ABSTRACT
BRAF V600E is the key oncogenic driver mutation in hairy cell leukemia (HCL). We report the efficacy and safety of dabrafenib plus trametinib in patients with relapsed/refractory BRAF V600E mutation-positive HCL. This open-label, phase 2 study enrolled patients with BRAF V600E mutation-positive HCL refractory to first-line treatment with a purine analog or relapsed after ≥2 prior lines of treatment. Patients received dabrafenib 150 mg twice daily plus trametinib 2 mg once daily until disease progression, unacceptable toxicity, or death. The primary endpoint was investigator-assessed objective response rate (ORR) per criteria adapted from National Comprehensive Cancer Network-Consensus Resolution guidelines. Secondary endpoints included duration of response (DOR), progression-free survival (PFS), overall survival (OS), and safety. Fifty-five patients with BRAF V600E mutation-positive HCL were enrolled. The investigator-assessed ORR was 89.0% (95% confidence interval, 77.8%-95.9%); 65.5% of patients had a complete response (without minimal residual disease [MRD]: 9.1% [negative immunohistochemistry of bone marrow {BM} biopsy], 12.7% [negative BM aspirate flow cytometry {FC}], 16.4% [negative immunohistochemistry and/or FC results]; with MRD, 49.1%), and 23.6% had a partial response. The 24-month DOR was 97.7% with 24-month PFS and OS rates of 94.4% and 94.5%, respectively. The most common treatment-related adverse events were pyrexia (58.2%), chills (47.3%), and hyperglycemia (40.0%). Dabrafenib plus trametinib demonstrated durable responses with a manageable safety profile consistent with previous observations in other indications and should be considered as a rituximab-free therapeutic option for patients with relapsed/refractory BRAF V600E mutation-positive HCL. This trial is registered at www.clinicaltrials.gov as #NCT02034110.
Subject(s)
Leukemia, Hairy Cell , Proto-Oncogene Proteins B-raf , Humans , Proto-Oncogene Proteins B-raf/genetics , Leukemia, Hairy Cell/drug therapy , Leukemia, Hairy Cell/genetics , Pyridones/adverse effects , Pyrimidinones/adverse effects , Oximes/adverse effects , Mutation , Antineoplastic Combined Chemotherapy Protocols/adverse effectsABSTRACT
Although monoclonal antibodies to the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) are known, B-cell receptor repertoire and its change in patients during coronavirus disease-2019 (COVID-19) progression is underreported. We aimed to study this molecularly. We used immunoglobulin heavy chain (IGH) variable region (IGHV) spectratyping and next-generation sequencing of peripheral blood B-cell genomic DNA collected at multiple time points during disease evolution to study B-cell response to SARS-CoV-2 infection in 14 individuals with acute COVID-19. We found a broad distribution of responding B-cell clones. The IGH gene usage was not significantly skewed but frequencies of individual IGH genes changed repeatedly. We found predominant usage of unmutated and low mutation-loaded IGHV rearrangements characterizing naïve and extrafollicular B cells among the majority of expanded peripheral B-cell clonal lineages at most tested time points in most patients. IGH rearrangement usage showed no apparent relation to anti-SARS-CoV-2 antibody titers. Some patients demonstrated mono/oligoclonal populations carrying highly mutated IGHV rearrangements indicating antigen experience at some of the time points tested, including even before anti-SARS-CoV-2 antibodies were detected. We present evidence demonstrating that the B-cell response to SARS-CoV-2 is individual and includes different lineages of B cells at various time points during COVID-19 progression.
