Molecular Cancer Therapeutics - Discovery of a novel EGFR ......2019/04/06 · EGFR mutations...

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Discovery of a novel EGFR targeting antibody-drug conjugate,

SHR-A1307, for the treatment of solid tumors resistant or

refractory to anti-EGFR therapies

Kaijie He1*, Jianyan Xu1, Jindong Liang 1, Jiahua Jiang2, Mi Tang2, Xin Ye 1, Zhebin

Zhang1, Lei Zhang1, Beibei Fu1, Yan Li 1, Chang Bai1, Lianshan Zhang1, Weikang Tao1

Author affiliation: 1Shanghai Hengrui Pharmaceutical Co., Ltd., 279 Wenjing Road, Shanghai, 200245,

China. 2Jiangsu Hengrui Medicine Co., Ltd., 778 Dongfang Road, Shanghai, 200122, China

Running Title:

Effect of EGFR-ADC SHR-A1307 in drug resistant cancer

Key words:

antibody-drug conjugate; EGFR inhibitor; drug resistance; EGFR C797S

*Corresponding author:

Kaijie He,

Department of Target Research,

Shanghai Hengrui Pharmaceutical Co., Ltd.,

279 Wenjing Road, Shanghai, 200245, China.

Phone: +86-21-23511543

Email: [email protected]

Disclosure of potential conflicts of interest:

K. He, J. Xu, J. Liang, Z. Zhang, L. Zhang, B. Fu, Y. Li, C. Bai, L. Zhang and W. Tao

are current employees of Shanghai Hengrui Pharmaceutical Co., Ltd.; J. Jiang and M.

Tang are current employees of Jiangsu Hengrui Medicine Co., Ltd.

on July 22, 2021. © 2019 American Association for Cancer Research. mct.aacrjournals.org Downloaded from

Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Author Manuscript Published OnlineFirst on April 8, 2019; DOI: 10.1158/1535-7163.MCT-18-0854

http://mct.aacrjournals.org/

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Abstract:

While inhibiting EGFR-mediated signaling proved to be effective in treating certain

types of cancers, a quickly evolved mechanism that either restores the EGFR

signaling or activates an alternative pathway for driving the proliferation and survival

of malignant cells limits the efficacy and utility of the approach via suppressing the

EGFR functionality. Given the fact that overexpression of EGFR is commonly seen in

many cancers, an EGFR-targeting antibody-drug conjugate (ADC) can selectively kill

cancer cells independently of blocking EGFR-mediated signaling. Herein, we describe

SHR-A1307, a novel anti-EGFR ADC, generated from an anti-EGFR antibody with

prolonged half-life, conjugated with a proprietary toxin payload which has increased

index of EGFR targeting-dependent vs. EGFR targeting-independent cytotoxicity.

SHR-A1307 demonstrated strong and sustained anti-tumor activities in

EGFR-positive tumors harboring different oncogenic mutations on EGFR, KRAS or

PIK3CA. Anti-tumor efficacy of SHR-A1307 correlated with EGFR expression levels in

vitro and in vivo, regardless of the mutation status of EGFR-signaling mediators and a

resultant resistance to EGFR signaling inhibitors. Cynomolgus monkey toxicology

study showed that SHR-A1307 is well tolerated with a wide therapeutic index.

Together, SHR-A1307 is a promising therapeutic option for EGFR-expressing cancers

including those resistant or refractory to the EGFR pathway inhibitors.




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Introduction:

Epidermal growth factor receptor (EGFR), a member of ErbB family of receptor

tyrosine kinase, plays a central role in tumorigenesis of many types of solid tumors (1).

EGFR mutations and/or up-regulations were frequently identified in patients with

non-small cell lung cancer (NSCLC), colorectal cancer (CRC), head & neck

squamous cell carcionma (HNSCC) and glioblastoma (2). Inhibiting EGFR activation

by tyrosine kinase inhibitors (TKIs) or monoclonal antibodies (mAbs) has shown huge

clinical success in suppressing tumor growth and improving patient survival (3-5).

