HCW Biologics : First-in-Human Clinical Readout of HCW9218, SITC 2023

IL-15Rα

IL-15

2*TGFβRII

2*TGFβRII

Pre-clinical and First-in-Human Studies of HCW9218, a Bifunctional TGF-βAntagonist/IL-15 Protein Complex, in Advanced Solid Tumors

Melissa A. Geller 1, Manish Patel 2, Hing C. Wong 3, Peter R. Rhode 3, Philip M. Arlen 3, Danieska Sandino 3, Karen L. Kage 3, Delsey G Gaetano3, Jack O. Egan 3, Gilles M. Leclerc 3, Pallavi Chaturvedi 3 ,

Varghese George 3, Niraj Shrestha 3, Xiaoyun Zhu 3, Martin Felices 2, Shannon Lunn 4, Bethany Hanke 4, Kelsey Mullen 4, Deepa Kolaseri 4, Rose Wangen 4, Jeffrey Miller 2

1 University of Minnesota School of Medicine, Department of Obstetrics, Gynecology and Women's Health, Division of Gynecologic Oncology, Minneapolis, MN, USA 2 University of Minnesota School of Medicine, Department of Medicine, Division of Hematology, Oncology, and Transplantation, Minneapolis, MN, USA

3 HCW Biologics, Miramar, FL, USA

4 Masonic Cancer Center, Minneapolis, MN, USA

BACKGROUND

HCW9218 is a bifunctional protein complex comprised of dimeric extracellular domains of the human transforming growth factor beta (TGF-β) receptor II (2*TGFβRII) and human interleukin-15(IL-15) (Liu et al., Mol Ther 2021; Chaturvedi et

al., Mol Ther 2022). The mechanisms of action of HCW9218TF are to 1) activate and promote tumor infiltration of effector NK

and CD8+ T cells and 2) sequester soluble immunosuppressive

TGF-β. Previous studies in mouse tumor efficacy models demonstrated the potent antitumor activity of HCW9218 monotherapy and combination therapy with chemotherapy and immune checkpoint inhibitors.

New preclinical data and the Phase 1 clinical trial are presented.

HCW9218 PRECLINICAL DATA

A

1. Biodistribution of HCW9218 in lymphoid and tumor tissues

						B16F10(pg/mg1βTGF lysates)									IL12 in Tumor
(pg/mgHCW9218lysates)	0	20	40	60	80				B16F10pg/mglysates		IL2 in Tumor	B16F10pg/mglysates	Saline 6h	24h 72h
				Saline 6h	24h 72h
	4000		HCW9218 in lymph node		40								15	✱
											8		✱✱		ns
	3000		HCW9218 in spleen			30					✱			ns
			HCW9218 in B16F10							6	ns			10
	2000						20				4
	1000														5
						10	**			2
	0						0				0				0
		Post-Dose Time (Hrs.)			0	24	48	72
					Post-Dose Time (Hrs.)		post HCW9218		post HCW9218

1. CD8+ T cell subsets in draining lymph nodes

Ag-experienced (CD44

+

)

Proliferating (Ki67) Ag-experienced

Progenitor exhausted (Tpex)

Proliferating (Ki67

+

) Tpex

CD8

+

T cells in dLN

CD8

+

T cells in dLN

+

CD8

T cells in dLN

+

CD8

T cells in dLN

✱

✱✱

80000

100

✱✱✱✱

100

✱

4000

✱✱✱✱

✱

+

✱✱✱

+

80

✱✱✱✱

PD1

+

Absolutecount

60000

✱✱✱✱

CD44

Absolutecount

CD44

80

ns

+

3000

%LiveCD8

CD8

%LiveCD8

✱

+

60

TCF1

ns

+

60

40000

+

2000

20000

40

CD44

40

ns

20

+

1000

20

0

0

Saline24h 48h 72h 120h

0

0

Saline24h 48h 72h 120h

Saline24h 48h 72h 120h

Saline24h 48h 72h 120h

days post HCW9218

days post HCW9218

days post HCW9218

days post HCW9218

1. Chemokine receptor expression on CD8+ T cell subsets in draining lymph nodes and blood

CXCR3+ Tpex CD8+ T cells in dLN

CX3CR1 expression on

CX3CR1 expression on

Ag experienced CD8 in blood

Ag-experienced CD8

+

T cells in dLN

+

50

✱✱✱✱

TCF1

2500

8000

PD1

40

ns

+

(MFI)

2000

✱

+

(MFI)