Subject(s)
COVID-19 , Genes, Immunoglobulin , Humans , COVID-19/genetics , SARS-CoV-2/genetics , Receptors, Antigen, B-Cell/genetics , B-Lymphocytes , Antibodies, ViralABSTRACT
Hairy cell leukemia (HCL) is a rare B-cell malignancy, and there is a need for novel treatments for patients who do not benefit from purine analogs. Ibrutinib, an oral agent targeting Bruton tyrosine kinase in the B-cell receptor signaling pathway, is highly effective in several malignancies. Its activity in HCL was unknown, so we conducted a multisite phase 2 study of oral ibrutinib in patients with either relapsed classic or variant hairy cell leukemia. The primary outcome measure was the overall response rate (ORR) at 32 weeks, and we also assessed response at 48 weeks and best response during treatment. Key secondary objectives were characterization of toxicity and determination of progression-free survival (PFS) and overall survival (OS). Thirty-seven patients were enrolled at 2 different doses (24 at 420 mg, 13 at 840 mg). The median duration of follow-up was 3.5 years (range, 0-5.9 years). The ORR at 32 weeks was 24%, which increased to 36% at 48 weeks. The best ORR was 54%. The estimated 36-month PFS was 73% and OS was 85%. The most frequent adverse events were diarrhea (59%), fatigue (54%), myalgia (54%), and nausea (51%). Hematologic adverse events were common: anemia (43%), thrombocytopenia (41%), and neutropenia (35%). Ibrutinib can be safely administered to patients with HCL with objective responses and results in prolonged disease control. Although the initial primary outcome objective of the study was not met, the observation of objective responses in heavily pretreated patients coupled with a favorable PFS suggests that ibrutinib may be beneficial in these patients. This trial was registered at www.clinicaltrials.gov as #NCT01841723.
Subject(s)
Adenine/analogs & derivatives , Leukemia, Hairy Cell/drug therapy , Leukemia, Hairy Cell/mortality , Piperidines/administration & dosage , Adenine/administration & dosage , Adenine/adverse effects , Administration, Oral , Adult , Aged , Disease-Free Survival , Female , Follow-Up Studies , Humans , Male , Middle Aged , Piperidines/adverse effects , Survival RateABSTRACT
Hairy cell leukemia (HCL) is an indolent B-cell malignancy characterized by high initial sensitivity to purine analog chemotherapy, minimal residual disease (MRD) frequently accompanying complete remission (CR), and relapses requiring additional treatment. Repeat chemotherapy shows decreasing efficacy and increasing toxicity with each course. Newer therapies targeting BRAF/MEK or Bruton's tyrosine kinase are effective but generally leave MRD. Rituximab has modest activity as a single agent and can achieve MRD-negative CR in combination with purine analogs, but there is significant toxicity from the chemotherapy. Moxetumomab pasudotox-tdfk (Moxe) is a biologic containing an antibody fragment (Fv) binding to CD22, attached to a portion of Pseudomonas exotoxin A. Binding to CD22 enables the toxin to enter and kill cells. Moxe is administered by 30-minute infusions on days 1, 3, and 5 of up to six cycles spaced 4 weeks apart. In phase I testing, 64% of 33 patients at the highest dose level achieved CR, most without MRD. Lack of MRD correlated with prolonged CR duration; of 11 MRD-negative CRs, 10 were still in CR after a median of 42 months of observation. In pivotal testing, 75% of 80 patients had a hematologic response, 41% with CR; 82% (27/33) of CRs were MRD-negative, and only 4 of the 27 MRD-negative patients relapsed during the follow-up period. Hemolytic uremic syndrome and capillary leak syndrome were each observed in 9% of patients, all reversible. In September 2018, the U.S. Food and Drug Administration approved Moxe for the treatment of relapsed/refractory HCL after ≥2 prior therapies. Moxe is undergoing further development in combination with rituximab. IMPLICATIONS FOR PRACTICE: Hairy cell leukemia (HCL) has effective treatments including purine analogs with and without rituximab, and oral inhibitors of BRAF, MEK and Bruton's tyrosine kinase (BTK). Despite these therapies, relapse occurs, and moxetumomab pasudotox has an important role in relapsed and refractory HCL because of its ability to achieve high rates of complete remissions (CRs) without chemotherapy; most of these CRs are without minimal residual disease (MRD). CR duration is enhanced in patients who achieve eradication of MRD. To improve the efficacy of this recombinant immunotoxin, a phase I trial is underway in combination with rituximab to reduce tumor burden and decrease immunogenicity.
