Alemtuzumab (Campath-1H) in the Treatment of Chronic Lymphocytic Leukemia

Home , Alemtuzumab, Monoclonal antibody, Rituximab

L Alinari, R Lapalombella, L Andritsos, RA Baiocchi, TS Lin and JC Byrd

Division of Hematology-Oncology, Department of Medicine, The Ohio State University, Columbus, OH, USA

Alemtuzumab (Campath-1H) is a humanizedIgG1 Introduction monoclonal antibody that targets the human CD52 antigen. CD52 is expressedby a variety of lymphoid B-cell chronic lymphocytic leukemia (CLL) is the most neoplasms andmost human mononuclear cell subsets. In common form of leukemia in Western countries with an 2001, alemtuzumab was approvedfor marketing in the annual incidence of 3–5 cases per 100 000. CLL is UnitedStates andEurope for use in patients with characterized by the clonal proliferation and accumula- fludarabine-refractory chronic lymphocytic leukemia tion of mature CD5 þ B lymphocytes in the blood, bone (CLL). In heavily pretreatedpatients with CLL, the marrow, lymph nodes and spleen. The median age of overall response rate (ORR) is approximately 35%, and patients at diagnosis is 65 years, with only 10–15 percent in previously untreatedpatients the ORR is greater than of those diagnosed under 50 years of age. Although it 80%, with a recent randomized study suggesting it is may have an indolent clinical course with survival in superior to alkylator-basedtherapy. Importantly, alemtu- some cases up to 20 years after diagnosis, once the zumab is effective in patients with high-risk del(17p13.1) disease progresses, death is almost unavoidable (Rozman and del(11q22.3) CLL. Alemtuzumab combination studies and Montserrat, 1995; Byrd et al., 2004). Although with fludarabine and/or monoclonal antibodies such as a number of treatment modalities, including alkylator rituximab have demonstrated promising results. Alemtu- therapy, purine analogs, monoclonal antibodies and zumab is also being studied in CLL patients as their combinations are available for CLL, none are consolidation therapy for treatment of minimal residual curative, making identification of new treatments quite disease, in preparation for stem cell transplantation and to relevant. Alemtuzumab is one such therapy. Alemtuzu- prevent acute andchronic graft versus host disease. mab is an unconjugated, humanized IgG1 kappa Alemtuzumab is frequently associatedwith acute ‘first- monoclonal antibody which targets the CD52 antigen dose’ reactions when administered intravenously, but is (Flynn and Byrd, 2000; Frampton and Wagstaff, 2003; much better toleratedwhen administeredsubcutaneously Robak, 2005). This review will encompass both the without loss of therapeutic efficacy. Additional potential biologic characteristics of the CD52 target and the adverse events associated with alemtuzumab administra- available clinical data regarding the use of alemtuzumab tion include myelosuppression as well as profound cellular for treatment of CLL. immune dysfunction with the associated risk of viral reactivation and other opportunistic infections. Additional studies detailing the mechanism of action of alemtuzumab as well as new strategies for prevention of opportunistic The CD52 antigen infections will aidin the future therapeutic development of this agent. The CD52 antigen is a glycoprotein with a short Oncogene (2007) 26, 3644–3653. doi:10.1038/sj.onc.1210380 sequence of only 12 amino acids and a molecular weight of 21–28 kDa. There is a large complex carbohydrate Keywords: B-cell chronic lymphocytic leukemia; CD52 attached to Asparagine-3 which terminates with nega- antigen; alemtuzumab; monoclonal antibody therapy; tively-charged sialic acid residues; this complex is 2–3 T-cell depletion; immunosuppression times larger than the peptide core (Hale et al., 1990; Xia et al., 1993a). At the C-terminus, a glycosylphosphatidylinositol (GPI) lipid anchor attaches the molecule to the outer layer of the cell membrane (Hale, 2001). CD52 is highly expressed on membrane lipid rafts of all B and T lymphocytes (Rossmann et al., 2001) at most stages of differentiation (except plasma cells), as well as on monocytes, macrophages, eosinophils, natural killer Correspondence: Dr JC Byrd, Division of Hematology-Oncology, cells and dendritic cells (Elsner et al., 1996; Buggins Department of Internal Medicine, The Ohio State University, B301 Starling Loving Hall, 320 West 10th Avenue, Columbus, OH, 43210, et al., 2002; Ratzinger et al., 2003). Radioisotopic USA. methods have estimated that peripheral blood lympho- E-mail: [email protected] cytes have approximately 500 000 CD52 molecules per Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3645 cell (Ginaldi et al., 1998), comprising 