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Autoantibodies in Chronic Lymphocytic Leukemia and Related Systemic Autoimmune Diseases

By Thomas J. Kipps and Dennis A. Carson

EUKEMIA B CELLS from patients with chronic lym- L phocytic leukemia (CLL) often make autoantibodies. Early studies by Preud'homme and Seligmann' showed that the leukemic cells from CLL patients often bear surface IgM (sIgM) that has rheumatoid factor (RF) activity, or binding activity for the Fc portion of human IgG. In another limited survey, the CLL cells from 4 of 13 patients (3 1 %) expressed sIgM with such RF activity.2 Using 12-myristate 13-acetate (PMA), Broker et a13 stimulated leukemia cells from 14 CLL patients to secrete IgM. Twelve (86%) of these cultures produced monoclonal IgM that reacted with one or more of a variety of different self antigens, including IgG, single- stranded DNA (ssDNA), double-stranded DNA (dsDNA), histones, cardiolipin and/or cytoskeletal determinants. Sthoeger et a14 instead used pokeweed mitogen (PWM) and/ or Staphylococcus aureus Cowan strain I (SAC) to induce neoplastic CLL lymphocytes to differentiate into antibody- secreting cells. Of the leukemia cell cultures from 17 CLL patients that had sufficient amounts of monoclonal Ig for testing, 9 (53%) had IgM RF and/or DNA-binding activity. Finally, Borche et a15 generated heterohybridomas with the leukemia cells from each of 23 CLL patients. Of the 12 CLL heterhybridomas that produced enough monoclonal antibody (MoAb) for testing, 7 (58%) produced IgM RF autoantibodies. Collectively, these studies indicate that the leukemia cells from most CLL patients express IgM autoantibodies, most notably RFs.

What is the significance of the frequent expression of autoantibodies in CLL? What implications, if any, does the type of antibodies expressed in CLL have for our understanding the cytogenesis and/or etiopathogenesis of this leukemia? What relationship do the autoantibodies produced by leukemia B cells have to the autoimmune diseases, such as autoimmune hemolytic anemia (AIHA) or immune thrombocytopenia (ITP)?' that frequently afflict patients with this disease? This report will address several of these questions and review some of the recently acquired information regarding antibody expression in CLL and associated systemic autoimmune diseases. First, it is helpful to review the mechanisms that contribute to antibody diversity.

GENERATION OF ANTIBODY DIVERSITY

Ig gene rearrangement. The antibody molecule consists of two polypeptide chains that are encoded by gene complexes located on different chromosome^.^^'' During B-cell development, discontinuous elements within these gene complexes undergo a series of Ig gene rearrangements to form the exons that ultimately may encode the heavy and light chains of the antibody molec~le '~- '~ (Fig 1). Generally, the first Ig gene rearrangements occur within the Ig heavy chain gene complex. More than two dozen minigenes, termed diversity segments (D),l6-I9 are positioned between six functional JH minigenes2'S2' and 100 to 200 heavy chain variable region genes (V, genes).22 Each VH gene belongs to one of at least

6 subgroups, with each subgroup comprised of V genes that share 280% nucleic acid sequence homology.23 Through Ig gene rearrangement, one or more D gene segments is joined with a single JH element.24s25 The resulting DJH complex may then rearrange with a VH gene17322,26,27 to form a VHDJH exon that may encode the variable portion of the antibody heavy hai in.^^,^^ After successful VHDJH rearrangement, one of approximately 70 K V genes (V, genes)28s34 rearrange to one of five J. minigenes:' thereby generating an exon that may encode a K light chain variable region.21,35,36 Should this rearrangement fail to generate a functional gene, then one of approximately 70 X light chain V genes (V, genes)37338 may rearrange to one of four functional JA-CA complexes3944 to generate an exon that may encode a X light chain variable region. The final products of such genetic gymnastics are the somatically generated genes that encode the two polypeptide chains of the antibody molecule.

After successful Ig gene rearrangement, a highly specialized process may introduce nu- merous mutations in the rearranged and expressed Ig V genes4548 (Fig 1). This process, called somatic hypermutation, is not triggered by the mere expression of Ig genes or B-cell p r o l i f e r a t i ~ n . ~ ~ ~ ~ ~ Rather, somatic hypermutation apparently operates only in a subset of B lymphocytes at discrete times during an immune response to antigen.48*5'v5Z Stru ctural analyses of the antibodies produced at different times in an immune response show that, at 6 to 14 days after antigenic stimulation, the Ig V genes expressed by antigen-reactive B cells may accumulate somatic mutations at rates nearly a thousand times higher than that of normal (eg, I 10-3/base pair/cell division).45,46,53-56 Thereafter, the mutation rates di- minish as the B cells differentiate into memory or plasma ~ e l l s . ' ~ - ~ ~ Selection for B cells expressing antibodies of higher affinity for antigen enhances the cumulative number of nonconservative mutations at sites that encode parts of the antibody that bind to antigen.47,52@64 Through this process, high-affinity antibodies can be generated against almost any antigen, including self antigen^.^^^^' As a consequence, B cells are always capable of making potentially pathogenic high- affinity autoantibodies and self tolerance is never assured. Thus, immunoregulatory mechanisms must perpetually o p erate to prevent production of pathogenic autoantibodies.

Somatic hypermutation.

From the Sam and Rose Stein Institute for Research on Aging, Department of Medicine, University of California, San Diego. La Jolla, CA.

Submitted August 5, 1992; accepted January 13, 1993. Supported in part by Grants No. CA-49870 and AG-04100 from

the National Institutes of Health. Address reprint requests to Thomas J. Kipps, MD, PhD, Sam and

Rose Stein Institute for Research on Aging, Department of Medicine, University of California, San Diego, La Jolla, CA 92093-0663.

0 1993 by The American Society of Hematology. 0006-49 71/93/81 10-0042$3.00/0

Blood, Vol81, No 10 (May 15). 1993: pp 2475-2487 2475

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2476 KlPPS AND CARSON

Fig 1. K light chain gene complex. (Top) The germline I light chain gene complex before lg gene rearrangement. (Middle) Somatic rearrangement of one of approximately 70 V. genes with any one of five J. joining gene segments generates an exon that may encode the K light chain variable region. (Bottom) lg somatic hypermutation may operate to introduce nucleic acid base point mutations (designated by the diamond headed lines) in and around the rearranged K variable region exon.

Ig variable region structure. Comparison of the amino acid sequences from different antibody variable regions shows three discrete segments of extreme hypervariabilit~.~~ Affinity labeling and crystallographic studies substantiated earlier contentions that the hypervariable regions on both chains fold together to form the antigen-combining site.66-70 Hence, these regions of hypervariability are designated the complementarity-determining regions (CDRs). The third CDR is generated through the recombinatorial process joining the antibody light chain V gene with the J segment, in the case of the light chain, or the VH gene with the somatically generated DJH segment of the antibody heavy hai in.'^,'^,'^,^^,^' The diversity in first and second hypervariable regions reflects, in part, differences between each of the various inherited antibody V genes.22,26,34,72-78 As discussed above, somatic hypermutation subsequent to antibody V gene rearrangement may also play an important role in increasing the amino acid sequence diversity noted within these regions.

Together, the CDRs of both the antibody heavy and light chains fashion the specificity of the Ig molecule. Owing to the need for antibodies to bind a diverse array of different pathogens, there exists great potential for diversity in the ter- tiary structures of these regions. This potential is reflected by the fact that the combining site of each antibody may possess determinants of unique specificity that, in turn, may be recognized by anti-idiotypic antibodies.

AUTOANTIBODY-ASSOCIATED CROSS-REACTIVE IDIOTYPES IN CLL

Despite the tremendous potential for diversity, the antibodies produced by leukemia B cells of unrelated CLL patients often share common idiotypic determinants. Initially identified using highly absorbed heterologous antisera,79 and then more recently using murine M O A ~ S , ~ ~ ' ~ these common

idiotopes, designated cross-reactive idiotypes (CRIs), were defined initially on IgM autoantibodies such as RFs. In an early study of more than 30 CLL patients, 5 of 20 (25%) with K light chain-expressing CLL had malignant cells that expressed a CRI defined by 17.109, a mouse MoAb raised against an IgM RF Furthermore, approximately 20% of both K and X light-chain-expressing CLL were found to react with G6,85,86 an MoAb specific for an Ig heavy- chain-associated CRI present on several RF paraproteins." Yet, an additional MoAb specific for a distinct RF heavy- chain-associated CRI, named Lc I ,86*88 labeled neoplastic Ig- expressing B cells from 7 of 56 (1 3%) patients with CLL and related B-cell lymphomas.86 Another 2 cases (4%) in this study were found to have malignant cells reactive with B6, a third MoAb specific for an autoantibody heavy-chain-associated CRI that is distinct from either the G6-CRI or L c I - C R I . ~ ~ , ~ ~ More recently, additional autoantibody-associated CRIs have also been found to be expressed frequently in CLL and related B-cell neoplasms.w391

Molecular characterization of several such CRIs shows that each may reflect expression of a conserved Ig V gene with little or no somatic mutation. For example, 17.109-reactive leukemia cells from unrelated CLL patients express Ig V, genes that are highly homologous with a conserved germline V,3 gene designated H ~ m k v 3 2 5 . ~ ~ This highly conserved Ig V, gene is but 1 of 70 Ig V, genes found present in the human haploid genome. Similarly, nucleic acid sequence analyses of the Ig heavy chain genes expressed by G6-reactive leukemic cells of unrelated CLL patients shows that each is homologous to a VH1 gene frequently used by human fetal splenocytes designated 5 lp l .93,94 5 l p l is but one of an estimated 20 to 60 VH genes that comprise the v H 1 gene subgroup.22 How- ever, despite expressing homologous VH l genes, G6-reactive leukemic cells have been found to use JH3, JH4, J H ~ , or JH6




AUTOANTIBODIES IN CLL 2477

gene segments and markedly different D segments. Further- more, comparisons of G6-reactive and G6-negative antibody heavy chains indicate that the G6-CRI may be relatively re- silient to substitutions within CDR3, but affected by differences in CDRl and CDR2 that are found encoded by other VH genes of the v H 1 subgroup. As such, the G6-CRI in CLL is an excellent marker for expression of 51pl with little or no somatic mutation. Accordingly, the frequent expression of such CRIs in CLL implies that the leukemia B cells in this disease express a highly restricted repertoire of Ig V genes that have not diversified significantly from the germline DNA.