Subject(s)
Antineoplastic Agents/therapeutic use , Bacterial Toxins/therapeutic use , Exotoxins/therapeutic use , Leukemia, Hairy Cell/drug therapyABSTRACT
Anti-CD22 moxetumomab pasudotox achieved 46% complete remissions (CRs) in previously reported phase 1 testing in relapsed/refractory hairy cell leukemia (HCL; n = 28). The importance of minimal residual disease (MRD) after CR in HCL is unknown. A 21-patient extension cohort received 50 µg/kg every other day for 3 doses in 4-week cycles. These patients plus 12 previously reported at this upper dose level received 143 cycles without dose-limiting toxicity. The combined 33-patient cohort achieved 64% CR and 88% overall response rates, with median CR duration of 42.4 months. Of 32 50-µg/kg patients evaluable for MRD by bone marrow aspirate flow cytometry (most stringent assessment), median CR duration was 13.5 (4.9-42.4) months in 9 MRD-positive CRs vs 42.1 (24.0-69.2) months in 11 MRD-negative CRs (P < .001). Among MRD-negative CRs, 10 patients had ongoing CR, 9 without MRD, at end of study. To our knowledge, moxetumomab pasudotox is the only nonchemotherapy regimen that can eliminate MRD in a significant percentage of HCL patients, to enhance CR duration. Repeated dosing, despite early neutralizing antibodies, increased active drug levels without detectable toxicity from immunogenicity. The activity and safety profiles of moxetumomab pasudotox support ongoing phase 3 testing in HCL. This trial was registered at www.clinicaltrials.gov as #NCT00586924.
Subject(s)
Antineoplastic Agents/therapeutic use , Bacterial Toxins/therapeutic use , Exotoxins/therapeutic use , Leukemia, Hairy Cell/drug therapy , Leukemia, Hairy Cell/pathology , Neoplasm, Residual/drug therapy , Adult , Aged , Antineoplastic Agents/pharmacology , Bacterial Toxins/pharmacology , Clinical Trials, Phase I as Topic , Drug Monitoring , Exotoxins/pharmacology , Female , Follow-Up Studies , Humans , Male , Middle Aged , Treatment OutcomeABSTRACT
AIMS: To characterize the pharmacokinetics (PK) of moxetumomab pasudotox, an anti-CD22 recombinant immunotoxin, in adults with relapsed or refractory hairy cell leukaemia, we examined data from a phase 1 study (Study 1001; n = 49) and from the pivotal clinical study (Study 1053; n = 74). METHODS: Data from both studies were pooled (n = 123) to develop a population PK model. Covariates included demographics, disease state, liver and kidney function, prior treatment, and antidrug antibodies (ADAs). Exposure-response and exposure-safety were analysed separately by study. A 1-compartment model with linear elimination from the central compartment and 2 clearance (CL) rates was developed. RESULTS: Moxetumomab pasudotox was cleared more rapidly after cycle 1, day 1 (CL1 = 24.7 L/h) than subsequently (CL2 = 3.76 L/h), with high interindividual variability (116 and 109%, respectively). In Study 1053, patients with ADA titres >10 240 showed ~4-fold increase in CL. Higher exposures (≥median) were related to higher response rates, capillary leak syndrome and increased creatinine (Study 1053 only), or grade ≥3 adverse events (Study 1001 only). Clinical benefits were still observed in patients with lower exposure or high ADA titres. CONCLUSION: Despite a high incidence of immunogenicity with increased clearance, moxetumomab pasudotox demonstrated efficacy in hairy cell leukaemia.
Subject(s)
Bacterial Toxins , Leukemia, Hairy Cell , Adult , Antibodies , Exotoxins , HumansABSTRACT
Novel therapies are needed to overcome chemotherapy resistance for children with relapsed/refractory acute lymphoblastic leukemia (ALL). Moxetumomab pasudotox is a recombinant anti-CD22 immunotoxin. A multicenter phase 1 study was conducted to determine the maximum-tolerated cumulative dose (MTCD) and evaluate safety, activity, pharmacokinetics, and immunogenicity of moxetumomab pasudotox in children, adolescents, and young adults with ALL (N = 55). Moxetumomab pasudotox was administered as a 30-minute IV infusion at doses of 5 to 50 µg/kg every other day for 6 (cohorts A and B) or 10 (cohort C) doses in 21-day cycles. Cohorts B and C received dexamethasone prophylaxis against capillary leak syndrome (CLS). The most common treatment-related adverse events were reversible weight gain, hepatic transaminase elevation, and hypoalbuminemia. Dose-limiting CLS occurred in 2 of 4 patients receiving 30 µg/kg of moxetumomab pasudotox every other day for 6 doses. Incorporation of dexamethasone prevented further dose-limiting CLS. Six of 14 patients receiving 50 µg/kg of moxetumomab pasudotox for 10 doses developed hemolytic uremic syndrome (HUS), thrombotic microangiopathy (TMA), or HUS-like events, exceeding the MTCD. Treatment expansion at 40 µg/kg for 10 doses (n = 11) exceeded the MTCD because of 2 HUS/TMA/HUS-like events. Dose level 6B (ie, 50 µg/kg × 6 doses) was the MTCD, selected as the recommended phase 2 dose. Among 47 evaluable patients, an objective response rate of 32% was observed, including 11 (23%) composite complete responses, 5 of which were minimal residual disease negative by flow cytometry. Moxetumomab pasudotox showed a manageable safety profile and evidence of activity in relapsed or refractory childhood ALL. This trial was registered at www.clinicaltrials.gov as #NCT00659425.
