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Keywords:

  • rituximab;
  • CD20+ plasma cells;
  • immunotherapy;
  • Y-90 ibritumomab;
  • I-131 tositumomab

Summary

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

CD20 is a particularly appealing target that is expressed on the surface of almost all B cells, with no significant shedding, secretion or internalization. In contrast to the demonstrated efficacy of anti-CD20 strategies in various B-cell lymphoproliferative disorders, the role of such therapy in multiple myeloma is undetermined and controversial. The expression of CD20 by myeloma cells is heterogeneous, and can be detected only in 13–22% of patients. However, there is increasing interest in testing anti-CD20 therapy in myeloma because of recent studies suggesting the existence of clonogenic CD20-positive precursor B cells in the disease. This article reviews the rationale, preclinical and clinical activity of anti-CD20 therapy in myeloma. Clinical trials show that anti-CD20 therapy with rituximab elicits a partial response in approximately 10% of CD20+ patients with multiple myeloma. In addition, there is preliminary evidence of disease stabilization in 50–57% of CD20+ patients for a period of 10–27 months. Further large-scale clinical trials are therefore needed to establish the role of this promising strategy in the treatment of myeloma.

The advent of monoclonal antibodies has ushered a period of rapid evolution and monumental changes in the landscape of therapy for many B-cell malignancies. Rituximab, a chimaeric mouse/human monoclonal antibody (mAb) against the CD20 antigen, is the first and arguably the most successful mAb approved for the treatment of cancer thus far (Grillo-Lopez, 2000). The initial success of rituximab in indolent follicular lymphomas was followed by positive results in diffuse large cell lymphoma. The R-CHOP regimen (rituximab, cyclophosphamide, adriamycin, oncovin, prednisolone) has now become the treatment of choice for most cases of diffuse large cell lymphoma (Coiffier et al, 2002; Campbell & Marcus, 2003; Feugier et al, 2005). Marked interest has also been generated by the objective response seen with this mAb in Waldenstrom Macroglobulinemia (WM; Dimopoulos et al, 2002; Gertz et al, 2004). Although rituximab and other strategies targeting CD20 show remarkable benefits in various B-cell malignancies, multiple myeloma (MM) is an exception.

CD20 surface antigen

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

CD20 is a transmembrane phosphoprotein expressed on committed B cells through all the stages of their development, but its expression is downregulated at the point of differentiation into plasma cells (Riley & Sliwkowski, 2000; Smith, 2003). The exact function of this pan-B-cell antigenic marker on the B cell or its malignant counterpart is not well known. It is probably involved in B-cell activation and differentiation, and regulation of transmembrane calcium conductance (Tedder & Engel, 1994; Grillo-Lopez, 2000; Riley & Sliwkowski, 2000). CD20 is particularly appealing for serotherapy by virtue of neither being shed from the cell surface nor secreted into the circulation. Consequently, after intravenous infusion, the anti-CD20 mAb reaches the target site in adequate amounts without any hindrance or competitive adsorption by soluble antigen in the circulation. Furthermore, the attachment of the mAb to CD20 does not lead to the downregulation or internalization of this antigen (Grillo-Lopez, 2000; Smith, 2003).

CD20 expression on plasma cells

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

The differential response of myeloma and non-Hodgkin lymphoma (NHL) to surface antigen-targeted therapy is noteworthy. The weak (<30% CD20+ plasma cells) and somewhat heterogeneous expression of CD20 on abnormal plasma cells, as opposed to predominantly uniform expression on lymphoma cells, is primarily responsible for this striking variation between the two malignancies (Fig 1).

image

Figure 1.  Immunohistochemical staining of bone marrow plasma cells demonstrates surface expression of CD20 (original magnification ×400).

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Although the majority of myeloma patients lack overexpression of this antigen on mature plasma cells (San Miguel et al, 1991; Gorschluter et al, 2001), initial studies showing complete absence of CD20 antigen on MM cells have been refuted by newer findings demonstrating heterogeneous (partial or complete) expression of this protein in 13–22% MM patients (Robillard et al, 2003). However, in this subpopulation, only a subset of patients expresses CD20 antigen on the majority of its myeloma cells (San Miguel et al, 1991; Leo et al, 1992; Li et al, 2002a; Treon et al, 2002; Robillard et al, 2003; Lin et al, 2004a).

A previous study noted that the expression of CD20 antigen on myeloma cells was associated with lower patient survival, and therefore portended poorer prognosis, as compared to CD20-negative myeloma patients (13 vs. 25 months; P = 0·05; San Miguel et al, 1991). More recent studies, however, demonstrated a correlation between CD20+ MM cells and a favourable prognostic marker, translocation t(11;14) (Mateo et al, 2005). In one study, this translocation was seen in 83% of CD20+ MM patients (Robillard et al, 2003). The CD20 phenotype was also found to be associated with lymphocytoid plasma cell morphology (Fonseca et al, 2002; Robillard et al, 2003; Matsuda et al, 2005). Additionally, a higher frequency of CD20 expression (50%) has been observed in de novo plasma cell leukaemia (Garcia-Sanz et al, 1999).

