HPV OncoTect® E6, E7 mRNA KitA highly specific molecular test for early detection of cervical cancer

Revolutionizing healthcare one cell at a time

®

Numerous studies confirm that the presence of HR HPV DNA is a necessary cause of cervical cancer.1 However, only a small percentage of HR HPV DNA infections result in pre-cancerous lesions (≥ CIN2).2 Additional data now demonstrates that E6 and E7 mRNA is a more specific indication of cellular transformation and the presence of ≥ CIN2.3,5,6,7,8,9,10,19

The HPV OncoTect® E6, E7 mRNA Kit is a unique detection method that measures both the number of transforming cells and the quantity of E6, E7 mRNA in each cell.5 These two measurements precisely assess the overexpression of E6, E7 mRNA in routine patient samples collected in ThinPrep® and Surepath® vials to further refine accuracy and specificity of HPV testing.

Clinical Performance of HPV OncoTect® E6, E7 mRNA Kit:• EquivalentclinicalsensitivitytoHRHPVDNATests(95%detectionof≥ CIN2)19

• Significantincreaseinspecificity

• Uniquespecimenadequacyfeature: > Quantifies number of ectocervical and endocervical cells> Quantifies the presence of obscuring inflammatory cells

Flow cytometry simplified:• Norequirementforextractionofnucleicacid,eliminatingcrosscontamination

• Resultsavailablein4hoursforfastturnaroundtime

• Variablebatchsizes(24or96specimens)forflexiblethroughput

• Resultoutputisclearlystatedtoindicatepositiveornegative

• Adaptabletomostflowcytometers

HPV OncoTect® E6, E7 mRNA KitA highly specific molecular test for early detection of cervical cancer

Performance of HPV OncoTect® E6, E7 mRNA Test

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90

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70

60

50

40

30

20

10

HR HPV DNA Positive HPV OncoTect® E6, E7 mRNA Test Positive Normal Biopsy CIN 3

Mature Squamous Layer

Squamous Layer

Para-Basal (non-dividing) Keratinocytes

Transit Layer

Cell Cell Nucleus APC Epithelial Basement Membrane

Hybrid Capture® 2 High-Risk cobas® HPV Test18 APTIMA® HPV Assay6 HPV OncoTect® E6, E7 HPV DNA Test® 6 mRNA Test

l Sens l Spec l PPV

Overexpression of E6, E7 within the cell makes the difference Many women with positive test results from a HR HPV DNA Test will have normal biopsy. The HPV OncoTect® E6, E7 mRNA Test is only positive if there is overexpression of E6, E7. In the life cycle of the human papillomavirus, the overexpression of E6, E7 mRNA in a cell is the molecular switch leading to cervical cancer.3,4

RE VOLUTIONIZING HEALTHC ARE ONE CELL AT A T IME

The HPV virus invades the mucosa of the basal membrane of the cervix. From there the infected cell divides and spreads out in a lateral fashion. Some of the virus migrates to the suprabasal layers where viral genes are activated.

Perc

enta

ge

E6, E7inside

nucleus

APC=Antigen Presenting Cell Developed from: N. Muñoz et al. / Vaccine 24S3 (2006)

Sens = Clinical Sensitivity in CIN2+Spec = Clinical Specificity in CIN2+

PPV = Positive Predictative Value

11,12,13,14,19

RE VOLUTIONIZING HEALTHC ARE ONE CELL AT A T IME

24 & 96 Test Procedure Aspirate Add Vortex Centrifuge Waterbath

1 l 1 mL PBS l l

2 l 300 uL R1

1 mL R2 l l

3 l 1 mL R3 l l

Make the master mix by combining 100 uL R4 and 3 uL R5 per sample, vortex.

4 l 103 uL R4/R5 mix l l 30 min

1 mL R6 l l

5 l 1 mL R7 l 15 min

l l

6 l 300 uL PBS l

The sample is ready for analysis on the flow cytometer.

Star

t To

Fini

sh: A

pp

rox.

