the NtRTI are analogues of endogenous 2´-deoxy-nucleoside and nucleotide. Drug-resistant viruses are an inevitable consequence of prolonged exposure of HIV-1 to anti-HIV drugs.

<span class="mw-page-title-main">Tenofovir alafenamide</span> Chemical compound

Tenofovir alafenamide, sold under the brand name Vemlidy, is an antiviral medication used against hepatitis B and HIV. It is used for the treatment of chronic hepatitis B virus (HBV) infection in adults with compensated liver disease and is given in combination with other medications for the prevention and treatment of HIV. It is taken by mouth.

<span class="mw-page-title-main">Sofosbuvir</span> Chemical compound

Sofosbuvir, sold under the brand name Sovaldi among others, is a medication used to treat hepatitis C. It is taken by mouth.

Janet Rideout is an organic chemist and one of the scientists who discovered that azidothymidine (AZT) could be used as an antiretroviral agent to treat Human Immunodeficiency Virus (HIV). She also played a key role in the development of acyclovir, the first effective treatment for herpes simplex virus.

<span class="mw-page-title-main">NS5B inhibitor</span> Class of pharmaceutical drugs

Non-structural protein 5B (NS5B) inhibitors are a class of direct-acting antivirals widely used in the treatment of chronic hepatitis C. Depending on site of action and chemical composition, NS5B inhibitors may be categorized into three classes—nucleoside active site inhibitors (NIs), non-nucleoside allosteric inhibitors, and pyrophosphate analogues. Subsequently, all three classes are then subclassified. All inhibit RNA synthesis by NS5B but at different stages/sites resulting in inability of viral RNA replication. Expression of direct-acting NS5B inhibitors does not take place in cells that are not infected by hepatitis C virus, which seems to be beneficial for this class of drugs.

<span class="mw-page-title-main">GS-441524</span> Metabolite of remdesivir

GS-441524 is a nucleoside analogue antiviral drug which was developed by Gilead Sciences. It is the main plasma metabolite of the antiviral prodrug remdesivir, and has a half-life of around 24 hours in human patients. Remdesivir and GS-441524 were both found to be effective in vitro against feline coronavirus strains responsible for feline infectious peritonitis (FIP), a lethal systemic disease affecting domestic cats. Remdesivir was never tested in cats, but GS-441524 has been found to be effective treatment for FIP.

Katherine Seley-Radtke is an American medicinal chemist who specializes in the discovery and design of novel nucleoside or nucleotide based enzyme inhibitors that may be used to treat infections or cancer. She has authored over 90 peer-reviewed publications, is an inventor of five issued US patents, and is a professor in the department of chemistry and biochemistry at the University of Maryland, Baltimore County. Her international impact includes scientific collaborations, policy advising and diplomatic appointments in biosecurity efforts.

References

  1. ↑ Mehellou Y, Rattan HS, Balzarini J (March 2018). "The ProTide Prodrug Technology: From the Concept to the Clinic". Journal of Medicinal Chemistry. 61 (6): 2211–2226. doi:10.1021/acs.jmedchem.7b00734. PMC   7075648 . PMID   28792763.
  2. ↑ McGuigan C, Pathirana RN, Mahmood N, Devine KG, Hay AJ (April 1992). "Aryl phosphate derivatives of AZT retain activity against HIV1 in cell lines which are resistant to the action of AZT". Antiviral Research. 17 (4): 311–321. doi:10.1016/0166-3542(92)90026-2. PMID   1642482.
  3. ↑ McGuigan C, Pathirana RN, Mahmood N, Hay AJ (July 1992). "Aryl phosphate derivates of AZT inhibit HIV replication in cells where the nucleoside is poorly active". Bioorganic & Medicinal Chemistry Letters. 2 (7): 701–704. doi:10.1016/S0960-894X(00)80395-9.
  4. ↑ McGuigan C, Pathirana RN, Balzarini J, De Clercq E (April 1993). "Intracellular delivery of bioactive AZT nucleotides by aryl phosphate derivatives of AZT". Journal of Medicinal Chemistry. 36 (8): 1048–1052. doi:10.1021/jm00060a013. PMID   8478904.
  5. ↑ McGuigan C, Sheeka HM, Mahmood N, Hay A (June 1993). "Phosphate derivatives of d4T as inhibitors of HIV". Bioorganic & Medicinal Chemistry Letters. 3 (6): 1203–1206. doi:10.1016/S0960-894X(00)80315-7.
  6. ↑ Cahard D, McGuigan C, Balzarini J (May 2004). "Aryloxy phosphoramidate triesters as pro-tides". Mini Reviews in Medicinal Chemistry. 4 (4): 371–381. doi:10.2174/1389557043403936. PMID   15134540.
  7. ↑ McGuigan C, Harris SA, Daluge SM, Gudmundsson KS, McLean EW, Burnette TC, et al. (May 2005). "Application of phosphoramidate pronucleotide technology to abacavir leads to a significant enhancement of antiviral potency". Journal of Medicinal Chemistry. 48 (10): 3504–3515. doi:10.1021/jm0491400. PMID   15887959.
  8. ↑ Procházková E, Šimon P, Straka M, Filo J, Májek M, Cigáň M, Baszczyňski O (January 2021). "Phosphate linkers with traceable cyclic intermediates for self-immolation detection and monitoring". Chemical Communications. 57 (2): 211–214. doi:10.1039/D0CC06928K. PMID   33300900. S2CID   228087795.
  9. ↑ Yan VC, Muller FL (14 May 2020). "Gilead should ditch remdesivir and focus on its simpler ancestor". STAT.
  10. ↑ Yan VC, Muller FL (July 2020). "Advantages of the Parent Nucleoside GS-441524 over Remdesivir for Covid-19 Treatment". ACS Medicinal Chemistry Letters. 11 (7): 1361–1366. doi:10.1021/acsmedchemlett.0c00316. PMC   7315846 . PMID   32665809.
  11. ↑ Keown A (22 August 2019). "Retrophin's Late-Stage PKAN Drug Fails Against Placebo". BioSpace.
This page is based on this Wikipedia article
Text is available under the CC BY-SA 4.0 license; additional terms may apply.
Images, videos and audio are available under their respective licenses.