Detection of a potentially therapeutic target like CD20 is easily possible by use of an informative, widely available and cost-effective procedure, such as flow cytometry (Davis et al, 2007). Indeed, the international consensus group for plasma cell immunophenotyping has recommended a panel of antibodies, including CD20, as useful for evaluation and characterization of neoplastic plasma cells in bone marrow aspirate material (Braylan et al, 2001). The aberrant expression of this antigen may not always be detected in a newly diagnosed case of MM, and it may be useful to reassess for its expression at the time of exploration of other therapeutic options in a relapsed or refractory case (Bergua et al, 2007). The intensity of CD20 expression (weak, moderate or strong) may also dictate the clinicians’ approach to such patients. Furthermore, a substantial number of MM patients have commonly demonstrated only a distinct subpopulation of CD20+ tumour cells, with the remainder of the tumour cells not expressing this antigen (Lin et al, 2004a). Consideration of all these factors prior to the initiation of treatment may be crucial to predicting a response to anti-CD20 therapy.

Anti-CD20 monoclonal antibodies: mechanism of action

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

Rituximab is a chimaeric mouse/human mAb that targets both normal and malignant CD20+ cells. The mechanism of action of anti-CD20 antibodies on myeloma cells has not been studied extensively. It is presumed to be similar to that in lymphoma patients in whom the binding of murine Fab region of rituximab to CD20+ cells leads to apoptosis, complement activation and antibody-dependent cell-mediated cytotoxicity (ADCC). ADCC, a potent mechanism of action, is promoted by the human Fc portion of the chimaeric mAb which recruits activated natural killer cells, granulocytes, macrophages and monocytes (Smith, 2003). The presence of the human Fc portion allows repeated use of rituximab and prolongs its half-life by reduction of the human anti-murine antibody (HAMA) response (Reff et al, 1994; Smith, 2003).

Y-90 ibritumomab tiuxetan and I-131 tositumomab are the two currently available anti-CD20 radioimmunoconjugates. Myeloma cells are exquisitely radiosensitive, and these new radiolabelled mAbs could potentially destroy both CD20+ and CD20− plasma cells.

Ibritumomab tiuxetan is a murine mAb against CD20 antigen. It is covalently attached to tiuxetan that, in turn, is chelated to a pure beta-emitter, 90-Yttrium. I-131 tositumomab, another murine anti-CD20 antibody, contains the radioisotope, iodine-131 which delivers both beta- and gamma-radiation. The purely murine structure of both radioimmunoconjugates makes them suitable for faster clearance from the body and prevents damage to normal tissues by shortening exposure to radiation. The ‘crossfire’ or the ‘bystander’ effect of these radiolabelled antibodies is due to the radioisotopes, 90-Yttrium and Iodine-131, which confer on them an advantage over the ‘cold’ antibody. They target not only the cells which express CD20, but also the surrounding CD20-negative tumour cells, as well as the microenvironment. The long beta path length (5–11 mm) and high energy of 90-Y potentially render this outpatient therapy suitable for a malignancy like myeloma with a heterogeneous CD20 antigen expression (Sharkey & Goldenberg, 2006; Witzig, 2006).

Rationale for Anti-CD20 therapy in myeloma

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

The identity of the cell of origin in MM remains to be established (Berenson et al, 1998; Ruffini & Kwak, 2001; Jones et al, 2004). Several theories, including presence of a proliferative and non-proliferative compartment have been proposed (Epstein, 1997; Berenson et al, 1998; Mitterer et al, 1999; Pilarski et al, 2000).

This dual compartment theory states that two types of cell populations co-exist in MM tumours; a small, discrete proliferative B-cell population with ability to replicate, and a larger, quiescent population of terminally differentiated MM plasma cells formed from the clonogenic cells (cells giving rise to a colony with identical genetic constitution). These clonogenic cells possess the property of self-renewal and differentiation into mature tumour cells. Such clonogenic cells with stem cell-like properties may be responsible for malignant growth of myeloma (Reya et al, 2001). The hedgehog (Hh) signalling pathway allows clonal expansion of CD138 negative myeloma stem cells without differentiation. Such cells have been shown to express high levels of smoothened (SMO), a protein essential for Hh pathway activation. On the other hand, Hh receptor, patched 1 (Ptch 1), which is responsible for tonic inhibition of Hh signalling, is highly expressed in the differentiated compartment of CD138+ myeloma cells, possibly explaining the low proliferative index of mature, neoplastic plasma cells (Peacock et al, 2007). These cells bear similarity to their normal counterparts, the terminally differentiated, end stage and non-dividing plasma cells (Calame, 2001).

