Naomi Aronson

Decade-Long Safety and Function of Retroviral-Modified Chimeric Antigen Receptor T Cells

Adoptively transferred chimeric antigen receptor T cells have stable stem cell–like persistence for at least a decade and more than 500 years of patient safety. Standing the Test of Time Retroviral vectors were once the mainstay of gene transfer because they could stably integrate into the host genome. However, some patients in early trials developed leukemia because of insertional mutagenesis. Now, Scholler et al. report that retroviral vector–mediated gene transfer in T cells may not have the same safety concerns, and that these cells may persist over a decade in patients. The authors followed patients from three clinical trials who received T cells transduced with gammaretroviruses carrying a chimeric antigen receptor. They found that these cells were present in recipients over a decade after infusion at levels higher than those induced by standard vaccines. These cells were still functional, had stable levels of engraftment, and did not require host immunosuppression before transplant. Moreover, the authors found no evidence of integration-induced immortalization, with no observable enrichment of integration sites near genes involved in growth control or transformation. Thus, the safety of retroviral vectors may be cell type–specific, opening up engineered T cells as a delivery platform for therapeutics. The success of adoptive T cell gene transfer for treatment of cancer and HIV is predicated on generating a response that is both durable and safe. We report long-term results from three clinical trials to evaluate gammaretroviral vector–engineered T cells for HIV. The vector encoded a chimeric antigen receptor (CAR) composed of CD4 linked to the CD3ζ signaling chain (CD4ζ). CAR T cells were detected in 98% of samples tested for at least 11 years after infusion at frequencies that exceeded average T cell levels after most vaccine approaches. The CD4ζ transgene retained expression and function. There was no evidence of vector-induced immortalization of cells; integration site distributions showed no evidence of persistent clonal expansion or enrichment for integration sites near genes implicated in growth control or transformation. The CD4ζ T cells had stable levels of engraftment, with decay half-lives that exceeded 16 years, in marked contrast to previous trials testing engineered T cells. These findings indicate that host immunosuppression before T cell transfer is not required to achieve long-term persistence of gene-modified T cells. Further, our results emphasize the safety of T cells modified by retroviral gene transfer in clinical application, as measured in >500 patient-years of follow-up. Thus, previous safety issues with integrating viral vectors are hematopoietic stem cell or transgene intrinsic, and not a general feature of retroviral vectors. Engineered T cells are a promising form of synthetic biology for long-term delivery of protein-based therapeutics. These results provide a framework to guide the therapy of a wide spectrum of human diseases.

Adoptive transfer of costimulated CD4 + T cells induces expansion of peripheral T cells and decreased CCR5 expression in HIV infection

To study the safety and feasibility of T-cell reconstitution in HIV-infected individuals, we adoptively transferred activated autologous CD4+ T cells. Polyclonal peripheral blood CD4+ cells were costimulated ex vivo and subjects were given infusions of up to 3 × 1010 activated CD4+ cells. Dose-dependent increases in CD4+ cell counts and in the CD4:CD8 ratio were observed. Sustained increases in the fraction of cytokine-secreting T cells and decreases in the percentage of CD4+CCR5+ cells were noted in vivo, suggesting enhanced function and resistance to HIV infection. The frequency of CD4+Ki-67+ cells increased whereas CD4+ T cells containing T cell–receptor rearrangement excision circles (TRECs) decreased. These findings indicate that expansion of the peripheral T-cell pool mediated the increase in CD4 counts and suggest that approaches to reconstitute CD4 helper cell activity and decrease CCR5 expression may augment natural immunity to HIV infection.

Methodological standards and patient-centeredness in comparative effectiveness research: The PCORI perspective

Rigorous methodological standards help to ensure that medical research produces information that is valid and generalizable, and are essential in patient-centered outcomes research (PCOR). Patient-centeredness refers to the extent to which the preferences, decision-making needs, and characteristics of patients are addressed, and is the key characteristic differentiating PCOR from comparative effectiveness research. The Patient Protection and Affordable Care Act signed into law in 2010 created the Patient-Centered Outcomes Research Institute (PCORI), which includes an independent, federally appointed Methodology Committee. The Methodology Committee is charged to develop methodological standards for PCOR. The 4 general areas identified by the committee in which standards will be developed are (1) prioritizing research questions, (2) using appropriate study designs and analyses, (3) incorporating patient perspectives throughout the research continuum, and (4) fostering efficient dissemination and implementation of results. A Congressionally mandated PCORI methodology report (to be issued in its first iteration in May 2012) will begin to provide standards in each of these areas, and will inform future PCORI funding announcements and review criteria. The work of the Methodology Committee is intended to enable generation of information that is relevant and trustworthy for patients, and to enable decisions that improve patient-centered outcomes.

