Avicentrix

HLA-Matched CTL and Tumor Cell Lines

We now have access to a unique set of HLA-matched CTL and tumor lines (colon cancer). These are well characterized lines and useful in confirming tumor lysis by drug candidates by specific/matched CD8 T cells.

We offer a number of assays including:

1. CTL assays for a quick evaluation of tumor killing
2. Flow cytometry and cytokine based readouts
3. Target expression and binding studies using tumor lines
4. Integrate additional immune cells i.e. NK, Tregs etc. to evaluate ability to suppress/proliferate CTL

Tumor-infiltrating lymphocytes are white blood cells that have left the bloodstream and migrated into a tumor. They are mononuclear immune cells, a mix of different types of cells (i.e., T cells, B cells, NK cells, macrophages) in variable proportions, T cells being the most abundant cells. In this assay, we will use a melanoma TIL and tumor from the same patient. This patient is non-responsive to KEYTRUDA (anti-PD-1) therapy. The purpose of this assay is to study how well a drug candidate will perform in a Keytruda non-responsive line.

Study Design

Melanoma TIL and tumor are co-cultured for 24 hours
Study arms will include
Anti-PD-L1 – positive control
Keytruda (anti-PD-1) – 10ug/ml concentration – negative control
Drug candidate
No drug candidate (control)
Assay run in triplicates
Readouts measured:

IFNγ
Number of drug candidates to be tested = 5

Client Provides

Drug candidate at 5ug/ml or equivalent concentration
Material handling instructions
Deliverable

Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, Flow cytometer etc.,)
Price does not include analysis of the raw data. Client will be responsible for analysis.

Cancer – HDAC inhibition assay on a KEYTRUDA non-responder TIL and tumor line

Study Rationale
Studies have recently shown that therapies targeting the epigenetic regulatory family of histone deacetylases (HDACs) have demonstrated success in the treatment of certain hematologic malignancies. HDAC inhibitors have shown to enhance anti-tumor immune responses. Studies have shown that certain HDAC classes may up-regulate the expression of PD-L1 in solid tumors.

Tumor-infiltrating lymphocytes are white blood cells that have left the bloodstream and migrated into a tumor. They are mono-nuclear immune cells, a mix of different types of cells (i.e., T cells, B cells, NK cells, macrophages) in variable proportions, T cells being the most abundant cells. In this assay, we will use a proprietary, HLA-matched melanoma TIL and tumor to study the effects of HDAC targeting drug candidates in the presence and abscence of Keytruda. We will also compare the results with two approved HDAC inhibitors – Zolinza (Vorinostat -approved for CTCL) and Kyprolis (Panobinostat – approved for multiple myeloma)

Cell Lines
We will use a TIL and tumor cell line derived from a patient that did not respond to Keytruda (anti-PD-1) therapy. The characterization profile of this unique cell line is as follows:

TIL expression: TIM-3, TIGIT, CD27, 4-1BB, GITR, PD-1, CTLA-4, LIGHT, CXCR1, CXCR2

Tumor expression: HVEM, B7-H4, CSF-1R, GITRL, 4-1BBL, CD70, CD155

Study Design
Melanoma TIL and tumor are co-cultured for 24 hours
Assay run in triplicates
Single dose study
Study arms will include:
Zolinza(Vorinostat) + TIL + Tumor
Zolinza(Vorinostat) +Keytruda (anti-PD-1) + TIL + Tumor
Kyprolis (Panobinostat) + TIL + Tumor
Kyprolis (Panobinostat) + Keytruda (anti-PD-1) + TIL + Tumor
Keytruda (anti-PD-1) + TIL + Tumor
[your hdac drug candidate] + Keytruda (anti-PD-1) + TIL + Tumor
TIL + Tumor
Tumor
Readouts measured:
Cytokines: IL-2, IL-4, IL-5, IL-9, IL-10, IL-13, IL-17, and IFN-γ.
Number of Drug Candidates Tested in this Assay Run
We plan to test 5 drug candidates in this assay. First reservations are prioritized.