Subject(s)
Bacterial Toxins/therapeutic use , Exotoxins/therapeutic use , Immunotoxins/therapeutic use , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Sialic Acid Binding Ig-like Lectin 2/immunology , Adolescent , Adult , Bacterial Toxins/adverse effects , Bacterial Toxins/immunology , Bacterial Toxins/pharmacokinetics , Capillary Leak Syndrome/prevention & control , Child , Child, Preschool , Dexamethasone/therapeutic use , Exotoxins/adverse effects , Exotoxins/immunology , Exotoxins/pharmacokinetics , Female , Glucocorticoids/therapeutic use , Hemolytic-Uremic Syndrome/chemically induced , Humans , Hypoalbuminemia/chemically induced , Immunotoxins/adverse effects , Immunotoxins/immunology , Immunotoxins/pharmacokinetics , Infant , Male , Maximum Tolerated Dose , Neoplasm Recurrence, Local/drug therapy , Neoplasm Recurrence, Local/immunology , Neoplasm Recurrence, Local/pathology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/immunology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Thrombotic Microangiopathies/chemically induced , Weight Gain/drug effects , Young AdultABSTRACT
Hairy cell leukemia is an uncommon hematologic malignancy characterized by pancytopenia and marked susceptibility to infection. Tremendous progress in the management of patients with this disease has resulted in high response rates and improved survival, yet relapse and an appropriate approach to re-treatment present continuing areas for research. The disease and its effective treatment are associated with immunosuppression. Because more patients are being treated with alternative programs, comparison of results will require general agreement on definitions of response, relapse, and methods of determining minimal residual disease. The development of internationally accepted, reproducible criteria is of paramount importance in evaluating and comparing clinical trials to provide optimal care. Despite the success achieved in managing these patients, continued participation in available clinical trials in the first-line and particularly in the relapse setting is highly recommended. The Hairy Cell Leukemia Foundation convened an international conference to provide common definitions and structure to guide current management. There is substantial opportunity for continued research in this disease. In addition to the importance of optimizing the prevention and management of the serious risk of infection, organized evaluations of minimal residual disease and treatment at relapse offer ample opportunities for clinical research. Finally, a scholarly evaluation of quality of life in the increasing number of survivors of this now manageable chronic illness merits further study. The development of consensus guidelines for this disease offers a framework for continued enhancement of the outcome for patients.
Subject(s)
Antineoplastic Agents/therapeutic use , Cladribine/therapeutic use , Leukemia, Hairy Cell/diagnosis , Leukemia, Hairy Cell/drug therapy , Pentostatin/therapeutic use , Disease Management , Humans , Neoplasm Recurrence, Local/diagnosis , Neoplasm Recurrence, Local/drug therapy , Neoplasm, Residual/diagnosis , Neoplasm, Residual/drug therapy , Treatment OutcomeABSTRACT
Hairy cell leukemia (HCL) is a chronic B-cell malignancy with multiple treatment options, including several that are investigational. Patients present with pancytopenia and splenomegaly, owing to the infiltration of leukemic cells expressing CD22, CD25, CD20, CD103, tartrate-resistant acid phosphatase (TRAP), annexin A1 (ANXA1), and the BRAF V600E mutation. A variant lacking CD25, ANXA1, TRAP, and the BRAF V600E mutation, called HCLv, is more aggressive and is classified as a separate disease. A molecularly defined variant expressing unmutated immunoglobulin heavy variable 4-34 (IGHV4-34) is also aggressive, lacks the BRAF V600E mutation, and has a phenotype of HCL or HCLv. The standard first-line treatment, which has remained unchanged for the past 25 to 30 years, is single-agent therapy with a purine analogue, either cladribine or pentostatin. This approach produces a high rate of complete remission. Residual traces of HCL cells, referred to as minimal residual disease, are present in most patients and cause frequent relapse. Repeated treatment with a purine analogue can restore remission, but at decreasing rates and with increasing cumulative toxicity. Rituximab has limited activity as a single agent but achieves high complete remission rates without minimal residual disease when combined with purine analogues, albeit with chemotherapy-associated toxicity. Investigational nonchemotherapy options include moxetumomab pasudotox, which targets CD22; vemurafenib or dabrafenib, each of which targets the BRAF V600E protein; trametinib, which targets mitogen-activated protein kinase enzyme (MEK); and ibrutinib, which targets Bruton tyrosine kinase (BTK).
