Biotechnology and drug discovery: from bench to bedside.

Abstract: New biotechnology and drug discovery technologies are facilitating the rapid expansion of the clinical drug chest, empowering clinicians with a better understanding of disease as well as novel modalities for treating patients. Important research tools and themes include genomics, proteomics, ligand-receptor interaction, signal transduction, rational drug design, biochips, and microarrays. Emerging drug classes include monoclonal antibodies, cancer vaccines, gene therapy, antisense strands, enzymes, and proteins. In this article, we review these topics and illustrate their potential impact by presenting an overview of promising drugs in the pipeline. Clinicians who use these novel treatments must become familiar with these trends.

Key Words: biotechnology, drug classes, drug development

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Biotechnology is introducing new capabilities to drug discovery, which were considered until recently to be impractical and futuristic. There has been continuous evolution in the integrated approach to the development of therapies in medicine. This effort relies on clinicians, basic scientists, and feedback from novel translational applications.

Initially, biotechnology was synonymous with the emerging recombinant deoxyribonucleic acid (DNA) technology and was used for the large-scale production of proteins, initially "replacement" proteins such as insulin and factor VIII. Later, developments were based on an understanding of ligand-receptor interactions, their impact on disease processes, and the ability to manufacture such large macromolecular proteins for therapeutic purposes. Today, signal transduction and cell signaling and their role in normal and disease states are taking center stage. Small-molecule drug (SMD) discovery, which uses and builds on organic molecules as starting materials, is also benefiting from the input of newer technologies such as combinatorial chemistry and high-throughput screening.

Although many physicians are not exposed to biotechnology, we think that it is valuable for clinicians to gain some fluency in the important trends in this field because the fruits of biotechnological research are reaching the clinic. The speed of events that are occurring in biotechnology is breathtaking and inspiring indeed. The younger generation of physicians has had the privilege of studying molecular biology as medical students. Even those physicians who did study molecular biology in medical school, however, must be excited but somewhat bewildered and uncomfortable about the advent of novel treatment modalities involving the use of antisense strands and monoclonal antibodies (MAb). This review is aimed at practitioners and specialists who are not closely involved in the process of drug discovery and intends to highlight the main developments in biotechnology and their impact on medicine.

Overview of Biotechnology

Drug discovery and development are costly and complicated processes. More than 99% of experimental compounds ultimately fail or are discarded as treatment regimens. Of the chemicals evaluated as part of drug discovery and preclinical testing, only a few proceed to human clinical trials and are approved for marketing. (1) To address this issue, new therapeutic approaches based on genomic and proteomic technology have been developed during the past several years. The -omic suffix is an example of the lexicon that has emerged to define the varied populations and subpopulations in the cell. These terms generally carry the -ome suffix, with an associated research topic denoted by the -omics appellation. The "genome"--that is, the full complement of an organism's genetic information that includes both coding and noncoding DNA sequences--provides a basis for defining the "proteome," which is a list of only the encoding DNA regions that result in protein products. (2) Genomics and proteomics enable the discovery of new genes and proteins and the comparison of their levels in diseased cells, normal cells, and cells treated with compounds that vary in their efficacy and toxicity. Thus, they could prove valuable in identifying new drug targets.

[FIGURE 1 OMITTED]

In drug discovery, the drug target is key. A target for pharmaceutical intervention is almost invariably a protein whose function or dysfunction is implicated in a disease process--for instance, growth factors and their receptors, which are frequently overexpressed in carcinomas. (3) A case in point is the epidermal growth factor (EGF) receptor family, which is the most studied growth factor receptor system. These receptors are composed of an extracellular binding domain, a transmembranous lipophilic segment, and an intracellular protein tyrosine kinase domain with a regulatory segment. The interaction of the extracellular growth factor with its receptor (ie, ligand-receptor interaction) results in the activation of cell signaling pathways that lead ultimately to cell division, the synthesis of new proteins, and tumor progression. This cascade of events is known as signal transduction. Figure 1 illustrates the different steps along the EGF signal transduction pathway that can serve as targets, some of which are being addressed already.

Manifold new drug targets are expected to sprout from the Human Genome Project, which has focused much attention on biotechnology. Indeed, the insight that the Human Genome Project provides with regard to the cell's genetic makeup as well as disease states can be used to understand the cell's protein makeup to generate new protein targets for intervention, such as the EGF family. However, the gene-protein-disease triangle is complex. Therefore, additional contributions from genomic and proteomic technologies is necessary to understand the genetic makeup and expression of diseased cells as well as how the resulting cellular proteins interact to cause disease. For example, biochips are one key technology that enables mutational analysis, gene sequencing, and protein expression testing. They consist of many small arrangements called microarrays that contain DNA, ribonucleic acid (RNA), or protein affixed to a small wafer such as that used in computers. Each microarray, or chip, contains thousands of different sequences of nucleotides or proteins. When a gene chip is reacted with a sample of unknown nature, only complementary sequences of DNA bind to the chip; unbound strands are washed away. One illustration of a gene chip's utility is that by using a gene chip with different tumor-associated genes, it is possible to determine whether a mutant gene, or oncogene, is present in a suspected cancer cell. Biochips are thus useful for identifying potential new drug targets. (4), (5)

Once a target with a pivotal role in disease is identified, the next step requires designing a drug that will interact with it and deliver a therapeutic effect. Understanding ligand-receptor interaction is a key element in designing a drug to interact with a target. To bind with the target, most drug molecules insert themselves into a functionally critical site of the target protein, like a key in a lock. The molecule then either induces or, more commonly, inhibits the protein's function. Thus, a better understanding of the target's structure and functionality is key to designing better therapeutics, or ligands, that bind to the target. In recent years, better understanding of protein structure and function has yielded sophisticated approaches to the generation and optimization of drug candidates. These methods are commonly referred to as rational drug design. In essence, rational drug design tailors drug candidates to their target proteins by first elucidating the three-dimensional structure, the binding site, and the active site of the target. (6) Next, medicinal chemists apply combinatorial chemistry techniques and high-throughput screening tools to generate large libraries of compounds whose structure corresponds to the target's strategic site. With the use of biologic assays that reflect the activity of the target protein, researchers can modify the drug candidate to achieve the ideal in vitro effect and test antitargets to determine the drug's specificity.

