Therapeutic AIDS vaccines.

Therapeutic HIV vaccines represent promising strategy as an adjunct or alternative to current antiretroviral treatment options for HIV. Unlike prophylactic AIDS vaccines designed to prevent HIV infection, therapeutic vaccines are given to already infected individuals to help fight the disease by modulating their immune response. The first immunotherapeutic trial in AIDS patients was conducted in 1983. Since then several dozen conventional therapeutic vaccine trials have been carried out. Unfortunately, the results have consistently shown that while HIV-specific immune responses were evident as a result of vaccination, the clinical improvement has been seldom observed. The instances of the apparent clinical benefit were invariably associated with unconventional vaccines that acted in accord with the principles of alloimmunization and/or autologous vaccination. All such vaccines were derived from the blood of HIV carriers or a cell culture and thus they inherently contained allo- or self-antigens unrelated to HIV. This intriguing observation raises the issue whether this clinically successful approach has been unduly neglected. The current strategy biased toward vaccines, which have shown little evidence of clinical efficacy, needs to be diversified and supplemented with research on alternative vaccine approaches geared toward immune tolerance induction.

Antioxidant-independent ascorbate enhancement of catecholamine-induced contractions of vascular smooth muscle.

Ascorbate reduces the oxidation rate of catecholamines and, by an independent mechanism, enhances rabbit aortic ring contractions initiated by catecholamines. The largest significantly different fractional increases in force produced by ascorbate enhancement of norepinephrine (NE), epinephrine, phenylpropanolamine (PPA), and ephedrine (Eph) are 5.5, 1.8, 1.6, and 1.3 times, respectively. In physiological salt solutions bubbled with 95% O(2) at 37 degrees C, NE, PPA, and Eph have oxidation rate constants of 1.24, 247, and 643 h, respectively. Ascorbate significantly enhances 100 nM NE contractions by at least twofold at all ascorbate concentrations >15 microM, including the entire physiological range of 40-100 microM. Ascorbate preloading and washout followed by NE exposure produces significantly greater contractions than NE without ascorbate preloading but significantly lower than NE simultaneously with ascorbate. Ascorbate does not enhance K(+)- or angiotensin II-induced contractions. Ascorbate enhancement of catecholamine contractions occurs in addition to the reduction in oxidation rate, because the increases in force occur faster than oxidation can occur, the increases occur with compounds that have negligible oxidation rates, and the increases occur when ascorbate and NE are not physically present together. These results are consistent with ascorbate acting on the adrenergic receptor. Ascorbate may play a role in shock and asthma treatments and potentiate the cardiovascular health consequences of PPA and Eph (Ephedra).

Insulin binds to glucagon forming a complex that is hyper-antigenic and inducing complementary antibodies having an idiotype-antiidiotype relationship.

We demonstrate using physico-chemical techniques that insulin binds to glucagon with a Kd of 0.89 micromolar. While such binding is of little significance physiologically, it has important immunological consequences. Hormone binding is mirrored by specific binding between insulin antibody and glucagon antibody to form idiotype-antiidiotype complexes observable by Ouchterlony immunodiffusion and ELISA. These complexes may provide new insights into the formation of circulating immune complexes in diabetes. The insulin-glucagon complex is hyper-antigenic, inducing antibody production at concentrations that do not elicit immune responses from the individual hormones. The resulting immune response is not primarily against the individual hormones, but against the complex. In fact, all so-called insulin antibodies tested (rabbit, guinea pig, mouse and human) show substantially higher affinity for insulin-glucagon complex than for insulin alone, suggesting that this complex is the primary antigen in most, if not all, cases. These results lead to several testable predictions, including the possibility that glucagon antibody will bind to insulin receptors to cause type 2 (antibody mediated) insulin resistance.

Natural electrophoresis of norepinephrine and ascorbic acid.