Subject(s)
Leukemia, Hairy Cell/diagnosis , Leukemia, Hairy Cell/therapy , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Biomarkers , Biopsy , Clinical Decision-Making , Humans , Immunohistochemistry , Immunophenotyping , Leukemia, Hairy Cell/genetics , Molecular Targeted Therapy , Neoplasm, Residual , Standard of Care , Treatment OutcomeABSTRACT
In this issue of Blood, Pettirossi et al, including Drs Tiacci and Falini, who led the effort in 2011 defining the BRAF-V600E driving mutation in hairy cell leukemia (HCL),provide extensive laboratory studies showing that inhibitors of BRAF-V600E and/or mitogen-activated protein kinase kinase (MEK) reach their targets and cause HCL cell death
Subject(s)
Antineoplastic Agents , Imidazoles , Indoles , Leukemia, Hairy Cell/drug therapy , Leukemia, Hairy Cell/genetics , Oximes , Pyridones , Pyrimidinones , Sulfonamides , Transcriptome/drug effects , Humans , VemurafenibABSTRACT
Nonhuman proteins have valuable therapeutic properties, but their efficacy is limited by neutralizing antibodies. Recombinant immunotoxins (RITs) are potent anticancer agents that have produced many complete remissions in leukemia, but immunogenicity limits the number of doses that can be given to patients with normal immune systems. Using human cells, we identified eight helper T-cell epitopes in PE38, a portion of the bacterial protein Pseudomonas exotoxin A which consists of the toxin moiety of the RIT, and used this information to make LMB-T18 in which three epitopes were deleted and five others diminished by point mutations in key residues. LMB-T18 has high cytotoxic and antitumor activity and is very resistant to thermal denaturation. The new immunotoxin has a 93% decrease in T-cell epitopes and should have improved efficacy in patients because more treatment cycles can be given. Furthermore, the deimmunized toxin can be used to make RITs targeting other antigens, and the approach we describe can be used to deimmunize other therapeutically useful nonhuman proteins.
Subject(s)
Epitopes, T-Lymphocyte/immunology , Immunotoxins/immunology , Neoplasms/immunology , Recombinant Fusion Proteins/immunology , ADP Ribose Transferases/genetics , ADP Ribose Transferases/immunology , Amino Acids/genetics , Amino Acids/immunology , Animals , Antibody Formation/immunology , Bacterial Toxins/genetics , Bacterial Toxins/immunology , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/immunology , Electrophoresis, Polyacrylamide Gel , Epitope Mapping , Exotoxins/genetics , Exotoxins/immunology , Female , Humans , Immunotherapy/methods , Immunotoxins/genetics , Immunotoxins/therapeutic use , Lymphocyte Activation/drug effects , Lymphocyte Activation/immunology , Mice , Mice, SCID , Models, Molecular , Neoplasms/pathology , Neoplasms/therapy , Peptides/genetics , Peptides/immunology , Point Mutation , Protein Structure, Tertiary , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/therapeutic use , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Virulence Factors/genetics , Virulence Factors/immunology , Xenograft Model Antitumor Assays , Pseudomonas aeruginosa Exotoxin AABSTRACT
Unconjugated monoclonal antibodies that target hematopoietic differentiation antigens have been developed to treat hematologic malignancies. Although some of these have activity against chronic lymphocytic leukemia and hairy cell leukemia, in general, monoclonal antibodies have limited efficacy as single agents in the treatment of leukemia. To increase their potency, the binding domains of monoclonal antibodies can be attached to protein toxins. Such compounds, termed immunotoxins, are delivered to the interior of leukemia cells based on antibody specificity for cell surface target antigens. Recombinant immunotoxins have been shown to be highly cytotoxic to leukemic blasts in vitro, in xenograft model systems, and in early-phase clinical trials in humans. These agents will likely play an increasing role in the treatment of leukemia.