The proliferative index of MM plasma cells is low, and several investigators have verified the existence of clonogenic B cells sharing immunoglobulin gene sequences with myeloma cells (Billadeau et al, 1993; Kosmas et al, 2000; Guikema et al, 2004). Such clonogenic cells appear to originate from preswitched, but somatically mutated B cells (Billadeau et al, 1993; Corradini et al, 1993; Bakkus et al, 1994). The data of Matsui et al (2004) suggest that the myeloma stem cells are distinct, postgerminal centre B cells that express CD20, and lack CD138 antigens. Unlike the well differentiated plasma cells, the stem cells may be a rare cell population (Chen & Epstein, 1996; Matsui et al, 2004). Due to their capacity of self-renewal, such cells could contribute to the initiation, progression and maintenance of the disease. This CD20+ cellular compartment therefore serves as an attractive target in halting the progression of myeloma. Despite intensified chemotherapy, the clonotypic (genetically identical progeny of a single precursor) CD19+/CD20+ cell fraction has been shown to persist in the peripheral blood of myeloma patients (Cremer et al, 2001). Therefore, targeting this clonotypic B-cell fraction, especially in the minimum residual disease stage, theoretically offers a potential to prevent relapse (Rottenburger et al, 1999).

In contradistinction to the above view, the experiments of Yaccoby and Epstein (1999) demonstrated the proliferative and self-renewal capacity of mature MM cells. They showed that mere inoculation of purified CD38+, CD45− MM plasma cells into the human bones of SCID-hu mice (severe combined immunodeficiency mice in which human haematolymphoid organs can be engrafted) results in the development of MM in these hospitable hosts, and that other preplasmacytic and accessory cells could not produce myeloma in SCID-hu hosts. In addition, Davies et al (1999, 2000) questioned the existence of a significant level of clonogenic cells in the B-cell compartment of MM patients, and argued that the demonstration of clonality in some studies is probably a result of contamination by the myelomatous cells that cannot be removed during the B-cell enrichment process. Rigorous sequential gating strategies are able to identify only a single stage of neoplastic plasma cell differentiation with even distribution of proliferative fraction between CD45-positive and CD45-negative cells. These findings reject the discrete stem cell compartment theory, and state that mature, neoplastic plasma cells represent an independent self-replenishing population (Rawstron et al, 2001). Furthermore, it was demonstrated that mature MM cells cocultured with human osteoclasts retain plasticity, and could dedifferentiate into the immature stem cell phenotype (Yaccoby, 2005).

Therefore, even if the existence of CD20+‘premyeloma’ cells in the B-cell compartment is questioned, several investigators continue to evaluate the role of anti-CD20 therapy due to the expression of CD20 antigen on the surface of differentiated plasma cells in a subset of myeloma patients.

Global gene expression profiling has shown spikes for MS4A2 (CD20) gene expression in five patients in MM1 or MM2 subgroups, out of a total of 74 newly diagnosed MM cases (Zhan et al, 2002). Subsequently, a larger sample of 414 newly diagnosed patients revealed seven subgroups in which a CD (Cyclin D)-2 subgroup was characterized by overexpression of MS4A1 (CD20) (Zhan et al, 2006). Similarly, another signature of more immature B cells, PAX5 (BSAP), which correlates with CD20 expression, has been found in 72% of CD20+ MM. It is believed to be a remnant of an earlier stage of development since repression of PAX5 is necessary for plasma cell development (Lin et al, 2004b; Shapiro-Shelef & Calame, 2004).

Rituximab: preclinical antimyeloma activity

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

It is ironic that the synthesis of monoclonal antibodies hinged on the principle of creating immortal cell lines/hybridomas to produce specific antibodies by fusion of mouse myeloma cells to splenic lymphocytes (Kohler & Milstein, 1975), and that today we are investigating the destructive effect of one such antibody on myeloma cells themselves.

A preclinical study showed that incubation of CD138−/CD34− clonogenic myeloma progenitors with rituximab resulted in significant inhibition of tumour growth as compared to the untreated myeloma marrow samples (Matsui et al, 2004). Moreover, the addition of complement, which facilitates the lysis of rituximab-tagged cells, further suppressed the in vitro MM colony formation (Matsui et al, 2004). Another in vitro study demonstrated the enhancement of CD20 expression on myeloma cells with thalidomide, and therefore, increased suppression of myeloma colony formation was noted with the combination of rituximab and thalidomide (Li et al, 2002b).

The expression of CD20 is dependent on PU.1, a haematopoietic transcription factor that is downregulated in myeloma plasma cells (Pettersson et al, 1995). In the presence of interferon consensus sequence binding protein (ICSBP) or Pip, a member of the interferon regulatory factor (IRF) family, PU.1, binds to the CD20 promoter, thereby activating it (Pettersson et al, 1995; Himmelmann et al, 1997). Not surprisingly, gamma-interferon is a potent inducer of CD20 antigen at pharmacologically achievable levels, and leads to upregulation of this antigen on the CD20-negative myeloma cell lines (Treon et al, 2002). Furthermore, Fc receptors on monocytes are augmented nearly 10-fold with gamma interferon (Guyre et al, 1983). ADCC is therefore potentiated when gamma-interferon is used with mAb therapy (Weiner et al, 1988). On the other hand, while it has been noted that interferon-alpha has induced in vitro upregulation of CD20 expression in CLL B cells (Sivaraman et al, 2000), it did not affect CD20 expression on MM plasma cells (Treon et al, 2002).To date, no clinical trials using a combination of rituximab and interferon in MM patients have been published.

