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The immune system plays an active role in both the prevention and

the promotion of cancer dependent upon its interaction with the

tumor cells. The roles of both macrophages and T-cells in cancer

progression have been heavily studied over the past few years.

Macrophages have been found to be either tumor promoting or

tumor preventing depending upon their differentiation status and the

tumor microenvironment while the homing and cell destroying

capabilities of T-cells have been manipulated to effect better, more

specific tumor cell cytotoxicity through the development of therapies

such as chimeric antigen receptor T-cells (CAR-T cells). Unfortunately

the majority of research in the area of immune-oncology has relied

upon either 2D cell culture or animal models. While a large amount of

information has been learned from these models, it has been well

established that 2D cell culture does not mimic in vivo biology and the

immune system of mouse models differs from that of humans in

numerous ways including T-cell subsets, cytokine receptors, and

costimulatory molecule expression. To overcome these limitations, we

have developed a number of 3D in vitro tissue models including multi-

cell type models of glioblastoma (GBM), breast, and ovarian cancer.

Complex, 3D Tissues for Modeling the Immune Response in Cancer and Predicting the Activity of ImmunotherapiesTeresa M. DesRochers1, Lillia Holmes1, Qi Guo1, Lauren O’Donnell1, Stephen Shuford1, Larry Puls2, Jeffrey Elder2, Jeff Edenfield2, Howland Crosswell1

1KIYATEC Inc.; Greenville, South Carolina 29605 USA | 2Greenville Health System; Greenville, South Carolina 29605 USA |

Background Ovarian – Immune Cells Affect Viability

Immune Tumor Models

Conclusions

Figure 4: The presence of neurospheres and huvecs affects the differentiation of

CD14+ PBMCs. Scaffolds containing NS/HUVEC or PBMCs were cultured together in

perfusion for 14 days. Both metabolism (A) and cell numbers (B) increased in the

NS/HUVEC scaffolds. However, while the PBMC scaffolds increased in metabolic activity,

they lost cell numbers. The PBMCs were labeled with PKH26. After 1 week in culture a very

small number of PKH26 cells were found in the NS/HUVEC scaffolds indicating a lack of

cellular migration by the PBMCs (C). At day 7 (D) and day 14 (E), the PBMCs were removed

from their scaffolds and examined for macrophage markers. At both days the cells were

positive for both CD163 and CD206 but the signal was reduced at day 14 compared to day

7. *p<0.05, **p,0.01, error bars = standard deviation, n=3.

Glioblastoma

• The presence of CD3+ cells negatively affects cell viability in ovarian cancer while CD14+

cells do not

•Ascitic fluid of ovarian cancer patients affects drug response with EpCAM+ cells from

ascitic fluid reflecting the response of the primary tumor.

• The different cell types of a tumor can affect the differentiation of CD14+ immune cells

•Future Directions: preclinical immuno-oncology agent testing and

predictive assay for checkpoint inhibitor responders

#5119

Resected

Tumor

Mince & Wash

ascitic fluid CD14+

CD3+

Ovari

an

BreastM1/M2 Macrophages

+ ECM

Neurospheres + ECM

GBM

Figure 1: Model Development. (Ovarian) Both tumors and matched ascitic fluid are

collected from patients. Tumor cells are then combined with immune cells in 3D spheroid

assays. (Breast) Tumors and matched ascitic fluid are collected from patients. Tumor cells,

endothelial cells, fibroblasts, and fat are then combined with immune cells in 3D

microtumors. (GBM) Tumors are collected from patients and neurospheres are established.

These are combined with differentiated macrophages from normal blood in 3D

microtumors.

0

500

1250

2500

5000

0

5 0 0 0

1 0 0 0 0

1 5 0 0 0

2 0 0 0 0

2 5 0 0 0

O V B 0 1 & M a tc h e d C D 3 +

C D 3 + c e lls

Bla

nk

ed

R

LU

T u m o r

T u m o r + C D 3 +

p < 0 .0 5

0

500

1250

2500

5000

0

5 0 0 0 0

1 0 0 0 0 0

1 5 0 0 0 0O V B 0 3 & M a tc h e d C D 1 4 +

C D 1 4 + C e lls

T u m o r

T u m o r + C D 1 4 +

Bla

nk

ed

R

LU

Figure 2: Immune cells co-cultured

with patient-matched tumor cells

can affect proliferation in a cell-

type dependent manner. (A) When

ovarian tumors cells were co-cultured

with matched CD3+ cells for 96 hours,

there was a significant drop if the

viability of the tumor cell/CD3+

combo that was inconsistent with the

viability of the CD3+ cells alone. (B)

When ovarian tumor cells were co-

cultured with matched CD14+ cells for

96 hours, viability of the tumor

cell/CD14+ combos were consistent

with the viability of the CD14+ cells

alone.

