Use Of Campath-1H Research Articles

The Effects of CAMPATH-1H on Cell Viability Do Not Correlate to the CD52 Density on the Cell Surface

Graft versus host disease (GvHD) is one of the main complications after hematological stem cell transplantation (HSCT). CAMPATH-1H is used in the pre-transplant conditioning regimen to effectively reduce GvHD by targeting CD52 antigens on T cells resulting in their depletion. Information regarding CD52 expression and the effects of CAMPATH-1H on immune cells is scant and limited to peripheral blood (PB) T and B cells. To date, the effects of CAMPATH-1H on cord blood (CB) cells has not been studied. Here we aimed to analyze CD52 expression and the effects of CAMPATH-1H on fresh or frozen, resting or activated, PB mononuclear cells (PBMC) and CB mononuclear cells (CBMC). In resting state, CD52 expression was higher in CB than PB T cell subsets (653.66±26.68 vs 453.32±19.2) and B cells (622.2±20.65 vs 612.0±9.101) except for natural killer (NK) cells where CD52 levels were higher in PB (421.0±9.857) than CB (334.3±9.559). In contrast, CD52 levels were comparable across all cell types after activation. CAMPATH-1H depleted resting cells more effectively than activated cells with approximately 80–95% of apoptosis observed with low levels of necrosis. There was no direct correlation between cell surface CD52 density and depleting effects of CAMPATH-1H. In addition, no difference in cell viability was noted when different concentrations of CAMPATH-1H were used. CD52 was not expressed on HSC but began to be expressed as the cells differentiate, implying that CAMPATH-1H could potentially affect HSC differentiation and proliferation. Our study provides insightful information, which contributes to the better understanding in the use of CAMPATH-1H as part of the conditioning regime in HSCT.

The risk of early and late CMV DNAemia associated with Campath use in stem cell transplant recipients.

The risks associated with in vivo and ex vivo use of Campath-1H and -1G in a cohort of 206 stem cell transplant recipients for cytomegalovirus (HCMV) DNAemia have been quantified. DNAemia showed a biphasic incidence pattern with an inflexion at day 60. The first phase had a linear risk rate for HCMV DNAemia of 0.3 % day−1 whilst the second phase had a substantially lower risk rate of 0.058 % day−1. In multivariable analyses, risk factors for early DNAemia were HCMV serostatus, radiotherapy based conditioning and CD34 stem cell dose, with the use of in vivo Campath-1H having the most significant risk (Hazards Ratio = 3.68 (95% CI 2.02-6.72; p<0.001). Ex vivo use of Campath was not associated with an increased risk for HCMV DNAemia. Patients receiving either in vivo Campath-1H or -1G experienced HCMV DNAemia earlier (27 and 33 days respectively) compared to patients receiving no Campath (time to DNAemia, 51 days; p = 0.0006). Multivariable analysis of risk factors for HCMV DNAemia occurring beyond 100 days after transplant were older age, acute GVHD > grade II and a lower CD34 stem cell dose whereas Campath-1H use was not associated with late HCMV DNAemia.

396: B Lymphocyte Reconstitution Following Allogeneic Stem Cell Transplantation for Haematological Disorders: Correlation with Pre- and Post-Transplant Factors

