1 / 24

The B7/CD28 pathway is much more complicated than expected

The B7/CD28 pathway is much more complicated than expected. Chambers C.A. et al. Immuunol. Rev. 153:27, 1996. B7/CD28/CTLA-4. Activated T cell. AP2. AP2. TCR. CTLA-4. CD28. P. P. peptide/ MHC. B7.1 (CD80). Activated Professional APC. B7.2 (CD86).

apu
Télécharger la présentation

The B7/CD28 pathway is much more complicated than expected

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. The B7/CD28 pathway is much more complicated than expected

  2. Chambers C.A. et al. Immuunol. Rev. 153:27, 1996.

  3. B7/CD28/CTLA-4 Activated T cell AP2 AP2 TCR CTLA-4 CD28 P P peptide/ MHC B7.1 (CD80) Activated Professional APC B7.2 (CD86)

  4. CTLA-4 is not readily detectable innaïve T cells but is rapidly upregulated upon T cell activation. CTLA-4 mRNA can be readily detected within 1 h of TCR engagementand peaks at about 24–36 h. However, CTLA-4 is not readilydetectable at the cell surface until 24–48 h after activation. An accurate assessment of the kinetics of CTLA-4protein expression is complicated by the fact that it is notprimarily expressed at the cell surface. Surface expression of CTLA-4 is tightly regulated as a result of the presence of a tyrosine-based intracellular localization motif in its cytoplasmic tail which allowsan association with the medium subunit of the clathrin-coated pit adaptor complex, AP2,thus providing a mechanism for CTLA-4cellular localization. CTLA-4 expression onthe T cell surface is stabilized and increased by tyrosine phosphorylationof the endocytosis motif, which inhibits AP2 association. Itis noteworthy that the intracellular portion of CTLA-4 is 100%conserved among many different species of animals, suggestingthat control of intracellular trafficking may be extremely importantfor its function. This motif results in both the rapid endocytosis of CTLA-4 from the cell surface to endosomalcompartments, as well as the targeting of at least some CTLA-4to the lysosomes for degradation. It is unclear whetherendocytosed CTLA-4 can recycle back to the surface or if thisrepresents a terminal pathway for the protein.

  5. Dynamic integration of TCR, CD28 and CTLA-4 As previously discussed, CD28 is constitutively expressed onT cells, whereas CTLA-4 appears after activation. Because ofthis, and perhaps as a result of our innate appreciation forsymmetry, the idea arose that CD28 engagement allowed initiation,while CTLA-4 provided for termination of immune responses. Surprisingly, the majority of the in vitro data has demonstratedan inhibitory role for CTLA-4 in the early stages of T cellactivation. IL-2 production, expression of early markers such asCD69 and CD25, and a number of other aspects of activation areinhibited upon CTLA-4 cross-linking. These events take place within hours of T cell activation with anti-CD3 and CD28. Infact, the inhibition of the induction of IL-2 transcriptionwas detected 4 h after stimulation. This suggests eitherthat there is a physiologically relevant intracellular poolof CTLA-4 present in naïve T cells or that protein expressionis induced rapidly upon activation. The possibility that CTLA-4 can inhibit earlystages of T cell activation has led to the development of modelsthat stress that the dynamic interplay of costimulatory andTCR signals depends on the activation state of the T cell aswell as the activation state of the antigen-presenting cell.

  6. The threshold model:When B7 levelsare low and TCR signals are weak, the amount of CTLA-4 induced is low but may be sufficient to minimize costimulation and preventactivation. Under these conditions CTLA-4 may set a thresholdfor activation. In other words, it plays a role in setting the stimulatory threshold required for a T cell to progress to full activation.

  7. As in the classicaltwo-signal model, an encounter of a naïve T cell with acell expressing appropriate MHC/antigen complex but lackingB7 does not result in activation of the T cell owing to lackof costimulation. The cells receiving a TCR signal in the absence of CD28-mediated costimulation may be rendered anergic. However,engagement of the TCR can lead to rapid induction and/or mobilizationof small amounts of CTLA-4. Under conditions where there isan incompletely activated APC expressing only low amounts ofB7, CTLA-4 could, by virtue of its higher affinity, outcompeteCD28 for B7 and/or deliver inhibitory signals. This could effectively raise the threshold of CD28 and/or TCR signals needed for fullactivation. Chambers, C.A. et al. Ann.Rev.Immunol. 19:565, 2001