Ovarian – Drug Response Profiling

NS

/HU

VE

C

PB

MC

0

1 0 0 0 0

2 0 0 0 0

3 0 0 0 0

4 0 0 0 0

5 0 0 0 0

P re s to B lu e

S c a ffo ld T y p e

Bla

nk

ed

RF

U

D a y 0

D a y 7

D a y 1 4

*

*

*

**

NS

/HU

VE

C

PB

MC

0

1 0

2 0

3 0

4 0

5 0

P ic o G re e n

S c a ffo ld T y p e

DN

A c

on

ten

t (u

g/m

L) D a y 0

D a y 7

D a y 1 4

*

*

-4 -2 0 2 4

0

5 0

1 0 0

1 5 0

A s c e t ic F lu id - P re -S u rg e ry

L o g D o s e (u M )

% V

iab

ilit

y

C is p la tin

T o p o te c a n

L Y 2 9 4 0 0 2

D o x o ru b ic in

G e m c ita b in e

C a rb o p la tin

T ra m e tin ib

A fa tin ib

P a c lita x e l

A. B.

C. D.

-4 -2 0 2 4

0

5 0

1 0 0

1 5 0

T u m o r

L o g D o s e (u M )

% V

iab

ilit

y

C is p la tin

T o p o te c a n

L Y 2 9 4 0 0 2

D o x o ru b ic in

G e m c ita b in e

C a rb o p la tin

T ra m e tin ib

A fa tin ib

P a c lita x e l

-4 -2 0 2 4

0

5 0

1 0 0

1 5 0

A s c e t ic F lu id - S u rg e ry

L o g D o s e (u M )

% V

iab

ilit

y

C is p la tin

T o p o te c a n

L Y 2 9 4 0 0 2

D o x o ru b ic in

G e m c ita b in e

C a rb o p la tin

T ra m e tin ib

A fa tin ib

P a c lita x e l-4 -2 0 2 4

0

5 0

1 0 0

1 5 0

A s c e tic F lu id - S u rg e ry

E p C A M + C e lls

L o g D o s e (u M )

% V

iab

ilit

y

C is p la tin

T o p o te c a n

L Y 2 9 4 0 0 2

D o x o ru b ic in

G e m c ita b in e

C a rb o p la tin

T ra m e tin ib

A fa tin ib

P a c lita x e l

IC50 (uM)

Drug

ascitic Fluid

Pre-SurgeryTumor

ascitic Fluid

Surgery

Unselected

ascitic Fluid

Surgery

EpCAM+

Cisplatin 22.45 13.91 30.34 6.716

Topotecan 3.493 3.964 3.097 3.976

LY 294002 20.47 60.35 23.14 34.82

Doxorubicin 0.3053 0.3359 0.2329 0.3238

Gemcitabine >100 >100 18.15 >100

Carboplatin 79.72 >100 >100 >100

Trametinib >100 >100 29 >100

Afatinib 0.654 1.901 1.664 0.2887

Paclitaxel 11.4 21.68 5.903 3.962

Figure 3: Drug response is affected by the source of the cells and the cell

types present. ascitic fluid (A, C, D) and tumor (B) from the same patient were

assessed for drug response in our EV3D DRP assay. The ascitic fluid was assessed

both before surgery and at the time of surgery. Prior to surgery it predicted

response to carboplatin however their was no response when the tumor and the

ascitic fluid at surgery was assessed. The ascitic fluid at surgery predicted response

to both Gemcitabine and Trametinib which was not predicted by the tumor.

When the surgical ascitic fluid was selected for EpCAM+ cells (D) which made 20%

of the cellular population, the drug response profile was more similar to the tumor

than either unselected ascitic fluid samples, n = 6.

A.

D.

C.B.

E.

0.49%

CD14 PKH26 CD163 CD206

CD14 PKH26 CD163 CD206

This work is supported in part by NCI SBIR Contract #: HHSN261201300043C

With Special Thanks to

the Biorepository