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Event: 718

Key Event Title

The KE title should describe a discrete biological change that can be measured. It should generally define the biological object or process being measured and whether it is increased, decreased, or otherwise definably altered relative to a control state. For example “enzyme activity, decreased”, “hormone concentration, increased”, or “growth rate, decreased”, where the specific enzyme or hormone being measured is defined. More help

Binding, Tubulin

Short name
The KE short name should be a reasonable abbreviation of the KE title and is used in labelling this object throughout the AOP-Wiki. The short name should be less than 80 characters in length. More help
Binding, Tubulin

Biological Context

Structured terms, selected from a drop-down menu, are used to identify the level of biological organization for each KE. Note, KEs should be defined within a particular level of biological organization. Only KERs should be used to transition from one level of organization to another. Selection of the level of biological organization defines which structured terms will be available to select when defining the Event Components (below). More help
Level of Biological Organization
Molecular

Cell term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help
Cell term
eukaryotic cell

Organ term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Key Event Components

Further information on Event Components and Biological Context may be viewed on the attached pdf.Because one of the aims of the AOP-KB is to facilitate de facto construction of AOP networks through the use of shared KE and KER elements, authors are also asked to define their KEs using a set of structured ontology terms (Event Components). In the absence of structured terms, the same KE can readily be defined using a number of synonymous titles (read by a computer as character strings). In order to make these synonymous KEs more machine-readable, KEs should also be defined by one or more “event components” consisting of a biological process, object, and action with each term originating from one of 22 biological ontologies (Ives, et al., 2017; See List). Biological process describes dynamics of the underlying biological system (e.g., receptor signalling). The biological object is the subject of the perturbation (e.g., a specific biological receptor that is activated or inhibited). Action represents the direction of perturbation of this system (generally increased or decreased; e.g., ‘decreased’ in the case of a receptor that is inhibited to indicate a decrease in the signalling by that receptor).Note that when editing Event Components, clicking an existing Event Component from the Suggestions menu will autopopulate these fields, along with their source ID and description. To clear any fields before submitting the event component, use the 'Clear process,' 'Clear object,' or 'Clear action' buttons. If a desired term does not exist, a new term request may be made via Term Requests. Event components may not be edited; to edit an event component, remove the existing event component and create a new one using the terms that you wish to add. More help
Process Object Action
tubulin binding tubulin increased
tubulin complex disrupted

Key Event Overview

AOPs Including This Key Event

All of the AOPs that are linked to this KE will automatically be listed in this subsection. This table can be particularly useful for derivation of AOP networks including the KE. Clicking on the name of the AOP will bring you to the individual page for that AOP. More help
AOP Name Role of event in AOP Point of Contact Author Status OECD Status
Tubulin binding and aneuploidy MolecularInitiatingEvent Francesco Marchetti (send email) Open for citation & comment EAGMST Under Review

Stressors

This is a structured field used to identify specific agents (generally chemicals) that can trigger the KE. Stressors identified in this field will be linked to the KE in a machine-readable manner, such that, for example, a stressor search would identify this as an event the stressor can trigger. NOTE: intermediate or downstream KEs in one AOP may function as MIEs in other AOPs, meaning that stressor information may be added to the KE description, even if it is a downstream KE in the pathway currently under development.Information concerning the stressors that may trigger an MIE can be defined using a combination of structured and unstructured (free-text) fields. For example, structured fields may be used to indicate specific chemicals for which there is evidence of an interaction relevant to this MIE. By linking the KE description to a structured chemical name, it will be increasingly possible to link the MIE to other sources of chemical data and information, enhancing searchability and inter-operability among different data-sources and knowledgebases. The free-text section “Evidence for perturbation of this MIE by stressor” can be used both to identify the supporting evidence for specific stressors triggering the MIE as well as to define broad chemical categories or other properties that classify the stressors able to trigger the MIE for which specific structured terms may not exist. More help

