Pathophysiological aspects of memory B-cell development

Pathophysiological aspects of memoryB-cell developmentSandrine Roulland1,2,3, Felipe Suarez4, Olivier Hermine4,5 and Bertrand Nadel1,2,3

1 Centre d’Immunologie de Marseille-Luminy (CIML), Universite de la Mediterranee, 13288 Marseille, France2 INSERM, U631, 13288 Marseille, France3 CNRS, UMR6102, 13288 Marseille, France4 Department of Adult Haematology, Assistance Publique - Hopitaux de Paris, Hopital Necker-Enfants Malades, 75730 Paris, France5 CNRS, UMR8147, 75743 Paris, France

Review TRENDS in Immunology Vol.29 No.1

Glossary

CSR: class switch recombination

DLBCL: diffuse large B-cell lymphomas

FL: follicular lymphoma

GC: germinal center

MALT: mucosa-associated lymphoid tissue

MCII: mixed cryoglobulinemia type II

MZL: marginal zone lymphoma

PLO: peripheral lymphoid organs

RF: rheumatoid factor

B cells follow two functionally distinct pathways ofdevelopment: a classical germinal center (GC) T-depend-ent pathway in which diversification and maturationgenerate a slow, but virtually unlimited high-affinityresponse to cognate antigens; and a marginal zone(MZ) T-independent pathway providing a first line of‘innate-like’ defense against specific pathogens. Cellspopulating these two distinct locations are the normalcounterparts of two clinically important pathologicalentities, follicular lymphoma (FL) and MZ lymphoma(MZL). FL and MZ represent paradigms of two risingconcepts of lymphomagenesis, protracted preclinicaland antigen-driven lymphoproliferation, respectively.Integrating the mechanisms and functions of MZ andGC B cells and the distinctive features of their patho-logical counterparts should provide essential clues tothe understanding of their malignant development, andshould offer new insights into the design of effectivetreatments for B-cell lymphomas.

Setting the stage for lymphomagenesis: the generationof memory B cellsRecent progress in the understanding of B-cell developmentin the mouse and human has allowed us to draw a clearerpicture of the complex, multistep pathways involved in thegeneration ofmemoryB cells (summarized in Figure 1), andto set the stage for a better understanding of associatedpathologies.Most of the key factors crucial for normal B-celldifferentiation and survival are also involved in the malig-nant growth of many mature B-cell lymphomas [1,2]. Atvirtually every B-cell developmental stage there is a corre-sponding subtype of lymphoid neoplasia, ‘frozen’ by malig-nant transformation at a given maturation step. Marginalzone lymphoma (MZL) and follicular lymphoma (FL), twoseparate and clinically important pathological entities,represent paradigms of two major and functionally distinctB-cell subtypes: marginal zone (MZ) and follicular B cells.Althoughmany aspects of their origin and pathogenesis aredistinct, there is accumulating evidence for a role of foreignand/or self-antigens in the initiation and promotion oflymphomagenesis [3,4].

Corresponding authors: Hermine, O. ([email protected]);Nadel, B. ([email protected]).

Available online 3 December 2007.

www.sciencedirect.com 1471-4906/$ – see front matter � 2007 Elsevier Ltd. All rights reserve

Antigen-driven lymphomagenesis and marginal zonelymphoma: T-independent responses, T-dependentresponses, or both?The current WHO classification of lymphoid neoplasmsrecognizes three entities that are thought to derive fromMZ B cells: splenic MZ lymphoma (SMZL), with itsleukemic subtype named splenic lymphoma with villouslymphocytes (SLVL); extranodal MZ lymphoma of mucosa-associated lymphoid tissue (MALT); and nodal MZ lym-phoma [5]. Although these different lymphoma subsetsdiffer in many aspects, they all share several commonpathogenic features, the most striking of which being theirpossible association with chronic antigenic stimulation bymicrobes and/or autoantigens [3]. Since the initial descrip-tion of the role of Helicobacter pylori in gastric MALTlymphoma [6], several other MZ-derived lymphomas havebeen associated with chronic infections: for instance,immunoproliferative small intestinal disease (IPSID),cutaneousMZL, and ocular adnexaMALT lymphoma havebeen associated with Campylobacter jejuni, Borelia burg-dorferi, and Chlamydophila psittaci, respectively [7–9].Similarly, a subset of SMZL, and particularly SLVL, mightbe associated with chronic hepatitis C (HCV) infection[10,11].

Infection-associated MZ lymphomas represent a modelof stepwise progression leading to indirect lymphoid trans-formation [3]. Unlike the classical transformation oflymphocytes by lymphotropic pathogens, epitomized bythe B-cell-immortalizing Epstein Barr Virus (EBV) [12],neoplastic MZ B cells in infection-associated lymphomasare not directly infected by the inducing pathogen. Rather,

SHM: somatic hypermutation

SLVL: splenic lymphoma with villous lymphocytes

SMZL: splenic marginal zone lymphoma

TD: T-dependent

TI: T-independent

d. doi:10.1016/j.it.2007.10.005

mailto:[email protected]

mailto:[email protected]

http://dx.doi.org/10.1016/j.it.2007.10.005

Figure 1. Summary of peripheral B-cell development. Early B-cell development occurs in the bone marrow (BM) [73]. Immature B cells then leave the BM and reach the

spleen [74,75], where a crucial cell-fate decision oriented toward either a T-independent (TI) or a T-dependent (TD) response occurs. Depending upon the type of signaling

[76,77], transitional B cells differentiate either into ‘preactivated’ marginal zone (MZ) B cells residing in the splenic MZ or into ‘primed’ follicular mature (FM) B cells residing

in the splenic follicle [74,78]. In humans, the origin of MZ B cells is still debated [79], and such cells might not directly originate from the T2 lineage. Splenic MZ B cells are

specifically educated to respond rapidly to blood-borne TI antigens, and they provide a first line of ‘innate-like’ defense against specific pathogens such as encapsulated

bacteria [13,42,80,81]. Most (if not all) human MZ B cells display low levels of somatic hypermutation (SHM) in their BCR, suggesting that receptor diversification can occur

outside of the classical TD immune response and before antigen encounter [42]. Upon antigen stimulation, preactivated, -diversified, and -expanded MZ B cells rapidly

migrate into the blood either as plasma cells or as circulating IgM ‘memory’ B cells [42,81]. By contrast, FM B cells are not yet effector cells when they exit the spleen and

recirculate in the bloodstream. These ‘naıve, mature’ B cells have not yet encountered cognate antigen, but they are primed to functionally respond to TD stimulation, a

reaction taking place in the follicles of peripheral lymphoid organs [82]. When naıve, mature B cells enter the peripheral lymphoid organs, they reach a primary follicle and

scan for their specific antigen. Antigen-activated B cells then rapidly relocalize to the B-T interfollicular area, where, upon antigen-primed T-cell signaling via cognate and