The electric field produced by cell membranes, extending only a few nanometers, is 1000 times stronger than the electric fields required to produce dissociation of molecular complexes. Using the complex formed by norepinephrine (NE) and ascorbic acid (AA), we have demonstrated the quantitative binding of AA to NE, the use of capillary electrophoresis to measure quantitative binding of nonelectrolyte complexes, the determination of a dissociation constant (Kd) from electric field-dissociation constants (Ke), and a model for natural dissociation of the NE-AA complex due to the electric field generated by a cell membrane. NE-AA dissociation constants show little effect of NE concentration or pH changes. NE-related compounds also bind AA: epinephrine > norepinephrine > tyrosine > histamine > phenylalanine. Serotonin does not bind AA. Phosphorylated AA and glucose also bind NE at 0.05 and 0.08 of the AA binding, respectively. Natural electrophoresis of molecular complexes allows compounds to travel through the body in a protected state and still be available for physiological activity upon reaching a membrane.

Toward a philosophy of cancer research.

Every theory and all research assumes, either explicitly or implicitly, a scientific philosophy. Making the philosophy of cancer research explicit is likely to improve our understanding both of what we know and what we still need to discover. The implications for cancer research of three different philosophies of science are therefore outlined here: 1) reduction-mechanism; 2) holism; and 3) complementarity. We show that each of these philosophies leads to a different notion of causation, a different expectation of what represents a valid explanation of cancer, and thus to different problems that are addressed by different types of experiments. We conclude that the development of an appropriate philosophy of science is not only a relevant but necessary element in research on carcinogenesis.

A simple stochastic model of development and carcinogenesis.

Game theory can be a powerful tool for generating testable hypotheses concerning biological systems. We present a simple game that has many features analogous to a developing cellular system. The game mimics random turning on of genes in cells. Despite the randomness explicit in the mechanism, the game nonetheless results in deterministic outcomes that are extremely resistant to perturbation. Analysis of the types of mistakes or rule changes that are necessary in order to alter the outcome of the game suggests that there are a very limited number of mechanisms by which the differentiation process can result in tumor formation or carcinogenesis. The most significant causes of altered outcomes are alterations in gene order or number, and alteration of the rule by which the gene sequences are traversed. These alterations correspond to chromosomal defects or rearrangements, changes in chromosomal number, and changes in the "orders" delivered by regulatory genes. Notably, most common mistakes, which correspond to simple forms of mutations, have no effect on the outcome of the game, suggesting that mutation is relevant to tumorigenesis and carcinogenesis only to the extent that it results in altered "rules" for reading other gene sequences.

Complementarity and contradiction in cancer research: the role of hierarchies in carcinogenesis.

The essays on carcinogenesis in this volume tend to fall into groups according to the level of complexity at which the theories focus. Many investigators focus on events and processes involving genetic mutations. Others, however, identify events or processes at the chromosomal, cellular, tissue, or organsimal level as being of primary importance. While such different approaches to carcinogenesis may, at first, seem to be contradictory, they may be seen to be complementary in light of the general theory of hierarchical organization. Hierarchy theory, which is a development of general systems theory, describes the manner in which organized processes are formed by subunits that have unique properties of their own but which can acquire new properties through their interactions. Hierarchy theory suggests that carcinogenesis can only be understood as a set of interactions between organizational processes at every level from genetic to organismal.

Antisense peptides: a critical mini-review.

Antisense peptides are defined as those generated from the non-coding strand of DNA, and represent a peptide analog to antisense RNA technologies. Peptides generated from both parallel and anti-parallel readings of the non-coding strand of DNA have displayed biological activity, although considerable controversy exists concerning the mechanism(s) by which these "anti-peptides" exert their effects. This paper provides a critical review of some of the key data and issues defining this emerging field and focuses on contradictions and discrepancies in the current studies. We also suggest some directions for future research such as more physico-chemical studies and the use of combinatorial chemistry techniques combined with solid phase binding studies to test, once and for all, the generality and specificity of antisense peptide interactions.

Augmentation of aortic ring contractions by angiotensin II antisense peptide.

