Proteasome inhibition with bortezomib: an effective therapy for severe antibody mediated rejection after renal transplantation.

Antibody-mediated rejection (AMR) following renal transplantation is less responsive to conventional anti-rejection therapies. Plasmapheresis (PP), intravenous immunoglobulin (IVIg), rabbit antithymocyte globulin (rATG) and rituximab deplete immature B-cells but not mature plasma cells. The proteasome inhibitor bortezomib has activity against mature plasma cell, the source of damaging donor-specific antibody (DSA).We present the successful use of bortezomib in 2 patients who developed AMR following kidney transplantation. The first patient was a 54-year-old white female who received living-unrelated kidney transplantation from her husband. She developed severe AMR early after transplantation with rising DSA titers consistent with an anamnestic immune response by memory cells to the donor antigens. Renal function deteriorated despite treatment with pulse methylprednisolone (MP), PP and IVIg. After initiation of therapy with bortezomib, DSA titers became negative and serum creatinine returned to baseline with histological resolution of AMR. The second patient was a 19-year-old white male who received deceased donor kidney transplantation and developed AMR within 2 weeks, refractory to therapy with pulse MP, PP and IVIg with rising DSA. Bortezomib use resulted in disappearance of DSA and renal function improvement. Both patients tolerated the treatment well with stable renal function at last follow-up. The novel mechanisms of action and preliminary results with bortezomib are encouraging, but require larger studies and longer follow-up.

Effects of assertiveness training and expressive writing on acculturative stress in international students: A randomized trial.

International university students often experience acculturative stress, and culturally appropriate techniques to manage stress are needed. This randomized trial tested the effects of group assertiveness training, private expressive writing, their combination, and a wait-list control on the acculturative stress, affect, and health of 118 international students at an urban North American university. Interventions were conducted at the start of a semester, and assessments were conducted at baseline and at the end of the semester. Group assertiveness training was rated positively by students and led to lower negative affect, whereas expressive writing was less well received and led to higher homesickness and fear, but also to higher positive affect. The combined intervention had no effects, perhaps because the 2 components negated each other. It is concluded that group assertiveness training improves emotional adjustment of international students but that expressive writing has mixed effects and needs further development and study. (PsycINFO Database Record

2-deoxy-D-glucose as a potential drug against fusogenic viruses including HIV.

Most enveloped viruses fuse with host cells and catalyze fusion among host cells by expression of specific patterns of N-glycosylation on their envelope proteins. In the 1970s, it was observed that 2-deoxy-D-glucose (2DOG) and 2-fluoro-2-deoxy-D-mannose (2F2DOM) inhibited N-glycosylation of asparagine (Asn) sites on the external domain of viral envelope proteins. This effect led to the virus particles being non-fusogenic with greatly reduced infectivity and reduced ability to pass from cell to cell by catalyzing cell--cell fusion. At that time, this observation was not particularly important because viral diseases were readily prevented by vaccines and there was no known link between fusogenic viruses and cancer. Today, we are faced with a chronic and lethal viral disease (AIDS) caused by a virus (HIV) that mutates so quickly that we have not been able to produce a vaccine. Moreover, it is spreading among millions of people unable to afford more than basic medications. In addition, cell--cell fusion has been identified as an important, if not essential, step in the progression of abnormal cell clones to clinically significant cancer and fusogenic viruses have been shown to cause progression of some tumors. Here, we reiterate the hypothesize (first made in 1986 by Blough et al. [Blough HA, Pauwels R, De Clercq E, Cogniaux J, Sprecher-Goldberger S, Thiry L. Glycosylation inhibitors block the expression of LAV/HTLV-III (HIV) glycoproteins. Biochem Biophys Res Commun 1986; 141:33-8]) that 2DOG, 2F2DOM and related compounds, which interfere with normal N-glycosylation of virus envelope proteins, are attractive candidates for anti-fusogenic drugs that can be used against chronic virus diseases and cancers. This analysis also supports the concept of blocking N-acetylglucosaminyl-transferases with chloroquine or other drugs (proposed by Savarino et al. [Savarino A, Lucia MB, Rastrelli E et al. Anti-HIV effects of chloroquine: inhibition of viral particle glycosylation and synergism with protease inhibitors. J Acquir Immune Defic Syndr 2004; 35:223-32]) as an anti-viral approach. These drugs may have broader utility and lower cost than drugs designed specifically to target the gp41 protein of HIV, which have become popular as viral-entry inhibitors for treatment of HIV/AIDS.