We have performed a detailed analysis of the kinetics of B cell reconstitution in patients who have undergone allogeneic stem cell transplantation for haematological disorders. Whole blood samples from 76 patients were analysed using monoclonal antibodies directed against CD19, CD27 and IgD by multicolour flow cytometry. Total (CD19+), naive (IgD+CD27-), IgD memory (IgD+CD27+) and class switched memory (IgD-CD27+) B cell populations were identified. Indications for transplantation were haematological malignancy (n = 74) and aplastic anaemia (n = 2). The stem cell source was from an unrelated donor in 29 cases or related/sibling donor in 47 cases. 39 patients received reduced intensity and 37 full intensity transplant conditioning regimens. In vivo T-cell depletion with Campath 1H was used for all patients with a matched unrelated donor and those receiving reduced intensity conditioning. The majority of patients (n = 57) have retained a predominantly naive B cell phenotype at 2 to 48 months (median 18.7 months). 7 patients have a normal percentage of memory B lymphocytes with a low percentage of class-switched B cells. 7 patients currently have a normal percentage of class-switched B cells. 8 patients achieved normal class-switched B cell numbers but returned to a naive B cell phenotype. In one patient this preceded relapse of underlying leukaemia and in two others was consequent on treatment with chemotherapy or corticosteroid. Only one patient who required treatment with additional immunosuppressive therapy, post transplant, subsequently achieved a normal percentage of class-switched B cells. The use of Campath 1H, stem cell source (marrow or peripheral blood), donor type (unrelated or related), use of full or reduced intensity conditioning or the administration of donor lymphocyte infusions post-transplant were not predictive of development of a normal percentage of class-switched B lymphocytes. B cell immune reconstitution phenotype does not appear to correlate with the incidence of chronic GvHD or in this cohort of patients with current disease status. The timing of development of a normal percentage of class-switched B cells cannot be reliably predicted in this patient population. We have performed a detailed analysis of the kinetics of B cell reconstitution in patients who have undergone allogeneic stem cell transplantation for haematological disorders. Whole blood samples from 76 patients were analysed using monoclonal antibodies directed against CD19, CD27 and IgD by multicolour flow cytometry. Total (CD19+), naive (IgD+CD27-), IgD memory (IgD+CD27+) and class switched memory (IgD-CD27+) B cell populations were identified. Indications for transplantation were haematological malignancy (n = 74) and aplastic anaemia (n = 2). The stem cell source was from an unrelated donor in 29 cases or related/sibling donor in 47 cases. 39 patients received reduced intensity and 37 full intensity transplant conditioning regimens. In vivo T-cell depletion with Campath 1H was used for all patients with a matched unrelated donor and those receiving reduced intensity conditioning. The majority of patients (n = 57) have retained a predominantly naive B cell phenotype at 2 to 48 months (median 18.7 months). 7 patients have a normal percentage of memory B lymphocytes with a low percentage of class-switched B cells. 7 patients currently have a normal percentage of class-switched B cells. 8 patients achieved normal class-switched B cell numbers but returned to a naive B cell phenotype. In one patient this preceded relapse of underlying leukaemia and in two others was consequent on treatment with chemotherapy or corticosteroid. Only one patient who required treatment with additional immunosuppressive therapy, post transplant, subsequently achieved a normal percentage of class-switched B cells. The use of Campath 1H, stem cell source (marrow or peripheral blood), donor type (unrelated or related), use of full or reduced intensity conditioning or the administration of donor lymphocyte infusions post-transplant were not predictive of development of a normal percentage of class-switched B lymphocytes. B cell immune reconstitution phenotype does not appear to correlate with the incidence of chronic GvHD or in this cohort of patients with current disease status. The timing of development of a normal percentage of class-switched B cells cannot be reliably predicted in this patient population.

519: Campath-1H therapy for resistant rejection in pediatric heart transplant patients

Purpose: Resistant rejection in pediatric heart transplant patients is a cause of significant mortality; we describe the first successful use of Campath-1H in this setting. Campath-1H is an anti-CD52 monoclonal antibody, primarily utilized for treatment of GVH disease. CD52 is found on T & B cells, macrophages, monocytes and NK cells.

Risk Factors for Predicting CMV Infection in Allogeneic-PBSCT Setting: MNC Dose, Campath Use, and Acute GVHD Grade ≥II.