Client Provides
Drug candidate concentrations for cellular assay use
Material handling instructions
Deliverable
Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, Flow cytometer etc.,)
Price does not include analysis of the raw data. Client will be responsible for analysis.

JAK inhibition assay on a KEYTRUDA non-responder TIL and tumor line

Study Rationale
Studies have recently shown that acquired resistance to PD-1 blockade in patients with advanced melanoma can be associated with loss-of-function mutations with loss of heterozygosity in JAK1/2 or in beta 2-microglobulin. Genetic studies on melanoma and colon cancer samples have shown loss-of-function mutations of JAK1 and JAK2 that may lead to defects in interferon gamma receptor signaling and in turn absence of PD-L1 expression. This assay is designed to evaluate the combination of JAK inhibitors with Keytruda (anti-PD-1) to determine whether this combination approach can overcome resistance/non-responsiveness.

Tumor-infiltrating lymphocytes are white blood cells that have left the bloodstream and migrated into a tumor. They are mono-nuclear immune cells, a mix of different types of cells (i.e., T cells, B cells, NK cells, macrophages) in variable proportions, T cells being the most abundant cells. In this assay, we will use a proprietary, HLA-matched melanoma TIL and tumor from the same patient to test JAK inhibitors in the presence and absence of Keytruda. We will compare the results to an approved JAK inhibitor – Jakafi® Ruxolitinib.

Cell Lines
We will use a TIL and tumor cell line derived from a patient that did not respond to KEYTRUDA (anti-PD-1) therapy. The characterization profile of this unique cell line is as follows:

TIL expression: TIM-3, TIGIT, CD27, 4-1BB, GITR, PD-1, CTLA-4, LIGHT, CXCR1, CXCR2

Tumor expression: HVEM, B7-H4, CSF-1R, GITRL, 4-1BBL, CD70, CD155

Study Design
Melanoma TIL and tumor are co-cultured for 24 hours
Assay run in triplicates
Single dose study
Study arms will include:
Jakafi® Ruxolitinib (JAK1/2) + TIL + Tumor
Keytruda (anti-PD-1) + TIL + Tumor
Jakafi® Ruxolitinib (JAK1/2) + Keytruda (anti-PD-1) + TIL + Tumor
[your JAK inhibitor drug candidate] + Keytruda (anti-PD-1) + TIL + Tumor
TIL + Tumor
Tumor
Readouts measured:
Cytokines: IL-2, IL-4, IL-5, IL-9, IL-10, IL-13, IL-17, and IFN-γ.
Number of Drug Candidates Tested in this Assay Run
We plan to test 5 drug candidates in this assay. First reservations are prioritized.

Client Provides
Drug candidate concentrations for cellular assay use
Material handling instructions
Deliverable
Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, Flow cytometer etc.,)
Price does not include analysis of the raw data. Client will be responsible for analysis.
Contact Us

Our calendared assay program offers our clients a cost effective approach in generating additional, value-add datasets for their immuno-modulatory programs beyond the current indication strategy. Immuno-modulatory drugs lend themselves to broad spectrum disease applications. As a result, our calendared assay program is designed to afford our clients an opportunity to cost effectively obtain additional data points without distracting internal resources.

Our team compiles a list of immune mediated assays each month and lists them on our online calendar. If you like or are interested in an assay, you may use our website or Science Exchange to place a reservation. By partnering with Science Exchange, we offer a streamlined approach for client to sign up and send their drug candidates - hassle free.

Description

Certain autoimmune diseases are a result of overacting T cells. The aim of this study is to compare effects of drug candidates on proliferation and apoptosis of human T cells stimulated with Concavalin A.