Subject(s)
Immunotoxins/therapeutic use , Leukemia/therapy , Animals , Antigens, CD19/metabolism , Clinical Trials as Topic , Humans , Immunotherapy/methods , Immunotoxins/chemistry , Leukemia/immunology , Models, Molecular , Molecular Targeted Therapy , Sialic Acid Binding Ig-like Lectin 2/antagonists & inhibitorsABSTRACT
Recombinant immunotoxins (RITs) are fusion proteins that join antibodies to protein toxins for targeted cell killing. RITs armed with Pseudomonas exotoxin A (PE) are undergoing clinical trials for the treatment of cancer. The current design of PE-based RITs joins an antibody fragment to the catalytic domain of PE using a polypeptide linker that is cleaved by the protease furin. Intracellular cleavage of native PE by furin is required for cytotoxicity, yet the PE cleavage site has been shown to be a poor furin substrate. Here we describe the rational design of more efficiently cleaved furin linkers in PE-based RITs, and experiments evaluating their effects on cleavage and cytotoxicity. We found that changes to the furin site could greatly influence both cleavage and cytotoxicity, but the two parameters were not directly correlated. Furthermore, the effects of alterations to the furin linker were not universal. Identical mutations in the anti-CD22 RIT HA22-LR often displayed different cytotoxicity from mutations in the anti-mesothelin RIT SS1-LR/GGS, underscoring the prominent role of the target site in their intoxication pathways. Combining several beneficial mutations in HA22-LR resulted in a variant (HA22-LR/FUR) with a remarkably enhanced cleavage rate and improved cytotoxicity against five B cell lines and similar or enhanced cytotoxicity in five out of six hairy cell leukemia patient samples. This result informs the design of protease-sensitive linkers and suggests that HA22-LR/FUR may be a candidate for further preclinical development.
Subject(s)
ADP Ribose Transferases/chemistry , ADP Ribose Transferases/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Bacterial Toxins/chemistry , Bacterial Toxins/pharmacology , Exotoxins/chemistry , Exotoxins/pharmacology , Furin/metabolism , Immunotoxins/chemistry , Immunotoxins/pharmacology , Virulence Factors/chemistry , Virulence Factors/pharmacology , ADP Ribose Transferases/metabolism , Amino Acid Sequence , Antineoplastic Agents/metabolism , Bacterial Toxins/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Exotoxins/metabolism , Humans , Immunotoxins/metabolism , Leukemia/drug therapy , Models, Molecular , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/metabolism , Recombinant Fusion Proteins/pharmacology , Virulence Factors/metabolism , Pseudomonas aeruginosa Exotoxin AABSTRACT
BACKGROUND: CD22 is a B-lineage differentiation antigen that has emerged as a leading therapeutic target in acute lymphoblastic leukemia (ALL). PROCEDURE: Properties of CD22 expression relevant to therapeutic targeting were characterized in primary samples obtained from children and young adults with relapsed and chemotherapy refractory B-precursor (pre-B) ALL. RESULTS: CD22 expression was demonstrated in all subjects (n = 163) with detection on at least 90% of blasts in 155 cases. Median antigen site density of surface CD22 was 3,470 sites/cell (range 349-19,653, n = 160). Blasts from patients with known 11q23 (MLL) rearrangement had lower site density (median 1,590 sites/cell, range 349-3,624, n = 20 versus 3,853 sites/cell, range 451-19,653, n = 140; P = <0.0001) and 6 of 21 cases had sub-populations of blasts lacking CD22 expression (22%-82% CD22 +). CD22 expression was maintained in serial studies of 73 subjects, including those treated with anti-CD22 targeted therapy. The levels of soluble CD22 in blood and marrow by ELISA were low and not expected to influence the pharmacokinetics of anti-CD22 directed agents. CONCLUSIONS: These characteristics make CD22 an excellent potential therapeutic target in patients with relapsed and chemotherapy-refractory ALL, although cases with MLL rearrangement require close study to exclude the presence of a CD22-negative blast population.
Subject(s)
Precursor Cell Lymphoblastic Leukemia-Lymphoma/immunology , Sialic Acid Binding Ig-like Lectin 2/analysis , Adolescent , Adult , Child , Child, Preschool , Chromosomes, Human, Pair 11 , Histone-Lysine N-Methyltransferase , Humans , Infant , Myeloid-Lymphoid Leukemia Protein/genetics , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Sialic Acid Binding Ig-like Lectin 2/antagonists & inhibitorsABSTRACT
Rationally designed anticancer agents that target cell-surface antigens or receptors represent a promising approach for treating cancer patients. However, antibodies that bind these targets are often, by themselves, non-cytotoxic. By attaching potent toxins we can dramatically improve the clinical utility of some anti-tumour antibodies. Here we describe the construction and clinical utility of several recombinant immunotoxins; each of which is composed of antibody Fv fragments fused to powerful bacterial toxins. Results from clinical trials indicate that recombinant immunotoxins and similar agents that are designed to combine antibody selectivity with toxin cell-killing potency will be useful additions to cancer therapy.