CD40-CD40L interaction, they undergo a limited expansion. A fraction of the cells differentiate into short-lived, ‘innate-like,’ unswitched and unmutated IgM plasma cells,

providing a first rapid response to invading pathogens, wheras others initiate the formation of a GC in a secondary follicle. In the dark zone of the GC, the activated B cells

downregulate surface BCR and BCL2 expression, proliferate vigorously as centroblasts (cb), and initiate the program of affinity maturation, including SHM of the

immunoglobulin genes. In most cases, the introduced mutations result in a reduction of the BCR affinity to the cognate antigen, whereas only a few generate a BCR of

higher affinity. The transition from centroblast to centrocyte (cc) is accompanied by cycling arrest, migration to the light zone, and by yet another crucial cell-fate decision.

The modified surface BCR is reexpressed and ‘checked’ by loaded follicular helper T cells (TFH) and DCs [83]. Depending on the new BCR affinity, centrocytes reenter cycles

of SHM as centroblasts (unchanged affinity), are eliminated by apoptosis (lower affinity), or undergo CSR and complete the final GC maturation stages (higher affinity).

Fully matured, switched, and hypermutated follicular B cells upregulate BCL2 and exit the GC either as ‘switched memory’ B cells bearing an IgG, IgA, or IgE of higher

affinity for the cognate antigen or, upon massive expansion and further differentiation, as short-lived IgG-, IgA-, or IgE-secreting plasma cells. A fraction of the plasma cells

recirculate in specific BM niches and acquire an extended life span (few months) [82]. Switched memory B cells, however, recirculate in the peripheral lymphoid organs (and

possibly the BM) [84,85] and are the key regulators of the long-term memory response to antigen recall. Upon rechallenge, two response modes take place: (1) in the

classical ‘antigen-dependent mode,’ antigen-specific, switched memory B cells are reactivated in a cognate fashion by the corresponding antigen-primed T cells and

undergo a new cycle of extensive proliferation and differentiation into plasma cells; (2) in a recently described ‘antigen-independent mode,’ polyclonal stimulation of

memory B cells is simultaneously triggered by bystander T-cell help provided in an antigen-independent fashion (e.g. via CD40-CD40L interaction and cytokine production),

ultimately leading to the (more limited) proliferation of memory B cells bearing unrelated specificity to the challenging antigen, and differentiation into a new generation of

secreting plasma cells [86]. Ongoing polyclonal activations of the entire pool of memory B cells during the successive antigenic challenges of an individual’s immunological

history are currently proposed to account for the observed long-term production of the broad spectrum of acquired antibody specificities, and thus to the maintenance of

serological memory for a human lifetime [26].

26 Review TRENDS in Immunology Vol.29 No.1

www.sciencedirect.com

Review TRENDS in Immunology Vol.29 No.1 27

protracted stimulation of antigen-specific B cells by patho-gens establishing a chronic infection leads to the emer-gence of transformed B cells, owing to the occurrenceof progressive oncogenic events in antigen-respondingB cells. The anatomical location of these B cells (i.e. theMZ of peripheral lymphoid organs, exposed to invadingpathogens), in addition to the functional similarities withMZ B cells (i.e. rapid proliferation after antigenic stimu-lation) [13], has led to the hypothesis that neoplastic cellsin MZ lymphomas derive from their normal MZ B-cellcounterparts [14]. As described in Figure 1, MZ B cellsparticipate in T-independent (TI) responses. However,several pathological aspects of infection-associated lym-phomas question the linear relationship between a TI-onlyresponse, MZ B cells, and lymphoma development, andthey suggest that the antigen-driven lymphomagenesiscould, in fact, involve either TI responses, T-dependent(TD) responses, or both.

H. pylori-associated gastric MALT lymphoma

H. pylori-associated gastric MALT lymphoma remains thearchetype of infection-associated indirect transformationof lymphocytes [15]. The majority of lymphomas involvingthe stomach are MALT lymphomas (which are also themost frequent MALT-type lymphomas) [16] and arestrongly associated with H. pylori [6]. The bacterium ispresent in biopsy samples in the majority of cases, andantibiotic eradication of H. pylori leads to completeregression of MALT lymphoma in over 80% of cases[17–19]. It is now clear that MALT lymphoma developson a background of chronic gastritis. Normal gastricmucosa is devoid of lymphoid tissue, but infection of thestomach withH. pylori leads to chronic gastritis in which alymphoid tissue develops [16]. Further evidence of thecausal relationship between H. pylori infection and theemergence of MALT lymphoma is obtained from the retro-spective detection of malignant B-cell clones in preexistingH. pylori-associated chronic gastritis [20]. Histologically,the neoplastic cells retain features of the MZ B cells fromwhich they are thought to derive [13]. The MZ is expandedby small neoplastic lymphocytes (centrocyte-like cells) thattend to extend into the adjacent epithelial mucosa, produ-cing characteristic lymphoepithelial lesions. However, ithas been shown that neoplastic MZ B cells from gastricMALT lymphomas are not specific for H. pylori antigens,but rather for gastric-mucosa-associated autoantigenspotentially exposed during chronic inflammation [21,22].Moreover, and in contradiction to a classical TI response,CD4 T cells isolated from gastric MALT lymphoma associ-ated with H. pylori directly respond in vitro to peptidesderived from H. pylori and provide proliferative signals tomalignant B cells through CD40L-CD40 interactions [23].If H. pylori-specific T cells are required, should MALTlymphoma development be considered a TI or a TDresponse? Are the MZL B cells truly malignant counter-parts of MZ B cells that are able to participate in a TDresponse, or are they germinal center (GC) B cells that arehoming in the MZ and display an MZ phenotype andfunction [24,25]? Lanzavecchia and colleagues haverecently provided a very elegant answer to this dilemma:similar to the bystander T-cell help provided to switched