4 H

ours

l Action No Action Required

Sample Collection Aspirate Add Vortex Centrifuge Analyze

(1 hr. wait)

Simple Workflow

Product Design FeaturesAnalytical Sensitivity 5 -10 E6, E7 mRNA copies per cell

Clinical Decision Point 2% of cells expressing

Specimens ThinPrep®, SurePath®

Specimen Adequacy Control Quantification of minimum level of endocervical cells and cells per microliter (FSC versus SSC)15,16,17

External Controls Preserved cell lines well characterized as positive or negative

Result Interpretation Positive or Negative, based on clinical decision cut-off

Input Volume 1 mL

HPV Products AvailableC12100 Kit includes all reagents and FL-labeled HPV probes (100-Test Kit)

C23008 HPV Positive Control Cells

C24008 HPV Negative Control Cells

1700 El Camino Real Menlo Park, CA 94027Phone: 1 650 777 7630 www.incelldx.com

OncoTect is a registered trademark of IncellDx®, Inc.

ThinPrep is a registered trademark of Hologic, Inc. SurePath is a trademark of Becton Dickinson APTIMA is a registered trademark of Hologic, Inc. Hybrid Capture 2 is a registered trademark of QIAGEN, Inc. Cobas is a registered trademark of Roche Diagnostics Corp.©2014 IncellDx® , Inc.

1 Wallboomers JM, et al. Human papilloma virus is a necessary cause of invasive cervical cancer worldwide. J Pathol 1999: 189:12. 2 Schiffman M, Kjaer SK, Chapter 2: Natural history of anogenital human papillomavirus infection and neoplasia . J Natl Cancer Inst Monograph 31 2003;14-9. 3 zur Hausen, H. Papillomaviruses and cancer: from basic studies to clinical application. Nat Rev Cancer 2002, 2:342-50. 4 Schwartz E, et al. Structure and transcription of the human papillomavirus sequences in cervical carcinoma cells. Nature 1985;314:111-114. 5 Grundhoefer, D and B.K. Patterson, Determination of liquid-based cervical cytology specimen adequacy using cellular light scatter and flow cytometry. Cytometry. 2001 46: 340-344. 6 Szarewski, et al, Comparison of predictors for high grade cervical intraepithelial neoplasia in women with abnormal smears. CEBP 2008:17 (ii). Nov 2008. 7 Ratnam, et al, Clinical correlation of APTIMA HPV assay in comparison with hc2 test in cervical cancer screening. ISSTDR Poster July 2009. 8 Dockter, et al, Clinical performance of APTIMA HPV assay for detection of E6/E7 mRNA from HR HPV types in liquid based cytology specimens. ISSTDR Poster July 2009. 9 S Dockter, J, et al. Clinical performance of the APTIMA® HPV Assay for the detection of high-risk HPV and high grade cervical lesions. J. Clin Virol 2009 Jul; 45 Suppl 1: 555-61. 10 Miralles, R., Editorial in ginecologia y obstetrician clinica. Volumen 9 Numero 3/2008, July/September. 11 Kottaridi C., Clinical performance of human papillomavirus E6, E7 mRNA flow cytometric assay compared to human papillomavirus DNA typing. AQCH 2011, 33:305-10. 12 Patterson B K, Quantification of intracellular HPV E6/E7 mRNA expression increases the specificity and positive predictive value of cervical cancer screening compared to HPV DNA. Gynecologic Oncology 2011, 120:89-93. 13 Pierry D, Intracellular human papillomavirus E6/E7 mRNA quantification predicts CIN 2+ in cervical biopsies better than papanicolaou screening for women regardless of age. Arch Pathol Lab Med 2012, 136:956-60. 14 Irwin D, et al Clinical Virology Symposium 2006 15 Kottaridi C, Use of flow cytometry as a quality control device for liquid-based cervical cytology specimens. Cytometry. 2010 78B:37-40. 16 Narimatsu, R and B K Patterson, 2005. High throughput cervical cancer screening using intracellular human paillomavirus E6 and E7 mRNA quantification by flow cytometry. Am J Clin Pathol 123 716-723. 17 Polina R, Sturgis C, Patterson J, Patterson BK. Rapid, high throughput determination of cervical cytology specimen adequacy using a capillary-based cytometer. Cytometry. 2008 Mar;74(2):133-6. 18 cobas® HPV Test, FDA package insert. 19 Kottaridi C, mRNA and DNA detection of human papillomaviruses in women of all ages attending two colposcopy clinics. PLOS One 2012, Volume 11 Issue 11:1-9.

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