5% of the cell rituximab in patients with follicular NHL, our group surface. The antigen is also found in the male (Lin et al., 2005b) has not confirmed this response to reproductive tract, where it is strongly expressed on alemtuzumab in CLL. epithelial cells lining the epididymis, vas deferens and Two recent studies have shown that alemtuzumab seminal vesicles (Hale et al., 1993). It is shed into may also induce enhanced apoptosis in vitro, alone or in seminal plasma, from where it can be transferred to combination with a cross-linking anti-Fc antibody, on other cells, particularly spermatozoa. Hematopoietic primary CLL cells in the absence of complement or stem cells, erythrocytes and platelets do not express this immune effector cells, leading to rapid cell death in vitro antigen and are thus spared a direct antibody effect through a nonclassic caspase-independent pathway (Gilleece and Dexter, 1993). (Stanglmaier et al., 2004; Mone et al., 2006). Mone The physiologic functions of CD52 remain to be et al. (2006) also proposed that crosslinking of CD52 by clarified, but it may mediate a variety of biological virtue of its GPI linkage could initiate CLL cell death effects such as signal transduction, promotion of through a membrane raft-dependent mechanism. In cell–cell adhesion and protection of the cell from fact, the authors hypothesized that the observed environmental insult. A recent paper suggested aggregation of glycolipid-enriched membrane domains that CD52 may contribute to the activation of in the CLL plasma membrane surface induced by T-regulatory cells (Watanabe et al., 2006). The CD52 alemtuzumab could mediate subsequent formation of antigen is also expressed on subsets of tumor cells, signaling complexes and induction of cell death (Deans particularly T-cell prolymphocytic leukemia followed et al., 2002; Garcia et al., 2003). There is also evidence to by CLL, hairy cell leukemia, non-Hodgkin lymphomas suggest that alemtuzumab may trigger caspase-depen- (NHL), and acute lymphoblastic leukemia (Ginaldi dent cell death (Nuckel et al., 2005; Smolewski et al., et al., 1998; Cheson, 2006). 2005). Direct cytotoxicity has also been reported in the B-lymphoid cell lines Wien 133 (Rowan et al., 1998) and Ramos (Stanglmaier et al., 2004). Contrasting with this, another study has shown that alemtuzumab alone did Alemtuzumab mechanism of action not induce apoptosis in serum-free medium (Zent et al., 2004). The in vivo relevance of such serum-free experi- Despite the frequent use of alemtuzumab in clinical ments is uncertain. trials, detailed mechanistic studies to elucidate specific Recently, Watanabe et al. (2006) proposed a novel pathways of cell killing have been hampered by the lack function of the CD52 antigen as a potential costimula- of cell lines expressing CD52. Thus, the mechanism of tory molecule for T cells, when they showed that CD52 action of alemtuzumab remains to be clarified. The stimulation of CD4 þ T cells by alemtuzumab con- epitope recognized by alemtuzumab consists of the tributed to the disruption of T-cell transmigration as C-terminal peptide and a portion of the GPI anchor. As well as to the induction of regulatory T cells. CD4 þ such, the alemtuzumab/CD52 complex approximates regulatory T cells are hyporesponsive cells that belong the cell membrane, favoring the deposition of activated to a specialized population of lymphocytes with complement molecules and facilitating the necessary immunosuppressive properties. It has been demon- contacts for cell-mediated killing (Xia et al., 1993b). strated that these cells are able to suppress the polyclonal Alemtuzumab can act through immunological mecha- responses of both CD4 þ and CD8 þ T cells when nisms, such as complement-mediated (CDC) (Xia et al., polyclonal or allogeneic stimulation is applied. These 1993b; Golay et al., 2004; Zent et al., 2004) and/or results suggest that activation of CD4 þ regulatory T antibody-dependent cellular cytotoxicity (ADCC) by cells by alemtuzumab combined with lymphocyte virtue of its IgG Fc region (Hale et al., 1983, 1985; depletion may contribute to the immunosuppressive Crowe et al., 1992). Clynes et al. (2000) showed that the effects of this drug. inhibitory Fcg receptor (Fcg R) IIB is a potent regulator of ADCC in vivo, modulating the activity of Fcg RIII (activation receptor) on effector cells. The authors demonstrated that knocking out the Fcg loci completely Dosage and schedule of administration of alemtuzumab in abrogated the response to the anti-CD20 monoclonal CLL antibody rituximab in a xenograft model of human NHL, whereas knocking out the inhibitory Fcg RIIb The dosing schedule of alemtuzumab was