memory B cells in the antigen-independent mode of anti-gen recall (see Figure 1), bystander T-cell help can also beprovided in a noncognate fashion to MZ B cells outside of aGC reaction (Figure 2) [26]. In the case of MALT lym-phoma, it is possible that autoreactive MZ B cells arestimulated at low levels by autoantigens, but additionalstimuli are required for their enhanced proliferation andultimate transformation.H. pylori-specific T cells raised intheGC-dependent TD response of theMALT interfollicularzonewould thenmigrate to theMZ and provide noncognatebystander T-cell help to the autoreactive MZ B cellsaccumulating in response to the chronic inflammation.Here, antigen-driven lymphoproliferation would thusresult from the integration of two enhanced signals, bothowing to the persistence ofH. pylori: (1) a chronic gastritis-mediated TI response generating autoreactive MZ B cells,owing to their frequent autoreactive and crossreactiverepertoire; (2) a prolonged TD response, generating asustained pool of H. pylori-specific T cells. The extendedintermingling of both TI and TD responses would, in thisscenario, be at the origin of H. pylori-associated MALTlymphomagenesis, illustrating the fine line between thetwo response modes. Interestingly, the antigen is notpermanently required to drive the lymphoproliferation.Indeed, antigen signaling through the B-cell receptor(BCR) can be bypassed by constitutive activation of theNF-kB pathway, mostly via BCL10 upregulation duringrecurrent genetic aberrations such as t(11;18)/API2-MALT, t(1;14)/BCL10-IGH, and t(14;18)/IGH-MALT1.Several excellent reviews have described in great detailthe mechanism and the role of such translocations inMALT lymphomagenesis and in the progression to anti-microbial-insensitive tumors [3,27–30].

HCV-associated SLVL

SMZLand its variant SLVLpresent as indolent lymphomascharacterized by the replacement of the normal splenicarchitecture by neoplastic B cells with a distinct phenotype(IgM+IgD+/�CD19+CD20+CD5�CD10�CD22+CD11c+/�),leading to an expansion of the splenicMZ and splenomegaly[31,32]. Autoimmune features and, particularly, a mono-clonal IgM, are frequently encountered.

A minority of SLVL are associated with chronic HCVinfection, and most of these patients present with sympto-matic mixed cryoglobulinemia type II (MCII), a chronicimmune-complex-mediated disease with underlying clonalB-cell expansion with rheumatoid factor (RF) activity[10,11]. The causal relationship between viral replicationand lymphoproliferation is supported by the parallelmodulation of viral load and neoplastic lymphocytosis.Complete virological response after antiviral treatmentis associated with the disappearance of circulating villouslymphocytes, and with the resolution of the splenomegalyand the MCII. Virological relapse is associated with thereappearance of abnormal lymphocytosis and splenome-galy. Additional antiviral treatment produces further hem-atological response if it is associated with a reduction ofHCV viral replication [10,11].

In the case of HCV-associated lymphomas, there issome evidence that malignant MZ B cells directly respondto HCV antigens. Indeed, the BCR of two lymphomas

Figure 2. Models of bystander T-cell help within TI and TD response pathways. (a) Antigen-driven MALT lymphomagenesis: a model of bystander T-cell help outside the

germinal center (GC). H. pylori-specific T cells (TFH) raised in the GC-dependent TD response of the MALT interfollicular zone migrate to the MZ and provide noncognate

bystander T-cell help to autoreactive MZ B cells stimulated at low levels by autoantigens and accumulating in response to chronic inflammation. (b) HCV infection: a model

of TI and TD response modes in antigen-driven lymphoproliferation. In the TI response, autoreactive MZ B cells are stimulated in response to HCV infection, owing to

molecular mimicry between the HCV E2 envelope protein and human immunoglobulin. Sustained crossreactive antibody production leads to lymphoproliferation, MCII,

and ultimately SLVL, which might ultimately transform to DLBCL of MZ origin. In parallel, a classical TD response occurs in the GCs. Polyclonal stimulation of follicular B

cells due to bystander help and/or direct HCV E2 protein binding to the CD81/CD19/CD21 complex might lead to preferential proliferation of t(14;18)+ clones and progression

to FL or DLBCL of GC origin.


associated with HCV were found to be specific for the E2glycoprotein of HCV [33]. Moreover, tumor-related immu-noglobulin preferentially uses the VH1-69 heavy-chaingene, similar to normal B cells responding to the viralE2 protein and to RF [34,35]. The direct relationshipbetween viral load, MCII, and tumor burden, as well assequence analysis of the BCR from HCV-associated lym-phomas [36], favors the hypothesis that cryoglobulinsare produced by the tumor clone. The fact that thereappears to be mimicry between the E2 protein and humanimmunoglobulin also supports this hypothesis [37],because antibodies against E2 probably crossreact with


immunoglobulins and display RF activity. Thus, HCV-associated SLVL could represent a true model ofantigen-driven lymphoma developing from a TI response.B-cell stimulating cytokines such as BAFF, produced bydendritic cells (DCs) andmacrophages in theMZ sinuses ofthe spleen, could provide additional proliferative impulseto MZ B cells during HCV infection, because HCV has beenfound to infect DCs through DC-SIGN [38].

Does HCV drive only TI lymphoproliferation? HCV in-fection triggers a complex immunological response invol-ving both TI and TD arms [39], and there are reasons tobelieve that the TD response is also involved in parallel


antigen-driven lymphoproliferation. First, HCV infectionis associated with several lymphoma types besides SMZL,including FL and de novo diffuse large B-cell lymphomas(DLBCL) of both MZ and GC origin [40]. Second, similarlyto SLVL [10], the immunomodulation of potential FL pre-cursors has also been observed in HCV patients uponvirological response to antiviral treatment (see below)[41]. The observed predominance of TI lymphoproliferationover the TD typemight partly be explained bya difference oflatency in the emergence of a malignant clone of MZ versusGC origin, in agreement with the seminal features of thecorresponding response modes. Interestingly, in contrast tothe antigen-independent prediversification of the MZ B-cellrepertoire [42], �30% of SMZLs harbor unmutated immu-noglobulin genes [43–45]. As previously observed in B-CLL,this suggests a possible heterogeneity in SMZL pathogen-esis and raises the important issue of a potential ‘borderline’origin of this unmutated subset [46]. Althoughmost reportsseem to indicate that HCV-associated lymphomas aremutated [36,47], further studies will be necessary to inves-tigate the TI or TD response mode of unmutated versusmutated SMZLs, and to clarify the involvement of eachsubset in antigen-driven lymphoproliferation.

