Kenneth A. Foon

Flow cytometric immunophenotyping for hematologic neoplasms

Flow cytometric immunophenotyping remains an indispensable tool for the diagnosis, classification, staging, and monitoring of hematologic neoplasms. The last 10 years have seen advances in flow cytometry instrumentation and availability of an expanded range of antibodies and fluorochromes that have improved our ability to identify different normal cell populations and recognize phenotypic aberrancies, even when present in a small proportion of the cells analyzed. Phenotypically abnormal populations have been documented in many hematologic neoplasms, including lymphoma, chronic lymphoid leukemias, plasma cell neoplasms, acute leukemia, paroxysmal nocturnal hemoglobinuria, mast cell disease, myelodysplastic syndromes, and myeloproliferative disorders. The past decade has also seen refinement of the criteria used to identify distinct disease entities with widespread adoption of the 2001 World Health Organization (WHO) classification. This classification endorses a multiparametric approach to diagnosis and outlines the morphologic, immunophenotypic, and genotypic features characteristic of each disease entity. When should flow cytometric immunophenotyping be applied? The recent Bethesda International Consensus Conference on flow cytometric immunophenotypic analysis of hematolymphoid neoplasms made recommendations on the medical indications for flow cytometric testing. This review discusses how flow cytometric testing is currently applied in these clinical situations and how the information obtained can be used to direct other testing.

Intensive Chemotherapy for Acute Myelogenous Leukemia

A complete remission rate of 82% was obtained in a group of 68 patients with acute myelogenous leukemia treated with a high-dose induction chemotherapy (TAD) consisting of 7-day courses of 6-thioguanine, cytarabine, and daunorubicin. The patients who achieved remission received intensive consolidation chemotherapy and were randomized to receive maintenance chemotherapy with or without immunotherapy. Median remission duration was 13 months and median survival, 21 months. Neither central nervous system prophylaxis nor the addition of immunotherapy to the maintenance regimen prolonged remissions or improved survival. Age, sex, and subclassification of acute myelogenous leukemia had no effect on the remission rate or survival. These data indicate that a large proportion of patients with acute myelogenous leukemia can achieve remission with intensive induction chemotherapy. Attempts to prolong remission have been less successful.

Controversies in the therapy of acute myelogenous leukemia

In the past 10 years, there has been substantial progress in the treatment of patients with acute myelogenous leukemia. Intensive induction chemotherapy and consolidation chemotherapy have increased complete remission rates from 25 percent to more than 70 percent and have extended median survival from six months to more than two years. Attempts to prolong remission with maintenance chemotherapy, immunotherapy, and central nervous system prophylaxis have been less successful. Recent data suggest that the use of intensification chemotherapy or bone marrow transplantation in patients in remission may further reduce or eliminate residual leukemia. As a result of one or more of these advances an increasing proportion of patients, up to 25 percent in some series, are alive and free of disease three to five years following diagnosis. Most data indicate that some of these patients may be cured. In this article, we review the therapeutic interventions responsible for this substantial increase in survival in what was previously a uniformly fatal disease. Recent advances are discussed as are controversies in management and future directions.

The Antigenic Characteristics of Hematopoietic Stem Cells

THE surface antigenic structure of hematopoietic cells changes during the course of differentiation from proliferating precursors to nondividing, functionally mature cells. In a given cell lineage, some antigens are expressed and others are lost during the maturation process. Knowledge of the antigenic structure of hematopoietic progenitor cells and their mature progeny can help us to understand the process of cell differentiation and the function of specific surface antigens. This knowledge can aid in the identification of cells and cell lineages, in the classification of hematologic neoplasia, and potentially in the diagnosis and treatment of hematologic diseases. We shall summarize current .xa0.xa0.

Type 1-polarized dendritic cells loaded with autologous tumor are a potent immunogen against chronic lymphocytic leukemia

Induction of active tumor‐specific immunity in patients with chronic lymphocytic leukemia (CLL) and other hematologic malignancies is compromised by the deficit of endogenous dendritic cells (DCs). In attempt to develop improved vaccination strategies for patients with CLL and other tumors with poorly identified rejection antigens, we tested the ability of ex vivo‐generated DCs to cross‐present the antigens expressed by CLL cells and to induce CLL‐specific, functional CTL responses. Monocyte‐derived DCs from CLL patients were induced to mature using a “standard” cytokine cocktail (in IL‐1β, TNF‐α, IL‐6, and PGE2) or using an α‐type 1‐polarized DC (αDC1) cocktail (in IL‐1β, TNF‐α, IFN‐α, IFN‐γ, and polyinosinic:polycytidylic acid) and were loaded with γ‐irradiated, autologous CLL cells. αDC1 from CLL patients expressed substantially higher levels of multiple costimulatory molecules (CD83, CD86, CD80, CD11c, and CD40) than standard DCs (sDCs) and immature DCs, and their expression of CCR7 showed intermediate level. αDC1 secreted substantially higher (10–60 times) levels of IL‐12p70 than sDCs. Although αDC1 and sDCs showed similar uptake of CLL cells, αDC1 induced much higher numbers (range, 2.4–38 times) of functional CD8+ T cells against CLL cells. The current demonstration that autologous tumor‐loaded αDC1 are potent inducers of CLL‐specific T cells helps to develop improved immunotherapies of CLL.