Risk factors for predicting CMV infection in allogeneic-PBSCT setting: MNC dose, Campath use, and acute GVHD grade ≥IIBackground: CMV infection remains an important complication of allogeneic stem cell transplantation (SCT). Campath-1H (anti-CD52) antibodies have been widely used in vivo for T-cell depletion following conventional and reduced intensity conditioning regimens and associated with a significant reduction in GVHD but at the cost of impaired immune reconstitution. We studied the risk factors, for the development of CMV infection after allogeneic SCT, including the use of Campath-1H.Methods: From Sep. 1998 to May 2006, 122 patients who received allogeneic peripheral blood stem cell transplantation were enrolled. CMV infection was routinely sought by at least weekly screening for CMV related matrix protein pp65 antigenemia after engraftment (WBC>1,500/uL) was achieved. CMV antigenemia was treated with ganciclovir 5mg/kg twice daily i.v. as preemptive therapy for at least 10 days.Results: The overall incidence of CMV infection was 56/122 (45.9%) cases with a median onset of 44 days post transplants. Risk factors for CMV infection in the multivariate analysis were acute graft-versus-host disease (GVHD) grade II-IV (p=0.015), use of Campath-1H in conditioning therapy (p=0.012), and mononuclear cell (MNC) doses in the transplants (p=0.010). Among conditioning therapies, the use of Campath-1H was associated with increased incidence of CMV infection (75.0%) compared with conventional conditioning therapy (40.9%)(p<0.01) and relatively early occurrence of CMV infection, median 22 days post transplant (p=0.016).Conclusion:The incidence of CMV infection was strongly associated with acute GVHD, MNC doses, and especially Campath-1H use in an allogeneic PBSCT setting.Multivariate Analysis for the Incidence of CMV infectionRR95% CIP-valueConditioning: use of Campath-1H5.1121.429–18.2850.012Acute GVHD, grade II-IV2.6961.209–6.0100.015Stem cell dose: MNC(×10^8/kg)1.1491.034–1.2760.010Stem cell dose: MNC ≥10 vs. MNC<102.6131.165–5.8590.020

Determining the Appropriate PCR Trigger for Pre-Emptive Anti-CMV Therapy in Allogeneic Stem Cell Transplant Recipients.

Background: CMV-related complications are a major cause of morbidity and mortality post allogeneic stem cell transplantation (ASCT). Where a pre-emptive strategy is used, early detection is crucial as this will minimise the risk of CMV infection progressing to CMV disease. As a result, PCR techniques are increasingly replacing pp65 antigen detection as a means of monitoring CMV viral load in at-risk patients. Initiating treatment at too low a cut-off may result in over-treatment since very low level viraemia may not necessarily represent live or replicating virus. However, using too high a cut-off might lead to unacceptable treatment delay. As a result of these uncertainties the optimal pre-emptive strategy remains unknown. We therefore sought to determine whether, using quantitative real time PCR, the manufacturer's detection cut-off or the validated lower quantitation limit (LQL) was the more appropriate treatment trigger.Methods: CMV surveillance was done by weekly (twice weekly for positive results) quantitative real time PCR (Artus RealArt™ CMV LC PCR Kit, Hamburg, Germany) until immunosuppression was stopped. The manufacturer's detection cut-off was 6.5 × 102 copies/ml and the validated LQL was 1 × 104 copies/ml. Treatment was initiated following two consecutive results above the LQL. Below the LQL, treatment was only started if symptoms of CMV infection (e.g. fever, cytopenia) were also present.Results: Between 01/04 and 12/05 fourty four patients, median age 43y (range 18–69), underwent ASCT from sibling (n=27) or unrelated (n=17) donors for AML/MDS (n=22), ALL (n=8), CML (n=5), NHL (n=5), myeloma (n=3) and osteogenesis imperfecta (n=1). The conditioning regimen was myeloablative (Cy-TBI, Bu-Cy) in 21/44 and reduced intensity (Flu-Mel, Flu-Cy) in 23/44. Campath-1H was used in 16/44. The recipient/donor CMV serostatus was R+/D+ (n=11), R+/D− (n=9), R−/D+ (n=4) and R−/D− (n=20). With a median follow up of 10 months (range <1–26), 12/24 (50%) patients at high risk of CMV reactivation (6 of 11 R+/D+, 6 of 9 R+/D−, 0 of 4 R−/D+) had CMV detected compared with 1/20 (5%) low risk R−/D− patients. CMV detection occurred at a median of 21d (range 3–128) post ASCT. 12/13 patients with detectable CMV had initial levels below the LQL. All required treatment, 5 because of co-existent symptoms of CMV infection despite consecutive readings below the LQL and 7 because the second reading was above the LQL. Conditioning therapy, donor type and use of CAMPATH-1H did not appear to influence risk of CMV detection. There was no CMV related death in this cohort.Conclusion: We conclude that, using the above kit, the detection cut-off of 6.5 × 102 copies/ml rather than the manufacturer's validated LQL of 1 × 104/ml is the more appropriate trigger for initiating pre-emptive anti-CMV therapy in patients undergoing ASCT. Since manufacturers do not consider quantification below the LQL to be reliable, we recommend that where the detection cut-off is significantly lower than the LQL, transplant centres seek to determine their own appropriate treatment trigger.