Early steps of follicular lymphomagenesis: a protractedpreclinical state?At the other end of the spectrum, FL pathogenesis standsas a paradigm of B-cell lymphoma derived from a TD

Figure 3. Early steps of FL genesis: a protracted preclinical state? In the current model of

early B-cell development in the BM [48]. Yet, the translocation does not prevent furthe

process from pre- to immature B cells, then exit the BM. It is probable that t(14;18)+ cells

become MZ B cells. However, virtually nothing is known about the fate of such cells.

classical TD response pathway. t(14;18)+ follicular cells are driven into secondary follic

normally downregulate BCL2 to allow for elimination of B cells bearing low-affinity rec

their rescue from apoptosis. In the absence of appropriate signals for further matura

secondary follicles, occasionally re-entering cycles of SHM and CSR, delaying GC wanin

follicular B cells are released from the GC and enter the bloodstream as so-called FL-lik

along with the existence of niches, in which the follicular microenvironment might provi

antigenic stimulation. Ultimately, extended survival of BCL2+ cells and intensive tra

transformation to FL.


response. FL is the second most common form of non-Hodgkin’s lymphoma and accounts for �20%–30% percentof all cases [5]. Although the median length of survival is�10 years, the clinical course is widely variable. Mostpatients have an indolent form of the disease that slowlyprogresses over a period of many years, with waxing andwaning lymphadenopathy. The patients are generallydiagnosed at an advanced stage (III or IV) with dissemi-nated presentation involving both lymph nodes (LN) andbone marrow (BM) [5].

FL is a mature B-cell neoplasm, resulting from themalignant transformation of follicular B cells. Pathogen-esis is initiated by the ectopic expression of the antiapop-totic BCL2 protein, owing in more than 85% of the cases toits juxtaposition to the strong immunoglobulin heavy-chain enhancer via the t(14;18)(q32;q21) chromosomaltranslocation [48]. Despite the clear follicular B-cell phe-notype and localization of the tumor [5], molecularanalyses have revealed that FL genesis is not initiatedin the GCs, and that progression to FL pathogenesis is acomplex, multistep pathway escalating along severalstages of B-cell differentiation and over extended periodsof time (detailed in Figure 3). The expansion of FLresembles, both morphologically and functionally, normalGC B-cell growth because centroblast-like and centrocyte-like tumor cells proliferate in networks including follicularDCs and follicular helper CD4+ T cells (TFH). FL cells alsoretain the features of normal follicular B cells, including

FL genesis, t(14;18) translocation occurs as an error of V(D)J recombination during

r stages of lymphoid differentiation, and t(14;18)+ cells continue their maturation

follow the splenic transit of normal B cells, and that a fraction will receive signals to

Upon antigenic challenge, naıve mature t(14;18)+ cells are activated and enter the

les, where they undergo proliferation, SHM, and CSR. In the GC, follicular B cells

eptors. However, in t(14;18)+ follicular B cells, enforced expression of BCL2 allows

tion, such frozen t(14;18)+ B cells with low-affinity receptors then accumulate in

g, and eventually leading to follicular hyperplasia over time. Occasionally, t(14;18)+

e cells. The persistence and clonal expansion of such cells over many years goes

de support for maintenance and proliferation in the context of polyclonal or chronic

fficking between follicles might favor additional genetic changes necessary for


the dependence on BCR expression and activation, as wellas the interaction with T cells and/or stromal cells in thefollicular microenvironment [1]. Because of the presence ofconstitutive activation-induced cytidine deaminase (AID)expression [49], FLs usually display high levels of somatichypermutation (SHM) and class switch recombination(CSR) both on the expressed immunoglobulin heavy-chainallele and on the translocated BCL2-immunoglobulinheavy-chain locus [50]. Interestingly, and despite thestrong selection pressure to form a niche in reactive GCsowing to their dependency on the microenvironment forsurvival and growth signals, FL clones are still prone tointense trafficking between follicles, including within theLN and BM [51–53]. Although the reason for this traffick-ing is not yet well understood, it is possible that thepremalignant clone might not be able to sustain appro-priate levels of signaling in the founder follicle, and that itneeds therefore to opportunistically invade other morerecently formed reactive follicles.

Although t(14;18) translocation, ectopic BCL2 expres-sion, and follicular hyperplasia are crucial early events inthe natural history of FL pathogenesis, the acquisition ofsynergistic oncogenic events is nevertheless clearlyrequired for full malignant transformation. Accordingly,FL samples usually display many additional chromosomalalterations. SHMandCSR activity [49] have been shown toconfer a high propensity for genomic instability [1], and thesustained survival and growth of frozen follicular B cellsthus provide highly favorable grounds for the acquisition ofsuch complementary oncogenic lesions [54]. Nevertheless,the random acquisition of synergistic oncogenic eventsseems to be a very lengthy process. In support of this view,Em-bcl-2 transgenic mice mimicking the t(14;18) translo-cation uniformly develop follicular hyperplasia, but only afew progress to aggressive monoclonal B-cell lymphomaswith a relatively long latency period and often in conjunc-tion with cooperating genetic lesions [55,56]. In agreementwith data from transgenic mice, t(14;18) is also detected ata low frequency in human tonsils in the absence ofmanifestclinical lymphoma [57]. t(14;18) clones are also present inthe peripheral blood of most healthy individuals [58,59],where they can persist for more than 10 years, with bothprevalence and frequency increasing significantly overtime [60]. Interindividual frequencies are, however, veryvariable in the natural human population, and they rangefrom less than 1 t(14;18)+ cell in a million B cells in someindividuals, to more than 1 in 4000 in others. Surprisingly,the high t(14;18) frequencies observed in some healthyindividuals do not result from transient accumulation ofclonally unrelated naıve B cells as previously thought, butfrom the clonal expansion of GC B cells bearing the phe-notypic and genotypic features of FL cells [61]. Althoughthe relationship between the presence of such ‘FL-like’ cellsin blood and the stepwise progression to disease is stillunclear, it is probable that t(14;18)+ cells represent earlyintermediates of the FL pathway that have not (yet?)acquired the malignant phenotype. Circulating FL-likecells were indeed probably rescued from apoptosis byBCL2, frozen as follicular B cells, selected in the GCs,and released in the bloodstream possibly in search of ad-equate niches. The persistence and clonal expansion of