✱

+

CD44

PD1

CD44

PD1

6000

+

ns

CD44

30

ns

+

+

1500

+

+

+

+

4000

CD8%Live

LiveCD8

20

TCF1

1000

TCF1

CD8

+

10

500

2000

%Live

0

0

0

Saline24h 48h 72h 120h

Saline 24h 48h 72h 120h

Saline24h

48h 72h 120h

days post HCW9218

days post HCW9218

days post HCW9218

1. CD8+ TIL subsets in tumor

Ag-experienced (CD44+)

Tpex CD8+ T cells in Tumor

Terminally exhausted (Tex)

CD8+ T cells in Tumor

(cells/gm)countAbsolute

1500

ns

CD8

+

T cells in Tumor

(cells/gm)countAbsolute

ns

CD8

CD8

(cells/gm)countAbsolute

ns

6000

+

10000

PD1

✱✱

+

✱✱✱

1000

8000

✱✱✱

+

TIM3

6000

ns

TCF1

ns

TCF1

4000

-

4000

ns

+

500

ns

2000

400

+

ns

CD44

300

CD44

1000

2000

ns

+

200

+

500

100

0

0

0

Saline 24h 48h 72h 120h

Saline 24h 48h 72h 120h

Saline 24h 48h 72h 120h

days post HCW9218

days post HCW9218

days post HCW9218

Figure 1. 6-wkold C57Bl6/j mice were injected subcutaneously with 0.5x106 B16F10 melanoma tumor cells. When tumor size reached approximately 200-400mm3, HCW9218 (3 mg/kg) was administered subcutaneously and mice were sacrificed at indicated timepoints following treatment (A). (B) Tumor draining lymph (dLN) nodes, spleens and tumors were processed and HCW9218, TGF-β1levels and IL2 and IL12 levels were measured by ELISA. (C) Antigen-experienced(CD44+) CD8+ T cells and T progenitor exhausted (TCF1+PD1+ Tpex) CD8+ T cells in dLN at various timepoints. (D) Frequencies of chemokine receptor CXCR3+ Tpex in dLN and CX3CR1 (MFI) expressing Ag experienced CD8+ T cells in blood and dLN at indicated timepoints respectively. (E) Absolute numbers of antigen-experiencedCD8+ T cells, Tpex and terminally exhausted (TCF1- TIM3+) TILs in tumors at various timepoints.

Rationale for Combining HCW9218 with ICIs for Cancer Treatment

HCW9218 boosts potency of

Immune Checkpoint Inhibitors by:

• Stimulating and activating immune cells
• Promoting immune cell infiltration into tumor
• Reducing immunosuppression of TGF-β

PHASE 1 CLINICAL TRIAL

PRIMARY OBJECTIVE

• The primary objective of this Phase I first-in-human clinical trial is to determine the maximum tolerated dose (MTD) of HCW9218 in patients with chemo-refractory/resistant advanced solid tumors (excluding pancreatic and brain tumors).

PATIENTS AND METHODS

• HCW9218 is administered subcutaneously in the outpatient setting once every 3 weeks for a minimum of 2 cycles (Fig 2).
• HCW9218 dose ranges are from 0.25 mg/kg (DL1) to 1.2 mg/kg (DL4).
• Correlative objectives include immunogenicity, pharmacokinetic (PK) profiles of HCW9218, lymphocyte number, phenotype and function by flow cytometry analysis.
• Failed at least 2 prior lines of therapy given either in the recurrent or metastatic setting and must be refractory to or intolerant of existing therapy.
• Measurable disease per RECIST v 1.1.

Fig 2. Schema/Dose Levels

= tumor biopsy

RESULTS

				Table 1. Demographics
PATIENTS and PATIENT
DISPOSITION							Patients( n=15)
Patient enrollment began 04/22, 22 participants signed		Age, years, median (range)		56 (39-70)
consent, 7 were deemed not eligible with 15 enrolled
	Sex, male/female (%)		8/7, (53%)
at the time of this report. Five patients remain on
HCW9218. Four solid tumors were represented.
			ECOG PS
Median number of cycles received was 3. Baseline
			0			7 (47%)
characteristics are summarized in Table 1.
			1			8 (53%)
TUMOR RESPONSES
			Disease sites, n(%)
Stable disease was observed in 4 heavily pretreated			Ovarian			6 (40%)
advanced solid tumor patients (2 ovarian, 1 rectal, 1			Colon			4 (27%)
liver). Repeated HCW9218 administration (up to 6				Rectal			3 (20%)
cycles) resulted in immune cell activation,				Liver			2 (13%)
proliferation, and infiltration into the tumor
microenvironment without causing unacceptable				# previous lines of therapy, n (%)
toxicity. HCW9218 treatment presents a promising				2			2 (13%)
			>4			13 (87%)
approach to enhancing the antitumor activity of
immune checkpoint inhibitors in patients with solid		Table 2. Most Frequent TRAE's in N=15 patients
tumors.
						Grade ≥ 3
TOXICITY			Toxicity Summary	Any Grade	N, (%)
	Total Number of TRAEs Experienced	402		40 (9.9%)
During the dose escalation phase of the trial,
Total % of TRAEs experienced by the	15 (100%)		14 (93%)
there were no DLT's encountered. At the 4th DL				patients
expansion, there was 1 DLT (Gr 3 ascites) that					Any Grade
did not trigger the stopping rules. Treatment					Count, (% of	Grade ≥ 3
related AE's at least possibly related to the			TRAE, % of total TRAEs	pts)	Count, (% of pts)
study medication are summarized in Table 2.			Injection site rxn (18.1%)	72 (100%)		1 (7%)
			Flu like symptoms (9.7%)	39 (87%)		0
	Lymphocyte count decreased (16.4%)	35 (93%)		21 (74%)
CORRELATIVE DATA