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) can be selected from an ontology. In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help
Term Scientific Term Evidence Link
mouse Mus musculus High NCBI
Homo sapiens Homo sapiens Moderate NCBI
rat Rattus norvegicus Moderate NCBI
Xenopus laevis Xenopus laevis High NCBI

Life Stages

The structured ontology terms for life-stage are more comprehensive than those for taxa, but may still require further description/development and explanation in the free text section. More help
Life stage Evidence
All life stages High

Sex Applicability

No help message More help
Term Evidence
Mixed High

Key Event Description

A description of the biological state being observed or measured, the biological compartment in which it is measured, and its general role in the biology should be provided. For example, the biological state being measured could be the activity of an enzyme, the expression of a gene or abundance of an mRNA transcript, the concentration of a hormone or protein, neuronal activity, heart rate, etc. The biological compartment may be a particular cell type, tissue, organ, fluid (e.g., plasma, cerebrospinal fluid), etc. The role in the biology could describe the reaction that an enzyme catalyses and the role of that reaction within a given metabolic pathway; the protein that a gene or mRNA transcript codes for and the function of that protein; the function of a hormone in a given target tissue, physiological function of an organ, etc. Careful attention should be taken to avoid reference to other KEs, KERs or AOPs. Only describe this KE as a single isolated measurable event/state. This will ensure that the KE is modular and can be used by other AOPs, thereby facilitating construction of AOP networks. More help

The site of action is the tubulin in the cytoplasm. Tubulins represent a large superfamily, and several isotypes are described for both α and β tubulin in mammalian cells [Luduena 2013]. At least six different isotypes of the α subunit are known, while eight isotypes are known for the β subunit. These subunits share a high degree of homology (90% similarity). In addition to α- and β-tubulin, other tubulin homologues have been identified (γ, δ and ε), but their roles in the life cycle of the cell are uncertain [Bhattacharya and Cabral 2009]. All available isotypes are incorporated within microtubules, although with different tissue distributions in normal cells [Berrieman et al. 2004]. The currently known microtubule-disrupting agents bind to all isotypes, having only a slight preference for one over another [Miller et al. 2010].

Binding sites on the α/β-tubulin heterodimer: Conventionally, microtubule-interfering agents are categorized into two main groups: (1) microtubule destabilizers, including colchicine and a variety of vinca alkaloids; and (2) microtubule stabilizers, including taxanes and epothilones. Most agents interact with known binding pockets of α/β-tubulin; however, there are compounds that bind to tubulin on undefined sites. Three distinct sites are well characterized in the literature [Marchetti et al., submitted; Botta et al., 2009]: (1) the colchicine-binding domain at the interface between the α- and β-tubulin dimers; (2) the vinca domain surrounding the GTP binding site on β- and α-tubulin; and, (3) the taxane domain located on β-tubulin [Botta et al. 2009].

Colchicine binding domain on tubulin: The colchicine binding domain is a deep pocket located at the α/β interface of tubulin heterodimers. Crystal structures for tubulin and different ligands are available, although their resolution is not high [Lu et al. 2012; Massarotti et al. 2012]. Notwithstanding its deep location, significant conformational changes in the protein are necessary for accommodating the inhibitors. Both the A and C rings of colchicine are necessary for high affinity binding, while the B ring may only function as a linker between the other two. Three methoxy residues are present in the A ring and all of them are involved in the high affinity binding to tubulin. The C ring of colchicine interacts through van der Waals contacts with Valα181, Serα178, and Valβ315. The carbonyl group behaves as a hydrogen bond acceptor, interacting with Val181a. The A ring is buried in a hydrophobic pocket delimited by Lysβ352, Asnβ350, Leuβ378, Alaβ316, Leuβ255, Lysβ254, Alaβ250, and Leuβ242, and the methoxy group at position 3 is involved in a hydrogen bond interaction within the thiol group of Cysβ241 [Marchetti et al., submitted]. Different ligands may compete with colchicine for the same binding site, even in the absence of high structural correspondence [Lu et al. 2012].

