Bronchoalveolar lavage cell immunophenotyping facilitates diagnosis of lung allograft rejection.

Supplementary methods to identify acute rejection and to distinguish rejection from infection may improve clinical outcomes for lung allograft recipients. We hypothesized that distinct bronchoalveolar lavage (BAL) cell profiles are associated with rejection and infection. We retrospectively compared 2939 BAL cell counts and immunophenotypes against concomitantly obtained transbronchial biopsies and microbiologic studies. We randomly assigned 317 subjects to a derivation or validation cohort. BAL samples were classified into four groups: infection, rejection grade ≥A1, both or neither. We employed generalized estimating equation and survival modeling to identify clinical predictors of rejection and infection. We found that CD25(+) and natural killer cell percentages identified a twofold increased odds of rejection compared to either the infection or the neither infection nor rejection groups. Also, monocytes, lymphocytes and eosinophil percentages were independently associated with rejection. A four-predictor scoring system had high negative predictive value (96-98%) for grade ≥A2 rejection, predicted future rejection in the validation cohort and predicted increased risk of bronchiolitis obliterans syndrome in otherwise benign samples. In conclusion, BAL cell immunophenotyping discriminates between infection and acute rejection and predicts future outcomes in lung transplant recipients. Although it cannot replace histopathology, immunophenotyping may be a clinically useful adjunct.

Feasibility of organ-preservation strategies in head and neck cancer in developing countries.

BACKGROUND: Chemoradiotherapy is an established strategy for organ preservation in head-neck cancer. These protocols are associated with added toxicity and need support infrastructure. Practice setup and availability of resources vary at the community level in developing countries. AIM: To evaluate the feasibility of organ-preservation strategies in different settings in developing countries. SETTINGS AND DESIGN: Survey. MATERIALS AND METHODS: In a questionnaire-based study, questions were directed to clinicians with varied practice setups to gather information regarding infrastructure, finance, and feasibility of organ-preservation protocols and their current practice trends. STATISTICAL ANALYSIS: Descriptive. RESULTS: Responses from 100 clinicians with focused practice in head-neck oncology were analyzed. Sixty-one percent clinicians were practicing organ preservation for advanced head-neck cancers in their practice. However, 65% centers lacked sufficient infrastructure to support organ-preservation protocols. Forty percent patients were treated on cobalt-radiotherapy machine. Fifty-nine percent of clinicians suggested that less than third of their patients were fit to undergo chemoradiation and 67% believed that adherence to treatment protocol was observed in less than two-thirds of cases. Based on their experience 82% clinicians felt that only one-third patients requiring salvage would actually undergo treatment. The majority of the patients (68%) used personal funds for treatment and less than one-third of the patients could afford complete treatment. CONCLUSIONS: The infrastructure needed to support organ-preservation protocols varies significantly between centers in developing countries. It may not be feasible to perform organ-preservation strategies in certain centers and feasibility guidelines should be made for their judicious use in developing countries.

Head and neck cancer in India: need to formulate uniform national treatment guideline?

BACKGROUND: In a large and diverse country like India, there is a wide variation in the availability of infrastructure and expertise to treat head-neck cancer patients. Lack of consistent adherence to evidence-based management is the biggest problem. AIMS: There is an unmet need to evaluate the existing treatment practices to form the basis for development of effective and uniform treatment policies. SETTINGS AND DESIGNS: Prospective case series. MATERIALS AND METHODS: A group of previously treated, potentially curable patients presenting to our institution (from April 2009 to March 2011) were evaluated for appropriateness of initial treatment based on National Comprehensive Cancer Network or Tata Memorial Hospital guidelines. Data regarding treatment center, protocol and accuracy of delivered treatment and their eventual outcome were analyzed. STATISTICAL ANALYSIS: Descriptive. RESULTS: Amongst 450 newly registered patients, 77(17%) were previously treated with curative intent and 69(89%) of them were inappropriately treated. Seventeen (25%) patients were treated in clinics while 12(17%) in cancer centers and 34(50%) in corporate hospitals. Fourteen (20%) patients received chemotherapy, 22(32%) received radiotherapy and 14(20%) underwent surgery while 19(28%) patients received multimodality treatment. Disease stage changed to more advanced stage in 40(58%) patients and curative intent treatment could be offered only to 33(48%) patients. Amongst 56 patients available for outcome review, 18(32%) patients were alive disease-free, 20(36%) had died and 18(32%) were alive with disease. CONCLUSION: Large numbers of potentially curable patients are inappropriately treated and their outcome is significantly affected. Many initiatives have been taken in the existing National Cancer Control Program but formulation of a uniform national treatment guideline should be prioritized.

VEGF levels in the alveolar compartment do not distinguish between ARDS and hydrostatic pulmonary oedema.