t(14;18)+ cells over periods of time of more than 10 yearsin healthy individuals support the existence of such niches,in which the combination of ectopic BCL2 expression,maintenance of the BCR, and factors provided from thefollicular microenvironment might support maintenanceand proliferation in the context of antigenic stimulation.Ultimately, thematuration arrest of follicular B cells proneto genomic instability, combined with BCL2-mediated pro-tracted survival and a capacity for intensive trafficking,might favor the acquisition of the additional geneticchanges necessary for transformation to indolent FL overtime.

An alternative pathway of antigen-driven follicularlymphomagenesis?Several groups of individuals have been associated withhigh rates of t(14;18) in peripheral blood. In particular,t(14;18) frequency has been shown to be increased inpopulations of healthy individuals occupationally exposedto risk factors, such as genotoxic agents (e.g. pesticides ormycotoxins) [62]. However, agricultural pesticide use hasalso been repeatedly associated with risk of non-Hodgkin’slymphoma, particularly of the t(14;18)+ subtype [63]. Thesedata modify the perception of the role of genotoxics in FLpathogenesis, because increased t(14;18) frequency inexposed individuals, thus far perceived as transitoryaccumulation of t(14;18)+ cells with restricted oncogenicpotential, could in fact reflect more advanced steps oflymphomagenesis. Although t(14;18) is found in a pro-portion of HCV patients, especially in the presence of MCII[64,65], BCL2 probably does not play a direct role in HCV-associated lymphomas, because it is not found in themajority of HCV+ SLVL [10,11]. However, and in a strikingparallel to the HCV-driven SLVL lymphoproliferation,Giannelli et al. reported the parallel disappearance oft(14;18) detection with complete virological response uponantiviral treatment of HCV infected patients [41]. Virolo-gical relapse was accompanied by the reemergence of thesame initial t(14;18) clone. Our recent data showing clonalexpansion of FL-like B cells in healthy individuals suggestthat the interindividual t(14;18) frequency modulation isdue to an associated antigenic stimulation process than toan increase in breakage, provoking the accumulation ofnew translocations.

Can a parallel then be drawn between antigen-drivenlymphomagenesis in MZL and FL genesis? The idea ofprotracted or chronic antigen stimulation is indeed appeal-ing when considering lymphomagenesis in which the mainfeature is an acquired loss of BCL2-dependent apoptosis.The current understanding of FL genesis seems, however,in apparent contradiction with this idea. As describedabove, BCR affinity is randomized by SHM in the GCs,and cells ending up with a lower-affinity BCR die byapoptosis. However, in the presence of enforced BCL2expression, such cells are selectively rescued, and anabnormal population of follicular B cells with a low-affinityBCR accumulates in the GCs [66]. This strongly suggeststhat t(14;18)+ B cells developing in the GC also bear BCRswith poor specificity for the original antigen, and, by thisrationale, their development cannot be due to prolonged orchronic antigen-specific stimulation. A conceptual paradox

Figure 4. A model of bystander T-cell help in preclinical FL development. During antigenic rechallenge, bystander T-cell help is indiscriminately delivered in follicles to

switched memory B cells and to accumulating BCL2-rescued follicular B cells (FO). Top: generation of long-term memory via polyclonal stimulation of switched memory B

cells through noncognate bystander T-cell help. Bottom: protracted or chronic antigenic stimulation of FL-like t(14;18)+ B cells through noncognate bystander T-cell help as

propitious events for the stepwise progression to FL. Somatic hypermutation of the FL BCR can lead to its ‘crippling’ (i.e. weakly reactive to the recall antigen), but FL

expansion can still be driven through noncognate stimulation by T cells.


then emerges: despite the accumulation of t(14;18)+ B cellswith low-affinity receptors, most, if not all, FL malignantcells still express a surface BCR [67]. This stronglysuggests that although high-affinity antigenic recognitionis dispensable, signaling through the BCR is absolutelyrequired to sustain FL or FL-like survival and/or growth.How then could this be achieved? One potential way topromote survival and proliferation through a low-affinityBCR would be to significantly decrease the signalingthreshold. This might occur through several mechanisms.One mechanism involves the presence of costimulatorymolecules in some immune responses. During chronicHCV infection, for example, crosslinking of the BCRthrough the interaction of the E2 viral envelope proteinwith the CD81/CD19/CD21 coreceptor complex has beenshown to mediate activation of naıve B cells [68], and itcould similarly lower the activation threshold of accumu-lating follicular t(14;18)+ B cells [69]. Another mechanisminvolves enhanced T-cell help via a noncognate CD40-CD40L interaction and cytokine production. The acceler-ated FL genesis observed in VavP-BCL2 transgenic mice,in which the pan-hematopoietic expression of the trans-gene allows for significant enlargement of the peripheralT-cell compartment, supports a crucial role of enhancedT-cell help in FL development [70]. In humans, the seminalobservation that, besides their classical role in the TDresponse, activated T cells also lead to the polyclonalproliferation of bystander memory B cells carrying unre-lated specificity provides a general framework bywhich the


maintenance of BCR signaling could be achieved forextended periods of time in follicular BCL2+ B cells witha low-affinity BCR (Figure 4): during antigenic rechal-lenge, bystander T-cell help would be indiscriminatelydelivered in follicles to switched memory B cells and toaccumulating BCL2-rescued follicular B cells. In thisscenario, persisting antigens would provide sufficient sig-naling through the BCR to promote accumulation andproliferation of BCL2+ B cells in reactive GCs, delayingtheir waning until the occurrence of the next antigenicboost. In the context of HCV infection, chronic inflam-mation might favor noncognate expansion of t(14;18)-bear-ing clones (Figure 2b). As previously proposed for theemergence of MALT lymphoma, this subversion of themechanism of maintenance of serological memory [26]suggests that protracted or chronic antigenic stimulationsare propitious events for the stepwise progression to FL.This, however, does not imply that the t(14;18)+ clone isdirected against the persisting antigen. In antigen-drivenMZLs, B-cell lymphomas rarely directly respond to thechronic antigen itself, but rather express opportunisticcross(auto)reactive specificities.