In harm's way: infections in deployed American military forces.

Hundreds of thousands of American service members have been deployed to Afghanistan and Iraq since 2001. With emphasis on the common infections and the chronic infections that may present or persist on their return to the United States, we review the data on deployment-associated infections. These infections include gastroenteritis; respiratory infection; war wound infection with antibiotic-resistant, gram-negative bacteria; Q fever; brucellosis; and parasitic infections, such as malaria and leishmaniasis.

Prevalence of genotypic and phenotypic resistance to anti-retroviral drugs in a cohort of therapy-naïve HIV-1 infected US military personnel.

ObjectiveWhile transmission of drug-resistant HIV-1 has been reported, estimates of prevalence of resistance in drug-naïve populations are incomplete. We investigated the prevalence of genotypic mutations and phenotypic antiretroviral resistance in a cohort of HIV-1 infected U.S. military personnel prior to the institution of antiretroviral therapy. DesignCross-sectional cohort study. MethodsPlasma was obtained from 114 recently HIV-1 infected subjects enrolled in an epidemiological study. Genotypic resistance was determined by consensus sequencing of a PCR product from the HIV-1 pol gene. Sequences were interpreted by a phenotypic–genotypic correlative database. Resistance phenotypes were determined by a recombinant virus cell culture assay. ResultsGenotypic mutations and phenotypic resistance were found at a higher than expected frequency. Resistance to non-nucleoside reverse transcriptase inhibitors was most common, with a prevalence of 15% of 95 subjects by genotype and 26% of 91 subjects by phenotype. Genotypic and phenotypic resistance respectively were found in 4% and 8% of subjects for nucleoside reverse transcriptase inhibitors and in 10% and 1% for protease inhibitors. One subject harbored virus with resistance to all three drug classes. ConclusionsA substantial frequency of resistance to antiretroviral drugs was identified in a therapy-naïve U.S. cohort. In most cases, the genotypic and phenotypic assays yielded similar results, although the genotypic assay could detect some protease inhibitor resistance-associated mutations in the absence of phenotypic resistance. These data suggest the need for optimization of treatment guidelines based on current estimates of the prevalence of drug resistance in HIV-1 seroconverters.

Old world leishmaniasis: an emerging infection among deployed US military and civilian workers.

Many veterans of Operation Iraqi Freedom are now returning to the United States after potential exposure to leishmaniasis. In the past year, large numbers of leishmaniasis cases of a magnitude not encountered in the United States since World War II have challenged clinicians in both the military and the civilian sectors. Many Reserve and National Guard troops were deployed to Iraq and are now back in their communities. Hundreds of leishmaniasis cases, which were managed by a few practitioners initially, permitted further appreciation of the epidemiology and diagnostic and treatment options for Old World leishmaniasis. We describe the current situation, with on-the-ground experience, complimented by a literature review, and we provide a practical list of options for the clinician likely to encounter this parasitic infection in the coming months and years.

Safety and Efficacy of Intravenous Sodium Stibogluconate in the Treatment of Leishmaniasis: Recent U.S. Military Experience

The efficacy and toxicity of sodium stibogluconate (SSG) at a dosage of 20 mg/(kg.d) for either 20 days (for cutaneous disease) or 28 days (for visceral, mucosal, or viscerotropic disease) in the treatment of leishmaniasis is reported. Ninety-six U.S. Department of Defense health care beneficiaries with parasitologically confirmed leishmaniasis were prospectively followed for 1 year. One patient was infected with human immunodeficiency virus; otherwise, comorbidity was absent. Clinical cure occurred in 91% of 83 cases of cutaneous disease and 93% of 13 cases of visceral/viscerotropic disease. Adverse effects were common and necessitated interruption of treatment in 28% of cases, but they were generally reversible. These included arthralgias and myalgias (58%), pancreatitis (97%), transaminitis (67%), headache (22%), hematologic suppression (44%), and rash (9%). No subsequent mucosal leishmaniasis was identified, and there were no deaths attributable to SSG or leishmaniasis.