lg GENE REARRANGEMENT IN CLL

Analyses of CLL B cells not selected for expression of antibodies with particular binding specificities or CRIs show a generalized bias in the rearrangement and expression of select Ig V genes. Several investigators have noted that the Ig VH genes of the relatively small VH subgroups, vH4 and VH5, and the single copy vH6 gene are rearranged in CLL at fre- quencies that are disproportionate to the relative sizes of these subgroups in the germline DNA.5,95-97 On the other hand, the largest VH gene subgroup, the vH3 gene family, seems underrepresented among the VH gene used in CLL.96-98

Furthermore, within an Ig VH gene subgroup there may exist a bias toward the rearrangement of certain individual VH genes. As discussed, this may be exemplified by the G6- encoding gene 5 1 p I , a member of the relatively large VH 1 gene s ~ b g r o u p . ~ ~ , ~ ~ Deane and Norton99 noted that 3 of the 9 Ig VH1 gene rearrangements they identified in CLL were homologous to 51pl. In contrast, these investigators did not find this gene used in a set of 9 VH1 gene rearrangements detected among pre-B acute lymphocytic leukemias, sug- gesting that the biased rearrangement of 5 lp 1 is peculiar to CLL. In another survey of 44 patients with CLL, two-thirds (10 of 15) of the 15 leukemia samples found to have functionally rearranged VH 1 genes were found to use VH 1 genes homologous to 5 1 p 1 .loo Two of the 15 ( 13%) were homologous to V35. This v H 1 gene is distinctive in that it is the closest VH1 gene to the D and J H gene complexes."' As 5 lp l apparently is rearranged and expressed more often than V35, proximity to the D and J H loci evidently does not determine the hierarchy at which v H 1 genes are expressed in CLL. Fi- nally, because 5 lp l is but only 1 of approximately 20 to 60 VH 1 genes in the Ig heavy chain gene these data indicate that VH1 gene rearrangement in CLL is not random.

At least two mechanisms may contribute to the frequent rearrangement and expression of certain Ig V genes in CLL. First, certain Ig V genes may undergo Ig gene rearrangement more frequently than other Ig V genes, possibly due to intemal or neighboring recognition sequences for enzymes involved in Ig gene rearrangement."' On the other hand, certain Ig V genes may encode antibodies that have peculiar binding specificities that are selected during leukemogenesis, resulting in a biased repertoire of Ig V genes used in CLL. These mechanisms need not be mutually exclusive.

To determine whether nonrandom Ig gene rearrangement occurs in CLL independent of selection for expressed Ig pro- tein, we examined the abortive V, gene rearrangements in X

light chain expressing CLL (X-CLL).Io3 Although most of these X-CLL had rearranged both K light chain alleles, 9 had rearranged only one, providing us with a total of 57 identified K light chain gene rearrangements in a sample group of 33 A-CLL. We found the V, gene encoding the 17.109-CRI, Humkv325, to be involved in 8 of these nonproductive rearrangements. Another conserved V,3 gene, designated Vg, was rearranged nonproductively in 3 of the A-CLL. Similar to Humkv325, this conserved V,3 gene is frequently found to encode the K light chain variable region of IgM, autoanti-

In theory, any one of 70 V, genes may undergo Ig rear- r a ~ ~ g e m e n t . ~ ~ If V, gene rearrangement were random, then the chance of finding nonproductive Humkv325 or Vg gene rearrangements should be 1 in 70, or 1.4% of the number of V, gene rearrangements. Thus, the expected number of Humkv325 or Vg gene rearrangements in the sample group of 57 nonproductive V, gene rearrangements should be less than 1 (or 0.8). X2 analyses show that the observed frequency of abortive Humkv325 or Vg gene rearrangements significantly exceeds that expected if Ig V, gene rearrangements were truly random (P = .OW1 for Humkv325 and P = .0147 for Vg). Therefore, independent of Ig expression, there exists a significant bias in the rearrangement of certain Ig V genes in CLL.

bodies.78,l04,l05

lg SOMATIC MUTATION IN CLL

Studies on the Ig V genes used by leukemia B cells have shown that the process of somatic hypermutation is not active in CLL. First, the Ig V genes expressed in any given CLL B- cell clone are homogeneous, or without significant "intraclonal d i~e r s i ty . "~* ,~~ , 'O~ '~~ This contrasts with non-Hodgkin's lymphomas (NHL) of follicular center cell origin. Analyses of these lymphomas show substantial intraclonal diversity in the expressed Ig variable regions suggestive of ongoing somatic hypermutation.'" Furthermore, the Ig V genes expressed in most CLL share extensive homology (>97%) with known germline Ig V genes.89~92~93~97~99.100,106,Lo9~110 Collectively, these studies indicate that CLL may express germline Ig V genes with little or no somatic mutation.

However, there are some exceptions to this generalization. Humphries et a195 noted that the leukemia cells from related CLL patients expressed extensively mutated Ig VH genes be- longing to the relatively small VH5 subgroup. In addition, they noted that 30% of the CLL patients in their survey had leukemia cells that had rearranged one Ig vH5 gene, designated VH25 1. These investigators also examined Ig VHS gene- expressing CLL B-cell populations from 11 different patients."' Because prior studies indicated that Ig VH genes of this small subgroup are highly conserved and nonpoly- morphic,"' they compared the primary nucleic acid sequences of the expressed VH genes with that of known VH5 genes. In 9 of the 11 CLL, they found the nucleic acid sequences of the expressed Ig VH25 1 genes to differ substantially (594% homology) from that of the known germline sequence. Importantly, the nucleic acid base differences resulted in amino acid substitutions were clustered primarily within the CDRs, regions that form the pocket of the antibody's antigen- combining site. As discussed, such substitutions often are




2478 KIPPS AND CARSON

noted in the Ig selected in an antigen-driven secondary immune response. Accordingly, it appears that these CLL express antibodies that had been selected for their ability to bind some unknown antigen(s).

Expression of mutated Ig VH genes by these CLL may reflect a unique property of the Ig VH25 1 gene. This Ig vH5 gene, for example, is distinctive in that it drives relatively high rates of transcription in the germline nonrearranged configuration.' l3

To evaluate whether the Ig vH5 genes in CLL are distinctive, we recently examined for vH5 Ig gene rearrangements in leukemia cells from 68 patients that satisfied clinical and diagnostic criteria for common B-cell CLL. 'I4 Southern blot hybridization studies with probes for VH25 1 and the J H locus showed that only 7 (10%) of the 68 monoclonal CLL cell populations had undergone Ig gene rearrangement involving vH5 genes. Two (3%) were found to have functionally rearranged vH5 genes that shared 298% sequence homology with 5-2R1, a VH25 1 gene isolated from a pre-B-cell acute lymphocytic leukemia. The other 5 CLL (7%) had functionally rearranged vH5 genes that each shared 299% nucleic acid sequence homology with a germline VH32 isolated from human sperm DNA. These data indicate that VH251 or VH32 also may be expressed by common CLL B cells with little or no somatic mutation. As such, these data conflict with those discussed above. Conceivably, the leukemia cells that express highly mutated Ig V genes may represent a unique subset of leukemias that differs from common CLL in ways other than just Ig V gene expression. Further work is necessary to determine whether there exist differences in the clinical and/or phenotypic characteristics of patients with leukemia cells that express mutated versus nonmutated Ig V genes. However, in general, CLL B cells that express highly mutated Ig V genes appear to be exceptional.

NORMAL CELL COUNTERPART TO THE CLL B CELL

CD5 B cells-"BI B cells." CLL B cells coexpress B- cell-specific surface antigens and CD5 (Leu 1, OKTl).1'5-''8 As such, the normal counterpart to these leukemia cells ar- guably is the "CD5 B cell.""9 These cells constitute a small subpopulation of human B lymphocytes in the lymphoid or- gans and peripheral blood of normal adults that coexpress B-cell differentiation antigens and CD5 .120-'25 Such cells are enriched for B cells that spontaneously may produce IgM autoantibodies'26'128 and frequently may express autoantibody-associated C R I S . ' ~ ~ , ' ~ ~ , ' ~ ~

Recently, a new nomenclature has been adopted for CD5 B ~e1ls.l~' Because the CD5 surface antigen (1) may not be detected on the surface of B cells that otherwise have other developmental and/or phenotypic traits of "CD5 B cells,"'z5~'32-134 (2) may be induced on non-"CD5 B

and (3) can be reduced on "CD5 B cells" by treatment with various c y t o k i n e ~ , ' ~ ~ . ' ~ ~ it was argued that the term "CD5" (or "Ly-I") was not adequate for this type of B cell. Therefore, participants at the New York Academy of Sciences meeting on "CD5 B cells," held in West Palm Beach, F'L (June 199 I), proposed that "CD5 B cells" be designated B-1 B cells, and that "conventional" B cells be termed B-2 B

ce~ls,~~135-138

In mice, "B-1 B cells" can be distinguished readily from "B-2 B cells" by their anatomic localization, phenotype, functional characteristics, gene expression, and developmental stage of origin'24~'32*'41-'4 (Table 1). Although demon- strable as a rare subpopulation in the spleen of most normal inbred mouse strains, murine B-1 B cells normally are not found in the lymph node (LN), blood, or bone marrow (BM).'45 In contrast, B-1 B cells constitute a major lymphoid subpopulation in the murine peritoneal cavity.'46 B- 1 B cells also share several physical and developmental properties with monocytes and macrophage^.'^^*'^^*'^^ Studies indicate that mouse B-1 cells frequently may produce IgM autoantibodies and use a restricted set of Ig variable region genes (V gene^).'^^^'^^-''' In addition, some investigators have noted that B- 1 B cells may influence the repertoire of other B and T lymphocyte^,'^"'^^ indicating that these cells have potential immunoregulatory function. Finally, most B- 1 B cells originate in early ontogeny and are l~ng- l ived ."~ , '~~ . '~~ This latter point may be shown through analyses of the rearranged Ig heavy chain variable regions expressed by B- 1 B cells in adult m i ~ e . ' ~ ' . ' ~ ~ These studies show a paucity of nontemplated N- region sequence insertions in the VH-DH and &JH junctions in the rearranged Ig heavy chain variable regions of B-1 B cells. Such Ig gene rearrangements are typical of those made by early fetal B cells before the developmental expression of terminal deoxytransferase (TdT), the enzyme responsible for the addition of N-region sequences during VDJ gene rearrangement?4*157 As such, the absence of N-sequence insertions serves as a developmental clock, implying that these adult B-1 B cells may have undergone Ig gene rearrangement and differentiation during early B-cell ontogeny.

