Cell therapies for treatment of human immunodeficiency virus infection.

After the serendipitous discovery of HIV eradication in the "Berlin patient", interest has grown in curing HIV infection by replacing the patient's replication-competent blood cells with infection-resistant ones. At the same time, induced pluripotent stem cell technologies and genetic engineering have boosted cell therapy transfer into the clinic. Currently available cell therapy approaches to attempt to cure HIV infection include the following: (1) Transplantation of autologous or allogeneic cells spontaneously resistant or edited to resist HIV infection; (2) Transplantation of autologous T-lymphocytes spontaneously targeting or redirected against HIV; and (3) Transplantation of autologous cells engineered to work as anti-HIV antibody factories. We review here the preliminary results and potential for future applications of these approaches.

Viral vectors: a look back and ahead on gene transfer technology.

No matter what their origin, strain and family, viruses have evolved exquisite strategies to reach and penetrate specific target cells where they hijack the cellular machinery to express viral genes and produce progeny particles. The ability to deliver and express genetic information to cells is the basis for exploiting viruses as "Trojan horses" to genetically modify the natural cell target or, upon manipulation of the viral receptor to retarget the virus, to genetically engineer different cell types. This process, known as transduction, is accomplished using viral vectors derived from parental wild type viruses whose viral genes, essential for replication and virulence, have been replaced with the heterologous gene(s) required for cell manipulation. Rearrangement of the viral genome to impede replication or generation of infectious virions but maintaining the ability to deliver nucleic acids has been the object of intense research since the early 1980s. Technological advances and the ever-growing knowledge of molecular virology and virus-host cell relationships have constantly improved the safety profile of viral vectors that are now used in vitro and in vivo to study cellular gene function, correct genetic defects (gene therapy), express therapeutic proteins, vaccinate against infectious agents and tumors, produce experimental animal models, and for other purposes. This review illustrates the strategies used to generate some of the most used viral vectors, and their advantages, limitations and principal applications.

Comparison of oncogenic HPV type-specific viral DNA load and E6/E7 mRNA detection in cervical samples: results from a multicenter study.

High-risk human papillomavirus (HR-HPV) genotype viral load and E6/E7 mRNA detection are proposed as surrogate markers of malignant cervical lesion progression. Currently, the use of commercially available DNA-based or mRNA-based tests is under investigation. In this study, the viral DNA load and E6/E7 mRNA detection of the five most common HR-HPV types detected in cervical cancer worldwide were compared in 308 cervical samples by using in-house type-specific quantitative real-time PCR assays and PreTect HPV-Proofer test, respectively. Sensitivity and negative predictive values were higher for the HPV-DNA assays combined (95.0% and 96.0%, respectively) than the RNA assays (77.0% and 88.0%, respectively); conversely, the mRNA test showed a higher specificity and higher positive predictive value (81.7% and 66.9%, respectively) than the DNA test (58.6% and 52.5%, respectively) for detecting histology-confirmed high-grade cervical intraepithelial neoplasia. A significantly higher association between viral DNA load and severity of disease was observed for HPV 16 and 31 (γ = 0.62 and γ = 0.40, respectively) than for the other HPV types screened. A good degree of association between the two assays was found for detection of HPV 16 (k = 0.83), HPV 18 (k = 0.72), HPV 33 (k = 0.66), and HPV 45 (k = 0.60) but not for HPV 31 (k = 0.24). Sequence analysis in L1 and E6-LCR regions of HPV 31 genotypes showed a high level of intra-type variation. HR-HPV viral DNA load was significantly higher in E6/E7 mRNA positive than negative samples (P < 0.001), except for HPV 31. These findings suggest that transcriptional and replicative activities can coexist within the same sample.

A lentiviral vector-based, herpes simplex virus 1 (HSV-1) glycoprotein B vaccine affords cross-protection against HSV-1 and HSV-2 genital infections.

Genital herpes is caused by herpes simplex virus 1 (HSV-1) and HSV-2, and its incidence is constantly increasing in the human population. Regardless of the clinical manifestation, HSV-1 and HSV-2 infections are highly transmissible to sexual partners and enhance susceptibility to other sexually transmitted infections. An effective vaccine is not yet available. Here, HSV-1 glycoprotein B (gB1) was delivered by a feline immunodeficiency virus (FIV) vector and tested against HSV-1 and HSV-2 vaginal challenges in C57BL/6 mice. The gB1 vaccine elicited cross-neutralizing antibodies and cell-mediated responses that protected 100 and 75% animals from HSV-1- and HSV-2-associated severe disease, respectively. Two of the eight fully protected vaccinees underwent subclinical HSV-2 infection, as demonstrated by deep immunosuppression and other analyses. Finally, vaccination prevented death in 83% of the animals challenged with a HSV-2 dose that killed 78 and 100% naive and mock-vaccinated controls, respectively. Since this FIV vector can accommodate two or more HSV immunogens, this vaccine has ample potential for improvement and may become a candidate for the development of a truly effective vaccine against genital herpes.

In vivo and in vitro evidence for an association between the route-specific transmission of HHV-8 and the virus genotype.