developed enhanced the response to rituximab in the same empirically using primarily clinical response as a xenograft model. Similarly, Zhang et al. (2006) reported surrogate endpoint in initial phase I studies. Further that engagement of Fcg R on effector cells is also the schedule optimization has been limited by a paucity of main mechanism of the in vivo ADCC responses pharmacologic assays due to the difficulty of generating mediated by alemtuzumab against HTLV-1-expressing high-affinity anti-alemtuzumab antibodies. Although a T cells that are CD52 positive. To date, the importance flow cytometric cell surface antigen pharmacologic assay of ADCC in alemtuzumab for B-cell diseases has not exists, it is quite expensive and laborious. To this day, been described. While it has been demonstrated that the the optimal dose and schedule of alemtuzumab remain high-affinity Fcg RIIIA genotype (Cartron et al., 2002) to be defined. It has been estimated that approximately correlates with clinical and molecular response to 125 mg of alemtuzumab are required to saturate all of

Oncogene Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3646 the CD52 binding sites in a healthy adult, considering response was not reported. The median OS was 11 that an adult has an estimated 1012 lymphocytes and months. A recent German CLL Study Group study each lymphocyte has approximately 500 000 binding (Stilgenbauer and Dohner, 2002) demonstrated similar sites for alemtuzumab (Ginaldi et al., 1998). CLL cells efficacy of alemtuzumab (30 mg SQ) three times weekly express similar amounts of CD52 antigen as normal for 12 weeks when compared to historical data where lymphocytes, but their total number may be 10 times the same total dose of alemtuzumab was administered greater. Therefore, more than 1 g of alemtuzumab could i.v. In the first 50 patients enrolled with fludarabine- be required to saturate all CD52 receptors in some CLL refractory CLL, injection-related toxicity was minimal patients. and the ORR was 36% (CR 2%, partial remission (PR) The intravenous (i.v.) dosing schedule currently used 34%), with a median progression free survival (PFS) of as the standard regimen for alemtuzumab therapy is 9.7 months. comprised of a 2-h i.v. infusion at a starting dose of 3 mg day 1, 10 mg day 2 and 30 mg three times weekly for a total of 8–12 weeks, once the infusion-related side effects are tolerable. Given that the majority of patients treated Alemtuzumab as initial therapy for symptomatic CLL with i.v. alemtuzumab experience infusional toxicity combined with the observation that subcutaneous (SQ) A pilot study of nine patients with advanced CLL using alemtuzumab in patients with rheumatoid arthritis had alemtuzumab as first-line therapy demonstrated the biological activity with diminished infusion-related feasibility of this agent in untreated patients (Osterborg events (Isaacs et al., 1992), interest in s.c. administration et al., 1996). On the basis of these findings, Lundin et al. has increased. Indeed, recent reports have demonstrated (2002) treated 41 CLL patients with SQ alemtuzumab as that s.c. alemtuzumab administration achieves blood first-line therapy for up to 18 weeks, with an observed levels equivalent to those seen following i.v. adminis- ORR of 87% including seven patients (19%) with a CR. tration, even if accumulation of alemtuzumab in the Although CLL cells were rapidly cleared from periphe- peripheral blood takes significantly longer (on average, ral blood, alemtuzumab had to be continued for 18 6weeks longer to reach the same concentration obtained weeks to achieve the maximum response in the bone with i.v. administration) (Hale et al., 2004). Thus, SQ marrow. The median time to treatment failure had not dosing is becoming the favored route of administration been reached at the time of publication (18 þ months, in order to decrease toxicity and to facilitate outpatient range 7–44 þ ). The treatment was generally well treatment of CLL patients. tolerated, with adverse events mainly comprised of local injection site reactions, with no episodes of febrile neutropenia or major bacterial infections. Cytomegalo- virus (CMV) reactivation occurred in 10% of patients. Alemtuzumab in previously treatedCLL On the basis of the findings of Lundin et al. (2002) a prospective international randomized phase III trial was The effectiveness of alemtuzumab in patients with performed comparing the efficacy and safety of alemtu- previously treated CLL has been reported in several zumab with chlorambucil as first-line therapy in patients studies. Osterborg et al. (1997) reported a phase II trial with progressive CLL. Patients were randomized to