Prolymphocytic Leukemias

T and B subtypes of prolymphocytic leukemias (PLLs) are rare, highly aggressive lymphoid malignancies with characteristic morphologic, immunophenotypical, cytogenetic, and molecular features. Recent studies have highlighted the role of specific oncogenes such as TCL1, MTCP-1, and ATM in the case of T-cell and p53 mutations in the case of B-cell PLLs. Despite the advances in the understanding of the biology of these conditions, prognosis for these patients remains poor with short survival and no curative treatment. The advent of monoclonal antibody therapy has improved treatment options for this group. In particular, the use of Campath-1H, in T-PLL has more than doubled median survival. The role of allogeneic transplant with nonmyeloablative conditioning needs to be explored further.

Steroid-free Tacrolimus Monotherapy After Pretransplantation Thymoglobulin or Campath and Laparoscopy in Living Donor Renal Transplantation

Living donor renal transplantation was performed under a regimen of recipient pretreatment and low-dose postoperative immunosuppression with subsequent weaning. From October 9, 2002, to December 31, 2004, 196 consecutive, unselected laparoscopic live donor nephrectomies resulting in 196 living donor renal transplantations were performed. Recipients were pretreated with rabbit antithymocyte globulin (thymoglobulin; 24 patients or [12%]) or Campath 1H (alemtuzumab; 166 patients [85%]), or were not in protocol (6 patients [3%]), and were given postoperative steroid-free low-dose tacrolimus immunosuppressive monotherapy with subsequent weaning. There was no donor mortality. Major and minor donor morbidities were 2.6% and 4.2%, respectively. Laparoscopic live donor nephrectomy recipient outcomes with a mean follow-up of 401 days included (1) recipient and graft survival of 99.0% and 97.4%, respectively; (2) no ureteral stenosis; (3) 0.5% delayed graft function, from recurrent focal segmental glomerulosclinosis; and (4) no vascular thrombosis. The incidence of acute rejection at 30, 90, and 401 days was 1.5%, 3.8%, and 11.2% (all 196 recipients), 0%, 25%, and 29.2% (thymoglobulin recipients), and 1.8%, 3.9%, and 8.4% (Campath 1H recipients), respectively. Sixty-six patients (33.7%) are receiving spaced-dose immunosuppressive monotherapy. The mean creatinine concentration in all recipients was 1.5 ± 1.1 mg/dL. There were no instances of cytomegalovirus tissue invasive disease or posttransplantation lymphoproliferative disease. The incidence of new-onset posttransplantation insulin-dependent diabetes was 0.5%. At current follow-up, the use of Campath 1H rather than thymoglobulin for pretreatment seems to have significantly improved the efficacy of our regimen.

Use of CAMPATH-1H as Graft-Versus-Host Disease Prophylaxis in Myeloablative Unrelated Donor Transplants.

