UMass Chem 423 Student Projects 2011-1

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Student projects continued below:
Student projects continued below:
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== '''p38 kinase''' ==
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{{STRUCTURE_1a9u | PDB=1a9u | SCENE= }}
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P38 kinase belongs to one of the four subgroups of mitogen-activated protein (MAP) kinases. MAP kinases respond to extracellular stimuli by a signaling cascade leading to intracellular responses. Therefore, MAP kinases function to regulate fundamental cellular processes [1-5].
 +
 +
The p38 subgroup consists of four isoforms: p38α, p38β, p38γ, and p38δ [1-3]. Of these isoforms, p38α and p38β have the most similar amino acid sequences and both forms are expressed in most cell types. P38γ is mostly found in skeletal muscle, while p38δ is only found in the skin, small intestine, pancreas, and kidney [1, 2]. Since p38α was first discovered, most publications focus on this isoform and refer to p38α as p38 [2]. However, all four isoforms have the Thr-Glu-Tyr dual phosphorylation site in the regulatory loop. The substrate specificity of p38 is controlled Glu residue in this dual phosphorylation motif and the length of the loop [1]. This specificity is important for the signal cascade generated in response to stimuli.
 +
 +
Operating as a signal transduction mediator, p38 is activiated by both stress and mitogen stimuli. Environmental stress, particularly UV radiation and osmotic shock, cause an increase activity level of p38. Also, p38 is activated by pro-inflammatory cytokines, especially tumor necrosis factor (TNF) and interleukin-1 (IL1) [3]. However, activation of p38 depends on both the stimulus and the cell type [1]. Dual phosphorylation on the Thr and Tyr is necessary for p38 activation. This dual phosphorylation motif is common in all members of the MAP kinase family. Upstream kinases, which are the MAP kinase kinases (mkks), are responsible for p38 activation [1-3]. Due to selective activation, each p38 isoform requires distinct mkks. Further upstream activators of the MKK/p38 pathway are widely diversified. This cascade accounts for the various stimuli that lead to activating the p38 pathway [1]. Dephosphorylation by dual phosphatases is responsible for the major of the downregulation of p38 [1,3].
 +
 +
The activation of p38 pathway leads to the activation of downstream substrates, such as protein kinases and transcription factors. The p38 pathway regulates close to a hundred genes. P38 is associated with the expression of many cytokines, transcription factors, and cell surface receptors [1]. Various proteins that control transcription and translation are targeted, either directly or indirectly, by p38 kinases. Biological results of p38 activation include inflammation, apoptosis, cell cycle, and cell differentiation [1-3]. However, the role of p38 is specific to cell type [1].
 +
 +
The role of the p38 pathway in cellular inflammation places p38 as a key therapeutic target for inflammatory diseases, cancer, and other diseases. Therefore, p38 inhibitors are key therapeutic agents for the treatment of such diseases. Pyridinyl imidazoles, especially SB203580 (ligand shown in the Jmol diagram), inhibit the catalytic activity of p38 by binding to the ATP site [5]. The ATP binding site provides specificity necessary for highly selective pyridinyl imidazole inhibitors. The inhibitors for p38 do not bind to structurally similar MAP kinases. Other structural factors, such an unique pocket in p38 for the fluorophenyl ring of some pyridinyl imidazole inhibitors, contribute to the selectively of the inhibitor [4]. SB203580 and related inhibitors binds with about equal affinity to both the activated and inactivated forms of p38 kinase. Therefore, binding of the inhibitor can lock p38 into an inactivated conformation.
 +
 +
Gleevec is a brand name drug that targets p38. Gleevec, which is imatinib mesylate, is an inhibitor of p38. Imatinib mesylate, chemically designated as 4-[(4-Methyl-1-piperazinyl)methyl]-N-[4-methyl-3-[[4-(3-pyridinyl)-2-pyrimidinyl]amino]-phenyl]benzamide methanesulfonate [6], is structurally similar to SB203580, 4-[5-(4-fluoro-phenyl)-2-(4-methanesulfinyl-phenyl)-3h-imidazol-4-yl]-pyridine [7]. Gleevec is used to treat certain cancers, including chronic myeloid leukemia, gastrointestinal stromal tumors, and myelodysplastic/myeloproliferative diseases. Gleevec inhibits p38, preventing the proliferation of cancer cells [6].
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=== Overall Structure ===
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Kinase <scene name='Sandbox713/P38_kinase/1'>p38</scene> is a single 351 amino acid polypeptide chain made up of 10 alpha helixes and 10 beta strands. Kinase p38 is composed of two domains. The first is a 135 residue N-terminal domain and the second a 225 residue C-terminal domain. The beta strands in light blue form antiparallel beta sheets and are located mainly towards the N-terminus while the alpha helixes in green are located mainly towards the C-terminus. In this rainbow representation of <scene name='Sandbox713/Termini/1'>Kinase p38</scene> the N and C termini are found at the top of the protein. The catalytic site where the drug binds is located between the two domains.
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=== Drug Binding Site ===