Biotechnology Drug Classes and Selected Drugs in the Pipeline

A snapshot of some drug candidates in development is beneficial to the understanding of how the technologies discussed in this article are used in drug development and in highlighting areas in medicine in which they may have a significant impact in the near future. We analyzed publicly available information, including the medical literature as well as U.S. Food and Drug Administration (FDA) and drug company reports, to generate a representative but by no means exhaustive list of drug candidates currently in clinical trials. To facilitate this discussion, we found it valuable to assign drugs into classes according to their chemical composition.

Small-Molecule Drugs. SMDs normally have limited biologic interaction capability and less specificity than other drugs for desired targets. In general, an SMD acts as a "spoiler," because its therapeutic effect is limited to the inhibition of an effector protein. For example, by interacting with a hormone receptor, the SMD can inhibit the binding activity of the respective hormone by occupying its docking site or by causing a change in the receptor's three-dimensional configuration. Both patients and the pharmaceutical industry favor the use of SMDs rather than other modalities because of their attractive pharmacokinetic properties, especially their suitability for oral administration and ease of development. (7) SMDs are well positioned to target intracellular proteins (ie, enzymes), because cell membrane penetration is often feasible.

The tyrosine kinase inhibitor ST1571 (imatinib mesylate, Gleevec; Novartis Pharmaceuticals Corp., East Hanover, NJ) is an SMD that has had an exceptional impact on the management of Philadelphia-positive chronic myelogenous leukemia (ie, Bcr-Abl-positive) and gastrointestinal stromal tumors with Kit mutations. (8), (9) ST1571 was first developed to target the platelet-derived growth factor receptor but then was found to be an inhibitor of a specific target protein: the Bcr-Abl protein kinase. Bcr-Abl had previously been implicated in the pathogenesis of Philadelphia mutation-positive leukemia, and ST1571 was then developed through a rational process of screening and refining potential small molecules. It therefore serves as a fitting example of the way in which rational drug design is effecting drug development and disease management. (10) Selected SMDs that are currently in clinical trials for different applications are listed in Table 1. (11-30)

Protein Drugs. Protein drugs can be subdivided according to their mechanism of action. This categorization scheme helps to create order in the world of protein drugs, yet overlaps exist between different subtypes. Until the early 1970s, proteins were derived from animals or were manufactured synthetically. This process greatly limited their use as pharmaceuticals because of availability and cost issues. Recombinant DNA technology significantly changed these circumstances. Scientists could now insert into a bacterial or yeast cell the human DNA comprising a gene and use the host to manufacture the human protein. With the use of that technology, the bottleneck in protein therapeutics shifted from protein manufacturing to the identification of drug targets and the generation of drug candidates.

Therapeutic Hormones and Enzymes. Hormones have been an important subject in pharmaceutical research, because the biology of various hormone deficiencies is relatively straightforward, and thus animal models can be created for research purposes. For instance, insulin was discovered in 1921, leading to the advent of replacement therapy with porcine insulin and a better understanding of diabetes. Furthermore, because hormones are circulating entities, they are more accessible than other proteins--especially intracellular proteins--for research purposes. Hence, early protein therapeutics targeted hormone deficiency states, and when recombinant DNA technology emerged, it was used to manufacture replacement proteins, whose role in disease was relatively clear. Enzymes have enormous potential to serve as pharmaceuticals because of their vast number and ubiquitousness. That many enzymes operate intracellularly poses practical problems, however, because protein delivery into the cell is currently next to impossible. More intracellular enzymes are continually being discovered, and methods of enabling oral and intracellular protein delivery are among the most burning challenges in biotechnology. (31) Notable protein drugs that are in the advanced development to launch stages include a T cell-modulating fusion protein for patients with psoriasis and various other autoimmune diseases, as well as recombinant natriuretic peptide for the treatment of patients with congestive heart failure (see Table 2) (33-47)

Monoclonal Antibodies. Antibodies (immunoglobulins) are proteins manufactured by B-lymphocytes. They consist of a highly diverse binding site known as the variable region (Fab), which sticks to a corresponding antigen, and a crystallizable fragment domain (Fc), which determines the antibody's functionality. MAb are products of a distinct clone of B cells and are usually derived by immunizing mice against the desired antigen. The reactive B cells are fused with myeloma cells to create hybridomas, which are essentially clones of immortalized B cells that share specificity for the same antigen. (48) The hybridomas' identical product antibody is called a monoclonal antibody, or MAb.

The ability to manufacture clones of identical MAb enables different strategies to create MAb therapeutics. By recruiting the immune system's lytic action, MAb lead to the destruction of antigens implicated in disease. MAb can be used to target toxic drugs such as chemotherapy directly at their site of action, thereby reducing side effects and the required dosage. MAb can be used as diagnostic agents to locate, for instance, residual tumor cells after surgery. (49)

In recent years, MAb have made the leap from promising investigational therapies to the clinic, and currently there are 12 approved MAb in the United States. Eight of those 12 MAb are approved for therapeutic indications, including cancer, autoimmune disease, viral infection, and myocardial infarction. In addition, four MAb have been approved for use as diagnostic agents. Most of these have had a significant impact on disease, as evidenced in the discussion of selected MAb in the next few paragraphs.

Rituximab (Rituxan; Genentech, Inc., South San Francisco, CA), the first MAb approved for the treatment of cancer patients, is indicated for non-Hodgkin's B cell lymphoma. It is directed against the cytosine deaminase 20 (CD20) protein, which is found on the surface of normal and malignant B cells. As a single agent, rituximab induces meaningful responses in approximately one-half of patients with relapsed indolent lymphomas and in approximately one-third of patients with relapsed aggressive lymphomas. Because it is a nonchemotherapeutic agent, it also presents a relatively benign side effect profile. Rituximab is currently being tested for other B cell disorders. (50), (51)

Infliximab (Remicade; Centocor, Inc., Malvern, PA) is a chimeric (human-mouse) MAb that neutralizes tumor necrosis [alpha], a proinflammatory cytokine. It is approved for the treatment of patients with rheumatoid arthritis and Crohn's disease. In clinical trials for rheumatic arthritis, infliximab produced significant improvements in all measures of disease, and treatment with infliximab combined with methotrexate was found to be superior to treatment with methotrexate alone. (52) In Crohn's disease, clinical trials showed infliximab to be effective in producing and maintaining a clinical response in patients with refractory moderate to severe disease. (53)