AIDS: caused by development of resistance to drugs in a non-target intracellular parasite.

The origin of acquired immune disorder syndrome (AIDS) has been the subject of substantial controversy both in the scientific community and in the popular press. The debate involves the mode of transmission of a simian virus (SIV) to humans. Both major camps in the argument presume that humans are normally free of such viruses and assume that once the simian virus was transmitted, it immediately infected some T-cells and caused the release of toxic agents that killed off bystander (uninfected) T-cells resulting in AIDS. The evolution of the Simian virus (SIV) into a human virus (HIV) is regarded as an artifact. In contrast, a fundamentally different hypothesis has been proposed [Parris GE. Med Hypotheses 2004;62(3):354-7] in which it is presumed that in hyper-endemic areas of malaria (central Africa), all primates (humans and non-human primates) have shared a retrovirus that augments their T-cell response to the malaria parasite. The virus can be called "primate T-cell retrovirus" (PTRV). Over thousands of years the virus has crossed species lines many times (with little effect) and typically adapts to the host quickly. In this model, AIDS is seen to be the result of the development of resistance of the virus (PTRV) to continuous exposure to pro-apoptotic (schizonticidal) aminoquinoline drugs used to prevent malaria. The hypothesis was originally proposed based on biochemical activities of the aminoquinolines (e.g., pamaquine (plasmoquine(TM)), primaquine and chloroquine), but recent publications demonstrated that some of these drugs definitely adversely affect HIV and other viruses and logically would cause them to evolve resistance. Review of the timeline that has been created for the evolution of HIV in humans is also shown to be qualitatively and quantitatively consistent with this hypothesis (and not with either version of the conventional hypothesis). SARS and Ebola also fit this pattern.

Why G3139 works poorly in cancer trials but might work well against HIV.

The antisense drug G3139 (oblimersen sodium, Genta, Inc.) is a phosphorothioate oligodeoxynucleotide (ODN) containing unmethylated CpG units, which is targeted to suppress Bcl-2. To date, its effectiveness in cancer clinical trials has been minimal. Some suggestions are provided for that disappointment and recent citations are provided that support the idea that G3139 may be effective at clearing viral infections, specifically HIV. At the time G3139 was conceived as an anti-cancer drug candidate, it was viewed optimistically because Bcl-2 was widely believed to be the most important protein blocking p53-dependent apoptosis caused by internal stress. Since that time, we have learnt that Bcl-2 is not the only protein that inhibits apoptosis and that p53 itself is frequently malfunctioning in tumors. Thus, the anti-cancer utility of suppressing Bcl-2 in cancer cells is limited. Moreover, Bcl-2 has a role in halting the cell cycle (though p27), which may slow down tumor growth; and Bcl-2 even has pro-apoptotic roles in the execution of apoptosis initiated by external death signals (via Fas/CD95 and caspase 3). Overall, in the clinical setting, G3139 usually has statistically significant but medically unimportant benefit. These results have greatly diminished the enthusiasm for the drug especially when the side effects are considered. Specifically, the unmethylated CpG ODN (and/or the phosphorothioate group) activates the immune system, but this potentially important anti-cancer effect is lost when the immune cells undergo premature apoptosis apparently because their Bcl-2 levels have been lowered by the antisense effect of G3139. While this effect on immune cells is usually undesirable, it is exactly what would be useful for activating immune cells, initiating provirus transcription in retrovirus-infected cells, and facilitating selective apoptosis of these infected cells. In general, G3139 might have benefit in clearing chronic infections by intracellular parasites including viruses (HIV, SIV, HTLV, HBV, coronavirus, etc.). Indeed, G3139 has been shown to cause apoptosis in EBV-infected cells leading to clearance of the virus.

Mechanism and history of evolution of symbiotic HIV strains into lethal pandemic strains: the key event may have been a 1927 trial of pamaquine in Leopoldville (Kinshasa), Congo.

