University of Pennsylvania researchers report that they have genetically altered immune cells to resist infection in patients with HIV, while also reducing the viral load in some of the subjects after they were taken off of antiretroviral drug therapy. The findings of their study, published in the 

, hint at the future potential for gene therapy to treat the virus without the use of drugs.

The Phase I study, which is the first published report of gene editing in humans, analyzes the safety of altering a patient's T cells to replicate a naturally occurring resistance to HIV while potentially limiting viral loads. In the nonrandomized uncontrolled study, a single dose of autologous CD4 T cells modified with zinc-finger nuclease (ZFN) to inactivate the CCR5 gene, which allows HIV to take hold, were infused in 12 patients with chronic aviremic HIV infection between May 2009 and July 2012.

“Six of them underwent an interruption in antiretroviral treatment 4 weeks after the infusion of 10 billion autologous CD4 T cells, 11 to 28% of which were genetically modified with the ZFN,” the authors write. “The primary outcome was safety as assessed by treatment-related adverse events. Secondary outcomes included measures of immune reconstitution and HIV resistance.”

At the end of week 1, the median CD4 T-cell count totaled 1517 per cubic millimeter, a significant increase from the pre-infusion count of 448 per cubic millimeter. The median concentration of the CCR5-modified CD4 T cells at 1 week was 250 cells per cubic millimeter, which constitutes 8.8% of circulating peripheral-blood mononuclear cells and 13.9% of circulating CD4 T cells, according to the researchers.

The modified cells also had an estimated mean half-life of 48 weeks, but during treatment interruption and the resultant viremia, the decrease in the modified cells (−1.81 cells per day) was significantly lower than the drop in unmodified cells (−7.25 cells per day). In 1 of 4 patients who could be evaluated, the HIV RNA became undetectable, while the blood level of HIV DNA in most of the other patients declined.

The subject whose viral load fell to an undetectable level was later discovered to be heterozygous for a mutation that leads to CCR5 deletion.

“Successful clinical application will most likely depend on biallelic knockout in persons who have two non-mutated CCR5 alleles,” the authors write.

One serious adverse event occurred with the infusion of the modified CD4 T cells, but the reaction was attributed to a transfusion reaction. The treatment was ultimately deemed safe and tolerable, with the modified cells persisting in patients during follow-up testing.

“The firm conclusion of this study is that genome editing of human cells was safe and associated with an acceptable adverse-event profile and that the cells persisted in vivo,” 

write Mark A. Kay, MD, PhD, and Bruce D. Walker, MD, in an accompanying editorial

. “The tantalizing question raised by the transient treatment interruption is whether it might actually have been partially effective. A definitive answer to this question will require additional studies.”

Articles

Researchers safely infuse engineered immune cells in groundbreaking gene therapy study. 

Gene Therapy Shows Promise in Neutralizing HIV

Scientists have figured out a protein structure of the hepatitis C virus that could bring researchers closer to developing a vaccine to prevent the disease.

Research published online in the “Letters” section of the journal 

 describes the viral structural protein E2, which helps the hepatitis C virus (HCV) evade the body’s natural immune system response, said Joseph Marcotrigiano, an author of the study and professor of chemistry and chemical biology at Rutgers University in New Jersey. The research resulted from a collaboration of scientists from Rutgers’ Center for Advanced Biotechnology and Medicine and the Emory University School of Medicine.

There currently is no vaccine for HCV. Finding out more about the structure of the virus could bring scientists closer to creating one, Marcotrigiano said.

“Viruses are smart and it is a constant battle to keep them out,” said Marcotrigiano in a 

 that accompanied publication of the study results. “That’s why the development of a vaccine is so important. It’s always better to prevent infection through an effective vaccine then to treat after a chronic infection has been established.”

Hepatitis C causes inflammation of the liver, can lead to cirrhosis of the liver, and is the leading cause of liver transplants in the United States. The Centers for Disease Control estimates that 

 in the U.S. are infected with HCV. Though it is the most common chronic blood-borne infection, many people don’t know they have it because symptoms may not appear until the virus has severely damaged the liver.

Hepatitis C mutates constantly, which allows the virus to infect a host cell and dodge immune responses, while spreading infection throughout the body. Two envelope glycoproteins, E1 and E2, are on the outside membrane of the virus and are responsible for binding to host liver cells, leading to virus entry. Knowing more about the E2 protein structure could assist with future research aimed at understanding the binding mechanism of HCV.

“For many other viruses, like influenza, the entry pathway is well worked out. But in HCV, there is much we do not yet understand,” Marcotrigiano said.

Previously it was thought that the HCV E2 protein was a member of what is known as the class II viral fusion proteins found in related viruses such as yellow fever, Dengue and West Nile. In fact, this new research reveals that E2 has very little in common with these proteins.