The study was performed to determine if there is an association between the genotype and transmission of HHV-8 types A and C. These HHV-8 subtypes are prevalent in the area of North of Sardinia, which is an island off west Italy's mainland that has a high HHV-8 seroprevalence (35%). Blood and saliva samples from 30 patients with classic Kaposi's sarcoma who were lifetime residents of North Sardinia were analyzed to identify the HHV-8 genotype and quantitate the viral load. Genotype A, especially A1 subtype, was found more frequently (9/30 patients) and had a significantly higher viral load in saliva compared to blood (P = 0.029), where type C was found more frequently but with a viral load lower than 10(3) copies/ml. To determine if there is a correlation between the viral genotype and cellular tropism, type A1 and C3 HHV-8 viral particles were obtained from cell lines BCBL1 and BC3 infected chronically with HHV-8 A1 and C3 genotypes respectively and used to infect HEK293 epithelial origin cells and PBMCs in vitro. The data indicate that the A1 HHV-8 genotype is tropic and replicates at higher levels in the epithelial cell lines.

Progressive multifocal leukoencephalopathy: what's new?

Progressive multifocal leukoencephalopathy (PML), a severe demyelinating disease that is caused by human JC polyomavirus, was first described as a complication of immune suppression 50 years ago and emerged as a major complication of HIV infection in the 1980s. The prognosis has remained dismal since then, with discouraging results from clinical trials of various therapeutic approaches, including immunomodulation and/or inhibition of viral replication. PML is caused by reactivation of latent JC virus, and serotonergic 5-HT(2a) receptors have been identified as being critical for viral infection of glial cells. In recent years, immunosuppressive therapeutic antibodies have been associated with an increased incidence rate of PML. Here, the authors review findings on the pathogenesis of PML and the encouraging case reports of novel treatments.

Role of hematopoietic cells in the maintenance of chronic human torquetenovirus plasma viremia.

Many aspects of the life cycle of torquetenoviruses (TTVs) are essentially unexplored. In particular, it is still a matter of speculation which cell type(s) replicates the viruses and maintains the generally high viral loads found in the blood of infected hosts. In this study, we sequentially measured the TTV loads in the plasma of four TTV-positive leukemia patients who were strongly myelosuppressed and then transplanted with haploidentical hematopoietic stem cells. The findings provide clear quantitative evidence for an extremely important role of hematopoietic cells in the maintenance of TTV viremia.

Torquetenovirus viremia kinetics after autologous stem cell transplantation are predictable and may serve as a surrogate marker of functional immune reconstitution.

BACKGROUND: It is common experience that retreating patients too early after a course of intensive chemotherapy predisposes to opportunistic infections despite apparently normal lymphocyte levels. OBJECTIVES: The extent of replication of persistent viruses that cause no obvious disease (and hence need no treatment) might better define when a patient has recovered from functional immune deficiency. STUDY DESIGN: We used real-time polymerase chain reaction to monitor the kinetics of plasma torquetenovirus (TTV) viremia in hematological patients undergoing autologous hematopoietic stem cell transplantation as support to high-dose chemotherapy (HSCT). RESULTS: Independently from underlying hematological disease and therapeutic regimen, TTV viremia increased post-HSCT, and this increase paralleled the increase of circulating CD8(+)CD57(+) T lymphocytes, known to represent an indirect marker of functional immune deficiency. Subsequently, within a matter of months, TTV levels returned to baseline values, at a pace that appeared to be constant over time. CONCLUSION: Monitoring of TTV viremia represents a unique opportunity to follow functional immune reconstitution in immunosuppressed patients. Also, the size of the TTV viremia increases resulting from immunosuppressive treatments might be of guidance in determining the appropriate time interval before delivery of a next course of therapy.

Torquetenovirus DNA drives proinflammatory cytokines production and secretion by immune cells via toll-like receptor 9.

Active infection with torquetenovirus (TTV) has been associated with an increased severity of diseases in which inflammation plays a particularly important pathogenetic role. Here, we report that cloned DNA of a genogroup 4 TTV (ViPiSAL) is an activator of proinflammatory cytokine production by murine spleen cells and that the effect is mediated via toll-like receptor (TLR)9. The same DNA also increased the levels of proinflammatory cytokines induced by two well-characterized TLR9 stimulants. Finally, in silico analyses of the genomes of ViPiSAL and other TTVs revealed marked differences in the representation of CpG motifs known to be most effective at activating immune cells via TLR9. These findings demonstrate for the first time that at least one TTV isolate has the potential to stimulate and co-stimulate inflammatory responses.

Hyperbaric oxygen therapy in BKV-associated hemorrhagic cystitis refractory to intravenous and intravesical cidofovir: case report and review of literature.

Hemorrhagic cystitis is a common complication in hematopoietic stem cell transplant recipients. We report here a case of severe BKV-associated hemorrhagic cystitis who did not respond to intravenous cidofovir. Overt hematuria successfully resolved after a few days on hyperbaric oxygen and intravesical instillations of cidofovir, while BK viruria dropped after a few weeks and remained low. We review the literature for therapeutic options in hemorrhagic cystitis and try to explain how hyperbaric oxygen stimulates mucosal repair in the urinary bladder.

Immunotherapy with internally inactivated virus loaded dendritic cells boosts cellular immunity but does not affect feline immunodeficiency virus infection course.

Immunotherapy of feline immunodeficiency virus (FIV)-infected cats with monocyte-derived dendritic cells (MDCs) loaded with aldrithiol-2 (AT2)-inactivated homologous FIV was performed. Although FIV-specific lymphoproliferative responses were markedly increased, viral loads and CD4+ T cell depletion were unaffected, thus indicating that boosting antiviral cell-mediated immunity may not suffice to modify infection course appreciably.