Trastuzumab (Herceptin; Genentech, Inc.) is a humanized MAb that targets the extracellular portion of the human epidermal growth factor receptor 2 (HER2)/Neu receptor. The latter is a member of the EGF receptor family, the blockade of which inhibits the growth of tumors that express it. HER2/Neu is overexpressed in 25 to 30% of breast cancers, increasing tumor aggressiveness. In HER2/Neu-positive patients, the use of trastuzumab with chemotherapy was associated with improved time until disease progression and with overall survival. (54) Several promising MAb currently in late-stage clinical trials are targeting non-Hodgkin's lymphoma, asthma, psoriasis, and different solid organ tumors (Table 2). (33-47)

Cytokines. Cytokines are proteins that regulate cells that belong to the immune system, such as lymphocytes and macrophages. Cytokines have a pivotal role in normal and disease mechanisms in which immune processes play a role, including chronic infectious, autoimmune, cancer, and coronary heart diseases. Many cytokines and cytokine inhibitors are available or are being developed as therapeutics. (55-59)

Interleukins (ILs) and interferons are a large and varied family of compounds produced by lymphocytes, macrophages, and monocytes. The FDA has approved the use of recombinant IL-2 for the treatment of patients with renal cell carcinoma, but IL-2 is highly toxic because of its central role in the immune system, which thus far has limited its impact. (60) The FDA has approved the use of recombinant interferons for patients with human immunodeficiency virus-related Kaposi's sarcoma, genital warts, hairy cell leukemia, and hepatitis B and C. (59)

Colony-stimulating factors (CSFs) stimulate bone marrow stem cells to differentiate toward a particular cell type. Recombinant versions of CSF, including granulocyte-macrophage CSF, granulocyte CSF, and erythropoietin have revolutionized the ability to treat myelosuppression. Most notably, these agents have had a significant impact in cancer treatment, in which myelosuppression is a common complication of chemotherapy. Proved and suggested effects of treatment include shortening the duration of febrile neutropenia after myelosuppressive treatment, mobilization of hematopoietic stem cells for ensuing transplantation, and reductions in chemotherapy-associated infections, antibiotic use, hospital stay, and mortality. (61), (62) Erythropoietin has had a profound effect on the treatment of patients with end stage renal failure-associated anemia. (63) Some investigational cytokines are listed in Table 2.

Gene Therapy. Gene therapy may be defined as the transfer of recombinant DNA into human cells to achieve the production of a desired protein. Depending on the strategy used, DNA may be introduced into cells removed from the body (ie, the ex vivo approach) or directly into cells in their normal location (ie, the in vivo approach). (64) Gene therapy has various potential applications, such as treating patients with enzyme deficiencies or cancer. Efficient gene transfer requires the use of a vector. All vectors contain, at a minimum, the transgene of interest linked to a promoter to drive its expression. (65) Increasingly wider ranges of viral and synthetic vectors are available, each of which has characteristic advantages and limitations. Generally, viral vectors achieve better transfection than other vectors but have other problems such as immunogenicity and complicated manufacturing. Liposomes are nonviral vectors that mitigate the immunogenicity problem but provide less efficient transfection and protein expression. Naked DNA is a third method that uses plasmids, which usually are administered by direct injection into tumor or muscle as opposed to systemic delivery. (66)

There are several strategies whereby gene therapy may be used to treat cancer. In the corrective gene therapy approach, when malignant transformation is associated with inactivity of tumor suppressor genes such as p53 and p21, supplying tumors with the intact gene may reverse malignant transformation by promoting apoptosis. (67-69) Another strategy is cytoreductive gene therapy, in which immunotherapy or cytolytic/proapoptotic approaches are used. Immunotherapy uses gene transfer to facilitate a dormant host immune response directed against the tumor. Evasion of autologous host cellular immunity is a common feature of tumor cell neoantigens, because tumor cells are poor antigen-presenting cells. Cancer vaccine strategies are based on optimization of the context in which tumor antigens or tissue-specific antigens are presented to the host immune system. When appropriately primed, the activated host immune system can then act against tumor cells systemically.

The ex vivo approach starts with inactivated cancer cells obtained from the patient. Different techniques are then used to enhance the immunogenicity of tumor-specific antigens, including growth in a cytokine-rich environment, coinjection of tumor cells along with cytokines back into the patient, or transfection of these cells in vitro with genes that encode immunostimulatory cytokines. A second approach is to administer an injection of a purified tumor-associated protein or peptide into the patient, without injecting the entire tumor cell. The third approach uses in vitro manipulation of host antigen-presenting cells such as dendritic cells, which are involved in initiating the T cell-mediated response against antigens. Confronting dendritic cells with the desired antigen in vitro stimulates an immune response upon injection into the patient. (70), (71) Enzyme/prodrug gene therapy, also referred to as suicide gene therapy, relies on the conversion of an inactive prodrug into a toxic drug with the use of an enzyme vectored only to the target tumor cells. In this way, active drug is limited spatially to the transduced cells and adjacent surrounding cells, facilitating higher tumor drug concentrations without increased normal tissue toxicity. Prodrug-activating enzymes that have been used in this approach include cytosine deaminase, which catalyzes the conversion of the nontoxic 5-fluorocytosine to the cytotoxic 5-fluorouracil, and herpes simplex virus thymidine kinase, which, together with cellular enzymes, facilitates the conversion of ganciclovir into the toxic ganciclovir triphosphate.