of 29 previously treated CLL patients which demon- receive alemtuzumab (30 mg i.v.) three times weekly for strated an overall response rate (ORR) of 42%, with one a maximum of 12 weeks or chlorambucil 40 mg/m2 complete remission (CR) (4% of patients). The median orally every 28 days for up to a maximum of 12 cycles. duration of response was 12 months in this study. The accrual was completed in July 2004 with 297 Keating et al. (2002) investigated the efficacy and safety patients enrolled. The preliminary results of this trial of alemtuzumab therapy in 93 patients with relapsed or (Hillmen et al., 2006) showed an ORR rate of 83.2% in refractory CLL whose disease was previously treated the alemtuzumab arm as compared to 56% in chlor- with alkyaltor therapy and refractory to fludarabine. In ambucil arm. Alemtuzumab resulted in a 24% CR rate, this study, the ORR was 33%, including two patients whereas there were only 2% CRs in those receiving who achieved a CR (2%). The median duration of chlormabucil. The alemtuzumab arm had slightly more response was 8.7 months and median overall survival infectious complications, predominately due to CMV (OS) was 16months, with a median survival for viremia (46%) and infection (11%) in alemtuzumab- responders of 32 months. Responses in this study were treated patients as compared to CMV viremia (3%) only infrequent among patients with large lymph nodes in the chlorambucil arm. Responses in high-risk (>5 cm) or poor performance status (ECOG perfor- del(17p13.1) patients were more frequent among those mance status X2). The results were confirmed by other randomized to alemtuzumab compared with chloram- phase II trials (Ferrajoli et al., 2003). bucil. These findings corroborate those of others In an attempt to reduce initial infusion-related (Lozanski et al., 2004) who found that alemtuzumab toxicity seen with i.v. alemtuzumab, Bowen et al. may be effective in high-risk CLL as measured by (1997) investigated the efficacy and tolerability of SQ response. Although alemtuzumab improved ORR and alemtuzumab for 6–12 weeks in six fludarabine-refrac- CR rates compared with chlorambucil, longer follow-up tory CLL patients. The ORR was 60%, but no complete is needed to evaluate PFS and OS. Similarly, it will be responses were observed. The median duration of important to assess if improved ORR in patients with

Oncogene Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3647 del(17p13.1) and del(11q22.3) translate to improved combination was well tolerated, but CMV antigenemia PFS and OS. was observed in 27% of patients (13 of the 48 patients), 15% of which were associated with infection that required therapy. This study showed that the combina- Alemtuzumab in combination with other therapies in CLL tion of rituximab with alemtuzumab is feasible and safe, however, the results also indicate that although combi- The efficacy of single agent alemtuzumab in initial nation of these two monoclonal antibodies may initially clinical trials has naturally led to interest in using yield higher ORR, remission duration may be short. alemtuzumab in combination with other chemo- or Future efforts are examining maintenance strategies immunotherapy. Recent studies have demonstrated that using dual antibody therapy. alemtuzumab can be combined with other therapies with an acceptable toxicity profiles as outlined below. Alemtuzumab with fludarabine, cyclophosphamide and rituximab Alemtuzumab andfludarabine Alemtuzumab has been combined with fludarabine, Kennedy et al. (2002) treated six patients with CLL that cyclophosphamide and rituximab with the goal of was refractory to both alemtuzumab and fludarabine as improving CRs and eliminating minimal residual disease single agents, and found that five of six patients (MRD). Wierda et al. (2004) reported the preliminary responded to combination therapy, including one CR. results of a phase II trial in which 31 patients with However, long-term follow-up is not yet available. The pretreated CLL were treated with cyclophosphamide toxicity of this regimen was acceptable, with none of the (250 mg/m2 day 3–5), fludarabine (25 mg/m2 day 3–5, patients developing serious infections. i.v.) alemtuzumab (30 mg day 1, 3, 5) and rituximab Elter et al. (2005) extended these observations to a (500 mg/m2 day 2), every 28 days for six cycles. Twenty- larger cohort of patients with relapsed or refractory one patients were evaluable for response and after a CLL. Thirty-six patients were treated with fludarabine median number of three cycles (range 1–6) the ORR rate (30 mg/m2) and i.v. alemtuzumab. Dose escalation of was 52%, with three patients achieving a CR (14%) and alemtuzumab was