scale-X Hydro

Stages

1. 1.
   Producer cell line generation and comparison(library_build)

   This stage identifies the better stable producer-cell-line background before committing to process optimization and scale-up.

   Selection: Generate stable producer cell lines from GPRG and GPRTG packaging cell lines and compare resulting LV titer and CD34+ transduction performance.

2. 2.
   Manufacturing technology comparison in continuous perfusion mode(broad_screen)

   This stage identifies the better production hardware platform after selecting the producer cell line.

   Selection: Compare traditional flatware systems, iCELLis Nano, and scale-X Hydro using the GPRTG stable producer cell line in continuous perfusion and recirculation mode.

3. 3.
   Scale-up production in scale-X Carbo(confirmatory_validation)

   This stage confirms that the selected process can be transferred to a larger manufacturing scale.

   Selection: Scale the selected process from scale-X Hydro to scale-X Carbo and measure total TU output across harvests.

4. 4.
   Functional transduction validation in CD34+ cells(confirmatory_validation)

   This stage checks that process optimization and scale-up preserve the intended functional output of the LV product.

   Selection: Test whether LV produced by the optimized continuous perfusion process efficiently transduces CD34+ cells and achieves measurable VCN at defined MOI.

Steps

1. 1.
   Evaluate transfection reagents to generate stable producer cell lines from GPRG and GPRTG packaging cell linesproducer cell line candidates

   Create stable producer cell lines expressing WAS or GFP transgenes from two Tet-off regulated packaging-cell-line backgrounds.

   Producer cell lines had to be generated before their LV output and downstream manufacturing suitability could be compared.

2. 2.
   Compare producer cell lines by LV titer and CD34+ transduction performancecompared producer cell line platforms

   Identify the better producer cell line for downstream process optimization.

   The study needed to choose the stronger producer background before investing in bioreactor process comparison and scale-up.

3. 3.
   Optimize continuous perfusion and recirculation LV production using the selected GPRTG producer cell line across flatware, iCELLis Nano, and scale-X Hydroselected producer line and compared manufacturing platforms

   Determine which production technology gives better LV productivity per surface area under the optimized process mode.

   After selecting the better producer cell line, the next decision was which manufacturing platform best supports scalable production.

4. 4.
   Scale the selected process from scale-X Hydro to scale-X Carbo and collect multiple harvestsscale-up manufacturing platforms

   Demonstrate that the selected continuous perfusion process can be transferred to a larger 10 m2 platform while maintaining high output.

   Scale-up follows platform selection because only the preferred process configuration is worth testing at larger manufacturing scale.

5. 5.
   Test LV from the optimized process for CD34+ cell transduction and VCN at MOI 10

   Confirm that the optimized and scaled manufacturing process still yields functionally active LV for the intended ex vivo application.

   Functional testing is done after process optimization and scale-up to ensure manufacturing improvements did not compromise biological performance.