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The p38 kinase-SB2 complex binding chemistry is analyzed in this section. The <scene name='Sandbox713/Hbonds/1'>ligand</scene> is connected to the binding site by a hydrogen bond with Met109 and the cyclopropylmethyl group binds to the phosphate-binding ribbon in a depression formed by Val30 and Val38. The complex is also stabilized by <scene name='Sandbox713/Other_interactions/1'>more distant bonds</scene> contributing from Lys53 and Val105. The conformation of the phosphate-binding ribbon of the p38 kinase changes significantly in order to bind with SB2.
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=== Additional Features===
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Kinase p38 has a <scene name='Sandbox713/Large_lobe_and_small_lobe/1'>large lobe and small lobe</scene> between (also known as domains), which the inhibitors can bind. Blocking p38 kinase may be an valuable way of treating many inflammatory diseases. The pyridinylimidazole inhibitors bind in the ATP binding site, making it competitive against ATP. There has also been observed an allosteric binding site in the Asp-Phe-Gly motif in the active site on p38 kinase for a “diaryl urea class of highly potent and selective inhibitors”. These inhibitors have a new binding mechanism, which does not directly compete with
 +
<scene name='Sandbox713/Allosteric_binding_pocket/1'>ATP</scene>. BIRB 796 is an inhibitor, which has a 12,000-fold increase in binding affinity compared to the previous inhibitor. Structural changes that made such an increase in binding affinity are: methyl substituent on pyrazole ring replaced with tolyl group, chlorophenyl group replaced with naphthyl moiety, and addition of ethoxymorpholine subsituent on paththyl ring. Tolyl has hydrophobic interactions with with side chains of Glu 71 side chain. Glu 71 has one hydrogen bond with the NH on urea. This new conformation is better for supporting the hydrophobic interactions on the tolyl group with the inhibitor. Information about the structure of p38 kinase and its interaction with inhibitors is useful for research about reducing inflammation and diseases such as rheumatoid arthritis. New inhibitors are being designed an optimized to help fight inflammation and other diseases such as rheumatoid arthritis.
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=== Credits ===
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Introduction - Sarena Horava
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Overall Structure - Robert Nathan
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Drug Binding Site - Nick Cadirov
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Additional Features - Inna Brockman
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=== References===
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1. Ono, K.; J. Han, The p83 signal transduction pathway activation and function, Cellular Signalling 12 (2000) 1-13.
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2. Min, L.; B. He; L. Hui, Mitogen-activated protein kinases in hepatocellullar carcinoma development, Seminars in Cancer Biology 21 (2011) 10-20.
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3. Raingeaud, J; S. Gupta, et al. Pro-inflammatory cytokines and environmental stress cause p38 mitogen-activated protein kinase activation by dual phosphorylation on tyrosine and threonine, The Journal of Biological Chemistry 270 (1995) 7420-7426.
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4. Wang, Z.; Canagarajah, et al. Structural basis of inhibitor selectivity in MAP kinases, Structure 6 (1998) 1117-1128.
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 +
5. Young, P. R.; M. M. McLaughlin, et al. Pyridinyl imidazole inhibitors of p38 mitogen-activated protein kinase bind in the ATP site, The Journal of Biological Chemistry 272 (1997) 12116-12121.
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6. Gleevec http://www.rxlist.com/gleevec-drug-center.htm
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7. Ligand Summary for SB2 http://www.pdb.org/pdb/ligand/ligandsummary.do?hetId=SB2&sid=1A9U
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http://www.pdb.org/pdb/explore/explore.do?structureId=1A9U
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S. Pav, D. M. White, S. Rogers, K. M. Crane, C. L. Cywin, W. Davidson, J. Hopkins, M. L. Brown, C. A. Pargellis & L. Tong. (1997). Crystallization and preliminary crystallographic analysis of recombinant human p38 MAP kinase. Protein Science, 6, 242-245.
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 +
L. Tong, S. Pav, D. M. White, S. Rogers, K. M. Crane, C. L. Cywin, M. L. Brown & C. A. Pargellis. (1997). A highly specific inhibitor of human p38 MAP kinase binds in the ATP pocket. Nature Struct. Biol. 4, 311-316.
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C. Pargellis, L. Tong, L. Churchill, P.F. Cirillo, T. Gilmore, A.G. Graham, P.M. Grob, E.R. Hickey, N. Moss, S. Pav & J. Regan. (2002). Inhibition of p38 MAP kinase by utilizing a novel allosteric binding site. Nature Struct. Biol. 9, 268-272.
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 +
J. Regan, S. Breitfelder, P. Cirillo, T. Gilmore, A.G. Graham, E. Hickey, B. Klaus, J. Madwed, M. Moriak, N. Moss, C. Pargellis, S. Pav, A. Proto, A. Swanimer, L. Tong & C. Torcellini. (2002). Pyrazole urea-based inhibitors of p38 MAP kinase: From lead compound to clinical candidate. J. Med. Chem. 45, 2994-3008.
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 +
Drug Binding Site:
 +
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Zhulun Wang, Bertram J Canagarajah, Jeffrey C Boehm, Skouki Kassisà, Melanie H Cobb, Peter R Young, Sherin Abdel-Meguid, Jerry L Adams and Elizabeth J Goldsmith. (1998). Structural basis of inhibitor selectivity in MAP kinases. Structure, 6, 1117-1128.
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Revision as of 22:04, 13 February 2012