In previous papers, I have rejected both the zoonosis and the serial transfer hypotheses of the origin and evolution of the current lethal pandemic strains of HIV. The hypothesis that fits the critical observations is that all the human and nonhuman primate species in central Africa (an area of hyper-endemic malaria) have shared (through inter-species transfers) a "primate T-cell retrovirus" (PTRV), which has adapted to each host species. This retrovirus is believed to assist primate T-cells attack the liver stage of the malaria infection. Each geographic region has a dominant primate host and a characteristic virus. Starting in 1955 and continuing into the late 1970s, chloroquine was provided by the WHO and used for prophylaxis against malaria. Chloroquine has a number of biochemical activities but two of the most important are blocking transcription of cellular genes and proviruses activated by NF-kappaB and blocking the glycosylation of surface proteins on viruses and cells. Concurrent with the development of resistance of the malaria parasite to chloroquine, HIV strains were quickly selected, which have enhanced transcription rates (by inclusion of multiple kappaB binding sites in their long terminal repeats by recombination) and enhanced infectivity (fusogenicity) (most likely by mutations in multiple viral genes that regulate glycosylation of Env). There also may have been mutations that enhanced activation of NF-kappaB in the host cell. These changes in the retrovirus genome were not manifest in effects of the HIV strains as long as the hosts were under the influence of chloroquine. But, when the virus infects people who are not protected by chloroquine, the virus multiplies more rapidly and is more communicable. Fortunately, most of these strains (i.e., HIV-2 groups, and HIV-1 O and HIV-1 N) self-regulate (i.e., infected cells kill infected cells) well enough that viral loads remain subdued and bystander cells of the immune system are not excessively attrited. In the case of HIV-1 group M, however, there is more going on. Following the work of Korber et al. on the phylogenetics of HIV-1 groups M, I reach the conclusion that the major subgroups giving rise to the worldwide pandemic, were founded in a 1927 clinical trial of pamaquine (plasmoquine) in Leopoldville (Kinshasa). This drug is much more toxic that chloroquine and appears to have strongly selected for resistance to apoptosis in infected cells, which allows these subgroups to attrite bystander cells leading to AIDS.

The cell clone ecology hypothesis and the cell fusion model of cancer progression and metastasis (II): three pathways for spontaneous cell-cell fusion and escape from the intercellular matrix.

The two-stage initiation-progression model of cancer is widely accepted. Initiation appears to result most often from accumulation of damage to the DNA expressed as multiple mutations in the phenotype. Unsymmetrical chromosome segregation during mitosis of normal or mutated cells produces aneuploid cells and also contributes to the evolution of neoplasia. However, it has been pointed out (Parris GE. Med Hypotheses 2005;65:993-4 and 2006;66:76-83) that DNA damage and loss of chromosomes are much more likely to lead the mutant clones of cells to extinction than to successful expansion (e.g., an example of Muller's Ratchet). It was argued that aneuploid neoplasia represent new parasite species that successfully evolve to devour their hosts by incorporating sex-like redistribution of chromosomes through spontaneous or virus-catalyzed cell-cell fusion into their life-cycle. Spontaneous cell-cell fusion is generally blocked by the intercellular matrix to which the cells are bound via surface adhesion molecules (frequently glycoproteins, e.g., CD44). In order for progression of matrix-contained neoplasia toward clinically significant cancer to occur, the parasite cells must escape from the matrix and fuse. Release from the matrix also allows the parasite cells to invade adjacent tissues and metastasize to remote locations. Both invasion and metastasis likely involve fusion of the migrating parasite cells with fusion-prone blast cells. There are at least three pathways through which parasite cells can be liberated from the confining matrix: (i) Their adhesion molecules may be modified (e.g., by hyper-glycosylation) so that they can no longer grip the matrix. (ii) Their adhesion molecules or matrix may be saturated with other ligands (e.g., polyamines). (iii) Their adhesion molecules may be cleaved from the cell surface or the matrix itself may be cleaved (e.g., by MMPs or ADAMs). It is hypothesized that mobilization of parasite cells and cell-cell fusion go hand-in-hand in the progression of neoplasia to clinically significant cancer through invasion and metastasis. The latency between tumor recognition and exposure to mutagens and the increased incidence of cancer with age can probably be related to slow breakdown of the intercellular matrix that provides a barrier to cell-cell fusion.