Although overexpression of vascular endothelial growth factor (VEGF) 165 in the lung causes pulmonary oedema, its role in human acute lung injury (ALI) is unclear. VEGF levels are reported to be lower in bronchoalveolar lavage from ALI patients compared with normals, but these studies did not include a comparably ill control group with noninflammatory pulmonary oedema. The current authors hypothesised that VEGF levels in pulmonary oedema fluid would be lower in ALI patients compared with control patients with severe hydrostatic pulmonary oedema. VEGF was measured in pulmonary oedema fluid and plasma from 56 patients with ALI and 46 controls with severe hydrostatic pulmonary oedema. Pulmonary oedema fluid levels of VEGF did not differ between patients with hydrostatic oedema (median 799 pg x mL(-1), interquartile range (IQR) 226-2,281) and ALI (median 507, IQR 0.8-1,031). Plasma levels were also the same (median 20.5 pg x mL(-1), IQR 0-152 versus 4.8, IQR 0-99.8). There was no association between plasma or oedema fluid VEGF levels and outcomes including mortality. Vascular endothelial growth factor levels in pulmonary oedema fluid were depressed both in acute lung injury and hydrostatic pulmonary oedema. The decrease in air space concentrations of vascular endothelial growth factor in acute lung injury may not be a function of the degree of lung injury, but rather may result from alveolar flooding.

Modulation of amplitude and direction of in vivo immune responses by co-administration of cytokine gene expression cassettes with DNA immunogens.

Immunization with nucleic acids has been shown to induce both antigen-specific cellular and humoral immune responses in vivo. We hypothesize that immunization with DNA could be enhanced by directing specific immune responses induced by the vaccine based on the differential correlates of protection known for a particular pathogen. Recently we and others reported that specific immune responses generated by DNA vaccine could be modulated by co-delivery of gene expression cassettes encoding for IL-12, granulocyte-macrophage colony-stimulating factor and the co-stimulatory molecule CD86. To further engineer the immune response in vivo, we investigated the induction and regulation of immune responses following the co-delivery of pro-inflammatory cytokine (IL-1 alpha, TNF-alpha, and TNF-beta), Th1 cytokine (IL-2, IL-12, IL-15, and IL-18), and Th2 cytokine (IL-4, IL-5 and IL-10) genes. We observed enhancement of antigen-specific humoral response with the co-delivery of Th2 cytokine genes IL-4, IL-5, and IL-10 as well as those of IL-2 and IL-18. A dramatic increase in antigen-specific T helper cell proliferation was seen with IL-2 and TNF-alpha gene co-injections. In addition, we observed a significant enhancement of the cytotoxic response with the co-administration of TNF-alpha and IL-15 genes with HIV-1 DNA immunogens. These increases in CTL response were both MHC class I restricted and CD8+ T cell dependent. Together with earlier reports on the utility of co-immunizing using immunologically important molecules together with DNA immunogens, we demonstrate the potential of this strategy as an important tool for the development of more rationally designed vaccines.

Molecular and immunological analysis of genetic prostate specific antigen (PSA) vaccine.

Nucleic acid immunization has been investigated as immunotherapy for infectious diseases as well as for treating specific types of cancers. In this approach, nucleic acid expression cassettes are directly inoculated into the host, whose transfected cells become the production source of novel and possibly immunologically foreign protein. We have developed a DNA vaccine construct which encodes for PSA by cloning a cDNA for PSA into a mammalian expression vector under control of a CMV promoter. We investigated and characterized the immunogenicity of PSA DNA expression cassettes in mice. PSA-specific immune responses induced in vivo by immunization were characterized by enzyme-linked immunosorbent assay (ELISA), T helper proliferation cytotoxic T lymphocyte (CTL), and flow cytometry assays. We observed a strong and persistent antibody response against PSA for at least 180 days following immunization. In addition, a significant T helper cell proliferation was observed against PSA protein. Using synthetic peptides spanning the PSA open frame, we identified four dominant T helper epitopes of PSA. Furthermore, immunization with PSA plasmid induced MHC Class I CD8+ T cell-restricted cytotoxic T lymphocyte response against tumor cell targets expressing PSA. The prostate represents a very specific functional organ critical for reproduction but not for the health and survival of the individual. Understanding the immunogenicity of PSA DNA immunization cassettes offers insight into the possible use of this tumor-associated antigen as a target for immunotherapy. These results demonstrate the ability of the genetic PSA to serve as a specific immune target capable of generating both humoral and cellular immune responses in vivo.

An HIV type 2 DNA vaccine induces cross-reactive immune responses against HIV type 2 and SIV.

We have previously reported on the generation of specific functional immune responses after inoculation of animals with expression vectors encoding HIV-1 genes. This article provides the details of the first application of this new technology to induce immune responses against HIV-2. This virus is molecularly and serologically distinct from HIV-1 and is in fact more closely related to the simian immunodeficiency virus (SIV). Anti-HIV-2 and SIV antibodies were induced in mice of three different haplotypes following a single intramuscular inoculation with an HIV-2/ROD envelope glycoprotein expression vector (pcEnv-2). Boosting of animals with pcEnv-2 induced both anti-HIV-2 neutralizing antibodies and T cell-proliferative responses against HIV-2 and SIVmac proteins. We compared the humoral and cellular immune responses of mice injected with pcEnv-2 and then boosted with either the homologous DNA construct or a recombinant Env protein. Animals boosted with pcEnv-2 generated B and T cell immune responses as strong as those of mice boosted with recombinant gp140 protein in adjuvant. Finally, cellular immune responses were significantly increased with the coadministration of pcEnv-2 and a plasmid expressing interleukin 12. We therefore conclude that DNA plasmid inoculation induces cross-reactive anti-HIV-2 and anti-SIVmac immune responses in mice. This technology should be further investigated as a potential vaccine component for this human pathogen.