Spring 2011 Chem423 Team Projects: Understanding Drug Mechanisms Instructions posted here: Student Projects for UMass Chemistry 423 Spring 2011 Student projects continued below:


Contents

p38 kinase

PDB ID 1a9u

Drag the structure with the mouse to rotate
1a9u, resolution 2.50Å ()
Ligands:
Resources: FirstGlance, OCA, PDBsum, RCSB
Coordinates: save as pdb, mmCIF, xml



P38 kinase belongs to one of the four subgroups of mitogen-activated protein (MAP) kinases. MAP kinases respond to extracellular stimuli by a signaling cascade leading to intracellular responses. Therefore, MAP kinases function to regulate fundamental cellular processes [1-5].

The p38 subgroup consists of four isoforms: p38α, p38β, p38γ, and p38δ [1-3]. Of these isoforms, p38α and p38β have the most similar amino acid sequences and both forms are expressed in most cell types. P38γ is mostly found in skeletal muscle, while p38δ is only found in the skin, small intestine, pancreas, and kidney [1, 2]. Since p38α was first discovered, most publications focus on this isoform and refer to p38α as p38 [2]. However, all four isoforms have the Thr-Glu-Tyr dual phosphorylation site in the regulatory loop. The substrate specificity of p38 is controlled Glu residue in this dual phosphorylation motif and the length of the loop [1]. This specificity is important for the signal cascade generated in response to stimuli.

