Surrozen : SZN-1326 13-week GLP Toxicity Studies

SZN-1326, a tetravalent, bispecific antibody developed for the treatment of ulcerative colitis, has a favorable profile in 13-week GLP toxicity studies

A. Diep1, J. Tibbitts1, S.A. Roberts2, P. Stathis1, M. Newman1,3, W. Nowatzke1, G.F. Vanhove1; 1Surrozen, Inc. South San Francisco, California, United States,

2SAR Safety Assessment, Northbrook, Illinois, United States, 3Current affiliation: Nautilus Biotechnology, Inc. San Carlos, California, United States (formerly1); *Presenting author

Introduction and Background

• WNT/β-cateninsignaling promotes intestinal stem cell renewal which is essential for intestinal epithelial homeostasis and regeneration. There is a clear unmet need for agents that directly repair and regenerate the intestinal epithelium for the treatment of ulcerative colitis as mucosal healing has been associated with lower steroid use, reduced hospitalizations and longer-term remission.
• SZN-1326is a tetravalent, bispecifi c antibody that binds specifi cally to low-density lipoprotein receptor-related protein 6 (LRP6) and Frizzled 5 (FZD5) and/or Frizzled 8 (FZD8). LRP6, FZD5 and FZD8 are expressed across many tissues, but FZD5 receptors are highly enriched on the intestinal epithelium including stem and progenitor cells (Figure 1). Therefore, treatment with SZN-1326 could improve mucosal healing in ulcerative colitis patients through Wnt agonism.

Figure 1: SZN-1326 targets intestinal stem and progenitor cells and induces transient expansion cell expansion and differentiation resulting in mucosal healing.

Migration	SZN-1326 MOA	Microbes
Damage to epithelial cell barrier
		Microbial invasion
	DC	Activated T cells
		Cytokines/Chemokines
	Goblet cells	T cells
Tuft cells
Endocrine cells
Colonocyte
cells

Fzd5	Wnt
Intestinal stem and	SZN-1326
progenitor cells
(ISCs)

Wnt ligand

Stromal cells

Rationale

• To further develop SZN-1326 as a potential treatment for patients with ulcerative colitis, the preclinical safety of SZN-1326 was investigated by conducting GLP toxicology studies in Sprague Dawley rats and cynomolgus macaques dosed weekly with SZN-1326 for 13 weeks with a Recovery phase of
  4 weeks for a subset of animals.
• SZN-1326is pharmacologically active in rats and cynomolgus macaques based on 98.5% (rat) and 99.5% (monkey) homology of the binding domains of FZD5, FZD8 and LRP6 to the human amino acid sequences. Additionally, target binding, potency and tissue receptor mRNA expression of FZD5, FZD8 and LRP6 in human and nonclinical species are comparable.

Table 1: Experimental design for SZN-1326 GLP Toxicology Study in rats and cynomolgus macaques

Group	Dose Route	Test Article	Dose (mg/kg)
1	IV/SC	Vehicle	0
2	IV		3
3	IV		10
		SZN-1326
4	IV	30
5	SC		30
6	IV		75

• GLP toxicology studies were designed to evaluate the toxicity and toxicokinetics of SZN-1326 when administered once weekly via intravenous (IV) injection and/or subcutaneous (SC) injection for
  13 weeks.
• To assess the reversibility or persistence of any adverse effects, a subset of animals receiving vehicle (20 mM histidine, 240 mM sucrose, 10 mM L methionine, 0.04% Polysorbate 80, pH 6.0) or the high dose of 75 mg/kg were continually evaluated and terminated after a 4-weekdrug-free Recovery phase.

Table 2: Toxicology endpoints evaluated in the GLP repeat dose studies

Parameter	Frequency
Mortality & cage side observations	Twice daily
Clinical observations
Body weight	Twice weekly
Food consumption (qualitative)
Cardiovascular safety pharmacology	Once Pretest and during Weeks 1, 4, 8 and 12
endpoints (ECG, HR) (males only)
	of the Drug phase
Functional observational battery
Clinical pathology	Once Pretest and on Day 9, 23, 51, 93 of the Drug
phase and at terminal necropsy
Histopathology	End of Drug phase and end of Recovery phase
Immunogenicity (anti-drug antibody	Pretest and on Days 1, 15, 43, 64, and 92 of the
[ADA] analysis)	Drug phase and end of Recovery phase