Viral vectors may themselves be designed to target and kill tumor cells without the insertion of a foreign transgene (eg, oncolytic viruses). The adenovirus life cycle includes a lytic phase, which can result in host cell death independent of entry into the cell cycle. Adenovirus has evolved a potent repertoire of gene products that may exert profound effects on the growth regulation of the host cell to facilitate viral replication. The ONYX-015 vector, a replication-competent adenovirus designed to preferentially replicate in p53 mutant cells, is currently in clinical trials (Table 3). (72) ONYX-015 is the first genetically engineered replication-competent virus to demonstrate selective intratumoral replication and necrosis in patients. (73)

Antisense Drugs. Although traditional drugs are designed to interact with protein molecules, antisense drugs are designed to inhibit the production of disease-causing proteins. During the transcription of information from DNA to messenger RNA (mRNA), two complementary strands of DNA partly uncoil such that one strand is used as a template for the transcribing enzymes, which assemble mRNA in a process called transcription. mRNA then migrates into the cell, where its encoded information is read by the ribosomes and translated to the specific protein. (95)

Antisense drugs are complementary strands of small segments of mRNA. To create antisense drugs, nucleotides are linked in short chains (ie, oligonucleotides). Each antisense drug is designed to bind to a specific sequence of nucleotides in its mRNA target to inhibit the production of the protein encoded by the target mRNA. Fomivirsen (Vitravene; Isis Pharmaceuticals, Inc., Carlsbad, CA) is an antisense strand complementary to the mRNA of a crucial cytomegalovirus protein. It is an FDA-approved medication indicated for patients with acquired immunodeficiency syndrome-related cytomegalovirus retinitis. Oblimersen sodium (Genasense; Genta, Inc., Berkeley Heights, NJ) is an antisense drug that binds to the Bc1-2 mRNA, which is expressed by different cancers (Table 3).

Conclusions

The development of a new drug requires the identification of a protein target, techniques for the generation of compounds that react with the target in a desired fashion, and innovative delivery mechanisms by which to lead the drug to its target. The tools of biotechnology are effecting advancements on all of these fronts. An abundance of new gene and protein targets that can be targeted by therapeutics are being investigated. Moreover, whereas in the past most drugs were randomly generated small-molecule compounds that were limited to the blockade of certain pathways, other drug classes have emerged, including recombinant protein drugs and MAb, DNA and cellular vaccines, gene therapy, and antisense therapy. SMDs, which remain a fundamental weapon against many diseases, can be engineered to provide a better therapeutic profile than before. Some of the newer drug classes, including protein drugs and MAb, already have exhibited proof of concept as approved drugs on the basis of several years of experience. The potential impact on disease of cancer vaccines, gene therapy, and antisense therapy remains to be determined, but there seems to be consensus regarding the eventual important role of these technologies.

As a result of these developments, physicians will be able to attack the same target with a mix of various drug classes, such as combinations of MAb or a cancer vaccine against a tumor-associated protein, a cytokine to increase the antitumor immune response, and gene therapy encoding for a suicide protein. Such an approach is not yet feasible, but the drugs that will allow experimentation with such combinations are at our doorstep.

Several drug candidates are far along the route to becoming FDA-approved drugs. These include a cellular vaccine for hormone-resistant prostate cancer, a cancer vaccine and a small-molecule antiangiogenic drug for renal cell carcinoma, a new immune modulator protein drug directed against psoriasis and other autoimmune diseases, an SMD directed against transitional cell carcinoma, and several MAb that target cancer, autoimmune disease, and graft versus host disease. Judging from the wealth of advanced clinical candidates in the pipeline, the impact of biotechnology on the practice of medicine will soon increase markedly, empowering clinicians with new ways to fight disease.

Better to be kind at home

Than to burn incense in a distant place.

--Chinese proverb

Table 1. Small-molecule drug candidates (a)

Drug brand name              Drug manufacturer/
(generic name)                developer, location

Tarceva (OSI-774)            OSI Pharmaceuticals, Inc.,
                               Melville, NY

Incel (biricodar dicitrate,  Vertex Pharmaceuticals,
  VX-710)                      Inc., Cambridge, MA

Eflornithine                             --
  (difluoromethylornithine,
  or DFMO)

No brand name                            --
  (CCI779)

No brand name                Abbott Laboratories,
  (atrasentan, ABT-627)        Abbott Park, IL

Velcade (bortezomib,         Millennium
  PS-341)                      Pharmaceuticals, Inc.,
                               Cambridge, MA

Apomine (SR-45023A)          Ilex Oncology, Inc., San
                                Antonio, TX
Flavopiridol (HMR1275)       Aventis Pharmaceuticals,
                               Inc., Bridgewater, NJ
No brand name                            --
  (CEP701)

No brand name                GlycoDesign, Inc.,
  (GD0039)                     Toronto, ON, Canada

No brand name (UCN-                      --
  01)

Iressa (gefitinib,           AstraZeneca
   ZD1839)                     Pharmaceuticals LP,
                               Wilmington, DE

Thalomid (thalidomide)       Celgene Corp., Warren,
                               NJ

No brand name (NV-06,        Marshall Edwards, Inc.,
  phenoxodiol)                 North Ryde, NSW,
                               Australia

Drug brand name
(generic name)                             Condition

Tarceva (OSI-774)            Pancreatic, breast, non-small cell lung,
                               and head and neck cancers

Incel (biricodar dicitrate,  Ovarian, liver, CAP, and small cell
  VX-710)                      lung cancers; sarcomas

Eflornithine                 Colon and nonmelanoma skin cancers;
  (difluoromethylornithine,    TCC; Barrett's esophagus
or DFMO)

No brand name                Glioma, glioblastoma multiforme,
  (CCI779)                     melanoma, small cell lung cancer

No brand name                CAP
  (atrasentan, ABT-627)

Velcade (bortezomib,         Ovarian, head and neck, and refractory
  PS-341)                      hematologic malignancies, including
                               multiple myeloma and breast cancer

Apomine (SR-45023A)          CAP

Flavopiridol (HMR1275)       Solid tumors and lymphoma

No brand name                CAP
  (CEP701)

No brand name                Colorectal and breast cancers and RCC
  (GD0039)

No brand name (UCN-          Advanced solid tumors
  01)

Iressa (gefitinib,           RCC
   ZD1839)

Thalomid (thalidomide)       Multiple myeloma, CAP

No brand name (NV-06,        RCC and other solid tumors
  phenoxodiol)

                             Clinical
Drug brand name                trial
(generic name)               phase (b)             Comments

Tarceva (OSI-774)            III        Selective, orally active
                                        inhibitor of ErbB1
                                          tyrosine kinase (an epidermal
                                          growth factor
                                          receptor), a key oncogene in a
                                          variety of
                                          cancers, including ovarian,
                                          pancreatic, non-
                                          small cell lung, breast, and
                                          head and neck
                                          cancers. (11)

