#780 ALTA-002, a SIRPa-Directed TLR9 Agonist Antibody Conjugate, Activates Myeloid Cells AndPromotes Anti-Tumor ImmunityOns Harrabi1, Amy Chen1, Emma Sangalang1, Danielle Fontaine1, Min Li 1, Jaume Pons2, Hong I. Wan1, Janet Sim1, Tracy C. Kuo1
1Tallac Therapeutics, Inc., Burlingame, CA; 2ALX Oncology, South San Francisco, CA

Experimental Results

101001000

10000

110100anti-SIRPα

SIRPα TRAAC

2000

SIRPα TRAAC
B cell directed TRAAC1000Media only220.1

11010001000 10000

0.1

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anti-SIRPαanti-SIRPα1001000 10000

anti-SIRPαanti-SIRPα

ALTA-002
SIRPα
TRAAC

ALTA-002
SIRPα TRAAC

Fig 6: SIRPa TRAAC promotes phagocytosis of SIRPa- and SIRPa+ tumors bymacrophages% Phagocytosis of SIRPα+ DLD-1% Phagocytosis of SIRPα- DLD-1

B cell directed TRAAC

500

Media only

15105Median Fluorescent
Intensity (MFI)

50.01

0.1

11010511010001000

1101001000

anti-SIRPαanti-SIRPαMediaMediaonly
only

ALTA-002
SIRPα TRAAC

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Fig 7: Localizing mSIRPa TRAAC to SIRPa+ tumor demonstrates superioranti-tumor activity in syngeneic model when administrated systemicallyanti-SIRPαALTA-002

SIRPα TRAACABParental MC38 (SIRPα-)

✱✱✱✱

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pg/mL

800

pg/mL

pg/mL

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0

IL10IL121500✱✱✱

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10200

51015200Figure 4. Human PBMCs were stimulated with anti-SIRPa or SIRPa TRAAC for 24hr and assayed for IRF7induction (A) and cytokine production (B) from 9 healthy donors following 72hr stimulation by flow cytometry.
P-values were calculated using unpaired t-test.

500anti-PD-1
1000*500mSIRPα TRAAC + anti-PD-1* p< 0.05

0102030mSIRPα TRAAC (0.3mg/kg)

51015202530Days Post Tumor Implantation

Figure 9. Mice bearing CT26 tumors were intraperitoneally (i.p.) treated with mSIRPa TRAAC at 1mg/kg or PBS (A)or mSIRPa TRAAC at 0.3mg/kg, anY-PD-1 at 10mg/kg, in combinaYon, or PBS control (B). Arrows indicate doses
administrated. P-values comparing combinaYon group vs. anY-PD-1 in were calculated using unpaired t-test.

Conclusions• SIRPa expression on tumors potentiates the activation of myeloid cells by SIRPa TRAAC.
• SIRPa TRAAC promotes phagocytosis of tumors independent of SIRPa expression.
• Localization of mSIRPa TRAAC to SIRPa+ tumors demonstrates robust and curative single agent activity

400
200in multiple models including anti-PD-1 refractory tumors.
05101520

Days Post Tumor Implantation

Days Post Tumor Implantation

PBS

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PBS

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leading to robust cellular activation and cytokine induction in cultured immune cells.

60000mSIRPα TRAAC (1mpk)
1500• SIRPa TRAAC specifically targets myeloid cells triggering TLR9 signaling via SIRPa and FcgR engagement

MC38 SIRPα+• mSIRPa TRAAC enhances tumor regression in combination with anti-PD-1 in syngeneic tumor model.
• ALTA-002, our SIRPa TRAAC candidate, is in preclinical development for the treatment of patients withmSIRPα TRAAC✱✱pg/mLpg/mL

340000TLR9600

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Mean Tumor Size(mm3)ns1000✱CT26 (SIRPα-)administration.

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IL6Anti-PD1• SIRPa TRAAC is a SIRPa-directed TLR9 receptor agonist antibody conjugate designed for systemic

800

IFNγ302000

PBS

Days post implantation

Media only
Media onlynM

ALTA-002

B

CT26 (SIRPα-)0ALTA-002
SIRPα TRAACPBSmSIRPαTRAACmALTA-002

0nMnM
anti-SIRPαanti-SIRPα

20

Days post-Tumor Implantation

Fig 9: The combination of suboptimal dose of mSIRPa TRAAC and anti-PD-1
elicits enhanced anti-tumor response in CT26, a SIRPa negative tumor model
2000001030Figure 8. Mice bearing RENCA tumors were dosed intraperitoneally (i.p.) three times, three days apartwith mSIRPa TRAAC at 10mg/kg or PBS (A). A separate cohort was dosed with anti-PD1 at 10mg/kg threetimes, three days apart or PBS (B). Arrows indicate doses administrated.

AFigure 6. Human monocyte derived macrophages were co-incubated for 2hrs with either parental DLD-1(A) or DLD-1 expressing SIRPa (B) in presence of anti-SIRPa, SIRPa TRAAC or media only. % Phagocytosis
was determined by flow cytometry.

250

100100 1000 10000

20PBSPBS

150500010Figure 5. Human PBMCs were co-cultured either in presence of parental DLD-1 (A) or DLD-1 expressing
SIRPa (B) and stimulated with anti-SIRPa, SIRPa TRAAC, or media only for 48hr. Surface marker
expression was assayed by flow cytometry. Data is presented as fold over media only control.

B

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Days post-Tumor Implantation

300

10500

0CD14+ Monocytes:IRF7CD11c Dendritic Cells:IRF71000nMnM

Fig 4: SIRPa TRAAC triggers TLR9 signaling in SIRPa+ DC and monocytes, eliciting
pro-inflammatory and cytotoxic cytokine production in human PBMC cultures% CD11c4001500

SIRPα TRAACFigure 3: Human PBMCs were sYmulated with SIRPa TRAAC, B cell directed TRAAC or media only for 24hr
and surface marker expression on monocytes (A) and B cells (B) was assayed by ﬂow cytometry.

