Philip Caron

Genetically engineered deglycosylation of the variable domain increases the affinity of an anti-CD33 monoclonal antibody.

M195 is a murine monoclonal antibody that binds to the CD33 antigen and is being tested for the treatment of myeloid leukemia. Surprisingly, a complementarity determining region (CDR)-grafted, humanized M195 antibody displayed a several-fold higher binding affinity for the CD33 antigen than the original murine antibody. Here we show that the increase in binding affinity resulted from eliminating an N-linked glycosylation site at residue 73 in the heavy chain variable region in the course of humanization. Re-introducing the glycosylation site in the humanized antibody reduces its binding affinity to that of the murine antibody, while removing the glycosylation site from the murine M195 variable domain increases its affinity. The removal of variable region carbohydrates may provide a method for increasing the affinity of certain monoclonal antibodies with diagnostic and therapeutic potential.

Murine and humanized constructs of monoclonal antibody m195 (anti‐cd33) for the therapy of acute myelogenous leukemia

Long‐term survival rates of patients with acute myelogenous leukemia treated with intensive chemotherapy are 15–20%, despite efforts to develop new treatment strategies. Murine M195 (131I‐M195), an anti‐CD33, immunoglobulin (Ig) G2a monoclonal antibody has reactivity restricted to early myeloid cells and myeloid leukemic blasts but not hematopoietic progenitors. Previous trials in patients with relapsed or refractory myeloid leukemia showed that 131I‐M195 rapidly targeted to the bone marrow and internalized into target cells.

Monoclonal antibody therapy of leukemia and lymphoma.

Publisher Summary Through careful clinical trials, a number of obstacles to effective therapy with monoclonal antibodies (mAbs) have been identified. New technologies, however, have continued to provide strategies to overcome many of these obstacles. Four approaches—mAbs can be used as mediators of immune effector function, anti-idiotype (anti-id) mAbs can both target unique surface immunoglobulins expressed on B-cell neoplasms and mimic an antigen triggering tumor immunity, mAbs can be directed against receptors that control tumor growth, and mAbs can serve as vehicles to carry cytotoxic agents—to the therapeutic use of mAbs have been developed. Among the most promising applications of mAb-based therapies are those that take advantage of multiple mechanisms of tumor killing, such as the use of radioconjugates in combination with high-dose chemotherapy and TBI prior to bone marrow transplantation (BMT), the use of mAb-based therapies to eliminate residual disease, and the use of mAbs to purge tumor cells from bone marrow prior to reinfusion after dose-intensive therapy. The immunophenotypic characterization of the various stages and lineages found during hematopoietic differentiation has formed the basis for the selection of therapeutic mAbs. Leukemia- and lymphoma-associated antigens, however, are neither tumor specific, nor are they always stage or lineage specific. Therefore, myelosuppression—that affects the treatment of leukemia greatly—is often associated with mAb therapy of hematologic malignancies.

Brentuximab vedotin and AVD followed by involved-site radiotherapy in early stage, unfavorable risk Hodgkin lymphoma

This multicenter pilot study assessed the safety and efficacy of brentuximab vedotin (BV) and AVD (adriamycin, vinblastine, and dacarbazine) followed by 30 Gy involved site radiation therapy (ISRT). Patients with newly diagnosed, early stage classical Hodgkin lymphoma (HL) with unfavorable-risk features were treated with 4 cycles of BV and AVD. Patients who achieved a negative positron emission tomography (PET) scan (Deauville score of 1-3) received 30 Gy ISRT. Thirty patients received treatment and were assessable for toxicity. Twenty-nine patients completed 4 cycles of BV + AVD, and 25 patients BV + AVD + 30 Gy ISRT. No clinically significant noninfectious pneumonitis was observed. Serious adverse events (≥grade 3) were reported in 4 patients, including febrile neutropenia, peripheral neuropathy, and hypertension. After 2 and 4 cycles of BV + AVD, 90% (26 of 29) and 93% (27 or 29) of patients achieved a negative PET scan, respectively. Two patients with biopsy-proven primary refractory HL were treated off-study. All 25 patients who completed BV + AVD + ISRT achieved a complete response. With a median follow-up of 18.8 months, by intent to treat, the 1-year progression-free survival is 93.3% (95% confidence interval, 84-102). Overall, the treatment was well-tolerated with no evidence of significant pulmonary toxicity. The majority of patients (≥90%) achieved negative interim PET scans after 2 and 4 cycles of BV + AVD. Excluding the 2 primary refractory patients, all patients are disease free, suggesting that this is a highly active treatment program even in patients with substantial disease bulk. This trial was registered at www.clinicaltrials.gov as #NCT01868451.