Graft-versus-host disease (GvHD) remains a frequent, morbid, potentially fatal, and costly complication of allogeneic stem cell transplantation (alloSCT) particularly when utilizing stem cells from unrelated HLA-matched donors (URD). Optimal GvHD prophylaxis in URD transplants remains uncertain. With standard post-alloSCT tacrolimus (FK) and minidose-methotrexate (MTX) chemoprophylaxis the incidence of acute GvHD approaches 50–75% in adult recipients of URD stem cells without T-cell depletion (TCD). Severe acute GvHD (grade III-IV) occurs in about 30% of these patients (pts), with an associated risk of mortality >80%. TCD is commonly employed but is associated with increased risk of disease recurrence and graft rejection. In this abstract we report observations from 20 pts who received CAMPATH-1H pre-alloSCT for GvHD prevention with a myeloablative (18/20 TBI-containing) conditioning regimen and URD stem cells. There were 2 cohorts of patients - Pts 1-10 (A) were treated with 5 days of CAMPATH; Pts 11–20 (B) were treated with 3 days of CAMPATH (20mg/d). All pts received FK and MTX GvHD prophylaxis. All pts had hematologic malignancies; 12 of 20 with >CR2 or relapsed/refractory disease at the time of conditioning. All pts received standard infection prophylaxis through at least day 100; CMV PCR was obtained weekly with any positive result prompting empiric therapy. Pts were evaluated for ANC recovery by day 30, sustained donor engraftment, presence of GvHD before and after day 100, and disease-free survival at day 100. ANC >500 for 3 consecutive days occurred in 18/20 patients by day 30 - one pt died before day 30 and one had primary engraftment failure (PEF). 17/20 pts were alive and evaluable at day 100 - 13/17 had full donor chimerism at day 100: 1 pt transplanted with refractory disease had disease recurrence, 2 pts had received suboptimal <2X106/kg CD34 cells, and 1 was the pt with PEF. No pts developed severe acute GvHD. There have been no deaths attributable to acute or chronic GvHD. At this report 7/20 pts are alive (6/7 from cohort B) at a median follow-up of 20 mos - 5 pts alive >1 yr and 1 pt > 2 yrs. Of note, the frequency and severity of viral infections, and host failure to antiviral therapy prompted a dose reduction of CAMPATH that defines cohorts A & B. 7/20 pts had reactivation of CMV - CMV as the cause of death in 4 and multiple viral infections (including CMV) in another. 9/20 patients had symptomatic BK viruria and 7/20 developed HSV infections requiring IV therapy. 4 of cohort A and 2 of cohort B had multiple co-existent viral infections. 7/10 in cohort A and 3/10 in cohort B succumbed to infections. Severe acute or chronic GvHD does not occur with this prophylaxis at either dose in our 2 cohorts. Use of CAMPATH-1H for GvHD prophylaxis is highly effective in preventing GvHD. The frequency and severity of viral infections require aggressive monitoring and empiric therapy. Consideration should be given to the lower dose use as GvHD prophylaxis.PtCMV PCR(+)BKV in urineHSV (+) siteother viral infexnscause of death1graft fx/sepsis2adenovirus3XXechovirusviral infexns4XXXrelapse5Xrelapse6XXXCMV7Xfungal/mucor8Xrelapse9XXXsepsis/ARDS10PE11XXCMV/sepsis12Xrhino/paraflu1314X15X1617XXXtoxoplasmosis18XCMV/ICH19graft fx/sepsis20

Severe early acute humoral rejection resulting in allograft loss in a renal transplant recipient with Campath-1H induction therapy

One of the major goals in organ transplantation is to reduce immunosuppression by reprogramming the immune system so as to encourage tolerance. This process has been shown in animal models to be facilitated by lymphocyte depletion at the time of the transplant, and the degree of lymphocyte depletion substantially impacts the efficacy of such treatment [1,2]. Campath-1H is a humanized CD52-specific depleting complement-fixing cytotoxic IgG1 monoclonal antibody. CD52 antigen is located on the surface of T and B lymphocytes, natural killer (NK) cells, and less densely on monocytes [3]. Campath-1H profoundly depletes T lymphocytes from the peripheral blood for several months as well as having less marked effects on B lymphocytes, NK cells, monocytes and CD34-positive immature stem cells [3]. Campath-1H was initially used to treat acute renal allograft rejection [4] and later used successfully together with low dose cyclosporin (CsA) as induction therapy in renal transplantation [3]. Several other centres have also now reported their experiences with the use of Campath-1H together with various combinations of immunosuppressive agents in renal transplantation [5–9]. In some of the series there was a higher than usual incidence of acute humoral or antibody-mediated rejection (AHR) among the acute rejection (AR) episodes [6,8]. We report a case of early severe AHR resulting in renal allograft loss in a patient treated with Campath-1H induction therapy. Case

CD52 Is Expressed on Human Mast Cells and Is a Therapeutic Target for the Anti-CD52 Monoclonal Antibody Campath-1H in Waldenstrom's Macroglobulinemia and Mast Cell Disorders.