Operating as a signal transduction mediator, p38 is activiated by both stress and mitogen stimuli. Environmental stress, particularly UV radiation and osmotic shock, cause an increase activity level of p38. Also, p38 is activated by pro-inflammatory cytokines, especially tumor necrosis factor (TNF) and interleukin-1 (IL1) [3]. However, activation of p38 depends on both the stimulus and the cell type [1]. Dual phosphorylation on the Thr and Tyr is necessary for p38 activation. This dual phosphorylation motif is common in all members of the MAP kinase family. Upstream kinases, which are the MAP kinase kinases (mkks), are responsible for p38 activation [1-3]. Due to selective activation, each p38 isoform requires distinct mkks. Further upstream activators of the MKK/p38 pathway are widely diversified. This cascade accounts for the various stimuli that lead to activating the p38 pathway [1]. Dephosphorylation by dual phosphatases is responsible for the major of the downregulation of p38 [1,3].

The activation of p38 pathway leads to the activation of downstream substrates, such as protein kinases and transcription factors. The p38 pathway regulates close to a hundred genes. P38 is associated with the expression of many cytokines, transcription factors, and cell surface receptors [1]. Various proteins that control transcription and translation are targeted, either directly or indirectly, by p38 kinases. Biological results of p38 activation include inflammation, apoptosis, cell cycle, and cell differentiation [1-3]. However, the role of p38 is specific to cell type [1].

The role of the p38 pathway in cellular inflammation places p38 as a key therapeutic target for inflammatory diseases, cancer, and other diseases. Therefore, p38 inhibitors are key therapeutic agents for the treatment of such diseases. Pyridinyl imidazoles, especially SB203580 (ligand shown in the Jmol diagram), inhibit the catalytic activity of p38 by binding to the ATP site [5]. The ATP binding site provides specificity necessary for highly selective pyridinyl imidazole inhibitors. The inhibitors for p38 do not bind to structurally similar MAP kinases. Other structural factors, such an unique pocket in p38 for the fluorophenyl ring of some pyridinyl imidazole inhibitors, contribute to the selectively of the inhibitor [4]. SB203580 and related inhibitors binds with about equal affinity to both the activated and inactivated forms of p38 kinase. Therefore, binding of the inhibitor can lock p38 into an inactivated conformation.

Gleevec is a brand name drug that targets p38. Gleevec, which is imatinib mesylate, is an inhibitor of p38. Imatinib mesylate, chemically designated as 4-[(4-Methyl-1-piperazinyl)methyl]-N-[4-methyl-3-[[4-(3-pyridinyl)-2-pyrimidinyl]amino]-phenyl]benzamide methanesulfonate [6], is structurally similar to SB203580, 4-[5-(4-fluoro-phenyl)-2-(4-methanesulfinyl-phenyl)-3h-imidazol-4-yl]-pyridine [7]. Gleevec is used to treat certain cancers, including chronic myeloid leukemia, gastrointestinal stromal tumors, and myelodysplastic/myeloproliferative diseases. Gleevec inhibits p38, preventing the proliferation of cancer cells [6].

Overall Structure

Kinase is a single 351 amino acid polypeptide chain made up of 10 alpha helixes and 10 beta strands. Kinase p38 is composed of two domains. The first is a 135 residue N-terminal domain and the second a 225 residue C-terminal domain. The beta strands in light blue form antiparallel beta sheets and are located mainly towards the N-terminus while the alpha helixes in green are located mainly towards the C-terminus. In this rainbow representation of the N and C termini are found at the top of the protein. The catalytic site where the drug binds is located between the two domains.

Drug Binding Site

The p38 kinase-SB2 complex binding chemistry is analyzed in this section. The is connected to the binding site by a hydrogen bond with Met109 and the cyclopropylmethyl group binds to the phosphate-binding ribbon in a depression formed by Val30 and Val38. The complex is also stabilized by contributing from Lys53 and Val105. The conformation of the phosphate-binding ribbon of the p38 kinase changes significantly in order to bind with SB2.