  8. There are two scenarios in which CTLA-4 may play a role in establishinga threshold for CD28 and/or TCR signals needed for activationof naïve T cells. Both presume that low levels of CTLA-4pre-exist or can be rapidly induced in naïve T cells upon engagement of the TCR and CD28. The first scenario deals with the regulation of the responseof T cells to tonic signaling by self-peptide/MHC interactions. Continuous TCR interactions with self-peptide/MHCprovide important signals for the survival of peripheral T cells. Some of these tonic interactions, under conditions of lowlevels of CD28/B7 interaction, might be sufficiently stimulatoryto lead to the activation of CD4+ T cells and the inductionand/or mobilization of CTLA-4. Based on the analysis of theCTLA-4-/- mice, it is speculated that CTLA-4might prevent the signals generated by these interactions fromleading to full activation of CD4+ T cells. Thus, CTLA-4may be involved in maintaining naïve CD4+ T cells and previouslyactivated T cells in a resting state. This model is supported by the phenotype of CTLA-4-/- mice. The expansion of T cells that occurs in these miceis polyclonal. Thissuggests that the expansion is not the result of a failureto terminate responses to few environmental pathogens. Thus, the phenotype of CTLA-4-/- mice results from the activation and expansion of T cells reactive to low-affinity self-ligands due to a decreased activation threshold.

  9. The second scenario suggests a role for CTLA-4 in maintainingperipheral tolerance of T cells with specificity for tissue-specificantigens that are not expressed in the thymus and have not beendeleted as a consequence of negative selection. CTLA-4 may provide an additional level of regulation toensure peripheral tolerance by preventing activation when aT cell encounters a normal self-antigen in the context of lowB7 expression. This scenario may explain the observationthat CTLA-4 blockade or deficiency accelerates the onset and severity of insulitis and diabetes in nonobese diabetic (NOD)mice expressing a transgenic ß TCR cloned from anislet-ß-cell-specific CD4+ T cell clone isolated froma NOD mouse. T cells bearing this TCR are efficientlyselected, rather than deleted in the thymus, demonstrating thatcentral tolerance is not effective for T cells with this specificity.The observation that blockade or loss of CTLA-4 dramatically accelerates disease in this model system indicates a role for CTLA-4 in maintaining an activation threshold for autoreactive T cells bearingTCRs specific to autoantigens.

  10. The attenuation model:When B7 levelsare high and TCR signals are strong, the higher levels of CTLA-4 induced after activationmay be able to attenuate the response of activated cells. This model suggest that after activation and subsequent entry into the cell cycle, CTLA-4 can limit the capacity of a T cell to divide.

  11. Under conditions where the APC is activated and expressing high levels of B7, CD28 costimulation may dominate and activation proceeds. However, this results in induction and/or mobilization of CTLA-4 that may be proportional to the strength of the TCR signal, resulting in differential inhibition. CTLA-4 preferentially attenuatesthe expansion of T cells that have been strongly activated. This notion is supported by the effect of CTLA-4-blockade on the proliferative capacity, or averagenumber of daughter cells per responder, for T cells primed withagonist or weak agonist peptides in adjuvant. Blockade significantlyincreased the proliferative capacity of T cells primed withthe agonist ligand but had a minimal effect on the cells responding to the weak agonist peptide.. Chambers, C.A. et al. Ann.Rev.Immunol. 19:565, 2001

  12. CTLA-4 would prevent this high-affinity population fromdominating the primed pool by restricting proliferation earlyin the response. Engagement of CTLA-4 would then serve to broadenthe pool of T cells by limiting clonal representation of thehigh-affinity population, thus allowing more equal representationof the cells bearing lower affinity TCRs in the early stagesof the clonal evolution of the response. It appears thatincreased CTLA-4 expression upon activation modulates T cellresponses differentially and might serve to limit the burstsize of responding T cells. Overall, the results suggest thatthe quality of the TCR signal is critical to determining ifand/or how dramatically CTLA-4 regulates the proliferative capacityof any antigen-specific clone selected from the T cell repertoire. While the TCR and CD28 might primarily determine the range ofT cells responding to antigen, CTLA-4 would limit clonal representationof T cells with high-affinity TCRs.