		TGF-β1				TGF-β2
	60000	0.25 mg/kg HCW9218	0.80 mg/kg HCW9218		2400	0.25 mg/kg HCW9218	0.80 mg/kg HCW9218
(pg/mL)		0.50 mg/kg HCW9218	1.2 mg/kg HCW9218
	(pg/mL)		0.50 mg/kg HCW9218	1.2 mg/kg HCW9218
			2000
TGF-β1	40000			2βTGF-	1600
			1200
Serum	20000			Serum	800
			400
	0				0

D1.

D

5

3

3h

h

h

2

3

5

8

5

1

h

2

5

8

1

0

1

1

1

D

2

2D

D15

D1

D2

D

1D51D8D

2D1

2

2

2 D

C

5

3

D

D D

D

1

3

D

D

.

h

h

3

15

15

1

C1

C

C1

C

C

D1

C2

C2

C

C1

0

C1 C1 C

C

C

1

C

C

C

-

-

e

C1D

C1

e

2

1

1D1

C1

D

2

C

-

-

1

C2

C

C

r

D

r

e

re

C2

P

P

Pr

C1D

C

P

Visit day - time

Visit day - time

IMMUNE DATA

AB

Fig 4. % of Ki67+ NK cells (A) and CD8+ T cells (B) by flow cytometry. All subjects had a robust proliferation of blood NK cells, ranging from 77% to 97% Ki67-positivityby Day 8 after dosing for each treatment cycle. HCW9218-mediatedincreases in blood NK cell percentages and counts were also observed. Treatment induction for blood CD8+ T cell proliferation was also observed. Responses were sustained through Day 15, a biological effect beyond that previously observed for other IL-15agonists.

ABC

Fig 5. Absolute Number of NK cells (A) and CD8+ (B) and CD4+ T cells (C). Absolute numbers reflect that of the Ki67+ proliferation levels shown in Fig above.

CD8+ T cells

Fig 6. Immune Cell Staining in Pre- and Post-Treatment Tumor Biopsy Specimens. HCW9218 treatment induced CD8+ T cells trafficking to tumor in an ovarian cancer patient with stable disease. Similar results were seen in tumor biopsies of two other patients (ovarian and rectal cancer) with stable disease.

AB

Fig 7. % Blood CD8+ T cells that are PD-1+ and TCF1+ Eomes+

The presence of exhausted (PD1+) (A) and Tpex (TCF1+) (B) CD8+ T cells was evaluated using time of flight mass cytometry (CyTOF) using Maxpar Direct Immune Profiling assay with a NK cell expansion panel (for dose escalation) or a custom CD8+ T cell expansion panel (for the extension cohort) and then analyzed with Maxpar Pathsetter software.

CONCLUSION and FUTURE STUDIES

• Repeated HCW9218 administration at up to 1.2 mg/kg was well tolerated by heavily pretreated advanced solid tumor patients.
• HCW9218 treatment resulted in NK cell and CD8+ T cell activation, proliferation, and infiltration into the tumor microenvironment which correlated with disease stabilization.
• HCW9218 also reduced TGF-β levels in tumors (mouse tumor models) and blood (mouse and human clinical studies).
• Based on its ability to activate, expand and induce tumor trafficking of progenitor exhausted stem-like and transitory CD8+ T cells, HCW9218 treatment presents a promising approach to enhancing the antitumor activity of immune checkpoint inhibitors in patients with solid tumors.
• Phase 2 studies are planned to combine chemotherapy and HCW9218 and checkpoint blockade as a maintenance strategy in recurrent ovarian cancer.

Fig 3. Neutralization of TGF-β1 and TGF-β2 by dose level. HCW9218 dose-dependentreduction in serum TGF-β1and TGF-β2levels (to baseline at >0.5 mg/kg HCW9218) were observed.