However, disease progression seems inevitable after initial responses (6, 7), possibly

due to either primary or acquired resistance mechanisms, such as an EGFR

secondary or tertiary mutation (8, 9), mutations on downstream mediators of the

EGFR signaling (10, 11) or RTK signal bypass (12). Overcoming resistance to

current EGFR targeting agents represents a clinical challenge and a huge unmet

medical need in cancer therapies.

Antibody-drug conjugates (ADCs) are a new class of drugs that uses antibodies

to selectively deliver cytotoxic payloads to tumors expressing high levels of tumor

associated antigens (13). The local accumulation of toxins in tumor minimized its

systemic toxicity, and increased cancer cell killing in situ, thus providing a higher

therapeutic index than chemotherapy or parental antibodies (14, 15). Moreover, ADCs

can bypass some of the resistance mechanisms by selective delivery of anti-mitotic or

DNA damaging toxins into cancer cells to induce apoptosis, a mechanism that is

independent of inhibiting oncogenic signaling. Recently, two ADCs specifically

targeting EGFR variant III (AMG-595 and ABT-414) demonstrated promising

monotherapy efficacy in recurrent glioblastoma patients (16-18), and the responses

were correlated with EGFR amplification or EGFRvIII presence, providing early

clinical validation for EGFR-targeting ADCs in EGFR amplified or mutated cancer

patients.

Nimotuzumab (a humanized IgG1 also known as hR3) is an EGFR targeting

antibody approved for the treatment of HNSCC and glioma in many developing




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countries, and it is also in clinical trials for treating various solid tumors including

NSCLC, CRC, gastric cancer and pancreatic cancers (19). Because of its unique

binding kinetics with EGFR receptor, hR3 related adverse events in human were only

mild to moderate in severity (20, 21). In particular, it showed much lower incidence of

severe acneform rash and gastrointestinal mucosa toxicities in patients; those side

effects were limiting toxicities typically observed with EGFR targeting agents (19, 22).

The excellent selectivity of hR3 for EGFR positive cancer cells provides a strong

rationale of designing a hR3 based ADC to enhance cancer cell killing while

minimizing normal tissue toxicity for treating patients who have failed anti-EGFR

therapies. Herein, we reported a novel EGFR-targeting ADC SHR-A1307 generated

by using a serum half-life extended hR3 antibody, coupled to a novel auristatin analog

with attenuated free drug cytotoxicity. Preclinical characterizations of SHR-A1307 in

multiple drug resistant cell lines and xenograft models demonstrated potent anti-tumor

activities and a correlation between efficacy and EGFR expression levels regardless

of oncogenic mutation status or drug resistance profiles in cancer cells. In addition,

toxicology studies in cynomolgus monkeys suggested good pharmacokinetics and

safety profiles, supporting the clinical development of SHR-A1307 in EGFR

expressing cancer patients who are resistant or refractory to current EGFR targeted

therapies.




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Materials and Methods

Cell lines and Reagents

HCC827, MCF10A, H1975 and Detroit562 cells were purchased from ATCC; A431,

HCT116, COLO-205, HT29, SW480, SW620, DiFi, Lovo and HepG2 cells were from

Cell Bank of the Chinese Academy of Sciences (Shanghai, China); Primary human

hepatocytes were purchased from Rild Biotech (Shanghai). Cells were maintained in

incubator at 37°C, 5% CO2 in cell culture media as indicated by provider. All cells

used in this study were routinely tested and verified as mycoplasma negative using

MycoAlert PLUS Mycoplasma Detection kit (Lonza). EGFR targeting agents

Nimotuzumab, Cetuximab, Panitumumab, Erlotinib and Osimertinib were prepared at

Shanghai Hengrui Pharmaceutical.