  13. Role played by CTLA-4 in regulating antigen-specific T cell responses. (a) The threshold and attenuation models make different predictions about the function of CTLA-4, depending on the stimulatory conditions encountered during activation. Shown are graphical representations that relate the effect of CTLA-4– mediated inhibition on T cell expansion to the degree of stimulation (TCR + CD28 signals). Threshold model: T cells that are stimulated below a given activation threshold (dashed lines) will not commit to entering the cell cycle. According to the threshold model, CTLA-4 may participate in setting a threshold for activation, thereby keeping cells that receive low amounts of stimulation from responding (red lines). In the absence of CTLA-4 signaling, the T cell activation threshold would be shifted, so that cells that receive weaker amounts of stimulation could now become activated and proliferate (green lines).

  14. Attenuation model: a certain amount of T cell stimulation may be required to up-regulate and/or mobilize CTLA-4, thereby allowing it to participate in the regulation of a T cell response. Thus, according to the attenuation model, CTLA-4 does not affect the threshold of stimulation required for a T cell to enter the cell cycle; rather, it affects the extent of subsequent expansion. Based on its expression and localization patterns, CTLA-4 may most significantly restrict the expansion of T cells that receive stronger amounts of stimulation. (b) A diverse population of cells may respond to a given antigen, but due to the process of antigen-driven selection, a greater number of T cells bearing TCRs of higher affinity will be present in this population. By preferentially restricting the expansion of cells that receive stronger TCR signals, CTLA-4 would prevent T cells bearing higher affinity TCRs from dominating the response (red line). Thus, in the absence of CTLA-4, the distribution of cells within the responding population would shift toward higher affinity TCRs, resulting in a polyclonal response of reduced diversity (green line).

  15. CTLA- regulates cell cycle progression, not cell death The process by which CTLA-4-induced negative regulation occurs is clearly distinctfrom activation-induced cell death. There is no evidence that CTLA-4 ligationin conjunction with TCR and CD28 cross-linking on resting murineT cells induces apoptosis. Further, CTLA-4ligation does not alter the CD28-mediated upregulation of survivalfactor bcl-2, arguing against apoptosis induction beingthe major mechanism of CTLA-4 inhibition. The fact that CTLA-4 deficiency is lethal relatively early inlife and that the Fas pathway is intact indicates that CTLA-4can play a critical role in limiting T cell expansion. CTLA-4has effects on at least two aspects of activation that havecritical relevance to proliferation. The first is on IL-2 production.CD28 costimulation enhances IL-2 production both at the levelof transcription and mRNA stabilization. Extensiveligation of CTLA-4 under suboptimal conditions of stimulationby TCR plus CD28 can result in inhibition of IL-2 production,probably at the level of transcription. CTLA-4ligation inhibits TCR-induced productionof cdk4, cdk6, and cyclin D3, all of which are required forG0/G1 progression. Thus CTLA-4 can limit expansion notonly by reducing production of an important growth factor, butalso by inhibiting TCR-mediated induction and assembly of essentialcomponents of the cell cycle machinery.

  16. CTLA-4 induces inhibitory cytokines? It has been reported that cross-linking of CTLA-4 may enhanceproduction of TGFß by activated T cells. This raises the possibility that CTLA-4 does not directly inhibitT cell activation but does so by the active induction of thisinhibitory cytokine. One observation taken as evidence for anindirect role for CTLA-4 in the inhibition of T cell responseswas the failure of mixed bone marrow chimeric mice generatedwith CTLA-4-/- and wild-type bone marrow to develop a phenotypeequivalent to the ctla-4 null mice. It was proposed thatthere is no primary defect in CTLA-4-/- T cells but that theCTLA-4-/- phenotype is due to a failure of T cells to secreteinhibitory cytokines such as TGFß. Recent studies showed that CTLA-4 cross-linking resulted inthe inhibition of proliferation of T cells from TGFß-/- mice or of T cells from mice lacking Smad3, a critical downstreamsignaling molecule in the TGFß pathway. This suggests that neither TGFßproduction nor its signaling pathway is required for CTLA-4-mediatedinhibition of T cell responses. These studies also failed toshow a role for CTLA-4 in regulating TGFß production,since it was produced by CTLA-4-/- T cells. Finally, CTLA-4ligation failed to induce production of TGFß by normalnaïve T cells.