How It Is Measured or Detected

One of the primary considerations in evaluating AOPs is the relevance and reliability of the methods with which the KEs can be measured. The aim of this section of the KE description is not to provide detailed protocols, but rather to capture, in a sentence or two, per method, the type(s) of measurements that can be employed to evaluate the KE and the relative level of scientific confidence in those measurements. Methods that can be used to detect or measure the biological state represented in the KE should be briefly described and/or cited. These can range from citation of specific validated test guidelines, citation of specific methods published in the peer reviewed literature, or outlines of a general protocol or approach (e.g., a protein may be measured by ELISA).Key considerations regarding scientific confidence in the measurement approach include whether the assay is fit for purpose, whether it provides a direct or indirect measure of the biological state in question, whether it is repeatable and reproducible, and the extent to which it is accepted in the scientific and/or regulatory community. Information can be obtained from the OECD Test Guidelines website and the EURL ECVAM Database Service on Alternative Methods to Animal Experimentation (DB-ALM). ?

Binding properties to tubulin are generally evaluated in vitro, typically on tubulin extracts derived from brain tissues [Miller and Wilson 2010]. To determine whether a compound can bind to tubulin, a competitive [3H]colchicine tubulin-binding assay is conducted in vitro to measure whether the binding of colchicine is inhibited by the presence of the test agent [Verdier-Pinard et al. 1998]. A reaction mixture containing tubulin, [3H]colchicine and a potential inhibitor is incubated and after the addition of the scintillation fluid, the radioactivity of [3H]colchicine-bound tubulin is measured using a scintillation counter. The reduction of [3H]colchicine-bound tubulin value is inversely proportional to the test agent binding affinity [Hamel and Lin 1981]. A reaction mixture with only tubulin and [3H]colchicine is generally used as an experimental control standard. The inhibition constant (Ki) of colchicine is 5.75 μM [Zavala et al. 1980] and the ability of new chemicals to interfere with colchicine binding to tubulin is benchmarked against this value.

 

Domain of Applicability

This free text section should be used to elaborate on the scientific basis for the indicated domains of applicability and the WoE calls (if provided). While structured terms may be selected to define the taxonomic, life stage and sex applicability (see structured applicability terms, above) of the KE, the structured terms may not adequately reflect or capture the overall biological applicability domain (particularly with regard to taxa). Likewise, the structured terms do not provide an explanation or rationale for the selection. The free-text section on evidence for taxonomic, life stage, and sex applicability can be used to elaborate on why the specific structured terms were selected, and provide supporting references and background information.  More help

Chemical binding to tubulin has been measured in somatic and germ cells in a variety of species, from rodents in vivo to human cells in culture. Theoretically, chemical binding to tubulin can occur in any cell type in any organism.

Evidence for Perturbation by Stressor

Overview for Molecular Initiating Event

When a specific MIE can be defined (i.e., the molecular target and nature of interaction is known), in addition to describing the biological state associated with the MIE, how it can be measured, and its taxonomic, life stage, and sex applicability, it is useful to list stressors known to trigger the MIE and provide evidence supporting that initiation. This will often be a list of prototypical compounds demonstrated to interact with the target molecule in the manner detailed in the MIE description to initiate a given pathway (e.g., 2,3,7,8-TCDD as a prototypical AhR agonist; 17α-ethynyl estradiol as a prototypical ER agonist). Depending on the information available, this could also refer to chemical categories (i.e., groups of chemicals with defined structural features known to trigger the MIE). Known stressors should be included in the MIE description, but it is not expected to include a comprehensive list. Rather initially, stressors identified will be exemplary and the stressor list will be expanded over time. For more information on MIE, please see pages 32-33 in the User Handbook.