Previous biochemical experiments have revealed two antisense peptide antagonists to human angiotensin II (Ang II), one encoded in the cDNA in the antiparallel reading, the other in the parallel reading. Neither peptide's ability to produce physiological antagonism has been demonstrated previously. Both peptides were tested for their ability to antagonize Ang II-induced contractions on rabbit aorta smooth muscle. Neither peptide had any direct contractile activity. The antiparallel Ang II peptide had physiological antagonism to Ang II contractions at a lower sensitivity than reported in biochemical studies, and its antagonist activity was partially blocked by Ang II antiserum, suggesting that it is not an antipeptide but an Ang II homologue. The parallel Ang II antipeptide also required high concentrations for physiological inhibition. Its contractile inhibition was not affected by Ang II antiserum and diminished the Ang II contraction at high micromolar concentrations, findings consistent with physicochemical data showing that it is an Ang II complement. The concentration of either peptide required to produce an antagonistic physiological effect was too high to predict any pharmacological usefulness. The parallel antipeptide, however, significantly increased the force of muscle contractions at high nanomolar concentrations, thus displaying a unique dual augmentation/antagonist activity. This antipeptide seems to have highly sequence-specific activity because other similar parallel antipeptides had no activity. The parallel antipeptide augmentation mimics the shift in the Ang II dose-response curve produced in hypertension studies of the slow pressor effect of Ang II and may be useful in deducing the currently unknown cause of the slow pressor effect. It may also have some uses in migraine studies.

The necessity of cofactors in the pathogenesis of AIDS: a mathematical model.

Current arguments for the role of cofactors in the initiation of a chronic HIV infection and progression of AIDS are given. The natural history of an HIV infection as affected by cofactors which provide additional stimulatory signals is explored through a mathematical model. The model demonstrates that "antigen load" plays a role in determining susceptibility to an HIV infection. It also suggests that certain individuals may not be able to be infected by small doses of HIV and that the identification and treatment of existing cofactors may be useful in treating early stages of HIV infection. Prevention of cofactor exposures may also protect against HIV infection.

Molecular complementarity I: the complementarity theory of the origin and evolution of life.

We assert that molecular complementarity is much more widespread than is commonly acknowledged in biological systems, if not actually ubiquitous. It creates the coupling necessary for non-equilibrium systems to form. It stabilizes aggregates against degradation, thereby increasing concentrations to levels adequate to foster the formation of prebiotic systems and represents the earliest form in which natural selection was manifested. Complementarity confers on all interacting parts of such systems in formation carrying capacity. RNA or DNA are not, therefore, necessary to the emergence of life, but represent specialized forms of complementary molecules adapted specifically to information storage and transmission. Non-genetic information exists in metabolic functions and probably preceded genetic information historically. Complementarity also provides the basis for homeostasis and buffering of such systems not only in a chemical, but also in structural and temporal terms. It provides a mechanism for understanding how new, emergent properties can arise, and a basis for the self-organization of systems. We demonstrate that such aggregates can have properties not predictable from their individual components, thus providing a means for understanding how new functions emerge during evolution. Selection is for modules rather than individual components. The formation of functional sub-systems that can then be integrated as modules greatly increases the probability of the emergence of life. The result of such modular evolution alters the standard view of evolution from a tree or bush-like image to an integrated network composed of alternating periods of integration (as molecules and molecular aggregates merge) and divergence (as molecules and aggregates undergo variations). This provides a mechanism for evolution by punctuated equilibria. Molecular complementarity puts strict limits on variations, however, preventing evolution from being random. The evolutionary, physiological and embryological consequences of this view of life are outlined, and various models and experiments described that further characterize it.

Molecular complementarity II: energetic and vectorial basis of biological homeostasis and its implications for death.