Incel (biricodar dicitrate,   II        Small molecule blocker of
  VX-710)                               multidrug-resistant
                                         pumps MDR1 gene and multidrug
                                         resistance-
                                         associated protein. Intended to
                                         sensitize
                                         tumors to chemotherapy. (12)

Eflornithine                 III        Irreversible inhibitor of
  (difluoromethylornithine,             ornithine
  or DFMO)                                 decarboxylase, an enzyme
                                           elevated in most
                                           tumors and premalignant
                                           lesions. (13)

No brand name                 II        Binds to the immunophilin
  (CCI779)                              FK506-binding
                                          protein 12, and the resultant
                                          complex
                                          inhibits the activity of
                                          mammalian target of
                                          rapamycin (mTOR). (14)

No brand name                III        Endothelin A receptor inhibitor
  (atrasentan, ABT-627)                 that inhibits
                                          endothelin-1 receptor-mediated
                                          effects,
                                          including stimulation of
                                          prostate cancer
                                          proliferation and apoptosis
                                          inhibition, and
                                          promotes osteoblast activity.
                                          (15) (Note: Abbott
                                          Labs stopped Phase III trial
                                          in February
                                          2003.)

Velcade (bortezomib,          II        Inhibitor of
  PS-341)                               ubiquitin-proteasome pathway
                                        that
                                          has an important role in cell
                                          cycle regulation
                                          by degradation of
                                          intracellular proteins. This
                                          results in shifts in favor of
                                          increased cell
                                          survival, metastasis, and
                                          angiogenesis. (16)
                                          Recent report (17) showed that
                                          PS-341 has
                                          activity against refractory
                                          multiple myeloma
                                          and possibly non-Hodgkin's
                                          lymphoma.

Apomine (SR-45023A)           II        Bisphosphonate ester that binds
                                        to the
                                          farnesoid X nuclear receptor.
                                          Induces
                                          apoptosis of cancer cells.
                                          (18)

Flavopiridol (HMR1275)        II        A potent inhibitor of
                                        cyclin-dependent kinases
                                         and others. (19)

No brand name                 II        Inhibitor of the receptor for
  (CEP701)                              nerve growth
                                          factor, a tyrosine kinase
                                          receptor implicated
                                          in cancer pathways. (20)

No brand name                 II        Inhibitor of Golgi enzyme
  (GD0039)                              mannosidase II,
                                          which is responsible for the
                                          synthesis of key
                                          carbohydrate structures
                                          involved in
                                          disease. (21)

No brand name (UCN-           II        Nonspecific inhibitor of many
  01)                                   kinases causing
                                          cell cycle arrest in G1 and G2
                                          phases in
                                          different cell types. (22)

Iressa (gefitinib,            II        Inhibitor of epidermal growth
   ZD1839)                               factor receptor. (23)

Thalomid (thalidomide)       III        Small molecule antiangiogenesis
                                        agent. Also
                                         being evaluated in early
                                         clinical trials for
                                         recurrent or malignant glioma,
                                         myelofibrosis
                                         with myeloid metaplasia,
                                         colorectal cancer,
                                         ovarian epithelial cancer,
                                         metastatic RCC,
                                         and non-small cell lung cancer.
                                         (24)

No brand name (NV-06,         II        Signal transduction inhibitor.
  phenoxodiol)                          Blocks enzymes
                                          involved in cell division,
                                          including
                                          topoisomerase-2 and protein
                                          tyrosine
                                          kinases. Restores apoptosis in
                                          cancer cells. (c)

Reminyl (galantamine         Janssen Pharmaceutica
  HBr)                         Products LP, Titusville,
                               NJ

Neovastat (AE-941)           AEterna Laboratories, Inc.,
                               Quebec City, QB,
                               Canada

17-N-allylamino-17-                     --
  demethoxy
  geldanamycin
  (17-AAG)

No brand name                Pfizer, Inc., New York,
  (indiplon, NBI-34060)        NY, and Neurocrine
                               Biosciences, Inc., San
                               Diego, CA

Rotigotine CDS               Aderis Pharmaceuticals,
  (SPM-962)                    Hopkinton, MA, and
                               Schwarz Pharma AG,
                               Monheim, Germany

No brand name (2-                      --
  methoxyestradiol)

No brand name                Ilex Oncology, Inc., San
  (ILX295501)                   Antonio, TX, and Eli
                                Lilly & Co.,
                                Indianapolis, IN

Reminyl (galantamine         Alzheimer's disease
  HBr)

Neovastat (AE-941)           Lung and breast cancers and RCC

17-N-allylamino-17-          Epithelial and hematologic
  demethoxy                    malignancies, and sarcomas
  geldanamycin
  (17-AAG)

No brand name                Insomnia
  (indiplon, NBI-34060)

Rotigotine CDS               Parkinson's disease
  (SPM-962)

No brand name (2-            Advanced solid tumors
  methoxyestradiol)

No brand name                Solid tumors
  (ILX295501)

Reminyl (galantamine            III     Inhibitor of
  HBr)                                  acetylcholinesterase that also
                                          acts agonistically on
                                          nicotinic receptors. In
                                          clinical trials, patients who
                                          took galantamine
                                          experienced benefits in
                                          cognitive function
                                          and activities of daily
                                          living. (25)

Neovastat (AE-941)              III     Antiangiogenesis drug (shark
                                        cartilage extract)
                                         that activates apoptosis of
                                         endothelial cells
                                         and inhibits the vascular
                                         endothelial growth
                                         factor-R2 pathway. (26), (27)

17-N-allylamino-17-              I      Heat shock protein 90 inhibitor.
  demethoxy                              It maintains the conformation,
  geldanamycin                           stability and function of
  (17-AAG)                               key oncogenic proteins
                                         involved in signal transduction
                                         pathways leading to
                                         proliferation, cell cycle
                                         progression and
                                         apoptosis, as well as other
                                         features of the
                                         malignant phenotype such as
                                         invasion,
                                         angiogenesis, and metastasis.
                                         (28)

No brand name                   III     Nonbenzodiazepine agent that
  (indiplon, NBI-34060)                  acts on a specific site
                         of the [gamma]-aminobutyric
                                         acid A receptor in an
                                         agonistic fashion. (c)

Rotigotine CDS                  III     Lipid-soluble D2 dopamine
  (SPM-962)                               agonist in a transdermal
                                          delivery formulation
                                          (ie, patch). Intended to
                                          decrease motor complications
                                          associated with chronic use
                                          of dopamine agonists,
                                          which usually exhibit
                                          intermittent levels of
                                          dopamine stimulation. (29)