Clinical trials with rituximab in multiple myeloma

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

Only a few case reports and studies of anti-CD20 therapy in MM have so far enriched the medical literature (Table I). Rituximab has been used in different settings in myeloma patients. A few case reports and small phase 2 trials have employed anti-CD20 therapy based on the degree of expression of CD20 surface antigen, both in newly diagnosed MM and refractory/relapsed cases (Table I). Others have directed this therapy against the clonogenic B cells to attempt eradication of the proliferative compartment (Hofer et al, 2003; Zojer et al, 2006). It has also been used in the minimal residual disease setting, postautologous stem cell transplant, as maintenance therapy (Gemmel et al, 2002; Musto et al, 2003; Lim et al, 2004).

Table I.   Studies and case reports of rituximab therapy in multiple myeloma.
ReferenceTumor typeNumber of patientsCD20 expressionDosing scheduleResponseSide effectsComments
  1. ASCT, autologous stem cell transplant; MM, multiple myeloma; dex, dexamethasone; CD, cluster designation; VAD, vincristine, adriamycin and doxorubicin; MRD, minimal residual disease; CR, complete remission; BMPCs, bone marrow plasma cells; SD, stable disease; MiR, minor response; PD, progressive disease; TTP, time to progression; PFS, progression-free survival; AIHA, autoimmune haemolytic anaemia; PCs, plasma cells; Hb, haemoglobin; BM, bone marrow; DS, Durie-Salmon; PR, partial remission; PB, peripheral blood; S/P, status post; BJ, Bence Jones; MP, melphalan-prednisone; MR, major response; ORR, overall response rate.

Bergua et al (2007)Post-ASCT, relapsed MM, Stage IIIa, refractory to bortizomib/dex 1Initially CD20 negative; upon relapse CD20+; CD20% not reported375 mg/m2/week for 4 cycles along with VAD in standard dose, prior to 2nd ASCTClinical improvement after 1st cycle; no MRD after the 4th cycle; patient remains in CR 5 months post- 2nd ASCTNot reportedPossibility of synergism of rituximab with concurrent chemotherapy in a patient with evolving immunophenotype from CD20 negative to CD20+
Greipp et al (2008)Newly diagnosed MM 190% CD20+ BMPCs375 mg/m2 q week for 4 weeks followed by a repeat 4-week course 3 months later. Then, maintenance therapy at 375 mg/m2 q 2 months for 6 monthsSD for 12 monthsMild infusion-related nausea, fever and rigors with first infusion onlyUnderscored the importance of patient selection and individualizing therapy. Substantial clinical improvement in a patient with CD20+ BMPCs
Gozzetti et al (2007)MM pretreated with double transplant 1100%375 mg/m2 q week for 4 weeks 3 months post-2nd ASCTCRNot reportedExcellent response to rituximab postdouble ASCT, illustrates its usefulness in MRD in a patient with 100% CD20+  BMPCs
Moreau et al (2007)Never pretreated MM (7); refractory or relapsed MM (7)14All patients had CD20+ MM; range of expression: 33–100%375 mg/m2 q week for 4 weeksMiR (1); SD (5); SD followed by progression (3); no response (5)Well toleratedA small, prospective phase II trial. Selection based on the level of CD20 expression with at least 33% of tumor cells expressing this antigen
Zojer et al (2006)Relapsed MM (9); Refractory MM (1)10CD20+ (2), CD20 expression 10% and 50% respectively375 mg/m2 on days 1, 8, 15 and 22SD at 6 months (2). 1 patient with CD20+ BMPC had SD but withdrew at 3 months. PD (7). Median TTP: 4·2 months Grade ≤2 chills and hypotension (4); infection (2) A small phase II study with inadequate response. CD20+ cells’ contribution to disease progression was questioned by the authors
Lim et al (2004)MM or refractory MM (4)10Not reported375 mg/m2 on day +30 post-ASCT. Continued q3 months for 2 years, or until MM progressedCR: 3 months postrituximab maintenance (2), CR: 6 months after therapy with rituximab (2). CR: pretreatment (2)Moderate–severe infections (6); fatal pneumonia (1)Another study using rituximab maintenance therapy in MRD. Severe IgM deficiency and increased risk of infection noted with post-transplant rituximab use
Musto et al (2003)Newly diagnosed MM (s/p ASCT) 6All CD20-negative375 mg/m2 q week for 4 weeks followed by α-interferon Relapsed (5); median PFS after ASCT: 11 months comparatively, PFS in a previous series of 21 MM patients with ASCT alone (without rituximab) was 38 monthsNot reportedHigh rate of early relapse with rituximab as maintenance therapy after ASCT but the number of patients was too low to draw definitive conclusions
Hofer et al (2003)MM with severe AIHA 1CD20-negative PCs375 mg/m2 q week for 4 weeks after failure of VADIgG reduced dramatically. Hb normalized. BM had no morphological evidence of MMNot reportedLikely eradication of any CD20+ myeloma precursor cells (proliferative compartment) in this CD20-negative MM
Treon et al (2002)Previously treated MM, DS stage II or III19CD20-negative (7); low CD20+ (4); mixed (4); high (2); unknown (2)Rituximab alone q week for 4 weeksPR (5%); SD (26%); median time to treatment failure: 5·5 months Moderate haemoptysis (1). Overall, therapy was well toleratedAll responders had CD20 expression. No response in 64% of patients, despite reduction of B cells, reflects a lack of B-cell involvement in disease progression in majority of MM patients
Gemmel et al (2002)Plasma cell leukaemia (MRD, s/p ASCT) 122·6% CD20+ cells in PB. In enriched CD20+ fraction: 0·093% clonotypic cells375 mg/m2 q week for 4 weeks for consolidation Day 90: tumor load increased; day 120: relapse with reappearance of BJ proteinuriaGrade I flu-like symptomsRole of clonotypic B cells as proliferative compartment is questioned in this report
Hsi et al (2001)Newly diagnosed MM41100% of responders (6 patients) co-expressed CD20375 mg/m2 q week for 4 weeks q 6 months for 6 cycles; later, MP was also added16% MR and MiR on rituximab alone. With addition of MP, 42% MR, 25% MiR, 28% SD, and 6% progressedGrade 3 myalgias (1); myelosuppressionWith addition of rituximab to MP, ORR improved but not the quality of response. Myelosuppression was more pronounced than seen with MP alone
Treon et al (1999) Refractory MM168% CD20+ BMPCs375 mg/m2 q week. After PR 4 more weekly coursesPR 3 months post-therapy but MM progressed 6 months after initiation of rituximab Not reportedCD20+ PCs and B cells disappeared after therapy but CD20-negative PCs remained. In vitro study with interferon, showed increased CD20 expression
Korte et al (1999)Progressive MM1Strong/diffuse expression 375 mg/m2 q week for 4 weeks (rituximab used as 4th line therapy)30% rise in M-spike plus new osteolytic lesionsNot reportedInconclusive about the effect of rituximab on tumor load progression, but close observation of M spike suggested during rituximab therapy