The energetic basis of molecular complementarity is presented. In biological systems requiring both homeostasis and non-equilibrium state maintenance, molecular complementarity provides a framework for the co-existence of these states within an organism. Smoothly changing homeostatic and thermodynamic systems, such as regulation of pH or an ensemble of asynchronous muscle crossbridges, are modeled using Liapunov functions. When biological systems undergo discontinuous state changes, such as the initiation of the heartbeat, life/death transition or the detachment of molecules, alternative analytical systems such as catastrophe theory provide information that continuous analytical methods cannot. Catastrophe theory produces a model of biology in which death can occur by two distinct mechanisms: loss of homeostatic control or loss of sufficient free energy. Molecular complementarity buffers molecules from temporal and physical changes. The usefulness of molecular complementarity is limited to association energies near the ambient energy, kT. Within this range, complementarity will alter molecular functions and will convert scalar biochemical reactions into vectorial physiological processes. Both thermodynamic and catastrophic models can be used to link energetic and homeostatic processes: the former providing quantitative information from continuous systems; the latter providing qualitative information from discontinuous systems involving state changes.

Dental and research transmission of acquired immune deficiency syndrome? Or, is there any evidence that human immunodeficiency virus is sufficient to cause acquired immune deficiency syndrome?

An unpublished report of three laboratory workers exposed to concentrated human immunodeficiency virus cultures and one anomalous case of putative dental transmission to six patients represent the only evidence of acquired immune deficiency syndrome developing in the absence of immunologic cofactors such as exposure to sexually transmitted diseases, drugs, malnutrition, and alloantigens. A review of the Centers for Disease Control (USA) files shows that all six dental patients had identified risks for human immunodeficiency virus infection other than dental treatment, and evidence of cofactor exposures. Several demonstrated immunologic abnormalities prior to dental treatment. These data were ignored by the Centers for Disease Control. Furthermore, known rates of human immunodeficiency virus transmission by blood clotting factor use (1 infection per 20 exposures) and percutaneous exposure (1 per 250) make the number of patients the dentist would have had to expose to get six infections unrealistic (circa 120-1500). Thus, no clear-cut evidence exists for human immunodeficiency virus infection in the absence of predisposing immunologic cofactors.

Five myths about AIDS that have misdirected research and treatment.

A number of widely repeated and factually incorrect myths have pervaded the AIDS research literature, misdirecting research and treatment. Five of the most outstanding are: 1) that all risk groups develop AIDS at the same rate following HIV infection; 2) that there are no true seroreversions following HIV infection; 3) that antibody is protective against HIV infection; 4) that the only way to treat AIDS effectively is through retroviral therapies; and 5) that since HIV is so highly correlated with AIDS incidence, it must be the sole necessary and sufficient cause of AIDS. A huge body of research, reviewed in this paper, demonstrates the falsity of these myths. 1) The average number of years between HIV infection and AIDS is greater than 20 years for mild hemophiliacs, 14 years for young severe hemophiliacs, 10 years for old severe hemophiliacs, 10 years for homosexual men, 6 years for transfusion patients of all ages, 2 years for transplant patients, and 6 months for perinatally infected infants. These differences can only be explained in terms of risk-group associated cofactors. 2) Seroreversions are common. Between 10 and 20 percent of HIV-seronegative people in high risk groups have T-cell immunity to HIV, and may have had one or more verified positive HIV antibody tests in the past. 3) Antibody, far from being protective against HIV, appears to be highly diagnostic of loss of immune regulation of HIV, and some evidence of antibody-enhancement of infection exists. 4) Non-retroviral treatments of HIV infection, including safer sex practices, elimination of drug use, high nutrient diets, and limited reexposure to HIV and its cofactors have proven to be effective means of preventing or delaying onset of AIDS. 5) Many immunosuppressive factors, including drug use, multiple concurrent infections, and exposure to alloantigens, are as highly correlated with AIDS risk groups as HIV. These data are more consistent with AIDS being a multifactorial or synergistic disease than a monofactorial one.

Semen alloantigens and lymphocytotoxic antibodies in AIDS and ICL.