With some exceptions, human B-1 B cells share many phenotypic and developmental characteristics with murine B-1 B cells (Table 1). However, the enzyme TdT apparently is active during the early human B-cell differentiation, resulting in human fetal Ig heavy chain gene rearrangements that have junctional N-sequence insertion^?^ As such, one cannot use the relative absence of N-sequence insertions to characterize Ig V genes that have rearranged during early human B-cell development. Also, human CD5 B cells coexpress higher levels of sIgD and CD23 than do their murine counterparts.1M Operationally then, expression of CD5 re-

Table 1. Proposed B-Cell Subpopulations

B-1 B cell Enriched for 8 cells that constitutively express CD5 Originate in early ontogeny Long-lived with slow cell turnover Low rates of lg V gene somatic mutation Enriched for cells that make "natural" IgM autoantibodies

CD5- unless induced Adult bone marrow derived Short-lived and cycling Capable of lg V gene somatic hypermutation Recruited in secondary humoral immune responses

8-2 B cell





mains one of the best markers for distinguishing human B- 1 B cells.

In fact, CD5 still may serve as a useful marker for iden- tifying human B-1 B-cell malignancies. More than 90% of patients with B-cell CLL have leukemia cells that express CD5. Patients with leukemia B cells that do not express this surface differentiation antigen may have a distinct disease that differs from common CLL in its etiopathogenesis and/ or its expression of Ig genes. For example, we examined the Ig V genes expressed by the neoplastic cells of an unusual CLL patient that did not express CD5 but still secreted an IgM RF autoantibody. The nucleic acid sequence of the V, gene expressed by this CLL differed substantially (<96% homology) from that of any known germline V, gene.'58s159 Fur- thermore, nucleic acid sequence analyses of independent VH gene clones showed substantial intraclonal variation in the VH genes expressed by this leukemia cell population." Sim- ilar intraclonal diversity recently was observed in a case of small lymphocytic lymphoma (SLL) that also failed to express CD5," and in another CLL cell population that also was CD5-.'Io These findings contrast with those made in analyzing conventional CD5 B-cell CLL,89,92,93,106,'07.110 as discussed above. Collectively, these studies suggest that the expression of CD5 may help distinguish conventional "B- I B-cell" CLL from other types of B-cell leukemias that differ physiologically in the ways they process their expressed Ig V genes.

To evaluate for normal B cells that also may express Ig V genes similar to those often observed in B-cell CLL, investigators have examined lymphocytes in nonmalignant human lymphoid tissue for reactivity with MoAbs specific for autoantibody-associated Flow cytometric analyses of tonsillar lymphocytes from 19 subjects, for example, showed that the lymphocytes binding 17.109 are a subpopulation of Ig K-bearing B cells comprising 1.3% to 6.3% (mean = 3.9% f 1.2%) of the total lymphocytes, 3% to 16% (mean = 10% f 3%) ofthe K light chain-bearing cells, and from 46% to 97% (mean = 77% rf: 16%) of the cells expressing K light chains of the VJIb subgroup. G6-positive cells expressed either K or X light chains and comprised SO. 1% to 4.5% (mean = 2.2% f 1.2) ofthe total lymphocytes. Cells expressing both the 17.109 and G6 CRI comprised a very small subpopulation constituting less than 0.1% to I % (mean = 0.4% f 0.3%) of the total lymphocyte population.

Molecular studies indicate that such CRI-expressing tonsillar lymphocytes may express Ig V genes similar to that used by CRI-expressing CLL B cells. G6-reactive tonsillar lymphocytes, for example, are found to express 5 1 p 1 with little or no somatic m~tation."~ In fact, in more than 5,600 nucleotide bases of the Ig VH genes from 13 independent G6- reactive tonsillar B-cell clones, there were only three base differences detected. The absence of even nonconservative base changes indicates that the conserved Ig VH gene sequences are not the mere consequence of selection for cells having reactivity for the G6 MoAb. Rather, these data indicate that G6-reactive tonsillar B cells generally express homologous VHl genes that have not diversified through somatic mutation.

B cells that express such autoantibody-qswciated CRIs are confined to the mantle zones surrounding the germinal cen-

Mantel zone B cells.

ters of secondary B-cell follicle^.^^^^^^ Tonsillar B cells can be delineated into subpopulations based on the differential expression of SI@, CD10, and receptors for peanut agglutinin (PNA). Mantle zone B cells bear SI@, but fail to bind PNA or MoAbs specific for CDlO (CALLA), whereas B cells within the germinal center generally lack sIgD, express low levels of CD10, and avidly bind PNA.'6L-163 Consistent with their mantle zone distribution, greater than 95% of CRI-positive B cells express sIgD but fail to bind PNA or MoAbs specific for CD 10.Lz5 Furthermore, multiparameter analyses indicate that human tonsillar B cells that express CD5 also have this phenotype, indicating that B-1 B cells may constitute a subpopulation of mantle zone 1ympho~ytes.l~~

It should be noted that sIgD-bearing cells constitute only 37% to 87% of the tonsillar B cells.L25 As such, the 17.109- reactive or G6-reactive cells, respectively, may constitute up to 17% or 12% of the mantle zone lymphocytes. These fre- quencies approach those noted for expression of these CRIs in B-cell CLL and argue that the Ig V gene repertoire of mantle zone B cells may be comparably restricted. Recent studies suggest that the expressed Ig VH gene repertoire of circulating B cells in adult peripheral blood also may be highly restricted. 164 It is conceivable then that transformation of lymphocytes such as those found in the mantle zone may account for the high frequency expression of autoantibody- associated CRIs in CLL.

However, it seems that CLL B cells may originate from only a subset of such B cells. For one, there is an apparent bias for coexpression of both 17.109 and G6 by leukemia cell populations that is not noted in most tonsillar lymphocyte populations s t ~ d i e d . ~ ~ , ' ~ ~ Biased coexpression of such CRIs reflects a predilection for CLL B cells to express certain pairs of heavy and light Ig chains.

Furthermore, there exists an apparent bias in the structure of the CDR3 of certain Ig heavy chains expressed in CLL.loo Nucleic acid sequence analyses of 13 G6-CRI-positive CLL cells that express 5 1 p 1 shows each to have an unusually long CDR3 of 30 codons (+ 4, SD). These CDR3 are encoded by germline D segments, N-sequence insertions, and JH segments. The JH segment use of G6-reactive CLL (JH3 [31%], JH4 [ 15%], J H 5 [8%], or JH6 [46%]) apparently favors the use of JH3 compared with that of normal G6-reactive tonsillar lymphocytes or normal circulating adult B cells.'65 D segment use also is skewed compared with that of normal circulating B cells,'65 with XP4, XP'I, and XP1 accounting for approximately three-fourths of the D segments used in G6-reactive CLL. However, the heavy chain variable regions expressed by 14 independent normal G6-reactive tonsillar B-cell clones did not show such a bias.lz5 Rather, the D segment use of G6-reactive tonsillar B cells mimics that observed for normal circulating B cells.165 Also, in contrast to the length of the CDR3 of G6-reactive CLL, CDR3s of the G6-positive tonsillar B cells are significantly shorter, averaging only 24 codons in length (+ 5, SD).

Conceivably, the bias in the composition of CDR3 may reflect the time in development when the normal cell counterpart to CLL had rearranged its Ig genes. Alternatively, apparent constraints in the Ig CDR3 and the bias in Ig chain pairing may reflect restrictions on the Ig repertoire of normal





B cells that are subject to malignant transformation in CLL. For example, certain pairs of heavy and light Ig chains that have appropriate CDR3s may generate antibody molecules with specificities that increase the likelihood for B-cell malignant transformation. If so, then the finding that the Ig expressed by CLL B cells so frequently have autoantibody activity may be relevant to the etiopathogenesis of this disease.

SOMATIC SELECTION OF CLL AUTOANTIBODIES

An important feature of the autoantibodies expressed in CLL is their “polyreactivity,” or binding activity for two or more seemingly disparate self antigens. RF autoantibodies made by CLL B cells, for example, generally also are found to bind other “self’ antigens, such as ssDNA, dsDNA, histones, cardiolipin, actin, thyroglobulin, and/or cytoskeletal c ~ m p o n e n t s . ~ - ~ J ~ ~ ~ ’ ~ ~ Such polyreactivity is a feature also noted for antibodies produced during early B-cell development, even in animals raised in apparently germ-free envi- ronment~ .~’”~*-’~’ Th ese types of autoantibodies are not a priori pathogenic in that they can be detected in the sera of all individuals and appear physiologic. Because of this, several investigators have used the term “natural autoantibodies” to describe these types of antibodies.

Polyreactive IgM autoantibodies may be encoded by Ig V genes that are present in the germline DNA.’72 Accordingly, the frequent expression of polyreactive IgM autoantibodies in CLL may be a direct consequence of the frequent use of such Ig V genes, with little or no somatic mutation. As noted, a high proportion of CLL patients have leukemia cells expressing Ig V genes that encode CRIs frequently found to be associated with IgM autoantibodies, particularly RFs.84-86~90 Moreover, more than 90% of the monoclonal IgM paraproteins that bear both 17.109 and G6 CRIs are noted to have RF a~ t iv i ty . ’~~ As such, the simple pairing of a 17.109-reactive Ig light chain with a G6-reactive Ig heavy chain may produce an Ig with polyreactive anti-self activity. If this were so, then frequent autoantibody activity of the Ig expressed by CLL B cells may be an epiphenomenon.

However, this model minimizes the contribution of the third complementarity determining region (CDR3) to the autoreactive binding activity of the Ig expressed in CLL. As discussed, this region of the Ig heavy chain is encoded by the D and JH gene segments that undergo recombination and N- terminal nucleic acid base insertion immediately before VH gene r e a ~ ~ a n g e m e n t . ~ ~ , ’ ~ ~ Accordingly, the sequence of the CDR3 is generally idiosyncratic to each Ig VH gene rearrangement, as has been noted with all G6-positive heavy chains sequenced to date93r’25 (T. Johnson, S.F. Duffy, T.J. Kipps, unpublished observations). In view of the large potential for diversity in the CDR3 of G6-positive heavy chains,’25 the random pairing of a 17.109-positive K light chain with any G6-positive Ig heavy chain may not be anticipated to form an autoantibody if the CDR3 is critical to autoantibody-binding activity (Fig 2).