Is then the persistence of the cognate antigen thatinitially mounted the t(14;18)+ response absolutelyrequired in FL progression? Probably not. The constantexposure to SHM of genes encoding the BCR could lead tothe acquisition over time of crossreactive specificities,such as auto- or poly-reactivity. One interesting recentobservation by Stevenson and colleagues is that in FL,


N-glycosylation sites are recurrently introduced by SHM inthe antigen-binding pocket of the BCR [71]. The density ofsuch sites in FL contrasts with the one found in normal GCB cells, suggesting that they are either counterselected innormal follicular B cells or positively selected in FL. Bio-chemical analyses have shown that the sites are occupiedby oligomannose glycans, possibly subverting conventionalantigen binding toward mannose-binding lectins presentat the surface of cells from the tumor microenvironmentsuch as DC, macrophages, or other stromal cells [71]. Inthis scenario, BCR signaling might acquire a completeautonomy from the initial cognate antigen over time.

In conclusion, the concept of antigen-driven lymphoma-genesis is more heterogeneous than initially anticipated: itcan involve a TI response, a TD response, or the integrationof both, and it can act through a direct BCR-mediatedstimulation of the malignant cell by the cognate chronicantigen or through polyclonal or noncognate bystanderactivation of crossreactive malignant B cells. In thisrapidly moving field, further understanding of the mech-anisms by which antigen-driven lymphoproliferation ariseshould, in the near future, offer insights into the design ofnew intervention strategies aimed at preventing themalig-nant evolution of preneoplastic lesions and controlling theclinical course of certain low-grade B-cell lymphomas [72].

AcknowledgementsWe are grateful to C. Schiff for helpful discussions. We thank C. Beziers-Guigue for help with illustrations. This work was supported by grants#7877, #3618, and #3157 from the Association pour la Recherche sur leCancer (ARC); by a grant from Le Conseil General des Bouches du Rhone;by grant #PL06-010 from the Institut National du Cancer (INCa); and byinstitutional grants from the Institut National de la Sante et de laRecherche Medicale (INSERM) and the Centre National de la RechercheScientifique. Work in the B.N. lab is supported by the AVENIR2003 grantfrom the INSERM. S.R. is a recipient of a fellowship from the INCa. B.N.is a recipient of a Contrat d’Interface INSERM/Assistance Publique-Hopitaux de Marseille.

References1 Kuppers, R. (2005) Mechanisms of B-cell lymphoma pathogenesis.Nat.

Rev. Cancer 5, 251–2622 Shaffer, A.L. et al. (2002) Lymphoid malignancies: the dark side of

B-cell differentiation. Nat. Rev. Immunol. 2, 920–9323 Suarez, F. et al. (2006) Infection-associated lymphomas derived from

marginal zone B cells: a model of antigen-driven lymphoproliferation.Blood 107, 3034–3044

4 Fisher, S.G. and Fisher, R.I. (2006) The emerging concept of antigen-driven lymphomas: epidemiology and treatment implications. Curr.Opin. Oncol. 18, 417–424

5 Harris, N.L. et al. (1994) A revised European-American classification oflymphoid neoplasms: a proposal from the International LymphomaStudy Group. Blood 84, 1361–1392

6 Wotherspoon, A.C. et al. (1991) Helicobacter pylori-associated gastritisand primary B-cell gastric lymphoma. Lancet 338, 1175–1176

7 Lecuit, M. et al. (2004) Immunoproliferative small intestinal diseaseassociated with Campylobacter jejuni. N. Engl. J. Med. 350,239–248

8 Goodlad, J.R. et al. (2000) Primary cutaneous B-cell lymphoma andBorrelia burgdorferi infection in patients from the Highlands ofScotland. Am. J. Surg. Pathol. 24, 1279–1285

9 Ferreri, A.J. et al. (2004) Evidence for an association betweenChlamydia psittaci and ocular adnexal lymphomas. J. Natl. CancerInst. 96, 586–594

10 Hermine, O. et al. (2002) Regression of splenic lymphoma with villouslymphocytes after treatment of hepatitis C virus infection. N. Engl. J.Med. 347, 89–94


11 Saadoun, D. et al. (2005) Splenic lymphoma with villous lymphocytes,associated with type II cryoglobulinemia and HCV infection: a newentity? Blood 105, 74–76

12 Kuppers, R. (2003) B cells under influence: transformation of B cells byEpstein-Barr virus. Nat. Rev. Immunol. 3, 801–812

13 Pillai, S. et al. (2005) Marginal zone B cells. Annu. Rev. Immunol. 23,161–196

14 Morse, H.C., III et al. (2001) Cells of the marginal zone–origins,function and neoplasia. Leuk. Res. 25, 169–178

15 Sagaert, X. et al. (2007) The pathogenesis of MALT lymphomas: wheredo we stand? Leukemia 21, 389–396

16 Cavalli, F. et al. (2001) MALT lymphomas. Hematology (Am. Soc.Hematol. Educ. Program), 241–258

17 Wotherspoon, A.C. et al. (1993) Regression of primary low-grade B-cellgastric lymphoma of mucosa-associated lymphoid tissue type aftereradication of Helicobacter pylori. Lancet 342, 575–577

18 Wotherspoon, A.C. et al. (1994) Antibiotic treatment for low-gradegastric MALT lymphoma. Lancet 343, 1503

19 Bayerdorffer, E. et al. (1995) Regression of primary gastric lymphomaof mucosa-associated lymphoid tissue type after cure of Helicobacterpylori infection. MALT Lymphoma Study Group. Lancet 345,1591–1594

20 Zucca, E. et al. (1998) Molecular analysis of the progression fromHelicobacter pylori-associated chronic gastritis to mucosa-associatedlymphoid-tissue lymphoma of the stomach. N. Engl. J. Med. 338,804–810