Study Design

Peripheral blood mononuclear cells (PBMC) are stimulated with concavalin A (Con A) (5μg/mL) for activation
PBMCs are then treated with the following over 5 days.
Control – Rapamycin 100uM and 200uM – Rapamycin is a member of a family of macrolide immunosuppressants that bind to and inhibit the FK506 binding protein (FKBP) proline rotamase.
Drug candidate – equivalent drug concentration to control
Cells will be examined using carboxyfluorescein succinimidyl ester (CFSE) assay for proliferation.
Readouts:

Cell apoptosis will be analyzed by FITC annexin V/PI staining and flow cytometry.
Number of drug candidates to be tested = 5

Client Provides

Drug candidate at 100uM and 200 um or equivalent concentration
Material handling instructions
Deliverable

Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, Flow cytometer etc.,)
Price does not include analysis of the raw data. Client will be responsible for analysis.

Description

Rheumatoid Arthritis (RA) is a chronic inflammatory disorder affecting many joints, including those in the hands and feet. It is an autoimmune disease in which the body’s immune system mistakenly attacks the joints. In this study, we will evaluate the alterations in RA PBMC cytokine production in response to a drug candidate.

Study Design

10 RA patients and 10 normal controls
Control stimulation via phytohemagglutinin (PHA)
PBMCs will be cultured for 48 hours in:
Medium alone
Medium with drug candidate per ml for 48 h
Medium with 5ug PHA per ml for 48 h
Culture supernatants are collected for analysis
Assay run in triplicates
Readouts measured:

Th1/Th2/Th17 8-Plex Human Panel (Multiplex)

Number of drug candidates to be tested = 5

Client Provides

Drug candidate at 5ug/ml or equivalent concentration
Material handling instructions
Deliverable

Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, )

Description

Multiple sclerosis (MS) is an autoimmune demyelinating disorder of the central nervous system (CNS). The disease is characterized by inflammatory lesions in the white matter of the CNS, consisting of a specific immune response to the myelin sheath. In this study, we will evaluate the alterations in MS PBMC cytokine production in response to a drug candidate

Study Design

10 MS patients and 10 normal controls
Control stimulation via phytohemagglutinin (PHA)
PBMCs will be cultured for 48 hours in:
Medium alone
Medium with drug candidate per ml for 48 h
Medium with 5ug PHA per ml for 48 h
Culture supernatants are collected for analysis
Assay run in triplicates
Readouts measured:

Th1/Th2/Th17 8-Plex Human Panel (Multiplex)

Number of drug candidates to be tested = 5

Client Provides

Drug candidate at 5ug/ml or equivalent concentration
Material handling instructions
Deliverable

Price includes the delivery of the raw data files obtained directly from the instrumentation (e.g. Luminex, Flow cytometer etc.,)
Price does not include analysis of the raw data. Client will be responsible for analysis

Bioanalytical Assays Services

Protein Purification and Quantification Services

KEYTRUDA (anti-PD-1) resistant TIL and tumor line

Tumor-infiltrating lymphocytes are white blood cells that have left the bloodstream and migrated into a tumor. They are mono-nuclear immune cells, a mix of different types of cells (i.e., T cells, B cells, NK cells, macrophages) in variable proportions, T cells being the most abundant cells. Studies have shown that certain immune checkpoint molecules are up-regulated in response to checkpoint therapy and may play a role in mediating resistance.
We use a TIL and tumor cell line derived from a patient that did not respond to KEYTRUDA (anti-PD-1) therapy. The characterization profile of this unique cell line is as follows:
TIL expression: TIM-3, TIGIT, CD27, 4-1BB, GITR, PD-1, CTLA-4, LIGHT, CXCR1, CXCR2
Tumor expression: HVEM, B7-H4, CSF-1R, GITRL, 4-1BBL, CD70, CD155

Studies conducted with this cell line:

1. Flow cytometry analysis of cell surface markers (i.e. checkpoints) in response to drug candidate
2. Cytokine readouts in response to drug candidate
3. Studying macrophages, Tregs sub-populations and their changes in response to drug candidate
4. Mechanisms of resistance to Keytruda (anti-PD-1)
5. 3D assays to study chemokines and chemokine receptors.
6. Other mechanism, function and biology based questions in IO

Human functional tissue assays Services