performed on consecutive days from eight patients achieving a PR (38%). CMV reactivation 3 to 10 to 30 mg. The combination was administered for was noted in five of 21 patients. Based upon these three consecutive days every 28 days for a total of six results, a larger phase II study in patients with high risk, cycles (four cycles in the first 14 patients). The ORR was previously untreated symptomatic CLL has been 83%, with 11 patients achieving a CR (30%) and 19 initiated at the MD Anderson Cancer Center. patients achieving a PR (53%). The median OS was 35.6 months for all patients, with a time to progression of 22 months in patients who achieved a CR and 13 months Alemtuzumab consolidation for patients who achieved a PR. The treatment was well tolerated with acceptable infectious morbidity. Based Alemtuzumab has been shown to work best against upon these results, a phase III study comparing fludara- blood, spleen and marrow CLL but is less effective bine to fludarabine plus alemtuzumab is currently underway against lymph node disease. Dyer et al. (1997) initially in Europe. demonstrated the feasibility of administering alemtuzumab as consolidation to eliminate MRD. A study by the CALGB was designed to confirm these results (Rai Alemtuzumab andrituximab et al., 2002). Fifty-six previously untreated, symptomatic CLL patients were treated with fludarabine for four Alemtuzumab and rituximab are monoclonal antibodies courses of therapy followed by a 2 month observation that target different antigens on CLL cells, and outside period and then i.v. alemtuzumab (30 mg) three times a of infusion-related events have nonoverlapping toxici- week for six consecutive weeks. Of 36evaluable patients, ties. Faderl et al. (2003) evaluated the safety and efficacy there were 15 CRs (42%) and 18 PRs (50%) with an of rituximab combined with alemtuzumab in 32 patients ORR of 92%. On the basis of the response rates in this with CLL. Patients received rituximab (375 mg/m2) study, the CALGB performed a phase II study of weekly for 4 weeks and i.v. alemtuzumab with the fludarabine and rituximab followed by consolidation escalating dose of 3 mg, 10 mg and 30 mg the first week alemtuzumab. The results of this study are greatly followed by a dose of 30 mg on days 3 and 5 of weeks 2, anticipated. 3 and 4. The ORR was 63% in the CLL patients, with A second study reported by O’Brien et al. (2003) only two (6%) achieving a CR. Almost all patients included 41 patients who had achieved a PR with experienced a complete resolution of circulating CLL conventional chemotherapy. Patients then received i.v. cells, but only 36% attained morphologic remissions in alemtuzumab three times weekly for 4–8 weeks. the bone marrow, possibly as a result of the short Responses were noted in nine of 23 patients (39%) duration of therapy. The median time to progression treated with 10 mg, whereas nine of 16patients (56%) was 6months and the median OS was 11 months. The who were treated with 30 mg responded. A trend toward

Oncogene Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3648 improved responses and remission durations was using alemtuzumab as consolidation and in vivo observed in the patients who received 30 mg. purging followed by autologous HSC collection and The German CLL Study Group (Wendtner et al., transplantation. 2004) reported the results of a phase III trial where patients responding to fludarabine alone or in combination with cyclophosphamide were randomized to receive Alemtuzumab in allogeneic stem cell transplant for CLL i.v. alemtuzumab 30 mg three times weekly for a maximum of 12 weeks, or observation. Of 21 evaluable Allogeneic HSCT is a potentially curative therapy for patients, 11 were randomized to receive alemtuzumab. CLL (Michallet et al., 1996), but its application has been This study was prematurely closed because of severe limited to younger, healthy patients because of the infections in seven of 11 patients in the alemtuzumab potential toxicity of the procedure. Myeloablative arm, whereas only two minor infections occurred in the allogeneic HSCT in CLL has a transplantation-related observation arm. The PFS was significantly improved mortality (TRM) rate of approximately 50% (Flinn and for patients receiving alemtuzumab (P ¼ 0.036) at a Vogelsang, 1998; Hale et al., 2002). Acute and chronic median follow-up of 21.4 months. Collectively, these graft-versus-host disease (aGVHD, cGVHD) remain studies along with other noncontrolled reports (Moreton significant causes of morbidity and mortality. In recent et al., 2005) provide support for this approach in the years, reduced intensity conditioning regimens have setting of a clinical