Anti-CD33 Monoclonal Antibody M195 for the Therapy of Myeloid Leukemia

Leukemia is well suited for monoclonal antibody therapy due to the accessible, differentiation antigens that characterize stages of maturation. In this paper, we describe the use of radio-labeled M195, a murine IgG2a, anti-CD33 monoclonal antibody, that can be used to effectively cytoreduce AML cells in relapsed patients when tumor burden is high; or to eliminate minimal residual disease and lengthen disease-free survival in patients with APL in remission. To decrease the likelihood of immunogenicity, a humanized IgG1 version of M195 was constructed that demonstrated a higher avidity and improved effector function than the parent murine antibody. Preliminary results of the first trial in AML using a humanized antibody showed specific bone marrow targeting without an immunogenic response.

Immunotherapy for acute leukemias.

Successful immunotherapy for cancer and, in particular, monoclonal antibody-based therapy are most likely to succeed first for the hematopoietic neoplasms because of their biology and because of the pharmacology of monoclonal antibodies. Ten years of steady development in the field, including identification of better antigen-antibody systems, genetic engineering of humanized monoclonal antibodies, construction of new toxin and radionuclide conjugates, and better understanding of the interactions of monoclonal antibodies with cytokines, eg, interleukin-2, has led to a series of recent trials showing significant activity of monoclonal antibodies in leukemias. This activity encompasses the ablation of large masses of cells prior to bone marrow transplantation, as well as the elimination of minimal disease in vivo and ex vivo.

The biological therapy of acute and chronic leukemia

With the increasing knowledge of the mechanisms of immune-mediated cytotoxicity, immunotherapeutic strategies are rapidly being incorporated into chemotherapy treatment schemes for acute and chronic leukemias. This includes the use of mAbs, immunotoxins, tumor-specific T cells, and, most recently, vaccines. Much of the new information is derived from bone marrow transplant data, where immune enhancement from IL-2 and donor T-cell infusions are being studied. Trials using humanized mAbs that permit prolonged and repeated dosing will allow better evaluation of the effectiveness of mAb therapy. More sophisticated molecular tests have been developed, allowing the detection of minimal residual disease to a greater degree. It is likely that biological and immunological therapy of leukemia will have its greatest impact here.

Acute myelogenous leukemia following irinotecan-based chemotherapy for adenocarcinoma of the small intestine

Irinotecan, a topoisomerase I inhibitor, is a camptothecan analogue with activity against gastrointestinal tumors [1]. Irinotecan is approved by the United States Food and Drug Administration for first line therapy of metastatic colorectal carcinoma in combination with fluorouracil and leucovorin, and as second line therapy of patients whose disease has progressed or recurred after fluorouracil-based therapy. Additionally, irinotecin is frequently used for a number of other malignancies including nonsmall-cell lung cancer, small-cell lung cancer, brain tumors, and other gastrointestinal malignancies [2 – 4]. The primary toxicities are diarrhea and myelosuppression [1]. No leukemogenic potential has been described for this drug. Currently we present a patient who developed acute leukemia after treatment of small bowel cancer with irinotecan. The patient was 43 years old when he presented to another institution in December 2000 with abdominal pain. The clinical impression was diverticulitis, which was managed medically. However, after several clinical exacerbations in 2001, in January 2002 the patient developed intestinal obstruction. At exploration, the patient was found to have generalized peritonitis and required emergency resection and ileostomy. He received postoperative antibiotics and improved. In March 2002, when the patient returned to the operating room for reversal of ileostomy, he was found to have an invasive moderate to poorly differentiated adenocarcinoma arising in the distal ileum with extension into the cecum. The tumor perforated the bowel with abscess formation, and biopsy of the abscess cavity revealed poorly differentiated adenocarcinoma. Limited resection of the terminal ileum and a right hemicolectomy were performed. Postoperatively, the patient received FOLFIRI (5-fluorouracil 2400 mg/m as a 48-h infusion, leucovorin 200 mg/m, and irinotecan 180 mg/m) every 2 – 3 weeks for eight courses with a dramatic response, and was then clinically without evidence of disease. In November 2003, the patient developed recurrent obstruction requiring surgery. He underwent further resection of the small bowel and all visible tumors were removed. Pathology revealed poorly differentiated adenocarcinoma pathologically identical to the primary lesion. His postoperative course was complicated by multiple abdominal infections. In March 2005, the patient was noted to have worsening fatigue due to anemia, and wound dehiscence, bleeding, and infection. After a bone marrow biopsy was performed in July 2005 the patient was referred to us. The physical exam was significant for an open abdominal wound. There was no lymphadenopathy or hepatosplenomegaly. Laboratories showed a white blood count of 10,100/mm with 24% neutrophils, 29% lymphocytes, and 47% monocytes. The hemoglobin was 9.8 g/dl and the platelet count was 203,000/mm. The bone marrow was markedly hypercellular with 20% blasts, 24% promonocytes, 26% monocytes, 1% promyelocytes, 2% metamyelocytes, 5% neutrophils, 6% lymphocytes, 2% plasma cells, and 14% normoblasts, consistent with acute monocytic leukemia (AML-M5). Flow cytometric analysis demonstrated