Colchicine is a prototypical spindle poison that has been extensively used to investigate binding to tubulin. The kinetics of colchicine binding are well established [Lambeir and Engelborghs, 1981; Engelborghs, 1998] and can be measured experimentally with high precision [Hamel and Lin, 1981]. Colchicine binds non-polymerized α/β dimeric tubulin by a two-step process. The first step is rapid but weak, resulting in the formation of an initial pre-equilibrium complex, which involves a low affinity binding of colchicine that is reversible. This is followed by slow conformational changes in tubulin, which finally lead to the formation of an irreversible final state tubulin–colchicine complex that has high activation energy [Garland 1978]. The conformational change in tubulin heterodimers, followed by the addition of the complex at the ends of microtubules, is responsible for the suppressed polymerization at microtubule ends leading to their depolymerization [Ravelli et al., 2004]. The binding kinetics have been studied at different temperatures. The standard enthalpy change of the first step (ΔH°1 = -33±8 kJ · mol–1) and the activation energy of the second step (ΔH°2 = 100±5 kJ · mol–1) were determined based on the temperature dependence [Lambeir and Engelborghs, 1981]. Using eight different analogues to study the binding mechanisms of colchicine, it was demonstrated that the C-ring of colchicine is responsible for the first step of the binding mechanism, while the second step involves the rearrangement of the initial complex to interact with the A-ring [Engelborghs, 1998].

Other chemicals are also known to bind to tubulin [Marchetti et al., 2016]. These chemicals can be grouped in two general classes: colchicine domain binders and vinca domain binders.

Known colchicine domain binders:

1. Podophyllotoxin ((5R,5aR,8aR,9R)-5-hydroxy-9-(3,4,5-trimethoxyphenyl)-5a,6,8a,9-tetrahydro-5H-[2]benzofuro[5,6-f][1,3]benzodioxol-8-one, POD) has a trimethoxybenzoic chemical structure similar to colchicine. It inhibits colchicine binding to the colchicine-binding domain of tubulin. However, although colchicine and podophyllotoxin bind in the same pocket on β-tubulin, their binding sites are not completely overlapping [Desbene and Giorgi-Renault 2002].

2. 2-methoxyestradiol ((8R,9S,13S,14S,17S)-2-methoxy-13-methyl-6,7,8,9,11,12,14,15,16,17-decahydrocyclopenta[a]phenanthrene-3,17-diol, 2ME) binds to the colchicine domain of tubulin [D'Amato et al. 1994].

3. Nocodazole (methyl (5-[2-thienylcarbonyl]-1H-benzimidazol-2-yl, NOC) belongs to the group of benzimidazole derivatives that were patented, as a class, for the treatment of cancer in conjunction with other pharmaceuticals. Nocodazole has been shown to bind in the colchicine domain [Xu et al. 2002].

4. Benomyl (methyl N-[1-(butylcarbamoyl)benzimidazol-2-yl]carbamate, BEN), another benzimidazole derivative, is the active compound in several agricultural fungicides. The benomyl-binding site is located in the core of β-tubulin at a site distinct from the colchicine domain [Clement et al. 2008].

5. Carbendazim (methyl N-(1H-benzimidazol-2-yl)carbamate, MBC) is a fungicide commonly used in agriculture for the control of a wide range of fungal diseases. Carbendazim is the methylbenzimidazolcarbamate product of the spontaneous hydrolyzation process incurred by benomyl in aqueous solution (it is the major metabolite). Therefore, it is at least partially responsible for the benomyl effects observed in vivo. The affinity of carbendazim for mammalian tubulin is less than that of benomyl, most probably because it lacks the position 1 side chain of benomyl. Like benomyl, carbendazim does not compete with colchicine for binding to tubulin [Yenjerla et al., 2008].

6. Thiabendazole (4-(1H-benzimidazol-2-yl)-1,3-thiazole, TBZ) is a benzimidazole-derived anthelmintic and an agricultural fungicide, structurally related to NOC, benomyl and MBC. TBZ competitively inhibits MBC binding to fungal tubulin [Davidse and Flach, 1978].

7. ABT-751 (N-(2-((4-hydroxyphenyl)amino)pyridin-3-yl)-4-methoxybenzenesulfonamide) has a scaffold based on a benzsulfamide group. It was identified as a potent antiproliferative agent and was subsequently found to be an antitubulin agent by targeting the colchicine binding site [Yoshimatsu et al., 1997].