Additional Features

Kinase p38 has a between (also known as domains), which the inhibitors can bind. Blocking p38 kinase may be an valuable way of treating many inflammatory diseases. The pyridinylimidazole inhibitors bind in the ATP binding site, making it competitive against ATP. There has also been observed an allosteric binding site in the Asp-Phe-Gly motif in the active site on p38 kinase for a “diaryl urea class of highly potent and selective inhibitors”. These inhibitors have a new binding mechanism, which does not directly compete with . BIRB 796 is an inhibitor, which has a 12,000-fold increase in binding affinity compared to the previous inhibitor. Structural changes that made such an increase in binding affinity are: methyl substituent on pyrazole ring replaced with tolyl group, chlorophenyl group replaced with naphthyl moiety, and addition of ethoxymorpholine subsituent on paththyl ring. Tolyl has hydrophobic interactions with with side chains of Glu 71 side chain. Glu 71 has one hydrogen bond with the NH on urea. This new conformation is better for supporting the hydrophobic interactions on the tolyl group with the inhibitor. Information about the structure of p38 kinase and its interaction with inhibitors is useful for research about reducing inflammation and diseases such as rheumatoid arthritis. New inhibitors are being designed an optimized to help fight inflammation and other diseases such as rheumatoid arthritis.

Credits

Introduction - Sarena Horava

Overall Structure - Robert Nathan

Drug Binding Site - Nick Cadirov

Additional Features - Inna Brockman

References

1. Ono, K.; J. Han, The p83 signal transduction pathway activation and function, Cellular Signalling 12 (2000) 1-13.

2. Min, L.; B. He; L. Hui, Mitogen-activated protein kinases in hepatocellullar carcinoma development, Seminars in Cancer Biology 21 (2011) 10-20.

3. Raingeaud, J; S. Gupta, et al. Pro-inflammatory cytokines and environmental stress cause p38 mitogen-activated protein kinase activation by dual phosphorylation on tyrosine and threonine, The Journal of Biological Chemistry 270 (1995) 7420-7426.

4. Wang, Z.; Canagarajah, et al. Structural basis of inhibitor selectivity in MAP kinases, Structure 6 (1998) 1117-1128.

5. Young, P. R.; M. M. McLaughlin, et al. Pyridinyl imidazole inhibitors of p38 mitogen-activated protein kinase bind in the ATP site, The Journal of Biological Chemistry 272 (1997) 12116-12121.

6. Gleevec http://www.rxlist.com/gleevec-drug-center.htm

7. Ligand Summary for SB2 http://www.pdb.org/pdb/ligand/ligandsummary.do?hetId=SB2&sid=1A9U

http://www.pdb.org/pdb/explore/explore.do?structureId=1A9U

S. Pav, D. M. White, S. Rogers, K. M. Crane, C. L. Cywin, W. Davidson, J. Hopkins, M. L. Brown, C. A. Pargellis & L. Tong. (1997). Crystallization and preliminary crystallographic analysis of recombinant human p38 MAP kinase. Protein Science, 6, 242-245.

L. Tong, S. Pav, D. M. White, S. Rogers, K. M. Crane, C. L. Cywin, M. L. Brown & C. A. Pargellis. (1997). A highly specific inhibitor of human p38 MAP kinase binds in the ATP pocket. Nature Struct. Biol. 4, 311-316.

C. Pargellis, L. Tong, L. Churchill, P.F. Cirillo, T. Gilmore, A.G. Graham, P.M. Grob, E.R. Hickey, N. Moss, S. Pav & J. Regan. (2002). Inhibition of p38 MAP kinase by utilizing a novel allosteric binding site. Nature Struct. Biol. 9, 268-272.