More than 90% of people with AIDS develop circulating immune complexes (CICs) and lymphocytotoxic antibodies (LCTAs). Animals infected with HIV, however, never display CICs or LCTAs, and remain healthy. Similarly, HIV-infected people who do not develop CICs or LCTAs also do not progress to AIDS. The appearance of CICs and LCTAs is, however, highly prognostic for AIDS and death. Since HIV infection does not, per se, lead to the development of CICs and LCTAs, other causes are likely. One such cause, for which both epidemiologic and experimental evidence exists, is semen. Semen components include sperm, seminal fluid, lymphocytes, and sometimes infectious agents, including HIV, mycoplasmas, and herpes and hepatitis viruses, all of which independently cause immune suppression. Extensive evidence demonstrates sperm (and various viruses) contains many proteins mimicking the CD4 protein of T-helper cells, while HIV, mycoplasmas, and seminal fluid mimic class II MHC proteins of other lymphocytes. We identify a large number of protein sequences that display such mimicry using computer homology searching, and demonstrate experimentally that sperm antibodies specifically precipitate antibodies against class II MHC mimics such as mycoplasmas, which in turn precipitate antibodies to lymphocyte antigens. These data prove that immunologic exposure to sperm and lymphocytes (as may occur in receptive anal intercourse, needle sharing, or blood transfusions) is theoretically capable of initiating lymphocytotoxic autoimmunity. Such autoimmunity may play a significant role in the pathogenesis of AIDS, and will need to be addressed clinically in high risk individuals regardless of HIV status and regardless of the success of anti-HIV prophylaxis and treatment.

Preliminary evidence for idiotype-antiidiotype immune complexes cross-reactive with lymphocyte antigens in AIDS and lupus.

Several investigators have proposed that autoimmunity may be induced by idiotype-antiidiotype antibody networks. It is generally assumed that the antiidiotype is produced in response to the idiotype, and therefore that autoimmune diseases have single antigenic initiators. The theory of multiple-antigen-mediated autoimmunity (MAMA) proposes, on the other hand, that idiotype and antiidiotype result from two primary immune responses to two chemically complementary antigens. Because of the complementarity of the antigens, and the complementarity of the antibodies for the antigens, the antibodies will themselves be complementary. They will thus form circulating immune complexes, the self-nonself distinction diffusion (DAD) experiments (a modification of Ouchterlony immunodiffusion), in which 1800 pairs of antibodies were screened for their ability to form precipitating complexes. Four sets of antibodies associated with AIDS (HIV + Staphylococcus; HIV + Mycoplasma; CMV + Mycoplasma; and HBV + Mycoplasma) specifically precipitated each other, and one of the antibodies in each set also precipitated monoclonal antibodies against one or more lymphocyte protein markers. These results therefore demonstrate that idiotype-antiidiotype antibodies can be elicited by independent antigens and may induce AIDS-related forms of autoimmunity directed at lymphocytes.

CD4 similarity to proteins of infectious agents in AIDS and their role in autoimmunity.

Lymphocytotoxic autoimmunity (LA) is ubiquitous in AIDS. Its causes are unknown. We report that significant amino acid sequence similarities exist between the proteins of infectious organisms associated with AIDS and the CD4 protein of T-helper lymphocytes. These included: HIV, cytomegalovirus (CMV), Epstein-Barr virus (EBV), herpes simplex viruses (HSV), Varicella Zoster virus (VZV), Escherichia coli, Mycobacteria, Mycoplasmas, Plasmodium, and Staphylococcus. It has been reported previously that HIV proteins have significant similarities with human class II MHC (HLA class II) proteins. Since CD4 and HLA class II proteins are chemically complementary, pairs of homologous antigens will also be complementary. It follows that concurrent infections with CD4 and HLA class II-homologous antigens will result in idiotype-antiidiotype antibody pairs that cannot distinguish 'self' from 'nonself', that acts as lymphocytotoxins, and form circulating immune complexes. Thus, combined HIV-CMV, HIV-EBV, HIV-HBV, HIV-mycoplasma, or other appropriate infectious pairs may suffice to trigger LA in AIDS.