Matt Gethers/20.380 HIV Project/Technical Paper/Matt Design

Matt Design

Stem Cell Collection

• How to collect/purify RBC progenitors?
  • Are there easy kit protocols to do this?
• What are concentrations of progenitors in blood?
• How many cells do you need to engineer?
• How do you store blood and cells during the engineering?
  • Will samples be engineered in the field or sent back to lab facilities?
• Keeping samples for later use/creation of a library?
  • Need storage facilities and permission from patients.
• Could these collected progenitors be infected?
  • Possible to cure them?

Progenitor Engineering

• Need to express CD4+ and Chemokine co-receptors on surface
  • Maybe express library of receptors, or create library of blood cells with different receptors.
  • What will genetic construct look like? Separate operons for everything? Same operon? Polycistronic? Post-transcriptional/translational regulation to further refine time expression profile?
• What's vector?
• How transfecting?
• Need to express RNAses
  • Post-transcriptional/translational regulation to refine expression profile?
• Protection of progenitor from infection through siRNA.

Reintroduction of Progenitors to Patient

• Paper to demonstrate that reintroduction of ex vivo cells works.
• How many cells need to be introduced to the patient in order to establish necessary presence?
• How to deliver?
  • Injection? Into where?
• Keep samples of the engineered cells in labs?
  • Need infrastructure and permission from patients.
• Expand and differentiate cells ex vivo to test before introducing to patient?
  • Need to test every batch of cells before giving to patient.
• Need FDA blanket approval?

Differentiation in vivo

• When/how will progenitors be induced to differentiate?
• Expression profile of genes
  • Modeling of transcription specifically with respect to RNAse expression
• Need alternate regulation mechanisms to refine time expression profile?
• Differentiation kinetics, resultant population of progenitors, sink-RBCs.

Therapeutic Action

• Virions bind RBCs, get caught in trap.
• What ratio of RBCs to T-cells/virions is necessary for viral ablation?
  • Model birth, death, infection of RBCs, T-cells, HIV.
  • Is appropriate metric surface area and receptor density?
• Concerns about selecting for anti-trap HIV?
• Possible to have RBCs evolve with virus through some sort of progenitor sensor system?
• Drug supersensitivity?

IP

• Dying, take it away.