The response to anti-CD20 therapy has been variable, ranging from rapid disease progression, analogous to the ‘flare’ seen in WM patients started on rituximab (Ghobrial et al, 2004; Treon et al, 2004), to stabilization of the disease, and even partial response. Rituximab has been used as a single agent or an adjunct to other myeloma therapies. Regardless, its place in the management of myeloma is far from established.

One study involving 19 MM patients included 10 patients with variable CD20 expression. A partial response (reduction in serum or urine M-protein of more than 50%) was seen in one (10%) of the 10 patients whose plasma cells expressed CD20 antigen (Treon et al, 2002). Another five (50% of patients with CD20+ myeloma cells) had stabilization of their disease. Two of these patients with progressive disease prior to rituximab therapy, had ongoing stable disease (<25% change in serum or urine M-protein, in the absence of new or other progressive signs or symptoms of MM) at 18+ and 27+ months.

In a more recent phase II trial (Moreau et al, 2007), 14 patients (seven treatment-naive stage I, and seven relapsed or refractory, stage III) were selected according to the level of CD20 expression (33–100% expression). All patients with stage I disease and one with Stage III relapsed myeloma showed stabilization of their MM for variable periods of time after monotherapy with rituximab. One patient in the Stage III group experienced an ongoing 18-month minor response. This patient was unique in that 100% of this patient’s tumour cells were CD20+, and this distinction probably explains the observed response. Despite the authors’ overall observation of a relatively low impact on outcome with rituximab in CD20+ MM, one could draw the conclusion that the percentage of tumour cells that are positive for CD20 determines response to rituximab monotherapy in MM.

The prolonged survival of mature, malignant plasma cells even after the elimination of clonogenic ‘premyeloma’ CD20+ stem cells could possibly skew the true response as seen by the persistence of paraprotein shortly after, or during therapy (Jones et al, 2004). The production of monoclonal protein by the surviving mature myeloma cells is not affected. Therefore, the laboratory and histopathological parameters, such as the M-spike and the percentage of the bone marrow plasma cells are slow to respond, and unlikely to decrease dramatically soon after or during the therapy (Jones et al, 2004). The lack of a rapid, meaningful response has unfortunately contributed to premature abandonment of this therapy, leading to a paucity of case reports and clinical experience with rituximab use in MM.

As can be inferred from Table I, the level of CD20 expression on plasma cells was not a criterion for patient selection in most of the reported studies and cases. The disappointing results may be partly attributable to random selection of the patients based on the postulation of invariable existence of a proliferative, clonogenic CD20+ compartment. It is plausible that extended treatment with rituximab may eventually demonstrate a response by severing the source of malignant plasma cells due to complete destruction of the proliferative CD20+ clonogenic B cells (Jones et al, 2004).

Adverse effects of anti-CD20 therapy

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

The favourable toxicity profile of monoclonal antibodies offers considerable attraction to the clinicians to explore and expand their use. Fever (53%), chills and rigors (33%) are usually encountered with the first infusion of rituximab, but these reactions are managed by slowing or interrupting the infusion (Boye et al, 2003). Headache (19%), abdominal pain (14%), myalgia (10%), rash (15%), nausea (23%) and vomiting (10%) are usually of grade 1 to 2 severity (http://www.rituxan.com/lymphoma/HCP/ISISafetyProfile.jsp).