To examine this, we generated murine transfectomas to pair the 17.109-positive K light chains of SMI, a 17.109/G6- positive CLL B-cell population, with each of several different G6-positive heavy chains expressed by normal or leukemic B cells.167 For this, we generated pRTM 1, a human p-chain

+ + Fig 2. Polyspecific autoantibody activity. Antibody A and B with

lg light and heavy chains AL and AH or B,and BH, respectively, have polyspecific binding activity. AL or AH may differ from BL or B,, respectively, only in the CDR3 (shaded area), the region ofthe variable region exon that is generated somatically during lg gene rearrangement. However, pairing of AL with B, fails to generate an antibody with polyspecific autoantibody activity. This indicates that such reactivity is dependent on the sequences encoded within the CDR3.

expression vector that contains the functionally rearranged SMI VH1 gene flanked by unique restriction enzyme sites. This enabled us to exchange several different functional VHDJH exons of other G6-positive leukemia cells or tonsillar lymphocytes. Each of the rearranged VH genes within these genes shared more than 99% nucleic acid sequence homology to 5 1 ~ 1 , a VH1 gene expressed during early fetal development.93.94x’25 In contrast, these exons differed markedly in the CDR3. We found that the myeloma cells cotransfected with the original pair of Ig heavy and light chain genes of SMI secrete polyreactive IgM, RF a~toantibodies.’~~ However, myeloma cells cotransfected with the SMI K light chain gene and any I of 10 different G6-encoding VHDJH exons produced 17.109/G6-reactive IgM, that failed to have such autoantibody activity (Fig 2 ) . This indicates that such polyreactive binding activity of these natural autoantibodies is dependent on the somatically generated CDR3.

These results imply that the polyreactive binding activity of natural autoantibodies is a selected specificity. In human fetal spleen, B cells frequently express autoantibody-associated CRIs, such as G6 and 17. 109.174 Genetic mechanisms may enhance rearrangement of Ig V genes that encode CRI- bearing IgM autoantibodies. IO3 However, random pairing of such Ig light and heavy chains only infrequently may give rise to polyreactive autoantibodies. Because more than 90% of Ig paraproteins that bear both CRIs have autoantibody activity, it appears that autoreactivity is a selected specificity.

In addition, several studies indicate that more than half of all CLL patients have leukemia cells that can be stimulated to secrete polyreactive aut~antibodies.~.~ This proportion greatly exceeds that noted for polyreactive B cells in normal embryonic tissues, cord blood, or adult peripheral blood. Even when analyzing B cells greatly enriched for B-1 type B cells (eg, neonatal or fetal B cells), Guigou et all7’ noted that only 1 1 % to 16% of all Epstein-Barr Virus (EBV)-transformed Ig- secreting B-cell clones expressed polyreactive antibodies. The high prevalence of CLL patients with leukemia cells that make polyreactive Ig thus may not be reflective of the repertoire expressed by the normal B- I B-cell population. Rather, this may reflect the possibility that expression of polyreactive Ig




AUTOANTIBODIES IN CLL 248 1

may enhance a B cell's risk for malignant transformation into CLL.

SYSTEMIC AUTOIMMUNE DISEASES ASSOCIATED WITH CLL

Patients with CLL frequently may develop intermittent autoimmune Soon after CLL popularly be- came recognized as a distinct clinical entity, investigators noted an association between CLL and various autoimmune diseases, such as AIHA'75,'78 and ITP.176 Less frequently, pa- tientswith CLL are noted to have pure red blood cell aplasia18' or neutropenia7 secondary to a presumed autoimmunity against precursor cells in the bone marrow.

Larger retrospective surveys confirmed the association between certain types of autoimmune disorders and CLL. Duhrsen et alls3 studied the records of nearly 1,000 patients with either myeloproliferative or lymphoproliferative diseases for clinical manifestations of autoimmune pathology. In- cluded in this survey were 104 patients with CLL. As a group, patients with CLL had the highest incidence of associated autoallergic hematologic diseases, accounting for more than 60% of all cases observed to have hemolytic anemia (9 of 14) or autoimmune thrombocytopenia (3 of 5). Similarly, Ham- blin et a17 studied the prevalence of autoantibodies in 195 patients with B-cell CLL. Fifteen patients (7.7%) tested positive in the direct antiglobulin test for antierythrocyte autoantibodies. One of these 15 had high titer anti-I IgM, MoAbs produced by the leukemia cell clone. The other 14 had polyclonal IgG anti-red blood cell autoantibodies, 10 of which produced clinically detectable hemolytic anemia. Four patients (2.1%) had ITP, l patient (0.5%) had pure red blood cell aplasia, and 1 patient (0.5%) had isolated neutropenia thought to be secondary to antineutrophil autoantibodies. The incidence of autoantibodies increased later in more advanced stages of CLL. The incidences of such autoantibodies were significantly higher than in a similarly sized group of age- and sex-matched control subjects.

ORIGIN OF PATHOGENIC AUTOANTIBODIES IN CLL

Although pathogenic autoantibodies occasionally may be made by the clone of malignant B cells,'04,i60,188 generally such autoantibodies appear to be made by bystander B cells. Similar to studies on autoantibodies that arise in systemic autoimmune disease, one can discern autoantibodies with Ig K light chains or Ig X light chains in any one patient. Fur- thermore, Ig isotypes of such autoantibodies usually differ from that of the Ig expressed by the malignant B-cell clone. In addition, the circulating levels and the clinical significance of such autoantibodies do not relate directly to the duration or severity of the underlying lymphoproliferative disease. Moreover, immunosuppressive therapy may result in clinical remission of pathologic autoimmunity without affecting the overall leukemia cell burden. Collectively, these observations suggest that the cells producing pathogenic autoantibodies may originate from bystander B cells that are not related to the leukemic cell clone.

Although difficult to detect, especially in patients with high leukemia cell counts, such bystander B cells may be present in the peripheral blood of patients with CLL. We noted hu-

man antierythrocyte autoantibodies in mice with severe combined immunodeficiency (SCID) that were engrafted with peripheral blood leukocytes (PBL) from patients with CLL.'" SCID mice fail to develop functional T or B lymphocytes due to an autosomal, recessive Leukemia cells injected into the peritoneal cavity of these animals may sur- vive for several weeks in vivo. Eight to 16 weeks after receiving PBL from patients with CLL, SCID mice develop human IgG autoantibodies to human red blood cells and/or high serum levels of human Ig. Soon thereafter, these animals develop lethal human B-cell tumors. In contrast to the original CLL cells, these human B-cell tumors are CDY, have genomic DNA of EBV, express antigens associated with latent EBV infection, and have distinctive Ig gene rearrangements by Southern. We conclude that bystander B cells, onginally present at 10.1% of the injected PBL, may generate tumors in CLL-reconstituted SCID mice that emulate the EBV-associated lymphoproliferations noted in SCID mice reconstituted with normal human PBL.'95-'97

Similar to the pathogenic autoantibodies that develop in patients with CLL, the human IgG antierythrocyte autoantibodies of such reconstituted mice apparently are produced by circulating bystander B cells that are not clonally related to the transferred CLL B-cell population. First, in addition to having different Ig isotypes, the antierythrocyte Ig may have Ig light chains that differ from that of the Ig expressed by the population of injected leukemia B cells. Second, although all mice received comparable numbers of PBL from patients with CLL, only a fraction of the mice developed antierythrocyte autoantibodies. Finally, the emergence of antierythrocyte autoantibodies in such SCID mice coincided with the development of high serum levels of human Ig. This invariably preceded the development of B-cell lymphomas that have clonal origins distinct from that of the original leukemia B-cell population. The data suggest that polyclonal activation and/or lymphoproliferation of bystander B cells may account for the antierythrocyte antibodies detected in these animals.

Conceivably, the occurrence of antierythrocyte or anti- platelet autoantibodies in CLL patients also may be associated with dysregulation of the cellular immune mechanisms that control latent EBV infections. Alternatively, the development of such autoantibodies may reflect a more generalized defect in the homeostatic mechanisms that prevent generation of antiself humoral immunity. According to this hypothesis, autoantibodies arise not from polyclonal B-cell activation, but rather from a pathologic immune response to self antigen.

Whatever hypothesis is advanced must take into consid- eration the peculiar predilection of CLL patients to develop autoantibodies primarily against hematologic cells (eg, erythrocytes, platelets, et^).^ This may reflect a requirement for the self antigen@) to be physically associated with or processed by either normal or leukemic B- 1 B cells. Clearly, more studies are required to define the mechanism(s) that accounts for the oftentimes pathogenic autoantibodies that may arise in CLL. Such studies may contribute to our understanding of normal B-1 B cells and how they potentially may contribute to the maintenance of immunologic tolerance and/or to the development of systemic autoimmunity.





CONCLUSIONS

Common CLL is a B- I B-cell malignancy. The leukemia cells often make polyreactive autoantibodies and use a restricted repertoire of nonmutated Ig V genes. Recent studies indicate that somatic selection of B-1 B cells for polyspecific autoreactivity may play an important role in shaping the repertoire of Ig expressed by these cells. However, chronic stimulation of such long-lived B-l B cells by ubiquitous self antigens also may increase their risk for neoplastic transformation into CLL.

Even though CLL B cells often make autoantibodies, the Ig expressed by leukemia B cells generally do not contribute directly to the types of autoimmune diseases that frequently are observed in patients with this disease. CLL-associated autoimmune diseases generally result from the intermittent production of pathogenic autoantibodies that most com- monly react with antigens present on human hematopoietic cells (eg, red blood cells or platelets). These autoantibodies are apparently produced by polyclonal bystander B cells as a consequence of an immune dysregulation associated with B-cell CLL.

The fact that CLL cells express a limited repertoire of Ig V genes that have not diversified substantially from the germline DNA implies that the surface Ig may be a convenient target antigen for passive or active immunotherapy. Current efforts to prepare additional anti-CRI MoAbs, to induce cy- totoxic T lymphocytes reactive with idiotypic antigens on CLL cells, and to develop improved delivery systems for Ig- directed immunotherapy may yield important advances in the treatment of this disease.