J. Regan, S. Breitfelder, P. Cirillo, T. Gilmore, A.G. Graham, E. Hickey, B. Klaus, J. Madwed, M. Moriak, N. Moss, C. Pargellis, S. Pav, A. Proto, A. Swanimer, L. Tong & C. Torcellini. (2002). Pyrazole urea-based inhibitors of p38 MAP kinase: From lead compound to clinical candidate. J. Med. Chem. 45, 2994-3008.

Drug Binding Site:

Zhulun Wang, Bertram J Canagarajah, Jeffrey C Boehm, Skouki Kassisà, Melanie H Cobb, Peter R Young, Sherin Abdel-Meguid, Jerry L Adams and Elizabeth J Goldsmith. (1998). Structural basis of inhibitor selectivity in MAP kinases. Structure, 6, 1117-1128.


Rituximab Fab



Introduction

PDB ID 2osl

Drag the structure with the mouse to rotate
2osl, resolution 2.60Å ()
Resources: FirstGlance, OCA, RCSB, PDBsum
Coordinates: save as pdb, mmCIF, xml


Rituximab Fab is used as a prescribed drug to stop transplant rejection, to help regulate autoimmune diseases and to fight cancer. Rituximab has become a key proponent in treatment for cancers such as lymphoma and leukemia and has become the most used chimeric (mouse/human) antibody of its kind. Developed in 1986 by Ivor Royston and Howard Birndorf, Rituximab became FDA approved for clinical use in 1997. Since then Rituximab has become the routine treatment for non-hodgkins lymphoma that otherwise proves to be resistant to certain forms of chemotherapy. Recently, in 2010, Rituximab was approved by the European Commission for use in fighting follicular lymphoma. Because of the wide range of effects that Rituximab causes through its interactions with the CD20 protein, many uses for this antibody have been discovered.


Rituximab is part of a family of antibodies called chimaric monochromal antibodies. These types of antibodies are developed from a single type of parent cell and specifically bind to a single host cell. These types of antibodies do not vary between each other and always interact in the same manner with the chosen cell. Each one of these antibodies is a mixture, chimera, of both rat and human cells both of which play the same role in the host organism. Being chimaric allows for testing on mice before implementation on human cells and combination of human antibodies with mice antibodies makes for simple transitioning between lab testing and clinical testing because the known antibodies have been linked together without making large assumptions with no research.


In brief, Rituximab acts on both benign and malignant lymphocytes, or B-cells. B-cells create antibodies when bombarded with foreign antigens. They are produced in the bone-marrow of humans and when acting normally, help the body fight off disease. If these B-cells are over-produced, under-produced, act abnormally, or are dysfunctional, many different diseases may arise as previously listed. Rituximab acts on the CD20 protein that is found on almost all B-cells. this CD20 protein is unchanged throughout the life of the B-cell.


Multiple proposed mechanisms of action may occur when Rituximab interacts with a lymphocyte (B-cell). The first is known as antibody-dependent cell-mediated cytotoxicity (ADCC) which allows Rituximab to trigger natural cell lysing mechanisms through the use of T-cells and other macrophages. In this proposed mechanism, Rituximab performs as a label to begin cell destruction. Upon further investigation, complement-dependent-cytotoxicity (CDC) became another possible mechanism of action. Through this mechanism, complement proteins are called upon to destroy cell membranes causing overall lysis of the cell. These complement proteins are again naturally occurring, yet usually rely on naturally occurring antibodies to be guided towards non-self antigens. Instead, rituximab acts as the antibody guiding these complements specifically to B-cells. The third and final possible mechanism of action occurs through apoptosis, a more direct route to cell death. Upon initial interaction with the B-cell, the cell triggers programmed cell death. Through this process the cell self-destructs and is immediately destroyed. All three of these mechanisms have been looked into with some uncertainty as to which most likely occurs. Scientists have witnessed many results from treatment such as the shedding of CD23, the down regulation of B-cell receptors, and many other effects that both prove and disprove all three proposed mechanisms.