trial. Off study, consolidation therapy been developed that are designed to be immunosup- with alemtuzumab is currently not supported by the pressive rather than myeloablative (Sorror et al., 2005). available literature. Preliminary studies suggest that the addition of alemtuzumab to nonmyeloablative conditioning regimens may reduce the severity of GVHD, presumably Alemtuzumab in autologous stem cell transplantation through in vivo T-cell depletion (Kottaridis et al., 2000; Pe´ rez-Simo´ n et al., 2002; Faulkner et al., 2004; Delgado As discussed previously, alemtuzumab is most effective et al., 2006). Kottaridis et al. (2000) treated 44 patients in the absence of bulky nodal disease. The elimination of with lymphoid and myeloid malignancies using i.v. MRD is crucial for the outcome of CLL patients alemtuzumab in combination with a fludarabine-based undergoing autologous hematopoietic stem cell trans- nonmyeloablative conditioning regimen. The engraft- plantation (HSCT) due to concerns regarding graft ment rate was 98% (42 of the 43 evaluable patients) and contamination. At the same time, it is important that 33 of 43 evaluable patients (75%) were alive without therapy not interfere with peripheral blood HSC evidence of progression/relapse after a limited follow-up mobilization and harvesting. The efficacy of alemtuzu- of 9 months. Interestingly, the incidence of aGVHD was mab for the treatment of MRD (O’Brien et al., 2003; very low – only two patients (5%) developed grade II Wendtner et al., 2004; Moreton et al., 2005; Montillo and no patients developed grade III–IV aGVHD. The et al., 2006) and the absence of CD52 on hematopoietic authors also reported a low incidence of cGVHD, stem cells (Gilleece and Dexter, 1993) support the although this could not be fully assessed because of the rationale for in vivo purging with alemtuzumab before short duration of follow-up at the time of publication. HSC collection. Pe´ rez-Simo´ n et al. (2002) retrospectively compared the Dyer et al. (1997) was the first to report six CLL results of two prospective trials carried out in the United patients treated with purine analogs to maximal Kingdom (Kottaridis et al., 2000) and Spain (Martino response followed by i.v. alemtuzumab for in vivo et al., 2001). The two groups used an identical purging of residual disease after chemotherapy. Five fludarabine-based nonmyeloablative regimen but used patients achieved MRD-negative CRs with no detect- different GVHD prophylaxis: Kottaridis et al. (2000) able CD5 þ /CD19 þ cells by flow cytometry in the used i.v. alemtuzumab as part of the conditioning peripheral blood or bone marrow. Moreover, the regimen at a dose of 20 mg/day on days À8toÀ4 and authors showed that alemtuzumab did not affect stem Martino et al. (2001) used methotrexate. Both groups of cell collection, and stem cell products were not patients received cyclosporine. The patients who re- contaminated with CD5 þ /CD19 þ cells in three cases. ceived alemtuzumab had a significantly lower incidence This preliminary report was recently built upon by of aGVHD that persisted even after donor lymphocytes Montillo et al. (2006), who treated 34 patients infusion (DLI). The authors found no significant with sequential therapy including fludarabine-based difference in terms of TRM rate between the two regimens followed by s.c. alemtuzumab for 6weeks. groups, but did note an association between develop- Overall, HSC collection after intermediate-dose ment of aGVHD and disease response. Patients who cytarabine (800 mg/m2 every 12 h for six doses) plus received alemtuzumab required DLI to achieve a similar granulocyte-colony-stimulating factor was successful in response rate when compared with those who received 24 (92%) of 26patients. Eighteen patients subsequently methotrexate. Furthermore, alemtuzumab significantly underwent HSCT after a median of 11 months from reduced the incidence of aGVHD incidence but was the end of alemtuzumab therapy, and 17 of these 18 associated with a higher rate of CMV reactivation. A patients were in CR at a median follow-up of 28 months recent study (Delgado et al., 2006) reported results of an after alemtuzumab therapy. The above study provides alemtuzumab-based reduced intensity regimen in 41 the first evidence demonstrating the feasibility of CLL patients. The engraftment rate was unexpectedly