REFERENCES 1. Preud’homme JL, Seligmann M Anti-human immunoglobulin

G activity of membrane-bound immunoglobulin M in lymphoproliferative disorders. Roc Natl Acad Sci USA 69:2132, 1972

2. Kipps TJ, Fong S, Tomhave E, Chen PP, Goldfien RD, Carson DA: Immunoglobulin V gene utilization in CLL, in Gale RP, Rai KR: (eds): Chronic Lymphocytic Leukemia: Recent Progress and Future Direction. New York, NY, Liss, 1987, p I15

3. Broker BM, Klajman A, Youinou P, Jouquan J, Worman CP, Murphy J, Mackenzie L, Quartey-Papafio R, Blaschek M, Collins P, Lal S, Lydyard P M Chronic lymphocytic leukemic (CLL) cells secrete multispecific autoantibodies. J Autoimmun 1:469, 1988

4. Sthoeger ZM, Wakai M, Tse DB, Vinciguerra VP, Allen SL, Budman DR, Lichtman SM, Schulman P, Weiselberg LR, Chiorazzi N: Production of autoantibodies by CD5-expressing B lymphocytes from patients with chronic lymphocytic leukemia. J Exp Med 169: 255, 1989

5 . Borche L, Lim A, Binet JL, Dighiero G: Evidence that chronic lymphocytic leukemia B lymphocytes are frequently committed to production of natural autoantibodies. Blood 76562, 1990

6. Kaden BR, Rose WF, Hauch T W Immune thrombocytopenia in lymphoproliferative diseases. Blood 53545, 1979

7. Hamblin TJ, Oscier DG, Young BJ: Autoimmunity in chronic lymphocytic leukaemia. J Clin Pathol 39:713, 1986

8. De Rossi G, Granati L, Girelli G, Gandolfo G, Arista MC, Martelli M, Conti L, Marini R, La Tagliata R, Leone R, Mandelli F Incidence and prognostic significance of autoantibodies against erythrocytes and platelets in chronic lymphocytic leukemia (CLL). Nouv Rev Fr Hematol30403, 1988

9. Malcolm S, Barton P, Murphy C, Ferguson-Smith MA, Bentley DL, Rabbitts TH: Localization of human immunoglobulin kappa light chain variable region genes to the short arm of chromosome 2 by in situ hybridization. Proc Natl Acad Sci USA 79:4957, 1982

IO. McBride OW, Hieter PA, Hollis GF, Swan D, Otey MC, Leder P: Chromosomal location of human kappa and lambda immunoglobulin light chain constant region genes. J Exp Med 155:1480, 1982

I I. Kirsch IR, Morton CC, Nakahara K, Leder P Human immunoglobulin heavy chain genes map to a region of translocations in malignant B lymphocytes. Science 2 16:30 I , 1982

12. Tonegawa S: Somatic generation of antibody diversity. Nature 302:575, 1983

13. Honjo T, Habu S : Origin of immune diversity: genetic variation and selection. Annu Rev Biochem 54:803, 1985

14. Seidman JG, Leder P: The arrangement and rearrangement of antibody genes. Nature 276:790, 1978

15. Seidman JG, Leder A, Nau M, Norman B, Leder P Antibody diversity. Science 202: 1 I , I978

16. Sakano H, Kurosawa Y, Weigert M, Tonegawa S: Identification and nucleotide sequence of a diversity DNA segment (D) of immunoglobulin heavy-chain genes. Nature 290562, 198 1

17. Siebenlist U, Ravetch JV, Korsmeyer S, Waldmann T, Mer P Human immunoglobulin D segments encoded in tandem multi- genic families. Nature 294631, 1981

18. Ichihara Y, Matsuoka H, Kurosawa Y: Organization of human immunoglobulin heavy chain diversity gene loci. EMBO J 7:4141, 1988

19. Buluwela L, Albertson DG, Shemngton P, Rabbitts PH, Spun N, Rabbitts TH: The use of chromosomal translocations to study human immunoglobulin gene organization: Mapping DH segments within 35 kb of the C mu gene and identification of a new DH locus. EMBO J 7:2003, 1988

20. Ravetch JV, Siebenlist U, Korsmeyer S, Waldmann T, Leder P: Structure of the human immunoglobulin mu locus: characterization of embryonic and rearranged J and D genes. Cell 27:583, 1981

21. Hieter PA, Max EE, Seidman JG, Maize1 JV Jr, Leder P: Cloned human and mouse kappa immunoglobulin constant and J region genes conserve homology in functional segments. Cell 22: 197, 1980

22. Berman JE, Mellis SJ, Pollock R, Smith CL, Suh H, Heinke B, Kowal C, Surti U, Chess L, Cantor CR, Alt W Content and organization of the human Ig VH locus: Definition of three new VH families and linkage to the Ig CH locus. EMBO J 7:727, 1988

23. Kabat E, Wu TT, Perry HM, Gottesman KS, Foeller C Se- quences of Proteins of Immunological Interest (ed 5). Bethesda, MD, US Department of Health and Human Services, 199 I

24. Yancopoulos GD, Alt FW: Regulation of the assembly and expression of variable-region genes. Annu Rev Immunol4:339, 1986

25. Reth MG, Jackson S, Alt FW: VHDJH formation and DJH replacement during pre-B non-random usage of gene segments. EMBO J 5:2131, 1986

26. Kodaira M, Kinashi T, Umemura I, Matsuda F, Noma T, Ono Y, Honjo T: Organization and evolution of variable region genes of the human immunoglobulin heavy chain. J Mol Biol 190529, 1986

27. Lee KH, Matsuda F, Kinashi T, Kodaira M, Honjo T: A novel family of variable region genes ofthe human immunoglobulin heavy chain. J Mol Biol 195:761, 1987

28. Bentley DL, Rabbitts TH: Evolution of immunoglobulin V genes: Evidence indicating that recently duplicated human V kappa sequences have diverged by gene conversion. Cell 32: 18 1, 1983

29. Pech M, Smola H, Pohlenz HD, Straubinger B, Gerl R, Zachau HG: A large section of the gene locus encoding human immuno-





globulin variable regions of the kappa type is duplicated. J Mol Biol 183:291, 1985

30. Lorenz W, Straubinger B, Zachau H G Physical map of the human immunoglobulin K locus and its implications for the mechanisms of VK-JK rearrangement. Nucleic Acids Res 15:9667, 1987

3 1. Pohlenz HD, Straubinger B, Thiebe R, Pech M, Zimmer FJ, Zachau H G The human V kappa locus. Characterization of extended immunoglobulin gene regions by cosmid cloning. J Mol Biol 193: 241, 1987

32. Straubinger B, Huber E, Lorenz W, Osterholzer E, Pargent W, Pech M, Pohlenz HD, Zimmer FJ, Zachau H G The human VK locus. Characterization of a duplicated region encoding 28 different immunoglobulin genes. J Mol Biol 199:23, 1988

33. Meindl A, Klobeck HG, Ohnheiser R, Zachau HG: The V kappa gene repertoire in the human germ line. Eur J Immunol 2 0 1855, 1990

34. Zachau H G The human immunoglobulin kappa locus and some of its acrobatics. Biol Chem Hoppe Seyler 371:1, 1990

35. Brack C, Hirama M, Lenhard-Schuller R, Tonegawa S: A complete immunoglobulin gene is created by somatic recombination. Cell l5:l, 1978

36. Seidman JG, Nau MM, Norman B, Kwan SP, Scharf M, Leder P Immunoglobulin V/J recombination is accompanied by deletion of joining site and variable region segments. Proc Natl Acad Sci USA 77:6022, 1980

37. Anderson ML, Szajnert MF, Kaplan JC, McColl L, Young BD: The isolation of a human Ig V lambda gene from a recombinant library of chromosome 22 and estimation of its copy number. Nucleic Acids Res 12:6647, 1984

38. Yamasaki N, Komori S, Watanabe T: Complementary DNA for a human subgroup IV immunoglobulin lambda-chain. Mol Im- munol 24:98 1, 1987

39. Dariavach P, Lefranc G, Lefranc M P Human immunoglobulin C lambda 6 gene encodes the Kem+Oz-lambda chain and C lambda 4 and C lambda 5 are pseudogenes. Proc Natl Acad Sci USA 84:9074, 1987

40. Udey JA, Blomberg BB: Intergenic exchange maintains identity between two human lambda light chain immunoglobulin gene intron sequence. Nucleic Acids Res 16:2959, 1988

41. Udey JA, Blomberg B: Human lambda light chain locus: Or- ganization and DNA sequences of three genomic J regions. Immu- nogenetics 25:63, 1987

42. Hieter PA, Hollis GF, Korsmeyer SJ, Waldmann TA, Leder P Clustered arrangement of immunoglobulin lambda constant region genes in man. Nature 294:536, 1981

43. Vasicek TJ, Leder P: Structure and expression of the human immunoglobulin lambda genes. J Exp Med 172:609, 1990

44. Bauer TR Jr, Blomberg B: The human lambda L chain Ig locus. Recharacterization of JC lambda 6 and identification of a functional JC lambda 7. J Immunol 146:2813, 1991

45. McKean D, Huppi K, Bell M, Staudt L, Gerhard W, Weigert M: Generation of antibody diversity in the immune response of BALB/c mice to influenza virus hemagglutinin. Roc Natl Acad Sci USA 81:3180, 1984

46. Clarke SH, Huppi K, Ruezinsky D, Staudt L, Gerhard W, Weigert M: Inter- and intraclonal diversity in the antibody response to influenza hemagglutinin. J Exp Med 161:687, 1985

47. Griffiths GM, Berek C, Kaartinen M, Milstein C Somatic mutation and the maturation of immune response to 2-phenyl ox- azolone. Nature 312:271, 1984

48. Rajewsky K, Forster I, Cumano A: Evolutionary and somatic selection of the antibody repertoire in the mouse. Science 238:1088, 1987

49. Manser T Mitogen-driven B cell proliferation and differentiation are not accompanied by hypermutation of immunoglobulin variable region genes. J Immunol 139:234, 1987

50. Wysocki W, Creadon G, Lehmann KR, Cambier J C B-cell proliferation initiated by Ia cross-linking and sustained by interleukins leads to class switching but not somatic mutation in vitro. Immu- nology 75:116, 1992

5 1. Berek C, Milstein C Mutation drift and repertoire shift in the maturation of the immune response. Immunol Rev 96:23, 1987

52. Wysocki L, Manser T, Gefter ML: Somatic evolution of variable region structures during an immune response. Proc Natl Acad Sci USA 83:1847, 1986