Rituximab has proven to be a very useful antibody in fighting many human disease, through looking closely at how Rituximab may interact with B-cells, further knowledge about its possible uses will result. This article put together by Chemistry Undergraduate Students at the University of Massachusetts Amherst, will analyze and compile evidence of the chemical mechanisms, binding sites, and applications of Rituximab, a highly used drug in medicine.

Structure

PDB ID 2osl

Drag the structure with the mouse to rotate
2osl, resolution 2.60Å ()
Resources: FirstGlance, OCA, RCSB, PDBsum
Coordinates: save as pdb, mmCIF, xml



Rituximab is made up of four amino acid chain subunits. Two of these are considered major subunits and two are minor subunits. The two major amino acid sequences are made up of 451 amino acids and the two minor chains contain 213 amino acids. This large protein binds to the surface the surface of B-cells and therefore is exposed primarily to the aqueous external environment. To have stable interactions with the environment, the protein is largely polar, composed almost entirely of beta sheets.

The beta sheets formed in the protein’s native structure form two distinct sides of this protein. Each of these sides is made of woven beta strands that form anti-parallel sheets. The binding site of the protein is found within one of these beta-strand sheets.

Rituximab interacts the surface of its cell membrane through a alpha helix region. This is actually done in two places, where alpha helices of non-polar amino acids are found. Each of these is found at the terminal end of a beta sheet, allowing the active site to bind to the substrate elsewhere on the cell surface.

Each of these structures can be seen in the Jmol native structure found to the right. The red and blue beta strands are the , while the light and dark green strands are the . Each of these has an alpha helix structure, which is shaded in a slightly different color to emphasize its position.

Drug Binding Site

PDB ID 2osl

Drag the structure with the mouse to rotate
2osl, resolution 2.60Å ()
Resources: FirstGlance, OCA, RCSB, PDBsum
Coordinates: save as pdb, mmCIF, xml


Rituximab (Rituximab Fab) is a chimeric antibody with human IgG1 used in the therapy of non-Hodgkin’s B cell lymphomas. This antibody targets B cells by binding to the cell-surface receptor, CD20. CD20 (human B-lymphocyte-restricted differentiation antigen, Bp35) is a hydrophobic transmembrane protein with a molecular weight of approximately 35 kD located on pre-B and mature B lymphocytes. Rituximab has a binding affinity for the CD20 antigen of approximately 8.0 nM, which is similar to the parent murine antibody, 2B8. The amino acid residues alanine (170) and proline (172) within the extracellular loop of CD20 are critical for rituximab binding. Selection of random libraries yielded 2 distinct peptides binding Rituximab: 1 peptide was homologous to alanine (170)-proline(172), the other was assumed to mimic the same epitope.



Of Rituximab Fab, the Fab (fragment antigen-binding) is the region in which the antibody is bound to the antigen. A heavy and light chain domain located at the terminal end of the monomer shapes the binding site of Rituximab Fab. Each one of these chains (heavy and light) has a constant and variable portion. The constant chain is identical in all antibodies; the variable chain is unique to each specific B-cell antibodies. The heavy chains of Rituximab Fab are made up of approximately 451 amino acids and the light chains by approximately 213 amino acids.


Rituximab can begin complement activation by binding of Clq to the Fc region (region of an antibody that interacts with cell surface receptors) of an antibody when inducing complement-mediated cell lysis. C1q is a 400kDa protein split into 3 subunits. Each subunit consists of Y-shaped triple peptide helices joined at the stem, forming a globular non-helical head at its end. The helical components of the structure contain varied strands of Glycine, proline, isoleucine and/or hydroxylysine. The globular heads of the structure (along with two serine proteases, Clr and Cls) are then responsible for the multivalent attachment of the C1q structure; forming the complex C1 which triggers the compliment cascade that performs cellular lysis.

