therascreen PIK3CA RGQ PCR Kit (CA)

• Reliable detection of clinically relevant mutations in the PIK3CA gene
• High sensitivity and specificity
• Results in less than two working days
• Optimized reagents and reaction mixes
• Automated data analysis using Rotor-Gene AssayManager v2.1 software

The therascreen PIK3CA RGQ PCR Kit is a real-time qualitative in vitro diagnostic PCR test for the detection of 11 mutations in the phosphatidylinositol 3-kinase catalytic subunit alpha (PIK3CA) gene using a sample of DNA extracted from either formalin-fixed, paraffin-embedded (FFPE) breast tumor tissue or K₂EDTA anticoagulated blood plasma, taken from a patient with breast cancer.

The therascreen PIK3CA RGQ PCR Kit is an in vitro diagnostic (IVD) test to help clinicians identify breast cancer patients that may be eligible for treatment, based on the detection of a PIK3CA mutation.

The SOLAR-1 study (CBYL719C2301) was a randomized, double-blinded, placebo-controlled, international, multicenter Phase III clinical trial that compared treatment with PIQRAY (alpelisib) plus fulvestrant with placebo plus fulvestrant in men and postmenopausal women with hormone receptor-positive, HER2-negative advanced breast cancer that had progressed on or after aromatase inhibitor treatment. A total of 572 breast cancer patients were enrolled into two cohorts, with or without a PIK3CA mutation. Patients were randomized to receive PIQRAY (alpelisib) 300 mg plus fulvestrant or placebo plus fulvestrant in a 1:1 ratio. The primary endpoint was progression-free survival (PFS) determined using RECIST v1.1 criteria, based on investigator assessment.

Results using FFPE tissue specimens
SOLAR-1 showed that in patients whose tumors harbored specific PIK3CA mutations, treatment with PIQRAY (alpelisib) plus fulvestrant prolonged median PFS by a clinically meaningful 5.3 months (11 months overall) compared with treatment with placebo plus fulvestrant, and conferred an estimated 35% risk reduction in disease progression or death.

Analysis of the PIK3CA mutant-positive patient subset identified by the test (347 patients) demonstrated that those receiving PIQRAY (alpelisib) plus fulvestrant had an estimated 36% lower risk of disease progression or death (HR = 0.64; 95% CI: 0.48, 0.85) than patients receiving placebo plus fulvestrant.

By contrast, PFS was also estimated in the therascreen PIK3CA RGQ PCR Kit-negative population and no PFS benefit was observed in those patients (HR = 0.85; 95% CI: 0.58, 1.25).

The importance of establishing PIK3CA mutation status when identifying patients for treatment with PIQRAY (alpelisib) plus fulvestrant is therefore clear; only patients whose tumors harbor actionable PIK3CA mutations are likely to experience a clinically meaningful increase in PFS.

Results using plasma specimens
K₂EDTA anticoagulated peripheral venous whole blood clinical plasma specimens collected from breast cancer patients randomized in SOLAR-1 prior to initiation of study treatment (baseline) were tested retrospectively with the therascreen PIK3CA RGQ PCR Kit to evaluate concordance between tissue and plasma results.

Of the 328 therascreen PIK3CA RGQ PCR Kit tissue-positive patients, 179 were therascreen PIK3CA RGQ PCR Kit plasma-positive. Of the 215 therascreen PIK3CA RGQ PCR Kit tissue-negative patients, 209 were therascreen PIK3CA RGQ PCR Kit plasma-negative. There were no invalid plasma results (Table 1).

Table 1. Correspondence table between therascreen PIK3CA RGQ PCR Kit tissue results and therascreen PIK3CA RGQ PCR Kit plasma results

Agreement (PPA, NPA and OPA) between the therascreen PIK3CA RGQ PCR Kit plasma and therascreen PIK3CA RGQ PCR Kit tissue results was calculated using the therascreen PIK3CA RGQ PCR Kit tissue results as reference (Table 2). The point estimates of PPA, NPA and OPA were 55%, 97% and 72%, respectively.

Table 2. Agreement between therascreen PIK3CA RGQ PCR Kit plasma results and therascreen PIK3CA RGQ PCR Kit tissue results using the therascreen PIK3CA RGQ PCR Kit tissue results as reference

The therascreen PIK3CA RGQ PCR Kit therefore enables reliable selection of breast cancer patients with PIK3CA alterations who may be eligible for treatment with PIQRAY (alpelisib).

The therascreen PIK3CA RGQ PCR Kit is comprised of six reaction mixes; one control reaction targeting exon 15 and five mutation-specific reactions utilized to detect 11 mutations in exons 7, 9 and 20 of the PIK3CA gene (Exon 7: C420R; Exon 9: E542K; E545A, E545D [1635G>T only], E545G, E545K, Q546E, Q546R; and Exon 20: H1047L, H1047R, H1047Y). Allele-specific technology allows accurate and highly reproducible detection of mutations; DNA is selectively amplified using ARMS primers, probes and PCR clamps, with sensitive signal detection using the Rotor-Gene Q MDx 5plex HRM (CA) instrument. Result reporting is fully automated. If both the positive and no template controls are valid and the sample internal controls are valid, the PIK3CA alteration status will be displayed in the software.

The simple and straightforward testing workflow begins with manual DNA extraction from either FFPE breast tumor tissue (using the QIAamp DSP DNA FFPE Tissue Kit) or from K₂EDTA anticoagulated plasma (using the QIAamp DSP Circulating Nucleic Acid Kit), followed by sensitive real-time PCR on the Rotor-Gene Q MDx 5plex HRM (CA) instrument. Rotor-Gene AssayManager software rapidly and accurately determines mutations and reports results, informing the system operator if one or more of the 11 mutations detected by the kit are present in each sample. The kit is a qualitative assay that can yield results in less than two working days.

The therascreen PIK3CA RGQ PCR Kit enables qualitative detection of 11 mutations in the PIK3CA gene for in vitro diagnostic use. It is an IVD assay to identify breast cancer patients that may be eligible for treatment, based on the detection of a PIK3CA mutation.