8. A compound base on m-ethoxyaniline group (2-(6-ethoxy-3-(3-ethoxyphenylamino)-1-methyl-1,4-dihydroindeno[1,2-c]pyrazol-7-yloxy)acetamide) showed noteworthy low nanomolar potency against cancer cell lines. In mechanistic studies, it inhibited tubulin polymerization and disorganized microtubule by binding to tubulin colchicine binding site [Liu et al., 2016].

Vinca domain binders:

The vinca alkaloids, a class of antimitotic compounds derived from the periwinkle plant, Catharanthus roseus [Cutts et al., 1960], bind near the GTP-binding site on the β-subunit of tubulin at a site distinct from the colchicine-binding one [Rai and Wolff 1996]. Vinblastine and vincristine are first-generation vinca alkaloids [Kingston, 2009]. At low concentrations, vincas bind to the plus ends of microtubules producing a conformational change of dimers from a straight ‘‘growing’’ vector to a curved ‘‘peeling’’ vector [Toso et al., 1993]. At higher concentrations, the vinca alkaloids have affinity for free tubulin heterodimers, again potentially forming an altered, curved geometry of the dimeric biological vector [Warfield and Bouck, 1974]. Although vincas do not share structural similarity with colchicine and bind to a different site on tubulin, they similarly act by destabilizing microtubules [Stanton et al., 2011].

Vinca domain binders include:

1. Vinblastine (dimethyl (2β,3β,4β,5α,12β,19α)-15-[(5S,9S)-5-ethyl-5-hydroxy-9-(methoxycarbonyl)-1,4,5,6,7,8,9,10-octahydro-2H-3,7-methanoazacycloundecino[5,4b]indol-9-yl]-3-hydroxy-16-methoxy-1-methyl-6,7-didehydroaspidospermidine-3,4-dicarboxylate, VBL) is an anticancer drug that is used extensively. The crystal structure of vinblastine bound to tubulin has been determined [Gigant et al., 2005]. In contrast with the binding site for colchicine, which is mostly embedded in β-tubulin subunit, the vinblastine binding site is shared equally between α/β-heterodimer [Marchetti et al., 2016]. In the β-subunit, vinblastine interacts through van der Waals contacts with residues Serβ174-Aspβ179, Asnβ206-Aspβ211, Pheβ214 and Tyrβ224; while in the α-subunit, Pheα351, Lysα352, Valα353 and Ileα355 delimit the pocket occupied by VBL. Amino acids Proβ222 and Asnα329 are also involved in hydrogen bond interactions with VBL [Marchetti et al., 2016]. Following a mechanism similar to colchicine, VBL binds to tubulin in two consecutive steps: formation of a rapid equilibrium complex followed by a slower rearrangement linked to changes in the structure of the heterodimer. A major effect of VBL is the formation of spiral-like tubulin aggregates [Weisenberg and Timasheff, 1970; Himes, 1991]. VBL binds to microtubule ends [Wilson et al., 1982] and at low concentrations suppresses the dynamic instability of plus ends [Toso et al., 1993]. When used at much higher concentrations, VBL depolymerizes microtubules, giving rise in particular to protofilament spirals and curls.

2. Vincristine (methyl (3aR,3a1R,4R,5S,5aR,10bR)-4-acetoxy-3a-ethyl-9-((5S,7S,9S)-5-ethyl-5-hydroxy-9-(methoxycarbonyl)1,4,5,6,7,8,9,10-octahydro-2H-3,7-methano[1]azacycloundecino[5,4-b]indol-9-yl)-6-formyl-5-hydroxy-8-methoxy-3a,3a1,4,5,5a,6,11,12-octahydro-1H-indolizino[8,1-cd]carbasole-5-carboxylate, VCR) is an anticancer drug that binds to tubulin in the vinca binding domain [Kingston, 2009].