53. Levy NS, Malipiero UV, Levecque SG, Gearhart PJ: Early onset of somatic mutation in immunoglobulin VH genes during primary immune response. J Exp Med 169:2007, 1989

54. Siekevitz M, Kocks C, Rajewsky K, Dildrop R Analysis of somatic mutation and class switching in naive and memory B cells generating adoptive primary and secondary responses. Cell 48:757, 1987

55. Rada C, Gupta SK, Gherardi E, Milstein C Mutation and selection during the secondary response to 2-phenyloxazolone. h o c Natl Acad Sci USA 88:5508, 1991

56. Weiss U, Zoebelein R, Rajewsky K Accumulation of somatic mutants in the B cell compartment after primary immunization with a T cell-dependent antigen. Eur J Immunol22:5 1 I , 1992

57. Manser T, Wysocki LJ, Margolies MN, Gefter M L Evolution of antibody variable region structure during the immune response. Immunol Rev 96:141, 1987

58. Kocks C, Rajewsky K: Stable expression and somatic hypermutation of antibody V regions in B-cell developmental pathways. Annu Rev Immunol7537, 1989

59. Weiss U, Rajewsky K: The repertoire of somatic antibody mutants accumulating in the memory compartment after primary immunization is restricted through affinity maturation and mirrors that expressed in the secondary response. J Exp Med 172: 168 1, 1990

60. Manser T, Huang SY, Gefter ML: Influence of clonal selection on the expression of variable region genes. Science 226:1283, 1984

6 1. Claflin JL, Beny J, Flaherty D, Dunnick W Somatic evolution of diversity among anti-phosphocholine antibodies induced with Proteus morganii. J Immunol 138:3060, 1987

62. Kocks C, Rajewsky K: Stepwise intraclonal maturation of antibody affinity through somatic hypermutation. Proc Natl Acad Sci USA 853206, 1988

63. Shlomchik MJ, Marshak-Rothstein A, Wolfowicz CB, Roth- stein TL, Weigert M G The role of clonal selection and somatic mutation in autoimmunity. Nature 328:805, 1987

64. Sharon J, Gefter ML, Wysocki LJ, Margolies M N Recurrent somatic mutations in mouse antibodies to pazophenylarsonate increase affinity for hapten. J Immunol 142:596, 1989

65. Diamond B, Katz JB, Paul E, Aranow C, Lustgarten D, ScharE M D The role of somatic mutation in the pathogenic anti-DNA response. Annu Rev Immunol 10:731, 1992

66. Padlin EA, Davies DR: Variability of three-dimensional stuc- ture in immunoglobulins. Proc Natl Acad Sci USA 722319, 1975

67. Segal DM, Padlan EA, Cohen GH, Rudikoff S, Potter M, Davies D R The three-dimensional structure of a phosphorylcholine- binding mouse immunoglobulin Fab and the nature of the antigen binding site. Proc Natl Acad Sci USA 7 1:4298, 1974

68. Poljak RJ: X-ray diffraction studies of immunoglobulins. Adv Immunol21:1, 1975

69. Poljak RJ: Three-dimensional structure, function and genetic control of immunoglobulins. Nature 256:373, I975

70. Alzari PM, Lascombe MB, Poljak RJ: Three-dimensional structure of antibodies. Annu Rev Immunol6:555, 1988





7 I . Early P, Huang H, Davis M, Calame K, Hood L: An immunoglobulin heavy chain variable region gene is generated from three segments of DNA: VH, D and JH. Cell 19:98 I , 1980

72. Rechavi G, Ram D, Glazer L, Zakut R, Givol D Evolutionary aspects of immunoglobulin heavy chain variable region (VH) gene subgroups. Proc Natl Acad Sci USA 80:855, 1983

73. Takahashi N, Noma T, Honjo T Rearranged immunoglobulin heavy chain variable region (VH) pseudogene that deletes the second complementary-determining region. Proc Natl Acad Sci USA 8 1 : 5194, 1984

74. Matthyssens G, Rabbitts TH: Structure and multiplicity of genes for the human immunoglobulin heavy chain variable region. Proc Natl Acad Sci USA 77:6561, 1980

75. Bentley DL: Most kappa immunoglobulin mRNA in human lymphocytes is homologous to a small family of germ-line V genes. Nature 307:77, 1984

76. Klobeck HG, Solomon A, Zachau H G Contribution of human V kappa I1 germ-line genes to light-chain diversity. Nature 309:73, 1984

77. Jaenichen HR, Pech M, Lindenmaier W, Wildgruber N, Za- chau H G Composite human VK genes and a model of their evolution. Nucleic Acids Res 12:5249, 1984

78. Pech M, Zachau H G Immunoglobulin genes of different subgroups are interdigitated within the VK locus. Nucleic Acids Res 12:9229, 1984

79. Kunkel HG, Agnello V, Winchester RJ, Capra JD, Kehoe JM Cross-idiotypic specificity among monoclonal IgM proteins with anti-globulin activity. Proc Natl Acad Sci USA 71:4032, 1974

80. Carson DA, Fong S A common idiotype on human rheumatoid factors identified by a hybridoma antibody. Mol Immunol 20:1081, 1983

81. Mageed RA, Dearlove M, Goodall DM, Jefferis R: Immu- nogenic and antigenic epitopes of immunoglobulins XVII-Mono- clonal antibodies reactive with common and restricted idiotopes to the heavy chain of human rheumatoid factors. Rheumatol Int 6: 179, 1986

82. Posnett DN, Wisniewolski R, Pemis B, Kunkel H G Dissection of the human antigammaglobulin idiotype system with monoclonal antibodies. Scand J Immunol 23:169, 1986

83. Pasquali JL, Knapp AM, Farradji A, Weryha A: Mapping of four light chain-associated idiotopes of a human monoclonal rheumatoid factor. J Immunol 139:818, 1987

84. Kipps TJ, Fong S, Tomhave E, Chen PP, Goldfien RD, Carson D A High-frequency expression of a conserved kappa light-chain variable-region gene in chronic lymphocytic leukemia. Roc Natl Acad Sci USA 84:2916, 1987

85. Kipps TJ, Robbins BA, Kuster P, Carson DA: Autoantibody- associated cross-reactive idiotypes expressed at high frequency in chronic lymphocytic leukemia relative to B-cell lymphomas of follicular center origin. Blood 72:422, 1988

86. Kipps TJ, Robbins BA, Tefferi A, Meisenholder G, Banks PM, Carson DA: CD5-positive &cell malignancies frequently express cross-reactive idiotypes associated with IgM autoantibodies. Am J Pathol 136:809, 1990

87. Ono M, Winearls CG, Amos N, Grennan D, Gharavi A, Peters DK, Sissons JGP: Monoclonal antibodies to restricted and cross- reactive idiotopes on monoclonal rheumatoid factors and their recognition of idiotope-positive cells. Eur J Immunol 17:343, 1987

88. Winearls CG, Sissons JGP Use of idiotype markers for cellular detection of monoclonal rheumatoid factor. Springer Semin Im- munopathol 10:67, 1988

89. Pratt LF, Szubin R, Carson DA, Kipps TJ: Molecular characterization of a supratypic cross reactive idiotype associated with IgM autoantibodies. J Immunol 147:204 1, 199 1

90. Swisher EM, Shawler DL, Collins HA, Bustria A, Hart S, Bloomfield C, Miller RA, Royston I: The expression of shared idiotypes in chronic lymphocytic leukemia and small lymphocytic lymphoma. Blood 77:1977, 1991

9 1. Lydyard PM, Quartey-Papafio R, Broker B, Mackenzie L, Jouquan J, Blaschek MA, Steele J, Petrou M, Collins P, Isenberg D, et al: The antibody repertoire of early human B cells. I. High frequency of autoreactivity and polyreactivity. Scand J Immunol 31:33, 1990

92. Kipps TJ, Tomhave E, Chen PP, Carson DA: Autoantibody- associated kappa light chain variable region gene expressed in chronic lymphocytic leukemia With little or no somatic mutation. Implications for etiology and immunotherapy. J Exp Med 167:840, 1988

93. Kipps TJ, Tomhave E, Pratt LF, DuRy S, Chen PP, Carson DA: Developmentally restricted VH gene expressed at high frequency in chronic lymphocytic leukemia. Proc Natl Acad Sci USA 86:5913, 1989

94. Schmeder HW Jr, Hillson JL, Perlmutter RM: Early restriction of the human antibody repertoire. Science 238:791, 1987

95. Humphries CG, Shen A, Kuziel WA, Capra JD, Blattner FR, Tucker PW: A new human immunoglobulin VH family preferentially rearranged in immature B-cell tumours. Nature 33 1:446, 1988

96. Mayer R, Logtenberg T, Strauchen J, Dimitriu-Bona A, Maya L, Mechanic S, Chiorazzi N, Borche L, Dighiero G, Mannheimer- Lory A, Diamond B, Alt F, Bona C CD5 and immunoglobulin V gene expression in B-cell lymphomas and chronic lymphocytic leukemia. Blood 75: 15 18, 1990

97. Deane M, Norton J D Immunoglobulin heavy chain variable region family usage is independent of tumor cell phenotype in human B lineage leukemias. Eur J Immunol 20:2209, 1990

98. Axelrod 0, Silverman GJ, Dev V, Kyle R, Carson DA, Kipps TJ: Idiotypic cross reactivity of immunoglobulins expressed in Wal- denstroms' macroglobulinemia, chronic lymphocytic leukemia, and mantle zone lymphocytes of secondary B-cell follicles. Blood 77:1484, 199 1

99. Deane M, Norton JD: Preferential rearrangement of developmentally regulated immunoglobulin VH 1 genes in human B-lineage leukaemias. Leukemia 5:646, 1991

100. Kipps TJ, Rassenti LZ, Duffy S, Johnson T, Kobayashi R, Carson DA: Immunoglobulin V gene expression in CD5 B-cell malignancies. Ann NY Acad Sci 651:373, 1992

101. Matsuda F, Lee KH, Nakai S, Sat0 T, Kodaira M, Zong SQ, Ohno H, Fukuhara S, Honjo T Dispersed localization ofD segments in the human immunoglobulin heavy-chain locus. EMBO J 7: 1047, 1988

102. Chen PP: Structural analyses of human developmentally regulated Vh3 genes. Scand J Immunol 31:257, 1990

103. Rassenti LZ, Pratt LF, Chen PP, Carson DA, Kipps TJ: Au- toantibody-encoding kappa light chain genes frequently rearranged in lambda light chain expressing chronic lymphocytic leukemia. J Immunol 147:1060. 199 I