No brand name (2-                I      Estrogen-related compound with
  methoxyestradiol)                       antiproliferative and
                                          antiangiogenic
                                          activities. (30)

No brand name                   II      Sulfonylurea with antitumor
  (ILX295501)                             activity against a
                                          broad spectrum of solid
                                          tumors. Inhibits the
                                          enzymes topoisomerase 1 and 2,
                                          causing
                                          DNA breaks and cell death in
                                          cancer cells. (c)

(a) CAP, carcinoma of prostate; TCC, transitional cell carcinoma; RCC,
renal cell carcinoma; DNA, deoxyribonucleic acid.
(b) Phase relates to clinical trial stage in the United States as of
April 2003. Some drugs may be in a more advanced clinical trial phase
in other countries.
(c) Unpublished data.

Table 2. Protein drug candidates (a)

Drug brand name            Drug manufacturer/
(generic name)             developer, location

Plenaxis (Abarelix-Depot,  Praecis Pharmaceuticals,
   PPI-149)                  Inc., Waltham, MA

Ampligen (AMP 719)         Hemispherx Biopharma,
                             Inc., Philadelphia, PA

No brand name              Abgenix, Inc., Fremont,
  (ABX-EGF)                  CA, and Immunex
                             Corp., Seattle, WA

Multikine                  CEL-SCI Corp., Vienna,
                             VA

Avicidin                   NeoRx Corp., Seattle, WA

Natrecor (nesiritide)      Scios, Inc., Sunnyvale, CA

Amevive (alefacept)        Biogen, Inc., Cambridge,
                             MA

No brand name              Abgenix, Inc., Fremont,
  (ABX-CBL)                  CA, and SangStat
                             Medical Corp., Fremont,
                             CA

Xolair (omalizumab)        Genentech, Inc., South San
                             Francisco, CA, Novartis
                             AG, East Hanover, NJ

No brand name              Amgen, Inc., Thousand
  (epratuzumab, AMG         Oaks, CA
  412)

Bexxar (tositumomab and    Corixa Corp., Seattle, WA,
[.sup.131]I-tositumomab)     and GlaxoSmithKline,
                             Research Triangle Park,
                             NC

Antegren (natalizumab)     Biogen, Inc., Cambridge,
                             MA, and Elan Corp.,
                             plc, Dublin, Ireland

Keyhole Limpet                    --
  Hemocyanin (KLH,
  DRG-0070)

Drug brand name                                   Clinical trial
(generic name)                  Condition            phase (b)

Plenaxis (Abarelix-Depot,  CAP                   NDA action date
   PPI-149)                                       extended 90 days
                                                  from August 27,
                                                  2003, by FDA

Ampligen (AMP 719)         RCC                         II/III

No brand name              RCC                           II
  (ABX-EGF)

Multikine                  CAP                           II

Avicidin                   Lymphoma, CAP,                II
                             ovarian, and colon
                             cancers

Natrecor (nesiritide)      Decompensated CHF     M (FDA approval
                                                  received 2001)

Amevive (alefacept)        Psoriasis             M (FDA approval
                                                  received January
                                                  30, 2003)

No brand name              GVHD                        II/III
  (ABX-CBL)

Xolair (omalizumab)        Asthma and seasonal   M (FDA approval
                             allergies            received June
                                                  2003)

No brand name              Non-Hodgkin's                III
  (epratuzumab, AMG         lymphoma
  412)

Bexxar (tositumomab and    Non-Hodgkin's         M (FDA approval
[.sup.131]I-tositumomab)    lymphoma              received June
                           30,     2003)

Antegren (natalizumab)     MS, Crohn's                  III

Keyhole Limpet             Superficial bladder          III
  Hemocyanin (KLH,           TCC
  DRG-0070)

Drug brand name
(generic name)                               Comments

Plenaxis (Abarelix-Depot,  Peptide antagonist to gonadotropin-releasing
   PPI-149)                  hormone, blocking its action on the
                             pituitary
                             gland. (32)

Ampligen (AMP 719)         Second-generation interferon that acts as an
                             immune system stimulator. (33)

No brand name              Humanized monoclonal antibody against the
  (ABX-EGF)                  epidermal growth factor receptor, which is
                             overexpressed in 70 to 90% of RCCs. (34)

Multikine                  Immunotherapeutic agent that is a mixture of
                             interleukins, interferons, and colony-
                             stimulating factors; intratumoral
                             administration. (35)

Avicidin                   Streptavidin/biotin monoclonal antibody
                             treatment. A multistep delivery system that
                             involves the use of an antibody to target
                             streptavidin to a tumor-associated antigen
                             receptor. Biotin is then used to target the
                             [.sup.90]Y radioisotope to the
                             tumor-localized
                             streptavidin. (36)

Natrecor (nesiritide)      Recombinant form of B-type natriuretic
                             peptide that produces hemodynamic and
                             symptomatic improvement through balanced
                             vasodilatory effects, neurohormonal
                             suppression, and enhanced natriuresis and
                             diuresis. (37), (38)

Amevive (alefacept)        Human fusion protein obtained by recombinant
                             DNA technology designed to modulate the
                             activity of T-cells that play a critical
                             role in the pathogenesis of psoriasis
                             and other autoimmune diseases. Alefacept
                             modulates the function of and selectively
                             induces apoptosis of [CD.sup.2+] human
                             memory effector T cells in vivo.
                             Psoriatic plaques are characterized by
                             infiltration with CD45RO+
                             memory effector T lymphocytes. The
                             recombinant protein alefacept binds to CD2
                             on memory effector T lymphocytes,
                             inhibiting their activation. (39), (40)

No brand name              A murine immunoglobulin M monoclonal
  (ABX-CBL)                  antibody that recognizes CD147 on the cell
                             surface. Human CD147, also known as
                             neurothelin or extracellular matrix
                             metalloproteinase inducer, is a member of
                             the immunoglobulin superfamily. On
                             activation, CD147 is upregulated on T and
                             B lymphocytes, which are then depleted
                             through a complement-dependent cytotoxic
                             mechanism. (41)