The cell clone ecology hypothesis and the cell fusion model of cancer progression and metastasis: history and experimental support.

The two-stage initiation-progression model of cancer is widely accepted. Although mutations explain initiation of neoplasia, the assumption that mutations are responsible for progression of neoplasia to cancer appears to have little experimental support. The "cell clone ecology hypothesis" explains why neoplasia evolve and the "cell fusion model of cancer progression and metastasis" describes how they evolve into clinically significant tumors. A brief history of important concepts and experiments is provided. Clinically significant cancers are effectively new parasite species that live, expand and evolve within the host. It is hypothesized that survival and fate of the parasite clones called "cancer" are governed by the principles of ecology. It is argued that while mutations or aneuploidy (asexual reproduction) can result in transient/self-limiting neoplasia, neither of these asexual modes of forming new karyotypes can maintain the ecologically fit parasites that develop into clinically significant cancer. Mutations and/or unstable genomes (aneuploidy) progressively degrade cell lines and if only these mechanisms were at work, neoplasia would spontaneously become extinct or benign (enfeebled) before reaching clinical significance (an example of "Muller's ratchet"). In the cell fusion model of (clinically significant) cancer progression and metastasis, cell-cell fusion is the essential element allowing normal cells or (transient) neoplasia to evolve into clinically significant cancers. Cell-cell fusion is required for producing and sustaining clinically significant cancer because it provides a sex-like mode of reproduction essential for an ecologically fit parasite organism. Cell-cell fusion provides the opportunity needed for tumors to rejuvenate cell lines containing abnormal genomes and rapidly evolve to acquire dramatically aggressive traits such as metastasis. Indeed, metastasis appears to require cell-cell fusion. Cell-cell fusion also partially overcomes erosion of teleomeres during clone expansion and allows the essential elements of a tumorigenic genome to hide from chemotherapy as recessive traits in cells with normal phenotypes and re-emerge (by cell-cell fusion) as a new cancer after the phenotypically cancerous cells have been eradicated by classical chemotherapy. Eradication of the cancer parasite cannot be routinely achieved by classical toxic chemotherapy alone or even by chemotherapy augmented with techniques needed to overcome anti-apoptotic traits of cancer cells. Direct chemical intervention against cell-cell fusion concurrent with classical toxic chemotherapy holds a promise of preventing re-lapse of the disease. Intervention against cell-cell fusion may also directly suppress metastasis based on the model presented here. The paper also summarizes work on the cell surface glycoprotein CD44 that implicates it as a key element in cell-cell fusion and hence cancer.

The role of viruses in cell fusion and its importance to evolution, invasion and metastasis of cancer clones.

The hypothesis described here is a logical extension of two areas of observation: First, it has been discovered that viruses (and perhaps other intracellular parasites) catalyze cell fusion as a means of cell-to-cell transmission. Effective cell-to-cell transmission appears to require: (i) induced expression of adhesion molecules on the cell surface; (ii) suppression of p53-dependent apoptosis; (iii) arrest of the cell cycle that would otherwise lead to cell death by "mitotic catastrophe". Suppression of apoptosis and cell death through "mitotic catastrophe" are important for formation of stable syncytia. Expression of Bcl-2 or a viral analogue of Bcl-2 (vBcl-2) is particularly useful to viruses because Bcl-2 both suppresses (p53-dependent) apoptosis and arrests the cell cycle through p27. Bcl-2 may also block any p53-independent cell death (e.g., mitotic catastrophe) that is initiated at the mitochondria. Second, it has been found that cell fusion plays a role in cancer clone evolution, invasion of normal cells in tissue adjacent to tumors and metastasis to remote normal tissues. Thus, it can be hypothesized that infection of cancer cells with viruses that spread by cell-to-cell transmission may coincidentally contribute to development of aggressive aneuploid clones and facilitate both invasion and metastasis of tumors. Regardless of the role of viruses, suppression of Bcl-2 may be an approach to preventing successful formation of syncytia and limiting the invasion and metastasis of tumors, thus, making surgical removal and radiation treatment more feasible.

