Current Medicinal Chemistry - Volume 9, Issue 24, 2002

One challenge associated with the clinical use of protein therapeutics destined for chronic administration is the potential for the development of unwanted anti-drug immune reactions. The molecular basis for this reactivity is the binding of peptide fragments (epitopes) derived from the breakdown of the protein drug to the HLA receptors expressed by the patient's immune cells. If these epitopes are recognized as “foreign” by the immune system, specific helper T lymphocytes (HTL), are activated, which initiate and direct the formation of antibodies against the protein drug. These antibodies can bind and neutralize the protein drug, resulting in either decreased efficacy or total ineffectiveness of the drug. Moreover, various safety concerns, such as allergic reactions and other adverse events, are also frequently associated with the formation of anti-drug antibodies. Herein, we describe the development of “ImmunoStealth”, an integrated bioinformatics, biochemical and cellular immunology approach that specifically addresses the issue of unwanted immune responses against protein therapeutics. Unwanted HTL epitopes are identified using in silico sequence analysis methods and high throughput in vitro biochemical evaluations and thereafter confirmed using cellular immunogenicity assays. The “offending” epitopes within the drug are then rationally modified to alter their HLA binding capacity, and thus render them non-recognizable by the immune system. This technology will ultimately facilitate the design of safer, more potent and more economical drugs.

Cytokines are powerful molecules that the body's immune cells secrete in response to an offending agent. Their main function is to direct the immune response into the most effective pathway that will eventually result in elimination of the offender. The last decade was marked by an enormous and ever growing interest that led to discovery of numerous cytokine molecules and their amazing influence on the body immune function. The more we are learning about the way cytokines modulate and direct the immune responses of the body, the interest in using them or their antagonist to change or enhance those responses is growing.Studies are currently underway showing the beneficial effect of TNFα antagonists on the cellular injury mediated by this cytokine in rheumatic diseases, inflammatory bowel disease and endotoxemia. Interferon therapies are also tested utilizing IFNα for treatment of Hepatitis B and C. The discovery of Th1 and Th2 cytokines had shown that the nature of the immune response is, in essence, directed by a few important cytokines. Which immune reactions will develop seems to depend on whether IL-2 and IL-12 are secreted (and the immune response becomes Th1 with secretion of IFNγ and efficient removal of some antigens such as viruses) or IL-4 is secreted in which case Th2 response results in down regulation of IFNγ and IL-2 secreting effectors.The discovery, isolation and purification of these molecules open the possibility to skew the immune response in order to facilitate better outcome. For example, studies have now being conducted aimed at using IL-2 as an adjuvant therapy in conjunction with HAART in HIV patients. Similarly, IL-12 seems to be beneficial in melanoma and has been used as a very potent adjuvant for eliciting immune responses to immunization. Furthermore, studies with IL-4 knockout mice and those utilizing IL-4 blocking agents have shown that this cytokine might play a crucial role in maintaining persistent viral infections and in mediating chronic, autoimmune diseases.Using body's own immunomodulators is becoming an exciting possibility to target inefficient or misdirected immune responses that result in disease. The potential benefits in terms of human disease are enormous and still largely unexplained. Thus, using cytokines and their antagonists as therapeutic agents is an emerging and growing area of research.

The most important natural sources of new leads are plant extracts, bacterial broths, animal venoms and peptides isolated from living organisms. However, only the three first have been used extensively in the development of new therapeutic agents. This is probably due to the low pharmacological profile exhibited by peptides, that requires a lengthy transformation to make them suitable as new leads. In contrast, bioactive compounds isolated from the other sources are regularly closer to be used as lead compounds. Nevertheless, the sources for compounds of this category are nowadays scarce. In contrast, there are new bioactive peptides discovered quite often and reported as ligands for different receptors. Under these circumstances peptides appear as an attractive source of prospective new leads. In order to reduce the time involved in the design of a potential lead from a peptide, molecular modeling tools have been developed in the last few years. The purpose of the present work is to review the different techniques available and to report various successful examples of design of new peptidomimetics published in the literature.

Diabetes mellitus refers to a spectrum of syndromes characterized by abnormally high levels of glucose in blood. These syndromes are associated with an absolute (Type 1 diabetes) or relative (Type 2 diabetes) deficiency of insulin, coupled with varying degrees of peripheral resistance to the actions of insulin. Clinical studies have shown that controlling hyperglycemia significantly reduces the appearance and progression of the vascular complications associated with diabetes.Insulin's regulation of glucose homeostasis is mediated by a cascade of signaling events that take place upon insulin binding to its cell surface receptor. Autophosphorylation of the receptor and activation of its intrinsic tyrosine kinase are critical processes for transmitting these intracellular signals. Type 1 diabetes patients depend on exogenous insulin to achieve these effects, whereas Type 2 diabetes patients can accomplish a similar response through oral medications that increase the production of endogenous insulin or enhance its actions on the target tissues. Current biochemical and clinical evidence suggests that defects within the insulin receptor itself may be a cause of insulin resistance leading to Type 2 diabetes.This review focuses on the insulin receptor as a target for therapeutic intervention, and describes the recent discovery of small molecules that act on the receptor and either enhance or directly emulate the actions of insulin both in vitro and in vivo.

Design and synthesis of metabolically stable peptide analogs that can either mimic or block the bioactivity of natural peptides or enzymes is an important constituent of bioorganic and medicinal chemistry research. Isosteric replacement of a scissile peptide bond represents a viable and popular approach in the rational design of peptidomimetics. Peptidomimetics find applications as drugs, in protein engineering and so on. This is evident from the wealth of therapeutically useful peptidomimetic leads incorporating any of the peptide isosteres that are currently available. In this review, we have given a brief account of the types of peptide isosteres widely known till date. With this background, we have described some of the recent developments in synthetic approaches. This includes methods involving a common intermediate to synthesize different possible isosteres and their peptide analogs, solid phase synthesis and combinatorial approach. One such method involving stereoselective nitrile oxide cycloaddition as the key step has been studied extensively in our research laboratory. Finally, we have also discussed about some of the recent reports on the design and inhibitory activities of peptidic or non-peptidic analogs against aspartic proteases (HIV-1, renin, ACE and pepsin) and peptide analogs of an immunomodulating hexapeptide.

The decision of stressed cells to die or to survive is made by integrating signals at different levels through multiple check points. However, initiation and continued progression toward cell death by apoptosis in cancer cells may be blocked by mutation of the tumor suppressor p53 or overexpression of members of the bcl-2 family of proteins. The existence of such mechanisms indicates that cancer cells lose the controls regulating their cell cycle. Therefore, the activation of their programmed cell death appears as a major therapeutic target. Oxidative stress can stimulate growth, trigger apoptosis, or cause necrosis depending upon the dose and the exposure time of the oxidizing agent. A large body of evidence supports the idea that oxidative stress induced by redox cycling of vitamins C and K3 in association surpasses cancer cellular defense systems and results in cell death. The molecular mechanisms underlying such a process are, however, still unknown. Indeed, several types of cell death may be produced, namely autoschizis, apoptosis and necrosis. Combined vitamin C and K3 administration in vitro and in vivo produced tumor growth inhibition and increased the life-span of tumor-bearing mice. CK3-treatment selectively potentiated tumor chemotherapy, produced sensitization of tumors resistant to some drugs, potentiated cancer radiotherapy and caused inhibition of the development of cancer metastases without inducing toxicity in the host. We propose the association of vitamins C and K3 as an adjuvant cancer therapy which may be introduced into human cancer therapy without any change in the classical anticancer protocols, and without any supplementary risk for patients.