Xolair (omalizumab)        A novel humanized monoclonal antibody
                             directed against the high-affinity
                             Fc[euro]RI portion of the immunoglobulin
                             E. It decreases serum immunoglobulin E
                             levels in a dose-dependent manner. It was
                             shown to significantly reduce the rate of
                             exacerbation and improve disease control
                             in patients at high risk for serious
                             asthma-related morbidity and
        mortality. (42)

No brand name              Humanized anti-CD22 monoclonal antibody.
  (epratuzumab, AMG          CD22 is commonly expressed by B-cell
  412)                       malignant cells. (43)

Bexxar (tositumomab and    Anti-CD20 murine monoclonal antibody
[.sup.131]I-tositumomab)     conjugated with radioactive iodine
                             ([.sup.131]I);
                             radioimmunotherapy. (44)

Antegren (natalizumab)     Humanized monoclonal antibody and the first
                             in a new class of potential therapeutics
                             known as [alpha]4 integrin inhibitors that
                             are designed to block immune cell
                             adhesion to blood vessel walls and
                             subsequent migration of lymphocytes
                             into tissue. (45), (46)

Keyhole Limpet             Immune modulator protein derived from the
  Hemocyanin (KLH,           mollusk Megathura crenulata (great keyhole
  DRG-0070)                  limpet), which is found along the coast of
                             California and Mexico. (47)

(a) CAP, carcinoma of prostate; NDA, new drug application to U.S. Food
and Drug Administration (FDA); RCC, renal cell carcinoma; CHF,
congestive heart failure; DNA, deoxyribonucleic acid; GVHD, graft
versus host disease; MS, multiple sclerosis; TCC, transitional cell
carcinoma; M, approved medication.
(b) Phase relates to clinical trial stage in the United States as of
April 2003. Some drugs may be in a more advanced clinical trial phase
in other countries.

Table 3. Cancer vaccines and gene therapy drug candidates (a)

Drug brand                      Drug
name (generic                manufacturer/
name)                     developer, location         Disease

Advexin (INGN 201,     Introgen Therapeutics,     Head and neck,
  Ad5CMVp53)             Inc., Austin, TX          breast, lung, and
                                                   pancreatic
                                                   cancers and TCC

OncoVAX                Intracel, LLC, Frederick,  Colorectal cancer
                         MD                        and RCC

Leuvectin              Vical, Inc., San Diego,    RCC and CAP
                         CA

Oncophage (HSPPC-      Antigenics, Inc., New      Lung cancer and
  96)                    York, NY                  RCC melanoma

Genasense (oblimersen  Genta, Inc., Berkeley      CLL, AML,
  sodium, G-3139;        Heights, NJ               melanoma, MM,
  formerly                                         and CAP
  augmerosen)

Provenge               Dendreon Corp., Seattle,   CAP
                         WA

GVAX                   Cell Genesys, Inc., South  CAP, lung, and
                         San Francisco, CA         pancreatic
                                                   cancers and AML

Avicine                AVI BioPharma, Inc.,       CAP
                        Portland, OR

Prostvac               Therion Biologics Corp.,   CAP
                         Cambridge, MA

CG7060 (formerly       Cell Genesys, Inc., South  CAP
  CV706)                 San Francisco, CA

CG7870 (formerly       Cell Genesys, Inc., South  CAP
  CV787)                 San Francisco, CA

TG4010 (MVA-MUC-       Transgene, S.A.,           CAP
 IL2)                    Strasbourg, France

INGN 201 AD-p53        Introgen Therapeutics,     CAP, TCC
                         Inc., Austin, TX

Prostate-specific      PSMA Development Co.,      CAP
  membrane antigen       LLC (Progenics
  (PSMA) vaccine         Pharmaceuticals, Inc.,
                         Tarrytown, NY, and
                         Cytogen Corp.,
                         Princeton, NJ, in
                         collaboration with
                         AlphaVax Human
                         Vaccines, Inc., Research
                         Triangle Park, NC)

Drug brand
name (generic          Clinical trial
name)                    phase (b)          Type

Advexin (INGN 201,     II/III (FDA         Gene
  Ad5CMVp53)              designated Fast   therapy
                          Track Drug
                          Product status
                          September
                          2003)

OncoVAX                       II           Cellular
                                             cancer
                                             vaccine

Leuvectin                     II           Gene
                                             therapy

Oncophage (HSPPC-             III          Peptide
  96)                                        cancer
                                             vaccine

Genasense (oblimersen       II/III         Antisense
  sodium, G-3139;
  formerly
  augmerosen)

Provenge                      III          Cellular
                                             cancer
                                             vaccine

GVAX                        II/III         Cellular
                                             cancer
                                             vaccine

Avicine                       II           Peptide
                                             cancer
                                             vaccine

Prostvac                      II           Gene
                                             therapy

CG7060 (formerly             I/II          Gene
  CV706)                                     therapy

CG7870 (formerly             I/II          Gene
  CV787)                                     therapy

TG4010 (MVA-MUC-              I            Gene
 IL2)                                        therapy

INGN 201 AD-p53               II           Gene
                                             therapy

Prostate-specific             II           Cellular
  membrane antigen                           cancer
  (PSMA) vaccine                             vaccine

Drug brand
name (generic
name)                                  Comments

Advexin (INGN 201,     Gene therapy applying the p53 gene using
  Ad5CMVp53)            an adenoviral vector. Most advanced
                        clinical trials are for head and neck
                        cancers, whereas the same approach is in
                        earlier trials for other oncologic
                        indications. (74), (75)

OncoVAX                Irradiated autologous cancer cells obtained
                        in surgery are attached to Bacille bilie de
                        Calmette-Guerin vaccine and returned to
                        patient to stimulate immune response. (76)

Leuvectin              Gene therapy based on the IL-2 gene.
                        Evaluated for metastatic RCC and local
                        CAP. Administered as direct intratumoral
                        injection. (77)

Oncophage (HSPPC-      Vaccine consisting of autologous heat
  96)                   shock protein-peptide complexes derived
                        from patient's own tumor tissue. (78)

Genasense (oblimersen  An antisense oligonucleotide specific for
  sodium, G-3139;       Bcl-2 that is expressed by most types of
  formerly              cancer. (79)
  augmerosen)