104. Silberstein LE, Litwin S, Carmack CE: Relationship of variable region genes expressed by a human B cell lymphoma secreting pathologic anti-Pr2 erythrocyte autoantibodies. J Exp Med 169:1631, 1989

105. Newkirk MM, Capra JD: Restricted usage of immunoglobulin variable-region genes in human autoantibodies, in Honjo T, Alt FW, Rabbitts T (4s ) : Immunoglobulin Genes. New York, NY, Academic, 1989, p 215

106. Ratt LF, Rassenti L, Lamck J, Robbins B, Banks P, Kipps TJ: Immunoglobulin gene expression in small lymphocytic lymphoma with little or no somatic hypermutation. J lmmunol 143:699, 1989

107. Meeker TC, Grimaldi JC, O'Rourke R, Loeb J, Juliusson G, Einhorn S: Lack of detectable somatic hypermutation in the V





region of the Ig H chain gene of a human chronic B lymphocytic leukemia. J Immunol 141:3994, 1988

108. Levy R, Levy S, Cleary ML, Carroll W, Kon S, Bird J, Sklar J: Somatic mutation in human B-cell tumors. Immunol Rev 96:43, 1987

109. Kuppers R, Gause A, Rajewsky K: B cells of chronic lymphatic leukemia express V genes in unmutated form. Leuk Res 15: 487, 1991

110. Ebeling SB, Schutte ME, Akkermans-Koolhaas KE, Bloem AC, Gmelig-Meyling FH, Logtenberg T: Expression of members of the immunoglobulin VH3 gene families is not restricted at the level of individual genes in human chronic lymphocytic leukemia. Int Immunol4:3 13, 1992

1 1 1. Cai J, Humphries C, Lutz C, Tucker PW: Analysis of VH25 1 gene mutation in chronic lymphocytic leukemia (CLL) and normal B e l l subsets. Ann NY Acad Sci 651:384, 1992

112. Sanz I, Kelly P, Williams C, Scholl S, Tucker P, Capra JD: The smaller human VH gene families display remarkably little poly- morphism. EMBO J 8:3741, 1989

11 3. Berman JE, Humphries CG, Barth J, Alt FW, Tucker PW. Structure and expression of human germline VH transcripts. J Exp Med 173:1529, 1991

1 14. Rassenti LZ, Kipps TJ: Lack of extensive somatic mutations in the VH5 genes used in common B cell chronic lymphocytic leukemia. J Exp Med (in press)

11 5. Boumsell L, Coppin H, Pham D, Raynal B, Lemerle J, Dausset J, Bernard A: An antigen shared by a human T cell subset and B cell chronic lymphocytic leukemic cells. Distribution on normal and malignant lymphoid cells. J Exp Med 152:229, 1980

116. Wang CY, Good RA, Ammirati P, Dymbort G, Evans RL: Identification of a p69,7 1 complex expressed on human T cells sharing determinants with B-type chronic lymphatic leukemic cells. J Exp Med 151:1539, 1980

117. Martin PJ, Hansen JA, Nowinski RC, Brown MA: A new human T-cell differentiation antigen: Unexpected expression on chronic lymphocytic leukemia cells. Immunogenetics 1 1 :429, 1980

118. Royston I, Majda JA, Baird SM, Meserve BL, Griffiths JC: Human T cell antigens defined by monoclonal antibodies: The 65,000-dalton antigen of T cells (T65) is also found on chronic lymphocytic leukemia cells bearing surface immunoglobulin. J Immunol 125:725, 1980

1 19. Gobbi M, Caligaris-Cappio F, Janossy G: Normal equivalent cells of B cell malignancies: Analysis with monoclonal antibodies. Br J Haematol 54:393, 1983

120. Caligaris-Cappio F, Gobbi M, Boa1 M, Janossy G: Infrequent normal B lymphocytes express features of B-chronic lymphocytic leukemia. J Exp Med 155:623, 1982

121. Gadol N, Ault KA: Phenotypic and functional characterization of human Leu1 (CD5) cells. Immunol Rev 93:23, 1986

122. Hardy RR, Hayakawa K: Development and physiology of Ly-1 Band its human homolog, Leu-1 B. Immunol Rev 9353, 1986

123. Kipps TJ, Vaughan JH: Genetic influence on the levels of circulating CD5 B lymphocytes. J Immunol 139:1060, 1987

124. Kipps TJ: The CD5 B Cell. Adv Immunol47:117, 1989 125. Kipps TJ, DufFy S F Relationship of the CD5 B cell to human

tonsillar lymphocytes that express autoantibody-associated cross reactive idiotypes. J Clin Invest 87:2087, I991

126. Casali P, Burastero SE, Nakamura M, Inghirami G, Notkins AL: Human lymphocytes making rheumatoid factor and antibody to belong to Leu-l+ B-cell subset. Science 236:77, 1987

127. Hardy RR, Hayakawa K, Shimizu K, Yamasaki K, Kishi- moto T: Rheumatoid factor secretion from human Leu- I + B cells. Science 2369 1, 1987

128. Burastero SE, C a d i P Characterization ofhuman CD5 (Leu- 1, OKTl)+ B lymphocytes and the antibodies they produce. Contrib Microbiol Immunol I 1:23 1, 1989

129. Lydyard PM, Quartey-Papafio R, Williams W, Feldman RF, Mackenzie L, Youinou PY, Isenberg DA: The antibody repertoire of early human B cells. 11: Expression of anti-DNA-related idiotypes. J Autoimmun 3:37, 1990

130. Inghirami G, Foitl DR, Sabichi A, Zhu BY, Knowles DM: Normal human B lymphocytes expressing cross reactive idiotypic immunoglobulins: Distribution and characterization in normal human lymphoid tissue, immunoglobulin VH gene expression and relationship to CD5 B lymphocytes. Blood 78:1503, 1991

13 1. Allison A, Alt F, Arnold L, Astrin S, Bishop GA, Casali P, Clark S, Cooper M, Coutinho A, Davidson WF, Gu H, Hardy R, Haughton G, Herzenberg LA, Howard M, Ikuta K, Kantor AB, Keamey JF, Kincade PW, Klinman NR, Kipps TJ, Kishimoto T, Kehrl JH, Kohler G, Koshland M, Kroese FG, Litman G, Lydyard P, Marcos M, McGrath M, Milstein C, Minoprio P, Nolan GP, Nossal GJ, OGarra A, Parnes JR, Plater-Zyberk C, Rajewsky K, Raveche ES, Reth M, Shirai T, Solvason N, Stall A, Takatsu K, Tala1 N, Tucker P, Vakil M, Waldschmidt TJ, Weill J-C, Witz IP: A new nomenclature for B cells. Immunol Today 12:383, 1991

132. Herzenberg LA: Toward a layered immune system. Cell 59: 953, 1989

133. Stall AM, Adams S, Herzenberg LA, Kantor AB: Charac- teristics and development of the murine B-lb (Ly-1 B sister) cell population. Ann NY Acad Sci 651:33, 1992

134. Kasaian MT, Casali P: Analysis of the human CD5- CD45RAlow B-cell subset. Ann NY Acad Sci 65159, 1992

135. Miller RA, Gralow J: The induction of Leu- 1 antigen expres sion in human malignant and normal B cells by phorbol myristic acetate (PMA). J Immunol 133:3408, 1984

136. Freedman AS, Freeman G, Whitman J, Segil J, Daley J, Nadler LM: Studies of in vitro activated CD5+ B cells. Blood 73: 202, 1989

137. Youinou P, Mackenzie L, Jouquan J, Le Goff P, Lydyard PM: CD5 positive B cells in patients with rheumatoid arthritis: Phor- bo1 ester mediated enhancement of detection. Ann Rheum Dis 46: 17, 1987

138. Cong YZ, Rabin E, Wortis HH: Treatment ofmurine CD5- B cells with anti-Ig, but not LPS, induces surface CD5: Two B-cell activation pathways. Int Immunol 3:467, 1991

139. Jyonouchi H, Voss RM, Good RA: Up-regulation and down- regulation of cell surface and mRNA expression of CD5 antigen by various humoral factors on murine 702/3 pre-B cell leukemia cell line: IL-4 down-regulates CD5 antigen expression. Cell Immunol 130:66, 1990

140. Caligaris-Cappio F, Riva M, Tesio L, Schena M, Gaidano G, Bergui L: Human normal CD5' B lymphocytes can be induced to differentiate to CD5- B lymphocytes with germinal center cell features. Blood 73: 1259, 1989

14 1. Kantor AB The development and repertoire of B- 1 cells (CD5 B cells). Immunol Today 12:389, 1991

142. Hayakawa K, Hardy RR: Normal, autoimmune, and malignant CD5+ B cells: The Ly-1 B lineage. Annu Rev Immunol6:197, 1988

143. Hardy RR, Hayakawa K Developmental origins, specificities and immunogiobulin gene biases of murine Ly-1 B cells. Int Rev Immunol8: 189, 1992

144. Waldschmidt T, Snapp K, Foy T, Tygrett L, Carpenter C: B-cell subsets defined by the Fc epsilon R. Ann NY Acad Sci 65 1: 84, 1992

145. Hayakawa K, Hardy RR, Parks DR, Herzenberg LA: The Ly- I B cell subpopulation in normal, immunodefective, and autoimmune mice. J Exp Med 157:202, 1983





146. Hayakawa K, Hardy RR, Herzenberg LA: Peritoneal Ly-1 B cells: Genetic control, autoantibody production, increased lambda light chain expression. Eur J Immunol 16:450, 1986

147. Kroese FG, Ammerlaan WA, Deenen GJ: Location and function of B-cell lineages. Ann NY Acad Sci 651:44, 1992

148. Davidson WF, Pierce JH, Holmes KL: Evidence for a developmental relationship between CD5+ B-lineage cells and mac- rophages. Ann NY Acad Sci 651:112, 1992

149. Forster I, Gu H, Rajewsky K: Germline antibody V regions as determinants of clonal persistence and malignant growth in the B cell compartment. EMBO J 7:3693, 1988

150. Hardy RR, Carmack CE, Shinton SA, Riblet RJ, Hayakawa K A single VH gene is utilized predominantly in anti-BrMRBC hy- bridomas derived from purified Ly- I B cells. Definition of the VH 1 1 family. J Immunol 142:3643, 1989