Subject(s)
Antineoplastic Agents/therapeutic use , Immunotherapy , Immunotoxins/therapeutic use , Neoplasms/therapy , Animals , Bacterial Toxins/immunology , Bacterial Toxins/therapeutic use , Humans , Immunoglobulin Variable Region/immunology , Immunoglobulin Variable Region/therapeutic use , Recombinant Proteins/immunology , Recombinant Proteins/therapeutic useABSTRACT
Recombinant immunotoxins (RITs) are hybrid proteins used to treat cancer. These proteins are composed of an Fv that reacts with cancer cells joined to a portion of Pseudomonas exotoxin A, which kills the cell. Because the toxin is a foreign protein, it can induce neutralizing antibodies and thereby limit the number of doses a patient can receive. We previously identified seven major mouse B-cell epitopes in the toxin, and subsequently silenced them using point mutations that converted large hydrophilic amino acids to alanine, yet retained full antitumor activity. Here we present results in which we identify and silence human B-cell epitopes in the RIT HA22. We obtained B cells from patients with antibodies to RITs, isolated the corresponding variable fragments (Fvs), and constructed a phage-display library containing Fvs that bind to the RITs. We then used alanine scanning mutagenesis to locate the epitopes. We found that human and mouse epitopes frequently overlap but are not identical. Most mutations that remove mouse epitopes did not remove human epitopes. Using the epitope information, we constructed a variant immunotoxin, HA22-LR-LO10, which has low reactivity with human antisera, yet has high cytotoxic and antitumor activity and can be given to mice at high doses without excess toxicity. The toxin portion of this RIT (LR-LO10) can be used with Fvs targeting other cancer antigens and is suitable for clinical development.
Subject(s)
Antineoplastic Agents/immunology , Epitopes, B-Lymphocyte/immunology , Immunotherapy/methods , Immunotoxins/immunology , Models, Molecular , Neoplasms/drug therapy , Recombinant Proteins/immunology , Amino Acid Sequence , Animals , Enzyme-Linked Immunosorbent Assay , Humans , Immunotoxins/chemistry , Immunotoxins/genetics , Immunotoxins/pharmacokinetics , Mice , Molecular Sequence Data , Mutagenesis , Neoplasms/immunology , Peptide Library , Protein Engineering/methods , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/pharmacokinetics , Single-Chain Antibodies/immunology , Statistics, NonparametricABSTRACT
Cladribine, 2-chloro-2'-deoxyadenosine, is a highly efficacious, clinically used nucleoside for the treatment of hairy cell leukemia. It is also being evaluated against other lymphoid malignancies and has been a molecule of interest for well over half a century. In continuation of our interest in the amide bond-activation in purine nucleosides via the use of (benzotriazol-1yl-oxy)tris(dimethylamino)phosphonium hexafluorophosphate, we have evaluated the use of O6-(benzotriazol-1-yl)-2'-deoxyguanosine as a potential precursor to cladribine and its analogues. These compounds, after appropriate deprotection, were assessed for their biological activities, and the data are presented herein. Against hairy cell leukemia (HCL), T-cell lymphoma (TCL) and chronic lymphocytic leukemia (CLL), cladribine was the most active against all. The bromo analogue of cladribine showed comparable activity to the ribose analogue of cladribine against HCL, but was more active against TCL and CLL. The bromo ribose analogue of cladribine showed activity, but was the least active among the C6-NH2-containing compounds. Substitution with alkyl groups at the exocyclic amino group appears detrimental to activity, and only the C6 piperidinyl cladribine analogue demonstrated any activity. Against adenocarcinoma MDA-MB-231 cells, cladribine and its ribose analogue were most active.