A Randomized, Double-Blind Study of the Efficacy of a 10- or 20-Day Course of Sodium Stibogluconate for Treatment of Cutaneous Leishmaniasis in United States Military Personnel

The recommended treatment for cutaneous leishmaniasis is pentavalent antimony at a dosage of 20 mg/kg/day for 20 days. Some studies conducted in locales in which Leishmania is endemic have suggested that shorter courses of treatment may be as efficacious. We conducted a randomized, double-blind, placebo-controlled study of 10 versus 20 days of sodium stibogluconate (SSG) in United States military personnel who contracted cutaneous leishmaniasis while serving overseas; 19 patients received SSG for 10 days (and placebo for 10 days), and 19 patients received SSG for 20 days. Cure rates were 100% (19 of 19 patients) in the 10-day group and 95% (18 of 19 patients) in the 20-day group. Side effects were more common among patients who received 20 days of therapy. In this group of otherwise healthy young adults, SSG at a dosage of 20 mg/kg/day for 10 days appears to have been therapeutically equivalent and less toxic than the standard 20-day course.

Devices for Rapid Diagnosis of Malaria: Evaluation of Prototype Assays That Detect Plasmodium falciparum Histidine-Rich Protein 2 and a Plasmodium vivax-Specific Antigen

ABSTRACT The ParaSight F test was developed as a pioneer industry effort in the large-scale, process-controlled production of a device for the rapid diagnosis of malaria. This device performed well in field settings but was limited to the detection of a single malaria species, Plasmodium falciparum. The ParaSight F+V assay advanced upon the ParaSight F test format by incorporating a monoclonal antibody directed against a proprietary Plasmodium vivax-specific antigen, in addition to the antibody directed against P. falciparum histidine-rich protein 2, which was used in the ParaSight F assay. The modified assay was developed to add the capability to detect P. falciparum and P. vivax in a single-test-strip format. The present study evaluated three distinct ParaSight F+V prototypes with samples from symptomatic patients in regions of Thailand and Peru where malaria is endemic. Over a 2-year enrollment period (1998 and 1999), a total of 4,894 patients consented to participation in the study. Compared with the results for duplicate microscopic examinations of Giemsa-stained blood smears as the reference diagnostic standard, each successive prototype showed substantial improvement in performance. The final ParaSight F+V prototype, evaluated in 1999, had an overall sensitivity for detection of asexual P. falciparum parasites of 98%. The sensitivity of the device was 100% for P. falciparum densities of >500 parasites/μl, with a sensitivity of 83% for parasite densities of ≤500/μl. The specificity for the exclusion of P. falciparum was 93%. For P. vivax, the overall sensitivity was 87% for the final 1999 prototype. The sensitivities calculated for different levels of P. vivax parasitemia were 99% for parasite densities of >5,000/μl, 92% for parasite densities of 1,001 to 5,000/μl, 94% for parasite densities of 501 to 1,000/μl, and 55% for parasite densities of 1 to 500/μl. The specificity for the exclusion of P. vivax was 87%. The areas under the receiver operating characteristic curves for the diagnostic performance of the assay for the detection of P. falciparum and P. vivax were 0.8907 and 0.8522, respectively. These findings indicate that assays for rapid diagnosis have the potential to enhance diagnostic capabilities in those instances in which skilled microscopy is not readily available.

Long-Term Efficacy of Routine Access to Antiretroviral-Resistance Testing in HIV Type 1–Infected Patients: Results of the Clinical Efficacy of Resistance Testing Trial

The long-term efficacy of making resistance testing routinely available to clinicians has not been established. We conducted a clinical trial at 6 US military hospitals in which volunteers infected with human immunodeficiency virus type-1 were randomized to have routine access to phenotype resistance testing (PT arm), access to genotype resistance testing (GT arm), or no access to either test (VB arm). The primary outcome measure was time to persistent treatment failure despite change(s) in antiretroviral therapy (ART) regimen. Overall, routine access to resistance testing did not significantly increase the time to end point. Time to end point was significantly prolonged in the PT arm for subjects with a history of treatment with > or =4 different ART regimens or a history of treatment with nonnucleoside reverse-transcriptase inhibitors before the study, compared with that in the VB arm. These results suggest that routine access to resistance testing can improve long-term virologic outcomes in HIV-infected patients who are treatment experienced but may not impact outcome in patients who are naive to or have had limited experience with ART.