Construction and characterization of DNA vaccines encoding the single-chain variable fragment of the anti-idiotype antibody 1A7 mimicking the tumor-associated antigen disialoganglioside GD2.

Anti-idiotype antibody, 1A7, functionally mimics the tumor-associated antigen disialoganglioside GD2, which is overexpressed on the surface of a number of neuroectodermal tumors such as melanoma, neuroblastoma, soft tissue sarcoma, and small cell carcinoma of the lung. Immunization of mice with 1A7 generated the production of anti-GD2 antibodies. In a phase I clinical trial, immunization of patients with 1A7, mixed with the adjuvant QS21, demonstrated that 1A7 could act as a surrogate antigen for GD2 and induce strong humoral immune responses in advanced stage melanoma patients. DNA vaccines have recently been shown to invoke humoral as well as cellular responses in injected hosts against the transgene product. To evaluate the efficiency of DNA vaccines encoding anti-idiotype antibodies, we constructed expression plasmids encoding the variable heavy (VH) and variable light (VL) chains of 1A7. The plasmids were made in two configurations, expressing either the VH (pc1A7VHLnVL) or the VL (pc1A7VLLnVH) chain of 1A7 at the amino terminus, linked together by a 15-amino acid linker (Ln). In vitro transcription/translation assays and transfection of CHO-K1 cells with the plasmids demonstrated that a ∼30-kDa protein was expressed by both configurations of the single-chain variable fragment. This protein can be specifically precipitated by monoclonal anti-GD2 antibody, 14G2a. Following intramuscular injection in mice, the plasmids were detectable in the injected tissues for at least 3 months and the injected plasmids actively transcribed the single-chain variable fragment 1A7 gene at the injected site. A single, intramuscular immunization of a group of C57BL/6 mice with pc1A7VLLnVH in phosphate-buffered saline induced humoral immune responses against 1A7 as well as GD2, the nominal antigen. Multiple immunizations, however, were required to elicit stronger immune responses. Cancer Gene Therapy (2000) 7, 1426–1436

Interleukin-15 enhances natural killer cell cytotoxicity in patients with acute myeloid leukemia by upregulating the activating NK cell receptors.

Interleukin-15 (IL-15) has a major role in NK-cell homeostasis. Modulation of the relative frequency and expression intensity of the NK-cell receptors by IL-15 may increase NK cell-mediated cytotoxicity in cancer patients. We investigated the receptor repertoire and measured NK-cell activity in newly diagnosed AML patients and evaluated the ex vivo effects of IL-15. The expression of the activating NK cell receptors was significantly decreased in the AML patients compared to that in NK cells of healthy donors. When NK cells obtained from AML patients were cultured with IL-15, expression of the activating receptors was significantly upregulated compared to pre-culture levels. Concomitantly, cytotoxic activity of NK cells against autologous leukemic blasts increased following IL-15 stimulation. This IL-15 induced increase in activity was blocked by neutralizing antibodies specific for the NK cell activating receptors. These pre-clinical data support the future use of IL-15 for NK cell- based therapies for AML patients.

Anti-idiotype vaccine against cancer

Immunization with anti-idiotype (Id) antibodies represents a novel new approach to active immunotherapy. Extensive studies in animal tumor models have demonstrated the efficacy of anti-Id vaccines in preventing tumor growth and curing mice with established tumor. We have developed and characterized several murine monoclonal anti-Id antibodies (Ab2) which mimic distinct human tumor-associated antigens (TAA) and can be used as surrogate antigens for triggering active anti-tumor immunity in cancer patients. Encouraging results have been obtained in recent clinical trials. In this article, we will review the existing literature and summarize our own findings showing the potential of this approach for various human cancers. We will also discuss where anti-Id vaccines may perform better than traditional antigen vaccines.

Therapy of acute myelogenous leukemia

Treatment of acute myelogenous leukemia (AML) is divided into remission induction and post-remission therapy. Remission induction is usually with cytarabine and an anthracycline. Daunorubicin is commonly used but recent data suggest idarubicin or mitoxantrone are equally effective, possibly better. High-dose cytarabine has also been used for remission induction but is not proven superior. Post-remission treatment is typically with two or more courses of drugs similar to those used for remission induction. Other studies use non-cross resistant drugs and/or high-dose cytarabine. Although some data favor use of high-dose cytarabine, no approach is clearly superior. There is considerable controversy whether persons in first remission and with an HLA-identical sibling should receive a bone marrow transplant immediately or after relapse. Although transplant results appear superior, especially in persons less than 20 years of age, the most effective strategy may be reserving transplants for persons failing chemotherapy. This strategy also applies to persons receiving autologous transplants or transplants from alternative donors, like HLA-matched related or unrelated persons.

Counterpoint. Cancer vaccines: Single-epitope anti-idiotype vaccine versus multiple-epitope antigen vaccine

Abstract Anti-idiotype (Id) vaccine therapy has been tested and shown to be effective, in several animal models, for triggering the immune system to induce specific and protective immunity against bacterial, viral and parasitic infections. The administration of anti-Id antibodies as surrogate tumor-associated antigens (TAA) also represents another potential application of the concept of the Id network. Limited experience in human trials using anti-Id to stimulate immunity against tumors has shown promising results. In this “counterpoint” article, we discuss our own findings showing the potential of anti-Id antibody vaccines to be novel therapeutic approaches to various human cancers and also discuss where anti-Id vaccines may perform better than traditional multiple-epitope antigen vaccines.