Hypotension, angioedema, hypoxia and bronchospasm may be a manifestation of a more serious infusion reaction, typically occurring within 30–120 min of the first infusion. Other potentially fatal, but rare events include severe hypersensitivity reactions, life-threatening cardiac arrhythmias and angina, tumour lysis syndrome associated with renal failure, progressive multifocal leukencephalopathy and severe mucocutaneous reactions.

It is recommended that patients be appropriately screened for hepatitis B virus prior to initiation of rituximab, as reactivation of hepatitis B virus has been reported with this immunosuppressive therapy (Westhoff et al, 2003; Niscola et al, 2005; Lalazar et al, 2007). In general, myeloma patients tolerate anti-CD20 therapy well, but rare reports of IgM suppression and increased risk of life-threatening infections necessitate caution with its use (Table I; Korte et al, 1999; Lim et al, 2004).

Cytopenias, primarily lymphopenia, which can be grade 3 or 4 in approximately 40% of patients, are common with rituximab. The median duration of lymphopenia is 14 d.

Severe cytopenias are commonly associated with both Y-90 ibritumomab tiuxetan and I-131 tositumomab. Hypothyroidism may be a complication in patients treated with I-131 tositumomab, and TSH-check prior to the therapy, and annually thereafter is recommended. The combination of the alkylating agent, melphalan and anti-CD20 therapy potentially exposes the patient to the risk of severe myelosuppression and myelodysplasia (Hsi et al, 2001).

Rituximab: mechanisms of resistance in myeloma

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

One of the mechanisms of action of rituximab is complement-dependent cytotoxity (CDC) which is initiated via the human Fc portion of rituximab. The presence of complement regulators/inhibitors on myeloma cells can antagonize this action of rituximab, conferring resistance to this drug.

Recent work has elucidated that the expression of complement regulatory proteins, CD55 and CD59, is higher in both normal and malignant plasma cells as compared to normal B cells (Alcindor et al, 2006). This may explain the reduced response rate to rituximab.

Treon et al (2001) showed that expression of the complement regulator, CD59 on MM and NHL cell lines, is associated with resistance to rituximab. However, these findings were refuted by Weng and Levy (2001) who found no significant difference in the level of expression of the complement inhibitors, CD46, CD55 and CD59 in the follicular lymphoma responders and non-responders to rituximab. They, however, cautioned that their data cannot be extrapolated to other B-cell malignancies. Another potential mechanism of resistance to anti-CD20 therapy is Fc receptor polymorphism, as it influences the binding of the effector cells to the rituximab-tagged cell, thereby severely compromising ADCC (Smith, 2003; Friedberg, 2005; Sharkey & Goldenberg, 2006; Moreau et al, 2007).

Limitations of rituximab use in myeloma

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

There are several limitations to the use of rituximab in MM. Unfortunately, the majority of MM patients have either dim or low frequency expression of CD20 antigen on their plasma cells, which substantially limits the efficacy of this mAb, particularly if one believes that MM plasma cells themselves are the proliferative cells with self-renewal potential, and that another clonogenic B-cell compartment does not exist. Moreover, plasma cells may lose CD20 expression with rituximab maintenance therapy. This finding gives credence to the idea that the initial response is seen when the CD20 expression is relatively better. It is typically followed by a period of progression of myeloma when the CD20+ plasma cells are either killed, or masked for expression of this antigen while the patients are maintained on anti-CD20 therapy (Table I).

Future directions

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

Despite significant advances in the management of MM over the past decade, complete cure continues to elude us, and the disease eventually relapses (Rajkumar & Kyle, 2005). It is easier and more economical to develop monoclonal antibodies than to engineer patient-specific anti-idiotype antibody (Campbell & Marcus, 2003). A number of phase I and II clinical trials are incorporating radioconjugates to exploit their ‘crossfire effect’ against the radiosensitive myeloma cells. A phase I trial testing the effect of ibritumomab on myeloma cells, and the safety of integrating 90-Yttrium ibritumomab with high dose melphalan chemotherapy and autologous stem cell transplant, is currently underway at the Mayo Clinic (http://clinicaltrials.mayo.edu/clinicaltrialdetails.cfm?trial_id=100217). Another trial is assessing the utility of Y-90 ibritumomab in patients with incomplete response to chemotherapy prior to ASCT (http://www.nemc.org/home/medicaleducation/Stem%20Cell%20News/Stem%20Cell%20Newsletter.pdf). A phase II trial is accruing stage II and III MM patients who have achieved partial remission with other therapies, to assess the role of consolidation therapy with I-131 tositumomab (http://clinicaltrials.gov/show/NCT00135200).

Another phase II trial is testing the combination of cyclophosphamide and rituximab in patients with relapsed or refractory disease (http://www.hopkinskimmelcancercenter.org/clinicaltrials/protocol.cfm?pID=J0478). The results of these trials and others will hopefully shed more light on the role of anti-CD20 therapy in this B-cell malignancy, and could possibly be just the beginning of what appears to be a promising approach in MM.