15 1. Pennell CA, Arnold LW, Haughton G, Clarke SH: Restricted Ig variable region gene expression among Ly- 1 + B cell lymphomas. J Immunol 141:2788, 1988

152. Okumura K, Hayakawa K, Tada T: Cell-to-cell interaction controlled by immunoglobulin genes of Thy-I-, Lyt-1+, Ig+ (B’) cell in allotype-restricted antibody production. J Exp Med 156:443, 1982

153. Marcos MA, de la Hera A, Pereira P, Marquez C, Toribio M, Coutinho A, Martinez C: B cell participation in the recursive selection of T cell repertoires. Eur J Immunol 18:1015, 1988

154. Raveche ES, Lalor P, Stall A, Conroy J: In vivo effects of hyperdiploid Ly-I+ B cells of NZB origin. J Immunol 141:4133, 1988

155. Hardy RR, Hayakawa K A developmental switch in B lym- phopoiesis. Proc Natl Acad Sci USA 88: 1 1550, 1991

156. Gu H, Forster I, Rajewsky K. Sequence homologies, N sequence insertion and JH gene utilization in VHDJH joining: Impli- cation for the joining mechanism and the ontogenetic timing of Lyl B cell and B-CLL progenitor generation. EMBO J 9:2 133, 1990

157. Desiderio SV, Yancopoulos GD, Paskind M, Thomas E, Boss MA, Landau N, Alt FW, Baltimore D: Insertion of N regions into heavy-chain genes is correlated with expression of terminal deoxytransferase in B cells. Nature 31 1:752, 1984

158. Jirik FR, Sorge J, Fong S, Heitzmann JG, Curd JG, Chen PP, Goldfien R, Carson DA: Cloning and sequence determination of a human rheumatoid factor light-chain gene. Proc Natl Acad Sci USA 83:2195, 1986

159. Chen PP, Robbins DL, Jirik FR, Kipps TJ, Carson DA: Iso- lation and characterization of a light chain variable region gene for human rheumatoid factors. J Exp Med 166:1900, 1987

160. Roudier J, Silverman GJ, Chen PP, Carson DA, Kipps TJ: Intraclonal diversity in the VH genes expressed by CD5-negative chronic lymphocytic leukemia producing pathologic IgM rheumatoid factor. J Immunol 144:1526, 1990

16 1. Hsu S-M, Jaffe ES: Phenotypic expression of B-lymphocytes. 2. Immunoglobulin expression of germinal center cells. Am J Pathol I14:396, 1984

162. Gadol N, Peacock MA, Ault KA: Antigenic phenotype and functional characterization of human tonsil B cells. Blood 7 1: 1048, 1988

163. Weinberg DS, Ault KA, Gurley M, Pinkus GS: The human lymph node germinal center cell: Characterization and isolation by using two-color flow cytometry. J Immunol 137:1486, 1986

164. Huang C, Stewart AK, Schwartz RS, Stollar B D Immuno- globulin heavy chain gene expression in peripheral blood B lymphocytes. J Clin Invest 89:1331, 1992

165. Yamada M, Wasserman R, Reichard BA, Shane S, Caton AJ, Rovera G: Preferential utilization of specific immunoglobulin

heavy chain diversity and joining segments in adult human peripheral blood B lymphocytes. J Exp Med 173:395, 1991

166. Youinou P, Mackenzie L, Broker BM, lsenberg DI, Drogou- Lelong A, Centric A, Lydyard PM: The importance of CD5-positive B cells in nonorgan-specific autoimmune diseases. Scand J Rheumatol Suppl 76:243, 1988

167. Martin T, Duffy SF, Carson DA, Kipps TJ: Evidence for somatic selection ofnatural autoantibodies. J Exp Med 175:983, 1992

168. Dighiero G, Lymberi P, Holmberg D, Lundquist I, Coutinho A, Avrameas S: High frequency of natural autoantibodies in normal newborn mice. J Immunol 134:765, 1985

169. Dighiero G, Lymberi P, Guilbert B, Temynck T, Avrameas S: Natural autoantibodies constitute a substantial part of normal circulating immunoglobulins. Ann NY Acad Sci 475:135, 1986

170. Guigou V, Guilbert B, Moinier D, Tonnelle C, Boubli L. Avrameas S, Fougereau M, Fumoux F: Ig repertoire of human polyspecific antibodies and B cell ontogeny. J Immunol 146: 1368, 199 I

171. Mackenzie LE, Youinou PY, Hicks R, Yuksel B, Mageed RA, Lydyard PM: Auto- and polyreactivity of IgM from CD5+ and CD5- cord blood B cells. Scand J Immunol 33:329, 1991

172. Sanz I, Casali P, Thomas JW, Notkins AL, Capra JD: Nu- cleotide sequences of eight human natural autoantibody VH regions reveals apparent restricted use of VH families. J Immunol 142:4054, 1989

173. Crowley JJ, Goldfien RD, Schrohenloher RE, Spiegelberg HL, Silverman GJ, Mageed RA, Jefferis R, Koopman WJ, Carson DA, Fong S: Incidence of three cross-reactive idiotypes on human rheumatoid factor paraproteins. J Immunol 140341 1, 1988

174. Kipps TJ, Robbins BA, Carson DA: Uniform high frequency expression of autoantibody-associated cross reactive idiotypes in the primary B cell follicles of human fetal spleen. J Exp Med 17 1 : 189, 1990

175. Rosenthal MC, Pisciotta AV, Komninos ZD, Goldenberg H, Dameshek W The autoimmune hemoltyic anemia of malignant lymphocytic disease. Blood IO: 197, 1955

176. Ebbe S, Wittels B, Dameshek W Autoimmune thrombo- cytopenic purpura ( “ I T P type) with chronic lymphocytic leukemia. Blood 19:23, 1962

177. Miller DG: Patterns of immunological deficiency in lymphomas and leukemias. Ann Intern Med 57:703, 1962

178. Dameshek W: Chronic lymphocytic leukemia-An accu- mulative disease of immunologically incompetent lymphocytes. Blood 29:566, 1967

179. Bergsagel DE: The chronic leukemias: A review of disease manifestations and the aims of therapy. Can Med Assoc J 96: 16 15, 1967

180. Hansen MM: Chronic lymphocytic leukaemia. Clinical studies based on 189 cases followed for a long time. Scand J Haematol Suppl 18:3, 1973

18 1. Abeloff MD, Waterbury L: Pure red blood cell aplasia and chronic lymphocytic leukemia. Arch Intern Med 134:72 I , 1974

182. Rustagi PK, Han T, Ziolkowski L, Farolino DL, Cunie MS, Logue GL: Granulocyte antibodies in leukaemic chronic lymphoproliferative disorders. Br J Haematol 66:461, 1987

183. Duhrsen U, Augener W, Zwingers T, Brittinger G: Spectrum and frequency of autoimmune derangements in lymphoproliferative disorders: Analysis of 637 cases and comparison with myeloproliferative diseases. Br J Haematol 67:235, 1987

184. Lischner M, Prokocimer M, Zolberg A, Shaklai M: Autoim- munity in chronic lymphocytic leukaemia. Postgrad Med J 64590, 1988

185. Chablani AT, Badakere SS, Bhatia HM: Incidence of antibodies to nuclear antigens, platelets, and circulating immune complexes in leukaemias. Indian J Med Res 88:348, 1988




AUTOANTIBODIES IN CLL

186. Bataille R, Klein B, Dune BG, Sany J: Interrelationship between autoimmunity and B-lymphoid cell oncogenesis in humans. Clin Exp Rheumatol 7:319, 1989

187. Koerner TA, Weinfeld HM, Bullard LS, Williams LC: An- tibodies against platelet glycosphingolipids: Detection in serum by quantitative HPTLC-autoradiography and association with autoimmune and alloimmune processes. Blood 74:274, 1989

188. Spatz LA, Wong KK, Williams M, Desai R, Golier J, Berman JE, Alt FW, Latov N Cloning and sequence analysis of the VH and VL regions of an anti-myelin/DNA antibody from a patient with peripheral neuropathy and chronic lymphocytic leukemia. J Immunol 144:2821, 1990

189. Kobayashi R, Picchio G, Kirven M, Meisenholder G, Baird SM, Carson DA, Mosier DE, Kipps TJ: Transfer of human chronic lymphocytic leukemia to mice with severe combined immune deficiency. Leuk Res 16:1013, 1992

190. Bosma GC, Custer RP, Bosma MJ: A severe combined immunodeficiency mutation in the mouse. Nature 301527, 1983

191. Dorshkind K, Keller GM, Phillips RA, Miller RG, Bosma GC, OToole M, Bosma MJ: Functional status of cells from lymphoid and myeloid tissues in mice with severe combined immunodeficiency disease. J Immunol 132: 1804, 1984

192. Malynn BA, Blackwell TK, Fulop GM, Rathbun GA, Furley AJ, Femer P, Heinke LB, Phillips RA, Yancopoulos GD, Alt FW: The scid defect affects the final step of the immunoglobulin VDJ recombinase mechanism. Cell 54:453, 1988

193. Schuler W, Weiler IJ, Schuler A, Phillips RA, Rosenberg N, Mak TW, Kearney JF, Perry RP, Bosma MJ: Rearrangement of antigen receptor genes is defective in mice with severe combined immune deficiency. Cell 46:963, 1986

194. Schuler W, Schuler A, Bosma MJ: Defective V-to-J recombination of T cell receptor gamma chain genes scid mice. Eur J Im- munol20:545, 1990

195. Mosier DE, Gulizia RJ, Baird SM, Wilson DB: Transfer of a functional human immune system to mice with severe combined immunodeficiency. Nature 335:256, 1989

196. Mosier DE, Baird SM, Kirven MB, Gulizia RJ, Wilson DB, Kubayashi R, Picchio G, Garnier JL, Sullivan JL, Kipps TJ: EBV- associated B-cell lymphomas following transfer of human peripheral blood lymphocytes to mice with severe combined immune deficiency. Curr Top Microbiol Immunol 166:317, 1990

197. Rowe M, Young LS, Cmker J, Stokes H, Henderson S, Rick- inson AB: Epstein-Barr virus (EBV)assoCiated lymphoproliferative disease in the K I D mouse model: Implications for the pathogenesis of EBV- positive lymphomas in man. J Exp Med 1 73: 147, 199 1




1993 81: 2475-2487

TJ Kipps and DA Carson autoimmune diseasesAutoantibodies in chronic lymphocytic leukemia and related systemic

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