“So E2 has some similarities, but is actually very different than class II fusion proteins,” Marcotrigiano said.

The Nature article describes the E2 core as a compact, globular domain structure, consisting mostly of β-strands and random coil with two small α-helices. The fold consists of many strands that are held together by an extensive hydrophobic core and disulphide bonds. Marcotrigiano explained that Sheet A has an IgG-like fold commonly found in viral and cellular proteins but sheet B represents a novel fold, possibly unique to HCV.

Knowledge of viral protein structures has been critical to development of new drugs. Finding out more about the structure of HCV could bring scientists closer to creating a vaccine to prevent the disease, Marcotrigiano said.

“Structure gives you a foundation for further studies… an unusual new look at basically how things work,” explained Marcotrigiano.

Scientists have mapped the structure of a hepatitis C virus surface protein that is responsible for binding to host liver cells, leading to virus entry. 

New Details about Hepatitis C Protein May Aid Development of Vaccine 

Although simple and convenient, saliva tests for hepatitis C infection may not be the best option for determining infection status, particularly in patients who may have a concurrent HIV infection.

According to the results of a study appearing in the July 2013 edition of 

, the tests are a passable option for screening at-risk populations for the virus; however, the increased chance for a false-negative result when a patient also has an HIV infection severely limits the tests’ efficacy.

“Hepatitis C oral fluid antibody detection methods have great compliance levels, low cost, and authorise sampling in any place, without highly qualified personnel,” the authors noted. “Yet, HCV antibodies saliva detection methods present insufficient performances for the general population … The relative lack of sensitivity in the HIV infected population found in the present work is, however, a serious limitation as many HIV-infected persons are also HCV infected.”

The researchers used a modified serological assay protocol and compared the results to participants’ HIV serological status, CD4 cell counts, HIV viral loads, and HCV replication status. Researchers included 30 patients with HCV but not HIV, 28 patients without HIV or HCV, 30 patients with both HCV and HIV, and 30 patients with HIV but without HCV. Researchers used the Salivette kit, a saliva testing kit manufactured by Sarstedt LTD, to obtain saliva samples.

In the HIV-negative groups of participants, the anti-HCV antibody tests had 90% and 100% sensitivity and specificity. In participants with an HIV infection, the tests registered 93% specificity, and 73% sensitivity.

“A recent study on HCV rapid screening assays presented also [sic] higher rates of false-negative results on HIV-positive patients,” the authors noted. “This decreased sensitivity in HIV-infected patients could be linked to a lower concentration of HCV-antibody oral fluids when compared to a non—HIV-infected control population … The observed sharp drop from 90 to 73.3% of salivary testing sensitivity in the HIV population is in accordance with the 2 previous observations.”

In the 22 patients with an HIV viral load below 200 copies per milliliter, 19 tested positive for salivary HCV-AB, which amounted to an 86% sensitivity for salivary detection of anti-HCV antibodies. Despite this, the sensitivity dropped to 37.5% for 8 patients with HIV replication that ranged from 1830 to 166,000 copies per milliliter.

Researchers noted that the small number of participants in the cohort prevented them from having a cut-off value for the viral load analysis.

At the time of saliva collection, HCV replication status was available for 26 patients in the HIV-negative and HCV-positive cohort, and 28 patients in the HIV-positive and HCV-positive cohort. 95% of participants in the HIV-negative and HCV-positive were correctly identified as having HCV by the saliva test.

In the HIV-positive, HCV-positive cohort, however, only 80% were considered HCV positive according to the saliva test. 

“Our study, in agreement with previous works, highlights the poor performances of such oral tests, particularly in co-infected patients,” the authors wrote. “Mostly, the poor sensitivity seems unacceptable and such tests should not be used even in populations with limited access to health care facilities. Alternatives do exist and may offer, through simple finger prick, a more reliable screening strategy.”

Saliva tests for detecting HCV infection may not be the best option for determining infection status.

Hepatitis C Saliva Tests May Show a False-Negative Result if HIV Is Also Present

Researchers from Yale University have measured the efficiency of cell-free and direct cell-to-cell transmission of HIV using an experimental system. Their study, which was published in 

, shows how antiretroviral treatment (ART) and other combinations of therapies are capable of suppressing HIV, even when the disease spreads through cell-to-cell contact.

In cell-free transmission, the virus particles move from one cell to another through intercellular space to find a target cell, to which it binds and subsequently infects. In direct cell-to-cell transmission, infected cells spread the virus through close contact and virological synapse: an organized contact area which concentrates virus particles and cellular HIV entry points.