Subject(s)
Antineoplastic Agents/chemical synthesis , Cladribine/chemical synthesis , Guanosine/chemical synthesis , Leukocytes, Mononuclear/drug effects , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cladribine/pharmacology , Guanosine/pharmacology , Humans , Inhibitory Concentration 50 , Leukemia, Hairy Cell/pathology , Leukemia, Lymphocytic, Chronic, B-Cell/pathology , Leukocytes, Mononuclear/pathology , Lymphoma, T-Cell/pathology , Organophosphorus Compounds/chemistry , Primary Cell Culture , Structure-Activity RelationshipABSTRACT
BACKGROUND: The primary objective of this study was to determine the safety and maximum tolerated dose (MTD) of the antimesothelin immunotoxin SS1(dsFv)PE38 (SS1P) (a recombinant antimesothelin immunotoxin consisting of a murine antimesothelin variable antibody fragment [Fv] linked to PE38, a truncated portion of Pseudomonas exotoxin A) in combination with pemetrexed and cisplatin in chemotherapy-naive patients with advanced malignant pleural mesothelioma (MPM). Secondary objectives included tumor response, SS1P pharmacokinetics, and serum biomarkers of response. METHODS: Chemotherapy-naive patients with stage III or IV, unresectable, epithelial or biphasic MPM and normal organ functions were eligible. Pemetrexed (500 mg/m(2) on day 1) and cisplatin (75 mg/m(2) on day 1) were administered every 3 weeks for up to 6 cycles with escalating doses of SS1P administered intravenously on days 1, 3, and 5 during cycles 1 and 2. Tumor response was evaluated every 6 weeks. RESULTS: Twenty-four patients received SS1P at 4 dose levels from 25 to 55 mcg/kg. Grade 3 fatigue was dose-limiting in 1 patient at 55 mcg/kg. The MTD of SS1P was established as 45 mcg/kg. Other grade 3 toxicities associated with SS1P included hypoalbuminemia (21%), back pain (13%), and hypotension (8%). Of 20 evaluable patients, 12 (60%) had a partial response, 3 had stable disease, and 5 had progressive disease. Of 13 patients who received the MTD, 10 (77%) had a partial response, 1 had stable disease, and 2 had progressive disease. Objective radiologic responses were associated with significant decreases in serum mesothelin (P=.0030), megakaryocyte potentiating factor (P=.0005), and cancer antigen 125 (P < .0001). CONCLUSIONS: SS1P given with pemetrexed and cisplatin is safe and well tolerated and exhibits significant antitumor activity in patients with unresectable, advanced pleural mesothelioma. Serum mesothelin, megakaryocyte potentiating factor, and cancer antigen 125 levels correlated with objective tumor responses.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Glutamates/administration & dosage , Guanine/analogs & derivatives , Lung Neoplasms/drug therapy , Mesothelioma/drug therapy , Pleural Neoplasms/drug therapy , ADP Ribose Transferases , Aged , Antibodies, Monoclonal/blood , Bacterial Toxins , CA-125 Antigen/blood , Cisplatin/administration & dosage , Exotoxins , Female , GPI-Linked Proteins/administration & dosage , GPI-Linked Proteins/blood , Guanine/administration & dosage , Humans , Lung Neoplasms/blood , Lung Neoplasms/pathology , Male , Maximum Tolerated Dose , Mesothelin , Mesothelioma/blood , Mesothelioma/pathology , Mesothelioma, Malignant , Middle Aged , Neoplasm Staging , Pemetrexed , Pleural Neoplasms/blood , Pleural Neoplasms/pathology , Virulence Factors , Pseudomonas aeruginosa Exotoxin AABSTRACT
B cell receptor (BCR) signalling plays a critical role in the progression of several B-cell malignancies, but its role in hairy cell leukaemia (HCL) is ambiguous. Bruton tyrosine kinase (BTK), a key player in BCR signalling, as well as B cell migration and adhesion, can be targeted with ibrutinib, a selective, irreversible BTK inhibitor. We analysed BTK expression and function in HCL and analysed the effects of ibrutinib on HCL cells. We demonstrated uniform BTK protein expression in HCL cells. Ibrutinib significantly inhibited HCL proliferation and cell cycle progression. Accordingly, ibrutinib also reduced HCL cell survival after BCR triggering with anti-immunoglobulins and abrogated the activation of kinases downstream of the BCR (PI3K and MAPK). Ibrutinib also inhibited BCR-dependent secretion of the chemokines CCL3 and CCL4 by HCL cells. Interestingly, ibrutinib inhibited also CXCL12-induced signalling, a key pathway for bone marrow homing. Collectively, our data support the clinical development of ibrutinib in patients with HCL.