Clinically significant cancer evolves from transient mutated and/or aneuploid neoplasia by cell fusion to form unstable syncytia that give rise to ecologically viable parasite species.

Following the idea of Duesberg and Rasnick (Cell Motil Cytoskeleton 2000; 47:81-107) that cancer is a separate species of organism, the ecology of cancer as a parasite is examined. The most important ecological feature of cancer is its ability to evolve. The mutation hypothesis and the "unstable genome" hypothesis of cancer evolution are considered but neither of these current hypotheses is believed to adequately explain how cancer successfully evolves. In particular, either of these processes alone should lead to extinction of the cell line before a clinically significant neoplasm is achieved. Moreover, the term "unstable genome" probably should be replaced by "labile genome" because cancer genomes must be stable enough to reproduce themselves through many generations if the clone is to expand. The key step in productive evolution of undetectable neoplasia into clinically significant cancer is hypothesized to be sex-like resorting of chromosomes from different cells (e.g., normal and abnormal cells). The sex-like process begins with cell fusion to form a syncytium, which may be stable (producing multinucleated giant cells seen in many tumors) or which may undergo "mitotic catastrophe" to produce polyploidy cells. The nuclei of polyploid cells may undergo a process called "neosis" in which they form buds and undergo karyokinesis followed by cytokinesis to yield karyoplasts (small cells with little cytoplasm) that found new cancer clone lines. Although both mutations and unstable (aneuploid) genomes are seen as dead ends in cancer evolution (i.e., using only these modes of genome modification, cancers would not likely advance to clinical significance before becoming extinct), they each produce transient genetic material, which can be incorporated into stable genomes with aggressive (i.e., ecologically fit) phenotypes by cell fusion. It is proposed that inhibition of cell fusion (or other steps in this sex-like process) concurrent with classical chemotherapy might prevent evolution of the clones and recurrence of the cancer. Similarly, active suppression of viruses or other conditions that catalyze cell fusion should also slow down evolution of cancer clones.

The possible use of arsine (AsH3) as therapy for malaria.

Arsine has long been recognized as a selective toxin that targets the membrane of erythrocytes and causes leakage of the internal contents of these cells. In acute poisoning episodes, the victims usually sustain kidney malfunction resulting from collection of debris from the erythrocytes. The hypothesis presented here is very simple. Namely, the hypothesis is to evaluate the use of arsine (AsH3) against erythrocytes that have become infected with malaria parasites. Once malaria parasites have become established in erythrocytes, it is extremely difficult to displace them. Erythrocytes do not spontaneously undergo apoptosis, which is a mechanism that many types of cells would use to deny intracellular parasites an immune privileged domain. Moreover, antibiotics that are powerful and selective enough to defeat malaria parasites once they are established in the erythrocytes have not been found. Thus, approaches for selective hemolysis of malaria-infected erythrocytes and evicting the parasites may be attractive. It is not known, but should be easily determined experimentally, what the relative toxicity of arsine (AsH3) is to normal erythrocytes and malaria-infected erythrocytes.

A Hypothesis Concerning the Biphasic Dose-response of Tumors to Angiostatin and Endostatin.

This manuscript proposes a hypothesis to explain the U-shaped dose-response observed for angiostatin and other high-molecular-weight drugs in various anti-cancer bio-assays. The dose-response curves for angiostatin and endostatin (measured as suppression of tumor growth) go through an optimum (i.e., minimum tumor growth) and then becomes less effective at higher doses. The literature suggests that at lower doses the primary action of these high-molecular-weight drugs is to counteract the angiogenic effects of vascular endothelial growth factor (VEGF). To do this, the drugs must pass out of the blood vessel and enter the extra-cellular matrix (ECM) where VEGF induces the growth and fusion of tip cells. Ironically, VEGF actually facilitates access of the drugs to the ECM by making the vascular endothelium leaky. At higher doses, the high-molecular-weight drugs seem to reverse VEGF-induced permeability of the endothelium. Thus, at high dose rates, it is hypothesized that the drugs are not able to enter the ECM and block the angiogenic effects of VEGF there. As a result, high doses of the drugs do not suppress vascularization of the tumor or tumor growth. Moreover, if the permeability of the vessels is suppressed, the VEGF released by the stroma is concentrated in the ECM where it amplifies the angiogenic activity around the tumor.

