ANIXA DIAGNOSTICS

LIQUID BIOPSY BASED ON FLOW CYTOMETRY AND AI TO DETECT THE EXISTENCE OF A SOLID TUMOR

Amit Kumar, Ph.D.

President and CEO

1---Subsidiary of ITUS Corporation

Technology uses the immune system as a signal amplifier

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IncreasingWBC Count

Solid TumorBacterial Infection

Healthy

Patients with solid tumors usually don’t have highly elevated WBCs

The frequency of certain Leukocytes sub-segments often change in tumor patients

Immuno-profiling can provide information regarding tumor status

WHITE BLOOD COUNT (WBC) INDICATES DISEASE

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• Malignancies do induce an immune response, though the increase in overall WBC count is modest• Tumors consist of cancer cells and immune cells• Research shows that high levels of a type of immune

cell called MDSC are found in malignant tumors• MDSC function is corrupted by the malignancy to

suppress general immune and T-cell functions.

Increased MDSC levels are also found in the circulation of cancer patients.

Myeloid Derived Suppressor Cell (MDSC)

MDSCS AND ONCOLOGYResearch supports the following conclusions

• Three phenotypically distinct MDSC populations

• E-MDSCs- Early

• M-MDSCs- Monocytic

• PMN-MDSCs- Poly-morphonuclear

• All MDSC are implicated in immune suppression

Published Literature

• Circulating MDSC levels correlate with Prognosis and Outcome• High MDSCs- poor outcome, poor response to therapy • Low MDSCs- better outcome, better response to therapy

• When Tumors are responding to therapy, circulating MDSC levels change• Monitoring of Tumor Response by MDSC analysis is being

conducted in multiple clinical trials• Other Lymphocytes also respond

• After a tumor has been resected, circulating MDSC levels change dramatically

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Hypothesis: Measuring MDSC levels could identify tumor-bearing patients

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BD LSR II High-speed Analyzer

How Do We Measure MDSCs? – Flow Cytometry

Increased MDSC Numbers in Peripheral Blood are Associated With Solid Tumors

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Angell et al. Thyroid 26(3) 2016.

Diaz-Montero et al. Cancer Immunol Immunother 58(1) 2009.

Zhang et al. PLoS ONE 8(2) 2013.

Dominguez et al. Clinical Cancer Research 23(12) 2017.

Thyroid BreastColorectal

Advanced Solid Tumors Lung

Chen et al. Oncology Letters 14 2017.

MDSCs can be measured in peripheral blood using flow cytometry – a process

known as immunophenotyping

Cancer Stage

MDS

C Co

unt

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ARTIFICIAL INTELLIGENCE ANALYSIS

Training Calls

Neural NetworkNormal Patient Samples

Cancer Patient Samples

Patient Sample

Malignancyor

Normal

Breast CancerPredicted

Naïve Testing All Samplesn = 56 n = 144

TB nTB TB nTB

Measured

TB 24 2 58 6nTB 4 26 11 69

Sens. (%) 92.31 90.62Spec. (%) 86.67 86.25Acc. (%) 89.29 88.19PPV (%) 85.71 84.06NPV (%) 92.86 92AUROC(95% CI)

0.8987(0.8047 to 0.9927)

0.9209(0.8705 to 0.9713)

ROC analysis: NN vs Manual Gating Vs Mammography/Ultrasound

All Naïve Samples were Early Stage (Stage 1 and 2)

Prostate CancerPredicted

Naïve Testing All Samplesn = 36 n = 146

TB nTB TB nTB

Measured

TB 16 2 68 5nTB 1 17 1 72

Sens. (%) 88.89 93.15Spec. (%) 94.44 98.63Acc. (%) 91.67 95.89PPV (%) 94.12 98.55NPV (%) 89.47 93.51AUROC(95% CI)

0.929(0.8418 to 1)

0.9797(0.9592 to 1)

ROC analysis: NN vs Manual Gating vs PSA Test

N = 73Predicted

Manual Gating Neural NetworkProstate Healthy Prostate Healthy

Measured

Prostate 16 19 33 3Healthy 10 28 3 34

Sens. (%) 45.71 91.67Spec. (%) 74.36 91.89Acc. (%) 60.81 91.78PPV (%) 61.54 91.67NPV (%) 60.42 91.89AUROC(95% CI)

0.597(0.4658 to 0.7282)

0.9354(0.8723 to 0.9985)

*Manual gating assumed a 75% specificity as the cutoff

Prostate Cancer Study:Memorial Sloan Kettering Data

Key Conclusions• Artificial Intelligence (AI) better than current methods used by scientists or physicians to make calls

• May work for all cancers

• To date, identified in 20 different types- May work for all tumor types

• Multiple samples:

• Single sample:

• Cost to run test is a few dollars12

Breast Prostate Lung Colon

Pancreatic Melanoma Ovarian Liver

Bladder Cervical Endometrial Thyroid

Gastric Head and Neck Testicular

Osteosarcoma (bone cancer)

Leiomyosarcoma (cancer of the soft tissue)

Liposarcoma (cancer of the connective tissue)

Vulvar

Appendiceal