Swap the envelope, keep the payload

Lentiviral vector pseudotyping

Virology · Colorado State University · 2016

Hu S, Mohan Kumar D, Sax C, Schuler C, Akkina R. Pseudotyping of lentiviral vector with novel vesiculovirus envelope glycoproteins derived from Chandipura and Piry viruses. Virology 488 (2016) 162–168.

PMID 26650691 · DOI 10.1016/j.virol.2015.11.012 · Dept. of Microbiology, Immunology & Pathology, Colorado State University

What the problem was

A lentiviral vector is a delivery mechanism: a hollowed-out virus that carries a genetic payload into a target cell. What decides which cells it can enter is not the payload but the envelope glycoprotein on its surface — the protein that meets the cell membrane.

Essentially the whole field used one envelope, VSV-G, because it works broadly. But it transduces some cell types poorly, and human complement inactivates it — which constrains where the vector can be used at all. One dominant interface, with the whole field’s limitations inherited from it.

What we did

We evaluated two serologically distinct envelopes from other vesiculoviruses — Chandipura (CNV-G) and Piry (PRV-G) — as alternatives. Both produced high-titre pseudotyped vectors and transduced a range of cell types efficiently across species.

The vector core did not change. Only the envelope did. That is the entire move: the payload and the machinery that packages it stay fixed while the surface that determines reach is swapped out, and the alternatives turn out to reach places the incumbent could not.

HIV-1 tissue reservoirs

CSU Journal · Colorado State University

Schuler C, Bennett M, Akkina R. HIV-1 localization and distribution within humanized Rag1^−/−γc^−/− mice organ tissue. CSU Journal.

Dept. of Microbiology, Immunology & Pathology, Colorado State University

What we did

A Rag-hu mouse is an immune-deficient mouse irradiated within days of birth and injected with human CD34+ hematopoietic stem cells, which rebuild it with a human immune system — human T and B cells, macrophages, dendritic cells. That matters because it can then be infected with HIV-1 itself rather than with the monkey analogue SIV, and it holds a chronic infection for over a year, the way an untreated patient does.

Three of these mice, twenty-three weeks into confirmed viremia, were necropsied and seven compartments taken: blood, brain, bone marrow, lymph nodes, thymus, liver, spleen. Each was quantified by PCR for viral copies and normalized against human β-actin, so the number reported is virus per human cell present in that tissue, not virus per gram of mouse. Human engraftment is uneven across organs, so without that denominator the brain and the spleen are not comparable measurements at all.

Why it mattered

Antiretroviral therapy turned HIV from fatal to chronic, but it does not cure it. Stop the drugs and viremia returns, because HIV integrates into a cell’s own genome and waits there as a provirus — invisible to the therapy, which only interrupts active replication. Clearing those reservoirs is the remaining problem, and you cannot clear what you have not located: latent virus is thought to sit in memory CD4+ T cells, but also in macrophages and dendritic cells in gut tissue, and in perivascular macrophages and microglia in the brain, which no blood draw will ever see.

The reason the question gets asked in a mouse is cost. Trialling candidate therapies in patients, or in rhesus macaques carrying SIV, is expensive and slow enough that only a handful of ideas ever get tested. A Rag-hu mouse is a fraction of that and can be run in numbers, which is what turns eradication work from a few one-off experiments into something closer to a screening programme.

So the measurement does double duty. It gives a reservoir map, and it tests the model itself — if the virus distributes across a Rag-hu mouse’s organs the way it does in a person, every cheap study built on that model inherits the credibility.

Emerald Cloud Lab

Senior Lab Scientist · 2015–2016

Emerald ran a laboratory as remote infrastructure — scientists submitted experiments as code and robots executed them. I designed and automated robotic DNA and PNA synthesis, cell culture, and molecular experimentation on that platform.

It is the earliest thing on this site that looks like the rest of it: a bench protocol is a script, an instrument is a driver, and a reproducible experiment is a deterministic pipeline with the sources of variance named and controlled. Automating a synthesis forces the same discipline as automating a deployment — every implicit step a human was quietly performing has to become explicit before anything runs unattended.

Clinical biomarker software

Health Restored · 2020–2022

Software that automated lab interpretation and physician reporting — taking raw biomarker panels and producing the structured read a clinician would otherwise assemble by hand.

This is the hinge between the two halves of the CV. It is also the direct ancestor of PrimeMaxing, which does the same work at a different scale and under a stricter privacy boundary — and of the Centennial Labs triage system, which handles the same documents at the moment they arrive.

Education

Colorado State University

• B.S. Biomedical Science — Highest Honors
• Dean’s List
• Commencement keynote speaker

The virology above was done in the Akkina lab in the Department of Microbiology, Immunology and Pathology.