Hypothesis links emergence of chloroquine-resistant malaria and other intracellular pathogens and suggests a new strategy for treatment of diseases caused by intracellular parasites.

Chloroquine and related anti-malarial drugs appear to promote apoptosis in T-cells by suppressing NF-kappa-B, which enhances the expression of anti-apoptotic proteins (e.g., Bcl-2). Thus, chloroquine has found applications in autoimmune diseases where it apparently facilitates apoptosis of abnormally persistent T-cell clones. The mode of action of chloroquine in prevention of malaria is not known, but it may be to minimize replication of the parasite in the liver cells, which occurs before invasion of the erythrocytes, by facilitating premature apoptosis of the infected host cells. After introduction of chloroquine in the 1950s world-wide for prophylactic use, chloroquine-resistant malaria emerged. Here it is hypothesized that concurrent with emergence of chloroquine-resistant malaria (presumably with enhanced anti-apoptotic capabilities), other intracellular parasites have evolved to enhance their ability to prevent apoptosis in host cells. Two examples of viral diseases that have emerged from areas of high incidence of chloroquine-resistant malaria are AIDS from HIV and SARS from coronavirus. The hypothesis holds that prophylactic exposure to pro-apoptotic chloroquine drugs caused natural selection for strains of viruses and other parasites that have enhanced anti-apoptotic abilities. When transmitted to host organisms that are not under the influence of the pro-apoptotic drug, the new "anti-apoptotic" strains may cause unexpected diseases. In the case of SARS, the coronavirus appears to have accessed a new niche where it proves to be lethal to its host. In the case of AIDS, the HIV (which has had a long-term symbiotic relationship with primates) has run amuck because the infected cells are now substantially more tolerant to the toxins (i.e., resistant to apoptosis) that they secrete than the uninfected bystander cells, which are not unusually resistant to apoptosis. A corollary to the hypothesis is that if the level of resistance to apoptosis in the infected cells were no higher than the level of resistance in the bystander cells, then the infected cells would preferentially kill themselves through apoptosis. It appears that in the case of HIV, the increased resistance to apoptosis is provided by expression of Bcl-2 and suppression of p53. Hence, drugs that suppresses Bcl-2 or restore p53 function might be effective in restoring the parity of resistance to apoptosis between infected and uninfected cells. Currently, an antisense drug targeting Bcl-2 (G3139/Genasense(TM), Genta, Inc.) is in late-stage cancer trials and may be on the market for those indications in months. It would be interesting to try these drugs against various intracellular parasites including HIV. This approach to prevent or eliminate active infections might be particularly attractive against a range of parasites (virus, bacteria, protozoa, fungus) when safe and effective vaccines are not available.

Historical perspective of cell-cell fusion in cancer initiation and progression.

This review discusses the topic of cell-cell fusion as it related to cancer causation, progression and metastasis. The relevant time period is over a century of research and analysis from the observation and hypothesis of Theodor Boveri in circa 1902 to the most recent publications in 2012. There are three main intertwined thrusts in this history: (i) progress in our overall understanding of cancer and the relative importance of mutations and aneuploidy; (ii) evidence that cell-cell fusion occurs and leads to aneuploidy in somatic cells, especially that cell-cell fusion allows rapid evolution of cancer cells that, most importantly, acquire a repertoire of traits from bone marrow-derived cells that facilitate metastasis; and (iii) evidence that viruses catalyze cell-cell fusion and that aneuploidy evolving from the hybrid cells is an important if not the principal basis for the association of viruses with certain cancers. There have been many competing hypotheses concerning all aspects of cancer development during this time period. Overall, the cell-cell fusion hypothesis has never gained preeminence. As a result, it has gone through long periods of obscurity only to be re-discovered because new supportive data appeared. Technical advances (especially in our ability to sequence genomes and establish the evolution of cell clones through cytogenetics) have recently made tools available that could help elucidate the role of cell-cell fusion, aneuploidy and viruses in cancer. Evidence seems to be increasing in support of the ideas stated by Boveri a century ago.