  17. CTLA-4 and regulatory T cells? Over the last several years, a considerable literature has documenteda role for CD25+CD45RBlowCD4+ regulatory T cells (Treg) in themaintenance of peripheral tolerance to organ-specific self-antigens. The observation that these cells constitutively expressCTLA-4 has raised the possibility that CTLA-4 may be directlyinvolved in their function. Moreover, some studies indicate that someCTLA-4 functions may not be necessarily T cell autonomous:(cf. bone marrow chimera with CTLA-4 -/- bone marrow cells or a mixture of CTLA-4 -/- and CTLA-4 +/+ bone marrow cells.) Administrationof either anti-CTLA-4 antibodies or anti-TGFß reversedthe inhibitory effects of transferred CD25+ Treg cells on theinduction of colitis by transferred CD4+ CD25- cells in SCIDmice. This was taken as evidence for a blocking of thesuppressive effects of Treg cells by preventing CTLA-4-mediatedinduction of TGFß production. However, it does not exclude the possibility that theeffect of anti-CTLA-4 was a result of enhancement of the effectorT cells. Other investigators have shown that CTLA-4 does not have a role In the function of Treg cells. CD25+CD4+ T cells from CTLA-4-/- mice retainedinhibitory activity in in vitro inhibition assays. One interestingcharacteristic of Treg cells is a failure to secrete IL-2 orto proliferate in response to ligation of the TCR and CD28. The hyporesponsiveness might be attributed to the inhibitoryproperties of CTLA-4. However, to date, attempts to releasethe block on proliferation in response to TCR engagement by CTLA-4 blockade have not been successful.

  18. T cell–mediated immunotherapy represents a promising treatmentfor human malignancies. In cancer patients, the absence of efficienttumor-specific immunity can be related to inadequate APC functionor to T cell tolerance/ignorance towards tumor antigens. Mice were injected intravenouslywith 5 x 104 or 105 B16-F10 melanoma cells. Treatment using irradiatedF10/g cells (GM-CSF transfected B16-F10 cells; Presumably,GM-CSF production at the site of vaccination might attract hostAPCs and enhance their function in vivo.) and antibodies was started after 24 h. These results indicatethat CTLA-4 blockade and GM-CSF–producing vaccines actsynergistically to cause rejection of poorly immunogenic tumors. van Elsas A. J. Exp. Med. 190:355, 1999

  19. No treatment Depletion of Treg Vacc. + CTLA-4 blocking Vacc. + depletion of Treg Vacc. + CTLA-4 blocking + depletion of Treg Figure 3. CD25+ T cell depletion before vaccination enhances efficacy of treatment. (A) Survival data of mice challenged subcutaneously with 2.5 x 103 B16-BL6 tumor cells. Mice received either no treatment (n = 6, ), or depleting anti-CD25 on day -4 (n = 6, ) or vaccination with GM-CSF–producing B16 on days 0, 3, and 6. The vaccinated mice were divided over three groups that received the following Ab: CTLA-4 blocking Ab on days 0, 3, and 6 (n = 8, •); depleting anti-CD25 Ab on day -4 (n = 8, x); or depleting anti-CD25 Ab on day -4 plus CTLA-4–blocking Ab on days 0, 3, and 6 (n = 8, ). (B) Survival data of mice challenged subsutaneously (day 0) with 5 x 103 B16-BL6 tumor cells. Mice received either depleting anti-CD25 Ab on day -4 (n = 6, ) or were vaccinated on days 0, 3, and 6 with GM-CSF– producing B16. The vaccinated mice were divided over two groups that received the following Ab: blocking anti–CTLA-4 Ab on days 0, 3, and 6 (n = 9, •); or depleting anti-CD25 Ab on day -4 combined with blocking anti-CTLA-4 Ab on days 0, 3, and 6 (n = 9, ).. Sutmuller, R.P. et al. J. Exp. Med. 194:823, 2001

  20. In conclusion, the data presented in this paper reveal thatcombination of CTLA-4 blockade and elimination of CD25+ Tregcells can result in more effective therapeuticantitumor immunity than when these intervention strategies areapplied separately. These findings support the notion that CD25+ Treg cells and CTLA-4 represent two alternative pathways forsuppression of autoreactive T cell immunity, but do not excludethat functional overlap between these pathways exists. Simultaneous intervention with both regulatorymechanisms appears to be a highly promising strategy for theinduction of T cell immunity against tumor-associated autoantigensin the immunotherapy of cancer.

  21. Therapy with anti-CTLA-4 and GM-CSF B16 melanoma vaccine leads to tumor rejection but also induces autoimmune responses (depigmentation).

More Related