3. Vinflunine (methyl (3aR,3a1R,4R,5S,5aR,10bR)-4-acetoxy-9-((4R,6R,8S)-4-(1,1-difluoroethyl)-8-(methoxycarbonyl)-1,3,4,5,6,7,8,9-octahydro-2,6-methanoazecino[4,3-b]indol-8-yl)-3a-ethyl-5-hydroxy-8-methoxy-6-methyl-3a,3a1,4,5,5a,6,11,12-octahydro-1H-indolizino[8,1-cd]carbazole-5-carboxylate) is a florinated Vinca alkaloid, like vinblastine and vincristine, and appears to interact at the Vinca binding domain [Kruczynski et al., 1998].

4. Vintafolide ((2R,5S,8S,11S,14S,19S)-19-(4-(((2-amino-4-oxo-3,4-dihydropteridin-6-yl)methyl)amino)benzamido)-5,8,14-tris(carboxymethyl)-2-(((2-((2-((3aR,3a1R,4R,5S,5aR,10bR)-3a-ethyl-9-((5S,7R,9S)-5-ethyl-5-hydroxy-9-(methoxycarbonyl)-1,4,5,6,7,8,9,10-octahydro-2H-3,7-methano[1]azacycloundecino[5,4-b]indol-9-yl)-4,5-dihydroxy-8-methoxy-6-methyl-3a,3a1,4,5,5a,6,11,12-octahydro-1H-indolizino[8,1-cd]carbazole-5-carbonyl)hydrazine-1-carbonyl)oxy)ethyl)disulfanyl)methyl)-11-(3-guanidinopropyl)-4,7,10,13,16-pentaoxo-3,6,9,12,15-pentaazaicosanedioic acid) is a drug conjugate consisting of a small molecule targeting the folate receptor and vinblastine. Vintafolide is designed to deliver vinblastine selectively to cells over-expressing the folate receptor such as ovarian cancer cells [Vergote and Leamon, 2015].

References

List of the literature that was cited for this KE description. Ideally, the list of references, should conform, to the extent possible, with the OECD Style Guide (https://www.oecd.org/about/publishing/OECD-Style-Guide-Third-Edition.pdf) (OECD, 2015). More help

Berrieman HK, Lind MJ, Cawkwell L. 2004. Do beta-tubulin mutations have a role in resistance to chemotherapy? Lancet Oncol 5:158-164.

Bhattacharya R, Cabral F. 2009. Molecular basis for class V beta-tubulin effects on microtubule assembly and paclitaxel resistance. J Biol Chem 284:13023-13032.

Botta M, Forli S, Magnani M, Manetti F. 2009. Molecular Modeling Approaches to Study the Binding Mode on Tubulin of Microtubule Destabilizing and Stabilizing Agents. In: Carlomagno T, editor. Tubulin-Binding Agents: Springer Berlin Heidelberg. p 279-328.

Clement MJ, Rathinasamy K, Adjadj E, Toma F, Curmi PA, Panda D. 2008. Benomyl and colchicine synergistically inhibit cell proliferation and mitosis: evidence of distinct binding sites for these agents in tubulin. Biochemistry 47:13016-13025.

Cutts JH, Beer CT, Noble RL. 1960. Biological properties of Vincaleukoblastine, an alkaloid in Vinca rosea Linn, with reference to its antitumor action. Cancer Res 20:1023-1031.

D'Amato RJ, Lin CM, Flynn E, Folkman J, Hamel E. 1994. 2-Methoxyestradiol, an endogenous mammalian metabolite, inhibits tubulin polymerization by interacting at the colchicine site. Proc Natl Acad Sci USA 91:3964-3968.

Desbene S, Giorgi-Renault S. 2002. Drugs that inhibit tubulin polymerization: the particular case of podophyllotoxin and analogues. Curr Med Chem Anticancer Agents 2:71-90.

Engelborghs Y. 1998. General features of the recognition by tubulin of colchicine and related compounds. Eur Biophys J 27:437-445.

