Predictive; Prognostic; Therapeutic management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Guidance for management

Patients with advanced solid and hematologic malignancies, considered for targeted therapies, unlikely to respond to conventional chemotherapies, or for whom no conventional therapies exist.

xT panel sample processing and nucleic acid extraction Expert pathologist assessment of overall tumor content and percent tumor cellularity as a ratio of tumor to normal nuclei verified specimens must meet a 20% threshold. Microdissection is utilized as required to enrich specimens below the 20% threshold. Solid tumor total nucleic … xT panel sample processing and nucleic acid extraction Expert pathologist assessment of overall tumor content and percent tumor cellularity as a ratio of tumor to normal nuclei verified specimens must meet a 20% threshold. Microdissection is utilized as required to enrich specimens below the 20% threshold. Solid tumor total nucleic acid was extracted from FFPE tissue sections using Chemagic 360 sample-specific extraction kits (Perkin Elmer) and digested by proteinase K. RNA was purified from the total nucleic acid by DNase-I digestion. xT panel DNA library construction and sequencing DNA sequencing of 648 genes are performed as previously described [Beaubier N, et al. Oncotarget. 2019; 10:2384-96]. Briefly, 50-300 nanograms (ng) of DNA for each tumor sample is mechanically sheared to an average size of 200 base pairs (bp) using a Covaris Ultrasonicator. DNA libraries are prepared using the KAPA Hyper Prep Kit, hybridized to the xT probe set, captured using Streptavidin-coated beads and amplified with the KAPA HiFi Library Amplification Kit. Between 50-300 ng of RNA for each tumor sample is heat fragmented in the presence of magnesium to an average size of 200 bp. Library preps are hybridized to the proprietary exome panel (Integrated DNA Technologies) and target recovery was performed using Streptavidin-coated beads, followed by amplification with the KAPA HiFi Library Amplification Kit. The amplified target-captured DNA tumor library is sequenced using 2x126bp PE reads to an average unique on-target depth of 500x (tumor) and 150x (normal) on an Illumina NovaSeq 6000. The amplified target-captured RNA tumor library is sequenced using 2x76bp PE reads to an average of 50M reads on an Illumina Novaseq 6000. Samples are further assessed for uniformity with each sample required to have 95% of all targeted bp sequenced to a minimum depth of 300x. Variant detection, visualization, and reporting are performed as previously described [Beaubier N, et al. Oncotarget. 2019; 10:2384-96]. View more

Tests performed
Entire test performed in-house

Single nucleotide variants in actionable genes were detected down to a variant allele fraction (VAF) of 5% with 98.2% Se, over 99.9% Sp in FFPE tissue specimens with at least 20% tumor fraction. Insertions/deletions up to 100bp in size were detected down to 5% VAF with 91.8% Se, 99.9% Sp. … Single nucleotide variants in actionable genes were detected down to a variant allele fraction (VAF) of 5% with 98.2% Se, over 99.9% Sp in FFPE tissue specimens with at least 20% tumor fraction. Insertions/deletions up to 100bp in size were detected down to 5% VAF with 91.8% Se, 99.9% Sp. Copy number variants (CNVs) were detected with 92% Se down to 30% tumor purity for copy number gains and 40% tumor purity for copy number losses. Fusions found in solid and hematological tumors were detected down to 10% tumor purity at 91.7% Se and 99.9% Sp. And finally, in specimens with at least 30% tumor fraction, the accuracy of MSI status call was 99.9%. View more

Tumor required to be at least 20%, but preferably 30% of the sample by a ratio of tumor nuclei to benign nuclei, to include MSI status report. Tumor normal match and 40% tumor content are required for Homologous Recombination Deficiency (HRD) testing.

Is proficiency testing performed for this test? Help
Yes

Method used for proficiency testing: Help
Formal PT program

PT Provider: Help
American College of Medical Genetics / College of American Pathologists, ACMG/CAP

Software used to interpret novel variations Help
N/A

Laboratory's policy on reporting novel variations Help
Tempus reports novel variants if detected somatically and classified as pathogenic, likely pathogenic, or VUS. Tempus reports novel germline variants if pathogenic and likely pathogenic for reportable disease conditions.