Results

• SZN-1326was well-tolerated with no drug-related adverse effects observed when administered once a week by IV or SV to Sprague Dawley rats or cynomolgus monkeys.
• The No Observed Adverse Effect Level (NOAEL) was the highest administered dose of
  75 mg/kg/week (IV) with no SZN-1326 related adversity observed at 30 mg/kg/week (SC) in both species.
• Minimal, non-adverse, and reversible decreases in reticulocytes, hemoglobin, hematocrit, red blood cells and platelets and minimal increases in blood urea nitrogen (BUN) were observed in Sprague-Dawley rats.
• Additionally, transient minimal to mild elevations in AST/ALT occurred in individual rats on Day 93, 107 and 121 at the end of Recovery phase. The changes lacked a dose relationship and had no histopathologic correlation; therefore, they were not considered related to SZN-1326 administration.
• No SZN-1326-related effects in hematology, coagulation, clinical chemistry, or urinalysis were identifi ed in the cynomolgus monkeys.
• Functional observational battery tests in rats and monkeys demonstrated no abnormalities in central nervous system parameters due to SZN-1326.
• The TK of SZN-1326 was consistent with the expected PK of human monoclonal antibodies (Figures 2-3).
• Serum exposure was proportional to dose in rat and monkey (Tables 3-4).
• The estimated SC bioavailability in the rat was poor (11%) but good (55%) in the monkey.
• The incidence of anti-drug antibodies (ADA's) ranged from ~10 to 60% in the monkey and the rat.

Table 3: Toxicokinetic profile of SZN-1326 in rats following weekly dosing

Dose		TK	AUC0-1	AUC0-1	AUC0-1 /D	AR	Cmax	Cmax / D	AR
Route	(µg-day/	(µg-day/	(µg-day/mL/	(µg/mL)/
(mg/kg)	Day	AUC	(µg/mL)	Cmax
			mL)	mL)	mg/kg)			mg/kg)
3	IV	0	32.9	84	28	1.18	58	19.3	1.13
92	38.9	NA	NA	65.8	21.9
10	IV	0	120	326	32.6	1	215	21.5	0.94
92	120	NA	NA	202	20.2
30	IV	0	360	901	30	0.805	608	20.3	1.01
92	290	NA	NA	617	206
75	IV	0	755	1680	22.4	0.768	1420	19	1.29
92	757	1290	NA	1830	24.5
30	SC	0	15.5	193	6.4	0.122	36.3	1.21	0.292
92	3.44	23.5	NA	10.6	0.35

Table 4: Toxicokinetic profile of SZN-1326 in monkeys following weekly dosing

Dose		TK	AUC0-1	AUC0-1	AUC0-1 /D	AR	Cmax	Cmax / D	AR
Route	(µg-day/	(µg-day/	(µg-day/mL/	(µg/mL)/mg/
(mg/kg)	Day	AUC	(µg/mL)	Cmax
			mL)	mL)	mg/kg)			kg)
3	IV	0	53.6	150	50	1.4	82.8	27.6	1.25
92	74.9	NA	NA	103	34.5
10	IV	0	206	471	47.1	0.898	365	36.5	0.795
92	185	NA	NA	290	29
30	IV	0	435	1120	37.33	1.73	662	22.1	1.54
92	753	NA	NA	1020	34
75	IV	0	98.1	627	20.9	1.48	167	5.57	1.32
92	159	926	NA	220	7.33
30	SC	0	1330	3060	40.8	1.08	2050	27.3	2.09
92	1650	3320	NA	4280	57.1

Figure 2. Mean (SD) Serum SZN-1326 Concentrations in Rats Dosed Weekly for 13 Week

3 mg/kg IV

10 mg/kg IV

30 mg/kg IV

30 mg/kg SC

75 mg/kg IV

Figure 3. Mean (SD) Serum SZN-1326 Concentrations in Cynomolgus Monkeys following 13 weeks of IV or SC Drug

3 mg/kg IV

10 mg/kg IV

30 mg/kg IV

30 mg/kg SC

75 mg/kg IV

Table 4: Summary of the ADA Positive Screening Results in Rat Serum

	Dose	Dose	% of ADA positive animals at the
Group	end of Drug and Recovery
Route	(mg/kg)
	Day 92	Day 121*
1	IV/SC	0	0%		0%
2	IV	3	11%		0%
3	IV	10	33%		22%
4	IV	30	39%		17%
5	SC	30	39%		11%
6	IV	75	61%		28%
*Day 121 corresponds to Day 29 of the Recovery Phase
Table 5: Summary of the ADA Positive Screening Results in Monkey Serum
	Dose	Dose	% of ADA positive animals at the
Group	end of Drug and Recovery
Route	(mg/kg)
	Day 92	Day 121*
1	IV/SC	0	10%		0%
2	IV	3	50%
3	IV	10	50%		NA
4	IV	30	33%
5	SC	30	30%		50%
6	IV	75	40%		100%

*Day 121 corresponds to Day 29 of the Recovery Phase

Conclusions

• SZN-1326has a favorable safety profile in 13-week rat and cynomolgus monkey GLP-compliant toxicology studies when given once a week by IV or SC.
• Minimal, non-adverse, clinical pathology findings (minimal decreases in HgB, minimal, transient elevations in BUN and
  AST/ALT) in rats were not correlated with any macro- or microscopic histopathological changes or clinical observations.
  Therefore, these transient clinical pathology changes were not considered adverse.
• The NOAEL is the highest IV administered dosage of
  75 mg/kg/dose with no SZN-1326-related adverse effects observed at 30 mg/kg dosed SC.

Presenting author: Anh Diep

email: [e-mail]

Surrozen Inc., South San Francisco, CA 94080 www.surrozen.com

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