Interferon-Inducible Gene Expression Kit As a Potential Diagnostic Test for Anifrolumab: Analytical Validation for Use in Clinical Trials

Philip Z. Brohawn¹, Brandon W. Higgs¹, Sabina Patel², Adrian Moody³, Peter Cooper³ and Koustubh Ranade⁴, ¹MedImmune LLC, Gaithersburg, MD, ²AstraZeneca, Cambridge, United Kingdom, ³QIAGEN Manchester Ltd., Manchester, United Kingdom, ⁴Translational Medicine, MedImmune LLC, Gaithersburg, MD

Meeting: 2018 ACR/ARHP Annual Meeting

Keywords: Interferons and systemic lupus erythematosus (SLE)

Session Information

Date: Tuesday, October 23, 2018

Title: Systemic Lupus Erythematosus – Clinical Poster III: Treatment

Session Type: ACR Poster Session C

Session Time: 9:00AM-11:00AM

Background/Purpose: Anifrolumab is a fully human monoclonal antibody that binds to the type I interferon (IFN) receptor. Its efficacy and safety in the treatment of patients with moderate-to-severe systemic lupus erythematosus (SLE) is being evaluated in Phase III clinical trials. Patients with a high IFN-inducible gene signature (IFNGS) respond better to anifrolumab than those with a low IFNGS.¹ AstraZeneca and QIAGEN have developed an in vitro, reverse transcriptase polymerase chain reaction (RT-PCR)-based diagnostic test for type I IFN-inducible gene expression (IFIGx) that enables the detection of expression of 4 IFN-inducible genes (IF127, IF114, IF114L, and RSAD2) relative to 3 housekeeping genes. The efficacy of anifrolumab will be evaluated in patients with high and low IFNGS. In the present study, we conducted analytical validation of the therascreen^® IFIGx RGQ RT-PCR kit (IFIGx kit) for use in clinical trials, as well as to potentially support future regulatory submissions.

Methods: Measurements were performed on mRNA extracted from whole blood from adults with SLE. Patients were identified as “type I IFNGS test‒high” or “type I IFNGS test‒low” based on a generated score. Analytical validation comprised 6 studies that measured lot interchangeability, linearity, repeatability, reproducibility, cross contamination, and system verification.

Results: Reproducibility and repeatability were >96% and 100%, respectively, with linearity of score observed at ±10% defined input concentration. No cross contamination in reverse transcription or PCR steps was observed. Results of all studies validated the IFIGx kit (Table).

Conclusion: The analytical validation of the therascreen^® IFIGx RGQ RT-PCR kit demonstrates this is a robust, reproducible diagnostic test for type I IFNGS. The IFIGx kit was shown to be valuable in a prior anifrolumab study,¹ and clinical utility is being further established in the anifrolumab Phase III study to potentially support regulatory filings.

Reference: 1. Furie et al. Arthritis Rheumatol. 2017;69:376-86.

Table: Summary of Analytical Validation of Anifrolumab IFIGx Test
Study	Description	Results
Lot interchangeability	Verification that scores and assay Ct values were robust when different lots of kit components were used Acceptance criterion: <0.58 Ct	Lot interchangeability verified Largest observed change=0.16 Ct
Linearity	Verification—using linear and quadratic regression analyses—that mRNA input concentration (10 ng/µL) is in the assay’s linear range	Linearity verified No change in score over the concentration range on either side of 10 ng/µL=0.0043 Ct
Repeatability	Verification of Dx result repeatability when the same operator tested 60 random samples using the same kit lot and instrument	Repeatability verified Observed repeatability=100%
Reproducibility	Verification of Dx result reproducibility when multiple operators tested 48 random samples using multiple kit lots and instruments at different sites For verification, overall rate of correct calls must be ≥95%	Reproducibility verified Overall rate=99.7% After 6 samples with values close to the cut-off were added for further confirmation, overall rate=96.5%
Cross contamination	Investigation of inter- and intra-run cross contamination	No cross contamination No cross contamination found in reverse transcription or PCR steps
System verification	Verification of functionality and utility of the IFIGx software and IFIGx assay package	System verification confirmed Software flags produced as expected
Ct, cycle threshold (PCR cycle at which fluorescence rises above background level); Dx, diagnostic; IFIGx, interferon-inducible gene expression; PCR, polymerase chain reaction; mRNA, messenger ribonucleic acid.

Disclosure: P. Z. Brohawn, MedImmune LLC, 3; B. W. Higgs, MedImmune LLC, 3; S. Patel, AstraZeneca, 3; A. Moody, QIAGEN Manchester Ltd, 3; P. Cooper, QIAGEN Manchester Ltd, 3; K. Ranade, MedImmune LLC, 3.

To cite this abstract in AMA style:

Brohawn PZ, Higgs BW, Patel S, Moody A, Cooper P, Ranade K. Interferon-Inducible Gene Expression Kit As a Potential Diagnostic Test for Anifrolumab: Analytical Validation for Use in Clinical Trials [abstract]. Arthritis Rheumatol. 2018; 70 (suppl 9). https://acrabstracts.org/abstract/interferon-inducible-gene-expression-kit-as-a-potential-diagnostic-test-for-anifrolumab-analytical-validation-for-use-in-clinical-trials/. Accessed .

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