Garland DL. 1978. Kinetics and mechanism of colchicine binding to tubulin: evidence for ligand-induced conformational change. Biochemistry 17:4266-4272.

Gigant B, Wang C, Ravelli RB, Roussi F, Steinmetz MO, Curmi PA, Sobel A, Knossow M. 2005. Structural basis for the regulation of tubulin by vinblastine. Nature 435:519-522.

Hamel E, Lin CM. 1981. Stabilization of the colchicine-binding activity of tubulin by organic acids. Biochim Biophys Acta 675:226-231.

Himes RH. 1991. Interactions of the catharanthus (Vinca) alkaloids with tubulin and microtubules. Pharmacol Ther 51:257-267.

Kingston DG. 2009. Tubulin-interactive natural products as anticancer agents. J Nat Prod 72:507-515.

Lambeir A, Engelborghs Y. 1981. A fluorescence stopped flow study of colchicine binding to tubulin. J Biol Chem 256:3279-3282.

Lu Y, Chen J, Xiao M, Li W, Miller DD. 2012. An overview of tubulin inhibitors that interact with the colchicine binding site. Pharm Res 29:2943-2971.

Luduena RF. 2013. A hypothesis on the origin and evolution of tubulin. Int Rev Cell Mol Biol 302:41-185.

Marchetti A, Massarotti A, Yauk CL, Pacchierotti F, Russo A. Submitted. The adverse outcome pathway (AOP) for chemical binding to tubulin in oocytes leading to aneuploid offspring. Environ Mol Mutagen.

Massarotti A, Coluccia A, Silvestri R, Sorba G, Brancale A. 2012. The tubulin colchicine domain: a molecular modeling perspective. ChemMedChem 7:33-42.

Miller HP, Wilson L. 2010. Chapter 1 - Preparation of Microtubule Protein and Purified Tubulin from Bovine Brain by Cycles of Assembly and Disassembly and Phosphocellulose Chromatography. In: Leslie W, John JC, editors. Methods Cell Biol: Academic Press. p 2-15.

Miller LM, Xiao H, Burd B, Horwitz SB, Angeletti RH, Verdier-Pinard P. 2010. Chapter 7 - Methods in Tubulin Proteomics. In: Leslie W, John JC, editors. Methods Cell Biol: Academic Press. p 105-126.

Rai SS, Wolff J. 1996. Localization of the vinblastine-binding site on beta-tubulin. J Biol Chem 271:14707-14711.

Ravelli RB, Gigant B, Curmi PA, Jourdain I, Lachkar S, Sobel A, Knossow M. 2004. Insight into tubulin regulation from a complex with colchicine and a stathmin-like domain. Nature 428:198-202.

Stanton RA, Gernert KM, Nettles JH, Aneja R. 2011. Drugs that target dynamic microtubules: a new molecular perspective. Med Res Rev 31:443-481.

Toso RJ, Jordan MA, Farrell KW, Matsumoto B, Wilson L. 1993. Kinetic stabilization of microtubule dynamic instability in vitro by vinblastine. Biochemistry 32:1285-1293.

Warfield RK, Bouck GB. 1974. Microtubule-macrotubule transitions: intermediates after exposure to the mitotic inhibitor vinblastine. Science 186:1219-1221.

Weisenberg RC, Timasheff SN. 1970. Aggregation of microtubule subunit protein. Effects of divalent cations, colchicine and vinblastine. Biochemistry 9:4110-4116.

Wilson L, Jordan MA, Morse A, Margolis RL. 1982. Interaction of vinblastine with steady-state microtubules in vitro. J Mol Biol 159:125-149.

Xu K, Schwarz PM, Ludueña RF. 2002. Interaction of nocodazole with tubulin isotypes. Drug Dev Res 55:91-96.

Zavala F, Guenard D, Robin JP, Brown E. 1980. Structure--antitubulin activity relationship in steganacin congeners and analogues. Inhibition of tubulin polymerization in vitro by (+/-)-isodeoxypodophyllotoxin. J Med Chem 23:546-549.