Campath-1H is a monoclonal antibody used in the treatment of CD52 expressing B-cell malignancies. Recently, we and others have observed that Campath-1H shows unusually high activity in patients with relapsed/refractory Waldenstrom's macroglobulinemia (WM), a B-cell malignancy with excess bone marrow mast cells (BMMC). Importantly, as we have recently shown, BMMC appear to provide direct support for WM tumor cell growth (Tournilhac et al, JCO 2004 22:571S), and are therefore a potential therapeutic target in WM. We therefore examined BMMC from patients with WM and other mast cell disorders for cell surface expression of CD52, and targeting by Campath-1H in preclinical studies. Multicolor flow cytometric analysis demonstrated CD52 expression on BMMC (FceRI+, CD117+) from 13/15 WM; 2/2 systemic mastocytosis (SM) patients; 2/4 healthy donors; as well as on the LAD and HMC MC lines. Moreover, RT-PCR analysis confirmed CD52 expression in sorted BMMC from 6/7 WM, along with 6/6 healthy donors. Importantly, Campath-1H induced high levels of antibody dependent cell mediated cytotoxicity (ADCC) actvity against LAD mast cells using activated NK effector cells. No direct cytotoxicity or antiproliferative activity by Campath-1H on LAD cells was observed. These studies demonstrate that CD52 is widely expressed on human MC and provide support for the use of Campath-1H in the treatment of WM and other systemic mast cell disorders.

Functional Helper T Cell Response to CMV Predicts Probability of CMV Vireamia Following Allogeneic Stem Cell Transplantation.

CMV reactivation after allogeneic stem cell transplantation remains problematic and is increasing in frequency with the use of reduced intensity conditioning regimens. A functional immune response is critical to control viral replication and hence disease. In this study, 19 allogeneic stem cell transplant recipients at risk for CMV reactivation were followed for a median of 185 days. Six patients received myeloablative conditioning and thirteen patients received reduced intensity conditioning therapy. Functional CMV specific immune responses were monitored by flow cytometry, measuring CD4 and CD8 specific intracellular interferon gamma production in response to stimulation with peptide pools of 15 mer overlapping peptides spanning the entire length of the immunodominant PP65 and IE1 proteins. In addition the CD4 response to whole CMV antigen, which is capable of presenting multiple viral antigens, was also determined. 53% of patients experienced a CMV reactivation with a median time to 1st reactivation of 53.5 days (range 20–134). In comparison with the CMV reactivation group, the mean day 50 CD4 whole CMV antigen response was approximately 2 logs lower in the reactivated group (0.065X106/L) than the non-reactivated group (3.620X106/L) suggesting that CMV specific CD4 function is an important predictor of patients at risk of CMV reactivation. The incidence of CMV reactivation in patients receiving Campath-1H was 71% (10 out of 14 patients) while none of the 5 patients in the non-Campath-1H group reactivated. The use of Campath-1H as part of the conditioning regimen was associated with a significantly lower day 50 CD4 whole CMV antigen response (0.06X106/L) vs (4.53X106/L) for the non-Campath-1H group, suggesting a mechanism for the observed increased frequency of CMV reactivation with the use of this agent. The relative contribution of the CD4 and CD8 responses to IE1 and PP65 has also been determined in this patient group. In summary, the absolute functional helper T cell response to CMV was predictive of viral reactivation following transplant.

Successful use of Campath-1H in the treatment of steroid refractory liver GvHD

Successful use of Campath-1H in the treatment of steroid refractory liver GvHD

High incidence of cytomegalovirus infection after nonmyeloablative stem cell transplantation: potential role of Campath-1H in delaying immune reconstitution

Nonmyeloablative conditioning is increasingly used for transplantation in a wide range of diseases, but little is known about its impact on the incidence of infections and immune reconstitution. We examined the pattern and outcome of cytomegalovirus (CMV) infections monitored by polymerase chain reaction-based assays and treated preemptively in 101 patients following nonmyeloablative conditioning containing in vivo Campath-1H. Fifty-one patients (50%) had a CMV infection at a median of 27 days after transplantation with a probability of 84.8% in patients at risk of CMV infection. The probability of recurrence of CMV infection before and after 100 days was 53.6% and 46.6%, respectively, and was more common in unrelated donor transplant recipients. All 3 patients who developed CMV disease died of this complication. The 2 patients with late CMV disease had grade III to IV graft-versus-host-disease (GVHD), which occurred de novo in only 4% of patients and in another 10% following donor lymphocyte infusions. The median time to CD4(+) T-cell count more than 200/microL was 9 months in the 48 patients studied. The probabilities of overall survival and nonrelapse mortality at 18 months were 65% and 27.8%, respectively, with no significant difference in survival between CMV-infected and -uninfected patients. The use of Campath-1H appeared to be associated with a low incidence of GVHD but a high incidence of CMV infections and prolonged immune paresis.