The scientists tested each drug in cell-free and cell-to-cell situations, including:

6 different nucleoside analog reverse transcriptase inhibitors (NRTIs)

4 non-nucleoside analog reverse transcriptase inhibitors (NNRTIs)

The results showed NNRTIs, ENT-Is, and PIs are highly effective at blocking the virus during cell-to-cell transmission, but NRTIs were unable to sufficiently block it. When the researchers combined 2 inactive NRTIs, they regained potent antiretroviral activity in the cell-to-cell experiment.

The multiplicity of infection (MOI), which is the ratio of the number of infectious particles to the number of target cells present in a defined space, is higher during cell-to-cell transmission, the investigators noted. When they changed the MOI in their experiments by adding more viruses to a defined space for their cell-free experiment, some NRTIs were ineffective when acting alone.

Higher drug concentration was required to suppress the elevated number of particles. Again, when 2 drugs were combined, they were effective.

During experiments with cell-to-cell and cell-free transmissions, researchers showed that existing drugs are effective in suppressing the spread of HIV. 

Combination Antiretroviral Therapy Effective for Preventing Spread of HIV between Cells 

Noting successful HIV/AIDS prevention efforts rely on early diagnosis, an analysis published in the March 2014 issue of 

 estimates that improved access to health insurance through the Affordable Care Act (ACA) will increase the number of patients getting tested for HIV by nearly 500,000 over a 5-year period, though that impact would be amplified if all US states opted to expand Medicaid coverage.

As part of an issue-wide series concerning the effects of the ACA’s implementation on HIV/AIDS patients, a team of researchers from the Leonard D. Schaeffer Center for Health Policy and Economics at the University of Southern California modeled the impact of health insurance expansion on testing, diagnoses, and HIV-unaware status between 2013 and 2017.

For their study, the investigators used 2 scenarios to offer upper and lower estimates of the ACA’s impact. The first anticipated only the 18 states and the District of Columbia that had committed to expanding Medicaid as of July 2013 actually did so, while the second assumed all 50 states and the District of Columbia participated in Medicaid expansion.

For both scenarios, the researchers used a macrosimulation model — which relied on data from reputable public- and private-sector sources, including the US Centers for Disease Control and Prevention (CDC) &mdash; to estimate the ACA’s impact on “the number of newly insured people who gained coverage as a result of the ACA; the number of the newly insured who were tested for HIV and who would not have been tested without coverage; and the number of the newly tested who were newly diagnosed as HIV-positive, and thus were newly aware of their HIV status.”

At the conclusion of their study, the authors predicted the ACA will “substantially increase the number of people being tested for HIV, with an additional 603,024 people being tested if all states expand Medicaid, (or) an additional 466,153 people if only the 18 committed states and the District of Columbia expand Medicaid,” which they noted is a 30% difference in impact. Although the study period was extended through 2017, the researchers’ model showed almost all of the increase in HIV testing will occur before 2016.

In terms of the effect of those presumed increases in testing, the authors found 2,598 additional new HIV diagnoses will be made by 2017 in the 18-state Medicaid expansion scenario, or 3,300 in the all-state scenario. In addition, HIV unawareness will be reduced by 22% between 2013 and 2017 among those unknowingly living with HIV/AIDS who become newly insured through the ACA.

“Making HIV-positive people aware of their status is at the forefront of HIV prevention. Yet, there are millions of people living with HIV/AIDS in the United States who are unaware of their HIV status,” the authors wrote. “A multi-prong approach is needed, (and) our analysis indicates that increasing health insurance might be an important part of such an approach.”

Since the ACA’s affect on HIV testing and diagnoses would be 30% greater if all states expanded eligibility for Medicaid coverage, the researchers advised that “policymakers should consider such epidemiological benefits when analyzing insurance expansion policies.”

Noting successful HIV/AIDS prevention efforts rely on early diagnosis, an analysis estimates that improved access to health insurance through the Affordable Care Act will increase the number of patients getting tested for HIV by nearly 500,000 over a 5-year period, though that impact would be amplified if all US states opted to expand Medicaid coverage. 

How Obamacare Will Affect HIV/AIDS Prevention

Hepatic decompensation was more common in people who have both human immunodeficiency virus (HIV) and hepatitis C virus (HCV) compared to those infected only with HCV, an indication that early HCV treatment among coinfected patients is needed, say authors of a recent study.

Unscreened and untreated, HCV can remain in a person’s blood stream for years without symptoms of illness. It can cause severe liver damage over time and lead to cirrhosis, liver cancer and the need for liver transplant.