Several next-generation anti-CD20 antibodies including veltuzumab (IMMU-106, hA20), ocrelizumab, ofatumomab (HuMaxCD20), AME-133v, GA-101, rhuMAb v114 and PRO70769 are currently being studied in NHL and autoimmune disorders. Their anti-myeloma activity is yet to be investigated, but with increased CDC and ADCC, and improved binding to the lower-affinity polymorphism FCγR3a receptor, they may be potentially useful in MM (Bello & Sotomayor, 2007; Maloney, 2007).

Emerging therapeutic monoclonal antibodies against multiple myeloma

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

That the quest for an ideal target on myeloma cells continues, is attested by the development of a panoply of monoclonal antibodies over the last few years (Maloney et al, 1999). While no mAb has been approved for the management of MM so far, significant preclinical activity by many provides grounds for conducting clinical trials, especially in the scenarios of low tumour burden, where immunotherapy is likely to be more effective (Chatterjee et al, 2006). In this section, we briefly discuss some other investigational mAbs with promising anti-myeloma activity.

Humanized monoclonal antibodies

HuLuc63, a mAb against cell surface CS1, a novel antigen highly expressed on MM cells, inhibits MM cell adhesion to bone marrow stromal cells (BMSCs), and triggers ADCC (Tai et al, 2007). Similarly, by induction of ADCC and CDC, AHM, a mAb against HM1·24 glycoprotein on both normal and neoplastic plasma cells, has demonstrated significant inhibition of tumour growth and cytolysis in MM xenograft mouse models (Ozaki et al, 1997; Kawai et al, 2006). Cytokines interleukin (IL)2, IL10, IL12 and macrophage colony-stimulating factor restore the diminished ADCC that can potentially limit the action of anti-HM1·24 antibody in refractory MM (Ozaki et al, 1997). HuN901, with high affinity towards CD56, is used to deliver a maytansine derivative, DM1, a potent tubulin-polymerization inhibitor, to the targeted MM cells. Its toxicity is limited because the active drug is released only in CD56+ cells upon attachment of the conjugated-mAb to such cells (Tassone et al, 2004a). Milatuzumab (hLL1), an anti-CD74 antagonist antibody is being studied as a carrier for toxins, chemotherapeutic drugs and radioisotopes (Burton et al, 2004; Stein et al, 2007). Notably, rituximab is reported to augment its antiproliferative effect on MM cells (Stein et al, 2004). Alemtuzumab, a mAb to CD52 epitope, is still under investigation in MM due to a high degree of heterogeneity in CD52 expression on myeloma cells (Kumar et al, 2003; Carlo-Stella et al, 2006). Its limitations included modest clinical activity and significant toxicities, including renal insufficiency and pancytopenia, with subsequent opportunistic infections (Gasparetto et al, 2004). A recent study reported significantly higher CD52 expression even on alemtuzumab-resistant cells as compared to MM cells (Rawstron et al, 2006). Thus, its potential value may be limited to highly selected myeloma cases with strong CD52 expression. The anti-CD40 mAb, SGN-40, demonstrated preliminary evidence of anti-myeloma activity in a phase I dose-escalation analysis involving 16 patients with relapsed/refractory myeloma. The first-dose related cytokine release reaction, characterized by severe headaches and aseptic meningitis, was also observed (Hussein et al, 2005). Tocilizumab is a genetically engineered humanized anti-IL6 receptor mAb that inhibits myeloma cell growth by interfering with IL-6 mediated actions (Katzel et al, 2007). A new receptor inhibitor (NRI) composed of VH and VL of tociluzumab in a single chain fragment, and dimerized by fusion to the Fc portion of human IgG1, has been developed for adenovirus gene-mediated delivery, and sustained in vivo production in SCID mice (Yoshio-Hoshino et al, 2007). Whether this adenovirus vector-related therapeutic strategy reduces the costs associated with the production of mAbs remains to be seen.

Chimaeric antibodies

Like rituximab, anti-CD70 and anti-CD54 (c-CUV3) are chimaeric mAbs that could prove to be effective agents with further clinical development, by interfering with regulation of antigen-primed B-cell differentiation, and blocking the interaction between CD54 and leucocyte function-associated antigen, respectively (Smallshaw et al, 2004; Bringhen et al, 2006; Coleman et al, 2006; McEarchern et al, 2007). A humanized variant of anti-CD70 mAB also shows potent anti-MM activity (McEarchern et al, 2005). Despite overcoming the safety concerns of HAMA associated with murine anti-IL6 antibody, its chimaeric mouse/human counterpart, CNTO 328 (CCLB IL6/8), has been ineffective in producing a meaningful response in MM (van Zaanen et al, 1998).