Additional Effects

Rituximab is used quite frequently to treat dysfunctional leukemias and lymphomas. Rituximab works extremely well to treat these diseases because of the CD20 binding site. This treatment can also lead to an increase in the number of circulating CD20+ B cells. Rituximab has also shown to be effective in the treatment of multiple sclerosis, rheumatoid arthritis, and anemias. However while effectiveness has been proven, there are also concerns about the safety of the treatment. There is current research being conducted in Norway which will research the effectiveness of rituximab to treat chronic fatigue syndrome. Rituximab is also being used to manage the recipients of kidney transplants. All of these treatments are because of the CD20 binding sites.

Credits

Introduction: David Peltier

Structure: Donald Einck

Drug Binding Site: Ethan Leighton

Additional Effects: Chris Coakley

Citations: Chris Coakley

All Green Screen Effects: David Peltier


Citations

  • Sieber, S, G Gydnia, W Roth, B Bonavida, and T Efferth. "Combination treatment of malignant B cells using the anti-CD20 antibody rituximab and the anti-malarial artesunate." Int J Oncol. 35.1 (2009): 149-158. Print.
  • RITUXAN® (Rituximab) full prescribing information, Genentech, Inc., 2008
  • DiJulio JE. Monoclonal antibodies: overview and use in hematologic malignancies. In: Rieger PT, ed.Biotherapy: A Comprehensive Overview. 2nd ed. Sudbury, Mass: Jones and Bartlett Publishers; 2001:283-316.
  • Maloney DG, Smith B, Rose A. Rituximab: mechanism of action and resistance. Semin Oncol. 2002; 29(suppl 2):2-9
  • Idusogie, Esohe, Leonard Presta, Helene Santoro, Pin Wong, and Michael Mulkerrin. "Mapping of the C1q Binding Site on Rituxan, a Chimeric Antibody with a Human IgG1 Fc." J Immunol. 164.4 (2000): 4178-4184. Print
  • Du, Jiamu, Hao Wang, Chen Zhong, Baozhen Peng, and Melian Zhang. "Structural Basis for Recognition of CD20 by Therapeutic Antibody Rituximab." J Biol Chem. 3.27 (2007): 15073-15080. Print
  • Edwards J, Szczepanski L, Szechinski J, Filipowicz-Sosnowska A, Emery P, Close D, Stevens R, Shaw T (2004). "Efficacy of B-cell-targeted therapy with rituximab in patients with rheumatoid arthritis". N Engl J Med 350 (25): 2572–81.doi:10.1056/NEJMoa032534.PMID 15201414
  • Braendstrup P, Bjerrum OW, Nielsen OJ, Jensen BA, Clausen NT, Hansen PB, Andersen I, Schmidt K, Andersen TM, Peterslund NA, Birgens HS, Plesner T, Pedersen BB, Hasselbalch HC. Rituximab chimeric anti-CD20 monoclonal antibody treatment for adult refractory idiopathic thrombocytopenic purpura. Am J Hematol 2005;78:275-80
  • Patel V, Mihatov N, Cooper N, Stasi R, Cunningham-Rundles S, Bussel JB,Long-term responses seen with rituximab in patients with ITP, Community Oncology Vol. 4 No. 2, February 2007:107
  • Polyak MJ, Ayer LM, Szczepek AJ, Deans JP (2003). "A cholesterol-dependent CD20 epitope detected by the FMC7 antibody". Leukemia 17 (7): 1384–9
  • Monoclonal antibody FMC7 detects a conformational epitope on the CD20 molecule: evidence from phenotyping after rituxan therapy and transfectant cell analyses. 2001
  • Depletion of B cells in vivo by a chimeric mouse human monoclonal antibody to CD20 Blood 1994 83:435-445


Drug Binding Site Green Screen Taken from This proteopedia page http://www.proteopedia.org/wiki/index.php/Rituximab

Proteopedia Page Contributors and Editors (what is this?)

Lynmarie K Thompson, Alexander Berchansky

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