The risk of hepatitis C-associated serious liver disease persists in HIV patients who otherwise benefit from antiretroviral therapy used to treat HIV, said Vincent Lo Re III, MD, of the Perelman School of Medicine at the University of Pennsylvania, lead author of the retrospective study published in the March 18 issue of 

“Our results suggest that serious consideration should be given to initiating hepatitis C treatment in patients coinfected with HIV and hepatitis C &#8209;&#8209; particularly among those with advanced liver fibrosis or cirrhosis &#8209;&#8209; in order to try to reduce the risk of serious potentially life-threatening liver complications,” Lo Re said in a 

. “By taking action sooner, we may be able to reduce the risk of advanced liver disease in coinfected patients.”

Decompensation occurs when the liver is severely damaged and no longer functions normally. The higher rates of serious liver disease found in coinfected patients in this study were classified as liver decompensation, the statement said.

Earlier studies have shown that liver fibrosis is accelerated in coinfected patients but this large study is the first that “head to head compared coinfected patients to monoinfected patients” and looked specifically at clinical liver complications, Lo Re said.

Researchers analyzed data from US Veteran’s Administration medical records of 4,280 patients infected with both HIV and chronic HCV and receiving antiretroviral therapy compared to 6,079 patients infected only with HCV who received care between 1997 and 2010. They measured incidence of hepatic decompensation defined by diagnosis of ascites, spontaneous bacterial peritonitis, or esophageal variceal hemorrhage. Most patients were men and any patients treated for HCV were excluded from the study.

A prevailing perception that effective antiretroviral treatment “will help to make an HIV coinfected person like a monoinfected person” was not supported by data from the study, Lo Re said.

Overall, there was a 56-to-83 percent higher rate of liver decompensation over time among patients who were coinfected and receiving highly active antiretroviral therapy. Among a subgroup that had virologic control in response to therapy, the risk decreased to 44-to-65 percent but was still significantly higher than patients with hepatitis C alone, Lo Re said. The lower range estimates were a result of data analysis that factored death as competing risk because coinfected patients died much more often than patients with HCV alone.

In the past, patients receiving highly active antiretroviral therapy have been reluctant to begin prolonged HCV treatment, which typically required interferon injections and ribavirin and can include serious side effects, Lo Re said. This study showed that while receiving antiretroviral therapy and having virologic control are important for decreased decompensation in coinfected patients, “It is still not enough,” he said.

“It emphasizes the importance for clinicians to consider hepatitis C treatments for their coinfected patients,” he said, especially now that new HCV medications are available that have been shown in clinical trials to have higher cure rates with fewer side effects.

The researchers concluded that despite receiving antiretroviral therapy, patients co-infected with HIV and HCV had higher rates of hepatic decompensation than HCV-monoinfected patients. Furthermore, rates of decompensation were higher for coinfected patients with advanced liver fibrosis, severe anemia, diabetes, and nonblack race. 

Study results show hepatic decompensation is more common in people who have both HIV and hepatitis C virus (HCV) compared to those infected only with HCV, an indication that early HCV treatment among coinfected patients is needed. 

Serious Liver Disease Rates Higher Among Coinfected Patients  

Men with HIV have greater risk for developing plaque in their coronary arteries than uninfected men, regardless of their individual cardiovascular (CV) risk factors, according to research published in the April issue of 

Researchers from Johns Hopkins Medicine evaluated 618 HIV-infected men and 383 uninfected men aged 40-70 years who participated in the Multicenter Aids Cohort Study (MACS) for coronary atherosclerosis using computed tomography (CT) scans. The men had no prior surgery to restore blood flow to their coronary arteries.

The scientists measured the presence and amount of plaque in the coronary arteries to determine whether plaques causing the stenosis were noncalcified, partly calcified (<50%), or calcified. Since noncalcified plaque is not visible using standard CT angiography, the researchers injected contrast dye to enhance visibility.

HIV-infected men were found to have greater prevalence of any plaque, especially noncalcified plaque, than uninfected men, which suggested an increased risk for heart attacks. This held true even after accounting for other coronary artery disease (CAD) risk factors, such as high cholesterol, smoking, and high blood pressure.

Coronary artery stenosis >50% was associated with more advanced HIV and longer treatment with highly active antiretroviral treatment among HIV-infected men.

“MACS participants are a very good population for this study, because the control group comprises uninfected men drawn from the same population of men who are infected,” lead study author Wendy S. Post, MD, MS, noted. “This means the uninfected men are also at risk for HIV and share similar environmental exposures and lifestyles.”

Traditional CV risk factors in HIV-infected men need to be modified, the authors wrote. They hoped to repeat the process to find out why HIV-infected men have more noncalcified plaque than their uninfected counterparts.

Men with HIV have more noncalcified plaque in their arteries than their uninfected counterparts. 

May 2014