Laboratory of Diana Huang, PhD

Our work

I have been the HIV research scientist of the VQA Program over the ~25 years on various projects focused on genotypic and phenotypic changes associated with antiretroviral resistance and proficiency testing of these protocols. I was an original member of the ACTG/PACTG working group that helped develop sequencing as a diagnostic tool, as well as designing the pilot study for the model of the current VQA genotyping proficiency testing program. Trugene® (discontinued), ViroSeq™ and in-house assays were evaluated. I directly worked with both assays and have set up in-house sequencing for other regions of the genome. I have set up in-house real-time assays and developed clones of interest with potential use as standards and proficiency samples. I have been the donor coordinator of the VQA since 1999 and a member of the Rush IRB since 1998. I was a member of other working groups that have addressed tropism, collection of subtypes, and other molecular approaches for diagnostics to measure antiretroviral resistance. I have worked with earlier versions of the SCA (single copy assay) originated by Dr. John Mellors and modified by Dr. Jonathon Li in preparation that this assay becomes used as a means to assess “Cure” of HIV in infected individuals, a current focus of the ACTG. I am interested in bringing assays considered traditional bench science to be applied clinically.
I was awarded a contract from CONRAD which ended when CONRAD stopped development of the compounds of interest. This contract examined the genotypic and phenotypic characteristics of HIV with CCR5 and CXCR4 affinities after long term growth in cultured cells in the presence of individual candidate compounds for microbicides. We were able to identify a subset of the panel that are NNRTI inhibitors, some of which have “tight binding” or “hypersensitive” phenotypes, based on their changes in IC50 in PBMC cells.
In the past, I have had significant working experience with influenza and other negative strand viruses. As a work study student for my Ph.D. thesis advisor, Dr, Hunein F. Maassab, who developed the master strain for the Flumist vaccine, I grew influenza in eggs and cells, characterized the strains virologically and generated reassortant strains. I also help to mentor a graduate student as a postdoc in Dr. Julius Youngner’s lab at the University of Pittsburgh. My early training fostered an interest in persistent viral infections.

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Impact of work

1. My role in the VQA program allowed active participation in the development of genotyping to detect mutations associated with HIV-1 anti-retroviral drug resistance for clinical management of HIV-1.
A collaborative working group was formed in 1994-1995 between practicing physician-scientists who were engaged in the ACTG virology laboratory workgroups, myself as a representative of the VQA to support and facilitate these activities, and companies with such as Applied Biosystems. Our activities included sharing and developing diagnostic tools for identifying “resistance” using basic molecular methods and protocols that were employed in all our labs. Eventually, two commercial FDA-approved kits were developed based on Sanger sequencing; one is no longer available. The VQA was involved in alpha and beta testing both these products. Some commercial and hospital laboratories have developed their own in-house tests based on the same principles. The assays are used routinely as a first line diagnostic test once a person has tested positive for HIV.

2. The VQA contract runs proficiency testing programs for the clinical laboratories associated with the ACTG and their foreign site partners, and less often, for WHO. As genotyping for antiretroviral drug resistance became widely used, I was involved in developing a pilot for the VQA proficiency program for diagnostic drug-resistance genotyping. This complex assay based on Sanger sequencing includes a skill set that is more subjective, requires discerning that the quality of the data acquired is good, and the ability to “edit” and correct anomalies. This step cannot be completely automated, as it requires user judgement calls. For drug resistance assays, proficiency in these combined skills is critical in order not to misinterpret data that directly affects subsequent treatment and management strategy. To date, these types of assays are still not CAP or CLIA certified. The VQA proficiency program has been in place for ~ 20 years for Sanger sequencing. A VQA pilot to evaluate in-house NGS for drug resistance is beginning.

3. Another ongoing project is to characterize the genomic sequence of the strains that the VQA acquires from outside the US as needed. Given the diversity and rapid evolution of HIV-1 through mutation and recombination, it is useful to have more efficient means of determining the full length genetic make-up of strains. We began a collaboration with an HIV clinic in based in The Medical City, Pasig City, the Phillipines where there has been an increase in HIV infections in the past 5 years. Our past studies allowed us to develop the methodology to do this molecular epidemiologic survey to determine what strains of HIV are prominent there to help guide their treatment strategy. Once the full length genome of the strains are characterized by Sanger seqeuncing, the ability to use NGS to develop the same information will begin with consultant/collaborator at RUMC, Dr. Lela Buckingham, Unit Director Molecular Pathology.

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Technology

 

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Funding

HHSN272201200023C NIH/NIAID Virology Quality Assurance Program
Principal Investigator James Bremer, PhD

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Our team

Faculty

Key personnel

  • Aleksandra Danilovic, research assistant 1
  • Wanda Girjatowicz, research tech 3
  • Austin Hodges, research tech 3
  • Cheryl Jennings, BS, project manager
  • Marcellus Johnson, research tech 3
  • Minerva Kinser, administrative assistant
  • Nina Reiser, research tech 3
  • Salvatore Scianna, BS, laboratory supervisor
  • Jakob Stricker, research tech 3
  • Alyss Wilkey, research tech 3

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Contact us

We welcome inquiries about our research, collaborations and funding. Please contact  Diana Huang, PhD.

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