21 Zucca, E. et al. (1998) Autoreactive B cell clones in marginal-zone B celllymphoma (MALT lymphoma) of the stomach. Leukemia 12, 247–249

22 Bende, R.J. et al. (2005) Among B cell non-Hodgkin’s lymphomas.MALT lymphomas express a unique antibody repertoire withfrequent rheumatoid factor reactivity. J. Exp. Med. 201, 1229–1241

23 Hussell, T. et al. (1993) The response of cells from low-grade B-cellgastric lymphomas of mucosa-associated lymphoid tissue toHelicobacter pylori. Lancet 342, 571–574

24 Weller, S. et al. (2005) Splenic marginal zone B cells in humans: wheredo they mutate their Ig receptor? Eur. J. Immunol. 35, 2789–2792

25 Willenbrock, K. et al. (2005) Human splenic marginal zone B cells lackexpression of activation-induced cytidine deaminase. Eur. J. Immunol.35, 3002–3007

26 Bernasconi, N.L. et al. (2002) Maintenance of serological memory bypolyclonal activation of humanmemory B cells.Science 298, 2199–2202

27 Isaacson, P.G. and Du, M.Q. (2004) MALT lymphoma: frommorphology to molecules. Nat. Rev. Cancer 4, 644–653

28 Farinha, P. and Gascoyne, R.D. (2005) Molecular pathogenesis ofmucosa-associated lymphoid tissue lymphoma. J. Clin. Oncol. 23,6370–6378

29 Bertoni, F. and Zucca, E. (2006) Delving deeper into MALT lymphomabiology. J. Clin. Invest. 116, 22–26

30 Du, M.Q. and Atherton, J.C. (2006) Molecular subtyping of gastricMALT lymphomas: implications for prognosis and management. Gut55, 886–893

31 Troussard, X. et al. (1996) Splenic lymphoma with villous lymphocytes:clinical presentation, biology and prognostic factors in a series of 100patients. Groupe Francais d’Hematologie Cellulaire (GFHC). Br. J.Haematol. 93, 731–736

32 Thieblemont, C. et al. (2003) Splenic marginal-zone lymphoma: adistinct clinical and pathological entity. Lancet Oncol. 4, 95–103

33 Quinn, E.R. et al. (2001) The B-cell receptor of a hepatitis C virus(HCV)-associated non-Hodgkin lymphoma binds the viral E2envelope protein, implicating HCV in lymphomagenesis. Blood 98,3745–3749

34 Chan, C.H. et al. (2001) V(H)1-69 gene is preferentially used byhepatitis C virus-associated B cell lymphomas and by normal B cellsresponding to the E2 viral antigen. Blood 97, 1023–1026

35 Carbonari, M. et al. (2005) Hepatitis C virus drives the unconstrainedmonoclonal expansion of VH1-69-expressing memory B cells in type IIcryoglobulinemia: a model of infection-driven lymphomagenesis.J. Immunol. 174, 6532–6539

36 De Re, V. et al. (2000) Sequence analysis of the immunoglobulinantigen receptor of hepatitis C virus-associated non-Hodgkinlymphomas suggests that the malignant cells are derived from therheumatoid factor-producing cells that occur mainly in type IIcryoglobulinemia. Blood 96, 3578–3584


37 Hu, Y.W. et al. (2005) Immunoglobulin mimicry by Hepatitis C Virusenvelope protein E2. Virology 332, 538–549

38 Cormier, E.G. et al. (2004) L-SIGN (CD209L) and DC-SIGN (CD209)mediate transinfection of liver cells by hepatitis C virus. Proc. Natl.Acad. Sci. U. S. A. 101, 14067–14072

39 Takaki, A. et al. (2000) Cellular immune responses persist and humoralresponses decrease two decades after recovery from a single-sourceoutbreak of hepatitis C. Nat. Med. 6, 578–582

40 Besson, C. et al. (2006) Characteristics and outcome of diffuse largeB-cell lymphoma in hepatitis C virus-positive patients in LNH 93 andLNH 98 Groupe d’Etude des Lymphomes de l’Adulte programs. J. Clin.Oncol. 24, 953–960

41 Giannelli, F. et al. (2003) Effect of antiviral treatment in patientswith chronic HCV infection and t(14;18) translocation. Blood 102,1196–1201

42 Weller, S. et al. (2005) Vaccination against encapsulated bacteria inhumans: paradoxes. Trends Immunol. 26, 85–89

43 Bahler, D.W. et al. (2002) Splenic marginal zone lymphomas appear tooriginate from different B cell types. Am. J. Pathol. 161, 81–88

44 Tierens, A. et al. (1998) Mutation analysis of the rearrangedimmunoglobulin heavy chain genes of marginal zone celllymphomas indicates an origin from different marginal zone Blymphocyte subsets. Blood 91, 2381–2386

45 Tierens, A. et al. (2003) Splenic marginal zone lymphoma with villouslymphocytes shows on-going immunoglobulin gene mutations. Am. J.Pathol. 162, 681–689

46 Traverse-Glehen, A. et al. (2005) Analysis of VH genes inmarginal zonelymphoma reveals marked heterogeneity between splenic and nodaltumors and suggests the existence of clonal selection. Haematologica90, 470–478

47 De Re, V. et al. (2006) HCV-NS3 and IgG-Fc crossreactive IgM inpatients with type II mixed cryoglobulinemia and B-cell clonalproliferations. Leukemia 20, 1145–1154

48 Marculescu, R. et al. (2006) Recombinase, chromosomal translocationsand lymphoid neoplasia: targeting mistakes and repair failures. DNARepair (Amst.) 5, 1246–1258

49 Greeve, J. et al. (2003) Expression of activation-induced cytidinedeaminase in human B-cell non-Hodgkin lymphomas. Blood 101,3574–3580

50 Vaandrager, J.W. et al. (1998) DNA fiber fluorescence in situhybridization analysis of immunoglobulin class switching in B-cellneoplasia: aberrant CH gene rearrangements in follicle center-celllymphoma. Blood 92, 2871–2878

51 Bognar, A. et al. (2005) Clonal selection in the bone marrowinvolvement of follicular lymphoma. Leukemia 19, 1656–1662