Fully human antibodies

CD40 is expressed on both myeloma cell and clonally related proliferating B-cell populations. CD40 activation contributes to myeloma cell migration and homing via activation of AKT/nuclear factor-kB signalling, and induces myeloma cell adhesion to fibronectin and BMSCs. Blocking these pathways by CHIR 12·12, a fully human anti-CD40 antagonist antibody, could potentially prove to be an effective treatment strategy (Tai et al, 2005). A fully human anti-HLA-DR antibody, 1D09C3, has activity in both in vitro myeloma cell culture studies, as well as in vivo murine studies with myeloma xenografts. Pretreatment with interferon-γ (IFNγ) is noted to enhance its therapeutic effect (Nagy et al, 2002; Carlo-Stella et al, 2007). Humax-CD38 antibody is also a human anti-CD38 IgG1 antibody in preclinical development (Stevenson, 2006). Another antibody derivative constructed by combining two monomeric domain antibodies, simultaneously targets both CD38 and CD138 antigens by avidly binding to CD38+/CD138+ MM cells (Stevenson, 2006).

Murine antibodies

β2-Microglobulin-specific mAbs have demonstrated ADCC and CDC-independent and caspase-dependent in vivo tumouricidal activity in xenograft mouse models of MM. In contrast to CD20, the blocking effect of soluble β2-microglobulin is an impediment to their use (Yang et al, 2006). Anti-syndecan-1 (CD138) mAb, B-B4, when conjugated with the cytotoxic, maytansine derivative, DM1, forms a novel immunoconjugate selectively targeting CD138+mature MM cell compartment, but not the clonogenic compartment (CD138−/CD20+). Although it demonstrates anti-myeloma activity in human-MM models in mice, the lack of CD138 expression on mouse tissues prevents further evaluation of its toxicity in such preclinical models (Tassone et al, 2004b). A recombinant single-chain Fv diabody (2D7-DB) constructed from murine mAb against the highly expressed HLA class I antigen on MM cells, bypasses the Fc-mediated effector mechanisms. It may therefore have clinical applicability in immunosuppressed myeloma patients (Sekimoto et al, 2007). The antiproliferative effects of the murine anti-IL-6 mAb, B-E8, have been studied in different clinical settings in MM with mixed results so far (Klein et al, 1991; Bataille et al, 1995; Moreau et al, 2000, 2006; Trikha et al, 2003; Rossi et al, 2005) . The clinical usefulness of murine anti-CD19 mAb, anti-B4, conjugated to modified ricin, targeting clonogenic CD19+ MM cell compartment, has not been demonstrated (Grossbard et al, 1993).

Familiarity with the effects of rituximab on human subjects has perhaps contributed to a relatively greater experience with this mAb in MM compared to many other mAbs currently under clinical development.

Conclusion

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References

From a practical standpoint, a clinician often faces multiple challenges in managing the newly diagnosed and the relapsed or refractory myeloma patients. A number of factors, including the prognostic indicators, presence of symptoms, age, quality of life, performance status and co-morbid conditions, determine the further course of management. The focus has also shifted towards tumour characteristics and biology as the molecular basis of myeloma continues to unravel. Regimens like melphalan-prednisone (MP), vincristine, adriamycin and dexamethasone (VAD), thalidomide or lenolidamide plus dexamethasone, and bortezomib-based therapies all show considerable activity against myeloma cells, albeit at the cost of significant side effects. Anti-CD20 therapy on the other hand, is relatively well tolerated, but its activity against myeloma cells is ill-defined, with limited objective responses seen in a few small studies.

A number of issues that still remain unanswered need to be addressed in the setting of larger clinical trials. The optimal dosage of rituximab, the duration of treatment with this drug, its integration with combination chemotherapy, or its place in sequential anti-myeloma therapy are still to be determined. In addition, the subset of myeloma patients that shows a durable response to anti-CD20 antibody therapy, and the level of CD20 expression that will translate to enhanced clinical activity with this immunotherapy will have to be unequivocally defined.

Clinically, it is important not to overlook the potential benefits of CD20-targeted therapy in individual patients with myeloma. Patients who harbour the t(11;14) translocation, or who demonstrate lymphoid morphology are more likely to express CD20. Such patients and all patients with limited options should be screened for CD20 expression on bone marrow plasma cells. If a high level and frequency of expression is demonstrated, one should consider anti-CD20 directed therapy, preferably alone, in order to assess the benefit. It would be premature to entirely dismiss the potential role of anti-CD20 therapy in the management of myeloma. With the availability of limited information in the current medical literature, the jury is still out on establishing its place in the therapeutic armamentarium against this incurable malignancy.

References

  1. Top of page
  2. Summary
  3. CD20 surface antigen
  4. CD20 expression on plasma cells
  5. Anti-CD20 monoclonal antibodies: mechanism of action
  6. Rationale for Anti-CD20 therapy in myeloma
  7. Rituximab: preclinical antimyeloma activity
  8. Clinical trials with rituximab in multiple myeloma
  9. Adverse effects of anti-CD20 therapy
  10. Rituximab: mechanisms of resistance in myeloma
  11. Limitations of rituximab use in myeloma
  12. Future directions
  13. Emerging therapeutic monoclonal antibodies against multiple myeloma
  14. Conclusion
  15. Acknowledgements
  16. References
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