Provenge               Therapeutic cellular vaccine for hormone-
                        resistant CAP. Based on sensitizing
                        patient's dendritic cells to a ubiquitous
                        CAP antigen. (80), (81)

GVAX                   Non-patient-specific cancer vaccine
                        composed of irradiated tumor cells
                        modified to secrete GM-CSF
                        Administered intradermally. (80), (82)

Avicine                Cancer vaccine designed to induce an
                         immune response against the hormone
                         human chorionic gonadotropin, which is
                         present in most prostate carcinomas.
                         Consists of a short peptide part of human
                         chorionic gonadotropin attached to the
                         diphtheria toxoid. (83)

Prostvac               DNA vaccine based on PSA gene with a
                        gene for an adjuvant attached, using a
                        vaccinia virus vector. (84)

CG7060 (formerly        Adenovirus engineered to grow in cancer
  CV706)                cells expressing PSA. (85)

CG7870 (formerly        Adenovirus engineered to grow in cancer
  CV787)                cells expressing PSA. (86)

TG4010 (MVA-MUC-        Gene therapy using the genes for tumor-
 IL2)                   specific antigen MUC-1 and for IL-2,
                        delivered by a vaccinia (ie, pox) virus
                        vector. (87)

INGN 201 AD-p53        P53 gene therapy using an adenoviral
                         vector. A similar product is in Phase III
                         clinical trials for head and neck cancer. (88)

Prostate-specific      Cancer vaccine based on the PSMA epitope
  membrane antigen       found in CAP cells using ex vivo
  (PSMA) vaccine         dendritic cell processing. (89)
Anti-CEA vaccine        Dendreon Corp., Seattle,    RCC, TCC, CAP,
                          WA                         and other

Vitravene (fomivirsen)  Isis Pharmaceuticals,       CMV retinitis in
                          Inc., Carlsbad, CA         patients with
                                                     AIDS

Angiozyme               Ribozyme                    Colon and breast
                          Pharmaceuticals, Inc.,     cancers
                          Boulder, CO, and
                          Chiron Corp.,
                          Emeryville, CA

Onyx-015                Onyx Pharmaceuticals,       Head and neck,
                         Inc., Richmond, CA,         pancreatic,
                         and XOMA (US)               colorectal, and
                         LLC, Berkeley, CA           lung cancers

Generx (Ad5FGF4)        Schering AG, Berlin,        Stable coronary
                          Germany                     artery disease

Anti-CEA vaccine        I                   Cellular
                                              cancer
                                              vaccine

Vitravene (fomivirsen)  M                   Antisense

Angiozyme               II                  Antisense

Onyx-015                III                 Gene/viral
                                              therapy

Generx (Ad5FGF4)        II/III              Gene
                                             therapy

Anti-CEA vaccine        Patient T cells modified in vitro with anti-
                          CEA immunoglobulin to serve as T cell
                          receptors and reinfused in patients with
                          CEA plus tumors. (c)

Vitravene (fomivirsen)  Antisense CMV inhibitor complementary to
                         mRNA of major immediate-early region
                         proteins of CMV. It was found to be a
                         potent and selective antiviral agent for
                        CMV retinitis. (90), (91)

Angiozyme               Inhibits the VEGF receptor, a key component
                          in angiogenesis. This ribozyme is a
                          catalytic RNA molecule that downregulates
                          VEGF receptor function by specifically
                          cleaving the mRNAs for the primary
                          VEGF receptors Flt-1 and KDR. (92)

Onyx-015                Recombinant adenovirus engineered to
                          replicate only in cells expressing a p53
                          mutation. The virus carries a loss-of-
                          function mutation that disables inactivation
                          of tumor suppressor protein p53. Thus, in
                          normal cells, p53 accumulates and inhibits
                          viral replication, whereas cancer cells with
                          a p53 mutation allow viral replication and
                          cell death. (72), (93) (Note: Drug development
                          discontinued in January 2003 pending
                          developers' ongoing partnership
                          discussions.)

Generx (Ad5FGF4)        Adenovirus vector containing human
                         fibroblast growth factor-4 planned to
                         trigger the production of new blood vessel
                         growth. (94)

(a) TCC, transitional cell carcinoma; RCC, renal cell carcinoma; CAP,
carcinoma of prostate; CEA, carcinoembryonic antigen; CLL, chronic
lymphatic factor; AML, acute myelogenous leukemia; MM, multiple
myeloma; GM-CSF, granulocyte monocyte colony stimulating
factor; mRNA, messenger ribonucleic acid; M,
approved drug; CMV, cytomegalovirus; VEGF,
vascular endothelial growth factor.
(b) Phase relates to clinical trial stage in the United States as of
April 2003. Some drugs may be in a more advanced clinical trial phase in
other countries.
(c) Unpublished data.

Acknowledgment

NJM is a recipient of a fellowship from The American Physicians Fellowship for Medicine in Israel.

Accepted January 21, 2003.

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RELATED ARTICLE: Key Points

* As a result of new biotechnological capabilities, the understanding of disease processes and the development of new treatments are expanding rapidly.

* Important tools and developments include genomics, proteomics, ligand-receptor interaction, signal transduction, rational drug design, biochips, and microarrays.

* There are several novel drug classes, each with its own structural architecture and mechanism of action, including monoclonal antibodies, cancer vaccines, gene therapy, antisense strands, enzymes, and proteins.

* A wealth of promising new drugs will enable better treatments for patients with cancer, autoimmune disease, neurologic disease, allergy, and transplant rejection, among other entities.

Yoav Avidor, MD, MBA, Nicola J. Mabjeesh, MD, PHD, and Haim Matzkin, MD

From Johnson & Johnson/Ethicon Endo-Surgery, Cincinnati, OH; The Winship Cancer Institute, Emory University School of Medicine, Atlanta, GA; and the Department of Urology, Sackler Faculty of Medicine, Tel Aviv Sourasky Medical Center, Tel Aviv, Israel.

Each of us contributed equally to this article. We have no commercial, proprietary, or financial interest in any drug, device, or equipment mentioned in this article.

Reprint requests to Nicola J. Mabjeesh, MD, PhD, Department of Urology, Tel Aviv Sourasky Medical Center, 6 Weizmann Street, Tel Aviv 64239, Israel. Email: [e-mail]

Copyright [c] 2003 by The Southern Medical Association

0038-4348/03/9612-1174