52 Cong, P. et al. (2002) In situ localization of follicular lymphoma:description and analysis by laser capture microdissection. Blood 99,3376–3382

53 Oeschger, S. et al. (2002) Tumor cell dissemination in follicularlymphoma. Blood 99, 2192–2198

54 Ramiro, A.R. et al. (2004) AID is required for c-myc/IgH chromosometranslocations in vivo. Cell 118, 431–438

55 McDonnell, T.J. and Korsmeyer, S.J. (1991) Progression from lymphoidhyperplasia to high-grade malignant lymphoma in mice transgenic forthe t(14; 18). Nature 349, 254–256

56 McDonnell, T.J. et al. (1989) bcl-2-immunoglobulin transgenicmice demonstrate extended B cell survival and follicularlymphoproliferation. Cell 57, 79–88

57 Limpens, J. et al. (1991) Bcl-2/JH rearrangements in benign lymphoidtissues with follicular hyperplasia. Oncogene 6, 2271–2276

58 Liu, Y. et al. (1994) BCL2 translocation frequency rises with age inhumans. Proc. Natl. Acad. Sci. U. S. A. 91, 8910–8914

59 Roulland, S. et al. (2003) BCL-2/JH translocation in peripheralblood lymphocytes of unexposed individuals: lack of seasonalvariations in frequency and molecular features. Int. J. Cancer104, 695–698

60 Roulland, S. et al. (2006) Long-term clonal persistence and evolution oft(14;18)-bearing B cells in healthy individuals. Leukemia 20, 158–162


61 Roulland, S. et al. (2006) Follicular lymphoma-like B cells in healthyindividuals: a novel intermediate step in early lymphomagenesis.J. Exp. Med. 203, 2425–2431

62 Roulland, S. et al. (2004) Characterization of the t(14;18) BCL2-IGHtranslocation in farmers occupationally exposed to pesticides. CancerRes. 64, 2264–2269

63 Chiu, B.C. et al. (2006) Agricultural pesticide use and risk of t(14;18)-defined subtypes of non-Hodgkin lymphoma. Blood 108, 1363–1369

64 Kitay-Cohen, Y. et al. (2000) Bcl-2 rearrangement in patients withchronic hepatitis C associated with essential mixed cryoglobulinemiatype II. Blood 96, 2910–2912

65 Zuckerman, E. et al. (2001) The effect of antiviral therapy on t(14;18)translocation and immunoglobulin gene rearrangement in patientswith chronic hepatitis C virus infection. Blood 97, 1555–1559

66 Smith, K.G. et al. (2000) bcl-2 transgene expression inhibits apoptosisin the germinal center and reveals differences in the selection ofmemory B cells and bone marrow antibody-forming cells. J. Exp.Med. 191, 475–484

67 Bende, R.J. et al. (2007) Molecular pathways in follicular lymphoma.Leukemia 21, 18–29

68 Rosa, D. et al. (2005) Activation of naive B lymphocytes via CD81, apathogenetic mechanism for hepatitis C virus-associated B lymphocytedisorders. Proc. Natl. Acad. Sci. U. S. A. 102, 18544–18549

69 Irish, J.M. et al. (2006) Altered B-cell receptor signaling kineticsdistinguish human follicular lymphoma B cells from tumor-infiltrating nonmalignant B cells. Blood 108, 3135–3142

70 Egle, A. et al. (2004) VavP-Bcl2 transgenic mice develop follicularlymphoma preceded by germinal center hyperplasia. Blood 103,2276–2283

71 Radcliffe, C.M. et al. (2007) Human follicular lymphoma cells containoligomannose glycans in the antigen-binding site of the B-cell receptor.J. Biol. Chem. 282, 7405–7415

72 Staudt, L.M. (2007) A closer look at follicular lymphoma. N. Engl. J.Med. 356, 741–742

73 Carsetti, R. et al. (2004) Peripheral development of B cells inmouse andman. Immunol. Rev. 197, 179–191

74 Mebius, R.E. and Kraal, G. (2005) Structure and function of the spleen.Nat. Rev. Immunol. 5, 606–616

75 Sims, G.P. et al. (2005) Identification and characterization ofcirculating human transitional B cells. Blood 105, 4390–4398

76 Loder, F. et al. (1999) B cell development in the spleen takes place indiscrete steps and is determined by the quality of B cell receptor-derived signals. J. Exp. Med. 190, 75–89

77 Allman, D. et al. (2004) Alternative routes to maturity: branch pointsand pathways for generating follicular and marginal zone B cells.Immunol. Rev. 197, 147–160

78 Su, T.T. et al. (2004) Signaling in transitional type 2B cells is critical forperipheral B-cell development. Immunol. Rev. 197, 161–178

79 Tangye, S.G. and Good, K.L. (2007) Human IgM+CD27+ B cells:memory B cells or ‘memory’ B cells? J. Immunol. 179, 13–19

80 Kruetzmann, S. et al. (2003) Human immunoglobulin M memory Bcells controlling Streptococcus pneumoniae infections are generated inthe spleen. J. Exp. Med. 197, 939–945

81 Weller, S. et al. (2004) Human blood IgM ‘memory’ B cells arecirculating splenic marginal zone B cells harboring a prediversifiedimmunoglobulin repertoire. Blood 104, 3647–3654

82 McHeyzer-Williams, L.J. et al. (2006) Checkpoints in memory B-cellevolution. Immunol. Rev. 211, 255–268

83 Vinuesa, C.G. et al. (2005) Follicular B helper T cells in antibodyresponses and autoimmunity. Nat. Rev. Immunol. 5, 853–865

84 Paramithiotis, E. and Cooper, M.D. (1997) Memory B lymphocytesmigrate to bone marrow in humans. Proc. Natl. Acad. Sci. U. S. A. 94,208–212

85 O’Connor, B.P. et al. (2002) Short-lived and long-lived bone marrowplasma cells are derived from a novel precursor population. J. Exp.Med. 195, 737–745

86 Lanzavecchia, A. et al. (2006) Understanding andmaking use of humanmemory B cells. Immunol. Rev. 211, 303–309

Pathophysiological aspects of memory B-cell development

Documents

Transcript of Pathophysiological aspects of memory B-cell development