A

60A

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B cells

Median Fluorescent
Intensity (MFI)Median Fluorescent
Intensity (MFI)

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Tumor destruction

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mTCpG

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RENCA (SIRPα+)

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8mSIRPα TRAAC

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IFNα2

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mALTA-002

Human B cell activation:CD86

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anti-SIRPα

SIRPα TRAACB

Human monocyte activation:CD86anti-SIRPα✔

10% Phagocytosis(CFSE+ Macrophages)

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Lymphoid organsp(DC)1Fig 3: SIRPa TRAAC specifically targets and activates SIRPa+ immune cells inhuman PBMC culturesnM

Myeloid0.1

Figure 2: Human, cynomolgus PBMCs, and mouse splenocytes were stimulated with either anti-SIRPa, SIRPaTRAAC, T-CpG, or SIRPa TRAAC conjugated to murine reactive mT-CpG (mSIRPa TRAAC) for 24hr or 48hrs andsurface marker expression was assayed by flow cytometry. DC: dendritic cells

0SIRPa+0.01

Unconjugated anti-SIRPαanti-SIRPαanti-SIRPαCell type010000nMnM0Tumor microenvironment

1000

EC50 : N.A

6EC50 : 3.34nM
Tumor volume (mm3),mean ± SEM

1SIRPa+ myeloid

SIRPa TRAAC
Tumor Targeted

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0015

SIRPa+ Tumors

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Mean Tumor Size(mm3)0Antibody: anti-SIRP a• Targets myeloid cells expressing SIRP a• Localizes to tumor microenvironment of SIRP a + tumors• Engages Fc gamma receptors• Blocks CD47-SIRPα myeloid checkpoint pathway

SIRPa TRAAC
Immune Cell Targeted

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008B
RENCA (SIRPα+)Tumor volume (mm3),
mean ± SEM

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Fig 8: mSIRPa TRAAC elicits potent single agent anti-tumor response inRENCA, a SIRPa+ model refractory to anti-PD-1
A

SIRPα+ DLD-1
DC activation:CD86

Fold over media

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SIRPα- DLD-1
DC activation:CD86% Phagocytosis(CFSE+ Macrophages)

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A

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Median Fluorescent
Intensity (MFI)Median Fluorescent
Intensity (MFI)

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Fig 1: SIRPa TRAAC, is a Toll-Like Receptor Agonist An<body Conjugate,
designed for systemic delivery of T-CpG, a potent TLR9 agonist

Immune Activator: T-CpG• Potent TLR9 agonist• Linear, monomeric, and non-aggregated• Sequence optimized for potency and stability• Innovative design enables efficient site-specific conjugation

Mouse splenocyte
DC activation:CD40

Cynomolgus Monkey
DC activation:CD40
1500

Fold over media

Human
DC activation:CD40
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Fig 5: SIRPa expression on tumors potentiates myeloid activation by SIRPa TRAAC

Tumor volume (mm3),
mean ± SEM

Fig 2: T-CpG is a potent TLR9 agonist with targeted activation across species

Median FluorescentIntensity (MFI)• Novel therapies engaging both innate and adaptive immune responses may
engender more robust and durable anti-cancer immunity [1].
• Activation of toll-like receptor 9 (TLR9) by unmethylated CpG oligonucleotides(ODNs) promotes innate inflammatory responses and the induction ofadaptive immunity [2]. Several CpG oligodeoxynucleotides havedemonstrated clinical response in patients with melanoma by intra-tumoral
injection [3].
• We developed a novel Toll-like Receptor Agonist Antibody Conjugate (TRAAC)
molecule comprised of a differentiated TLR9 agonist (T-CpG) conjugated to an
antibody against SIRPα termed SIRPa TRAAC.
• Signal regulatory protein α (SIRPα) is a myeloid inhibitory receptor thatsuppresses immune activation following binding of its ligand CD47 [4].
• Blockade of CD47-SIRPα myeloid checkpoint pathway has demonstrated
clinical response in patients with solid tumors [5].
• We present preclinical data demonstrating that SIRPa TRAAC delivers T-CpG
to myeloid cells via SIRPa and FcgR engagement, triggering TLR9 signaling, cell
activation and immune modulation, resulting in robust anti-tumor activity.

Tumor volume (mm3),
mean ± SEM

Introduc-on

Figure 7. Mice bearing either parental SIRPa- MC38 tumors (A) or expressing SIRPa (B) were dosed
intraperitoneally (i.p.) twice, three days apart with mSIRPa TRAAC conjugated with murine reactive mT-CpG
at 1mg/kg or PBS control. Arrows indicate doses administered.
.

various types of malignancies

References:
1. Kobold et al., Innate and adaptive immunity combined for cancer treatment. PNAS 2019, 116 (4) 1087-1088.
2. Dowling et al., Toll-like receptors: the swiss army knife of immunity and vaccine development.
Clin. Transl. Immunology. 2016 5(5), e85–10.
3. Hamid et al., Intratumoral Immunotherapy—Update 2019. The Oncol. 2019; 25:343–359.
4. Kuo, T et al., Targeting the myeloid checkpoint receptor SIRPα potentiates innate and adaptive immune
responses to promote anti-tumor activity. 2020. J Hematol Oncol 13, 160.
5. Lee KW et al., ALX148, a CD47 blocker in combination with standard chemotherapy and antibody regimens inpatients with gastric/gastroesophageal junction (GC) cancer and head and neck squamous cell carcinoma(HNSCC); ASPEN-01. SITC 2020.