Oncogene Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3649 low (85%) when compared with those previously Hematologic andimmunologic toxicity reported for NHL patients who received the same conditioning regimen (Morris et al., 2004). Delgado The majority of CLL patients experience transient et al. (2006) reported excellent results with a condition- cytopenias during initial therapy with alemtuzumab, ing regimen consisting of alemtuzumab 20 mg/day on especially if pre-existing cytopenias are present or days À8toÀ4 and combined with fludarabine and significant prior therapy has been administered. Throm- melphalan, with an ORR of 100% in chemosensitive bocytopenia is most common during the first 2 weeks of patients and 86% in chemorefractory patients. These therapy and may necessitate platelet transfusion, parti- responses were durable in 74 and 43% of chemosensitive cularly in patients with either pre-existing cytopenias or and chemorefractory patients, respectively. Acute significant infusional toxicity. Neutropenia is most GVHD was observed in 17 patients (41%), but only common during weeks 5 and 6(Keating et al., 2002). four patients (10%) developed grade III or IV aGVHD. Grade IV neutropenia in the absence of infection or Thirteen patients (33%) developed extensive cGVHD. progressive disease is not an absolute contraindication The 2-year TRM rate was 26%, mostly due to to therapy. Prophylactic filgrastrim for neutropenia is infections. Only one study has compared alemtuzumab not generally effective (Lin et al., 2005a) and may result with other agents for in vivo T-cell depletion. Juliusson in prolonged periods of neutropenia if administered for et al. (2006) found that patients who received alemtu- extended periods of time. Monocytes, natural killer cells zumab as part of the conditioning regimen initially had and peripheral blood (but not tissue) antigen-presenting less initial toxicity with fewer fevers and lower transfu- dendritic cells are also profoundly depleted by alemtu- sion requirements, the patients who received 30 mg/day zumab (Buggins et al., 2002; Klangsinsirikul et al., for three consecutive days were more likely to have 2002). T- and B lymphocytes subpopulations are heavily mixed chimerism post-transplant requiring DLI, and depleted, with the nadir usually occurring between had more serious opportunistic infections which trans- weeks 3 and 6of therapy (Lundin et al., 2004). These lated to greater non-relapse mortality. At the present often do not recover until 1 year or greater after therapy time, the role of alemtuzumab in non-myeloablative (Keating et al., 2002). The route of administration does stem cell transplant for CLL is uncertain. not seem to affect the hematologic and immunologic activities of alemtuzumab. Autoimmune hemolytic anemia has been reported following alemtuzumab administration but the pathogenesis remains unclear. Toxicity of alemtuzumab

Alemtuzumab is associated with predictable toxicities that may be prevented or abrogated, such as infusion- Infectious complications with alemtuzumab related events and infectious complications. As the experience with alemtuzumab increases and larger The stage of the disease, number of prior therapies and numbers of patients are treated, previously unrecog- degree of underlying myelo- and immunosuppression nized toxicities are being reported as summarized below. increase the susceptibility to opportunistic and other severe infections irrespective of treatment with alemtuzumab (Perkins et al., 2002). The incidence of infection Infusion-relatedevents appears to be augmented further by alemtuzumab. Rai et al. (2002) observed major infections in 10 (42%) of 24 The most common adverse events during alemtuzumab pretreated CLL patients during alemtuzumab therapy. therapy are infusion-related, including rigors, fever, In the pivotal study by Keating et al. (2002), 51 (55%) of nausea, vomiting, skin rash, dyspnea and hypotension. the 93 pretreated CLL patients experienced one or more These symptoms are a consequence of cytokine release infections: septicemia occurred in 14 patients (15%), 11 (TNF-a, IFN-g IL-6and IL10) likely derived from experienced opportunistic infections, including seven natural killer cells (Wing et al., 1996). Such infusional CMV reactivations, six herpes simplex reactivations, events generally occur with the first dose, with signi- four herpes zoster reactivations, two patients developed ficant variability in the severity of symptoms. In aspergillus pneumonia and one patient each developed general, the intensity and frequency of infusional events Pneumocystis jiroveci (previously Pneumocystis carinii), usually decrease over the course of therapy. Systemic rhinocerebral mucormycosis, systemic candidiasis, cryp- infusional events can be reduced by initial dose tococcal pneumonia and Listeria monocytogenes escalation and premedication with diphenhidramine meningitis. Infections are usually observed during alem- and acetaminophen. Steroids (hydrocortisone) may be tuzumab treatment or within 30 days after completing added in cases of prior severe adverse reactions therapy, and only occasionally seen after discontinuation (Osterborg et al., 2006), and generally are effective. of therapy (Lundin et al., 2004). Infusional events may also be significantly reduced by The most frequently observed opportunistic infection SQ administration. SQ administration does induce a local during alemtuzumab therapy is CMV reactivation, injection site inflammatory reaction (erythema/edema) which is seen in between 15 and 25% of patients that in most patients is self-limited (Lundin et al., 2002; (Nguyen et al., 2002). CMV reactivation is usually Hale et al., 2004). observed shortly after the T-cell nadir, between weeks 4

Oncogene Alemtuzumab (Campath-1H) in the treatment of CLL L Alinari et al 3650 and 6of therapy. Antiviral prophylaxis (acyclovir, Sterility with alemtuzumab famciclovir, valaciclovir) is effective for the prevention of herpes simplex and varicella zoster virus reactivation Epithelial cells lining the epididymis, vas deferens and and is recommended in all patients receiving alem- seminal vesicles strongly express the CD52 antigen (Hale tuzumab. However, these agents do not prevent et al., 1993) and shed it into seminal plasma, from where CMV reactivation. Fortunately, most cases of CMV it can be transferred to other cells, particularly reactivation respond rapidly to i.v. ganciclovir spermatozoa. Although alemtuzumab has been shown (Keating et al., 2002; Lundin et al., 2004). A randomized to agglutinate and inactivate sperm in vitro,todateno study of prophylactic versus pre-emptive therapy reproductive complications with systemic therapy have with valgancyclovir (O’Brien et al., 2005) demonstrated been reported, probably because alemtuzumab does not CMV reactivation could virtually be eliminated by reach the seminal fluid in significant concentrations. prophylaxis. Despite this single study, prophylactic treatment for CMV is controversial as is weekly monitoring for CMV antigenemia or viral DNA copy Conclusions numbers. Our approach consists of weekly monitoring during therapy and for 2 months afterward, with Alemtuzumab is a humanized monoclonal antibody that pre-emptive treatment if evidence of CMV viremia is targets the pan-lymphocyte antigen CD52 and has detected. Anti-Pneumocystis jiroveci prophylaxis is also demonstrated activity in CLL. Importantly, several recommended for these patients. recent studies have demonstrated that alemtuzumab is active against high-risk del(17p13.1) CLL which generally does not respond to other standard therapies in this setting. The widespread use of alemtuzumab has in Cardiac toxicity with alemtuzumab part been limited by concerns regarding toxicity, including infusional events, myelosuppression and im- Controversial data exists concerning the possibility of mune suppression. Recent data suggests that adminis- cardiovascular toxicity with alemtuzumab (Keating et al., tration of SQ alemtuzumab may diminish serious 2002; Ferrajoli et al., 2003; Lenihan et al., 2004). Cardiac infusional toxicity. In addition, a phase III study events, including arrhythmias and development of con- comparing alemtuzumab to chlorambucil as primary gestive heart failure, have been reported in both T-cell therapy found that fewer opportunistic infections (Lenihan et al., 2004) and B-cell malignancies (Keating occurred in patients treated with alemtuzumab, indicat- et al., 2002; Ferrajoli et al., 2003). In the randomized ing that some infections observed in patients with CLL phase III trial comparing alemtuzumab with chlorambucil may in fact be related to underlying immune dysfunc- as first-line therapy (Hillmen et al., 2006), serious cardiac tion related to the disease itself. Future studies with events were observed in four patients treated with alemtuzumab should focus on efforts to better under- alemtuzumab compared with no cardiac complications stand the mechanism of action of this antibody and the observed in the chlorambucil arm. function of CD52 in both normal and transformed T- The possible mechanism of cardiac events with and B cells. alemtuzumab remains unknown. CD52 is not expressed on the endothelial cell membrane, making direct injury Acknowledgements to the coronary endothelial cells an unlikely explana- tion. It has been hypothesized that cardiac toxicity may This work supported by the Leukemia and Lymphoma be the result of cytokine release leading to coronary Society, D Warren Brown Foundation, and National Cancer artery spasm (Lenihan et al., 2004). Institute.

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Oncogene