Generating lentivirus and stable expression cell lines

Full length EGFR cDNA was cloned from pcDNA3.1-EGFR (Origene) into

pCDH-CMV-MCS-EF1-puro plasmid, and EGFR exon19 deletion, L858R, T790M and

C797S mutations were introduced by site directed mutagenesis. Virus was generated

by co-transfection of pCDH-EGFR mutant constructs with helper plasmids (△ R8.9

and VSVG) into 293T cells (ATCC, CRL-3216). Supernatant was collected 48 hours

after transfection, filtered by 0.45 μm strainer, and then concentrated for 100 fold

using 50,000g centrifugation. HCC827 or H1975 cells were infected with 5mL

EGFR-mutatnt lentivirus supplemented with 8 μg/ml polybrene. After 24 hours

incubation at 37°C, virus-containing media was replaced and viral infected cells were

selected for at least 1 week with 2 μg/ml puromycin before experiment.

Flow cytometry

Membrane EGFR binding and antibody internalization were analyzed using

fluorescent-activated cell sorting (BD FACSVerse). For binding experiment, 3x105

cells were incubated with 5ng EGFR antibodies or normal human control IgG (R&D

systems, #1-001-A) for 1hr at 4°C, washed with cold FACS buffer twice, then mouse




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anti-human IgG Fc APC-conjugated antibody (R&D systems, #FAB110A) was added

to ice-cold media resuspended cells for 40min. Propidium iodide (Sigma-Aldrich) was

added to the cells before FACS analysis. For internalization experiment, protocol was

similar to binding experiment, except that after EGFR antibodies binding, cells were

maintained in complete media at 37°C for 0h, 0.5h, 1h, 2h, 4h to allow internalization.

Afterwards, cells were put on ice and 2nd antibody was added to detect the remaining

EGFR antibody on cell surface by FACS. Internalization efficiency was calculated as:

Internalization signal (MFI0h – MFItime point) / Maximal binding signal (MFI0h) x100%

Generation of EGFR-ADC

After buffer-exchange and purification, antibody was stored in pH=4.3, 100mM

sodium acetate buffer with a reaction concentration of approximate 10mg/ml. ATPPA

(β-Acetylthiopropionaldehyde) was dissolved in acetonitrile (10% volume) and then

mixed with antibody for 5 minutes. NaBH3CN (25mM) was added and reductive

amination reaction continued for 2 hours. A second ultrafiltration and purification step

with 100mM sodium acetate buffer containing 10% acetonitrile was applied to

concentrate and remove low-molecular-weight impurities (free ATPPA and NaBH3CN

and related products). Following exchange of six-volume PBS buffer (containing

50mM phosphate, 50mM NaCl and 2.5mM EDTA), NH2OH.HCl was added to remove

the acetyl deprotection group of sulphydryl. After a third ultrafiltration, MMAF or

MMAF derivatives with Maleimidocaproyl linker (8.0eq.) were dissolved in acetonitrile

(10% volume) were mixed with deprotected intermediates for 3 hours to generate the

thioether-linked antibody drug conjugate. Newly synthesized ADCs were further

concentrated and purified using G-25 by size exclusion chromatography (ÄKTA pure,

GE healthcare) to remove macromolecular aggregates and free drugs.

Drug-to-antibody ratio (DAR) was determined using Agilent 6530 Q-TOF LC/MS as

previously described (23). All procedures were performed at room temperature

25±2℃.




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In vitro cytotoxicity studies

Cells were seeded at density of 2,000~5,000/well with 90μl assay medium in 96-well

plates, added with 10μl of medium diluted test compounds and incubated in incubator

(5%CO2, 37℃) for about 72h. 50μl/well CellTiter-Glo agents (Promega) were added

into each well and incubate for additional 10min. Read luminescent signal with

microplate reader (Victor3, PerkinElmer) and analyze data using GraphPad Prism6.0

software.

In vivo efficacy studies

In vivo experiments were conducted under an approved protocol in accordance with

the institutional guidelines for the use of laboratory animals. For cell line derived

xenograft models, 1~2x106 tumor cells (suspended with 50% matrigel) were

inoculated subcutaneously into right flank of female Balb/c nude mice (Shanghai

Super B&K Laboratory Animal Corp. Ltd). Tumor sizes were measured by caliper

twice weekly and tumor bearing animals were randomized when average tumor

volumes [(length x width2)/2] reached ~150 mm3. Patient-derived xenograft (PDX)

models (LU1429, LU2503, LU2512, LU3075, CR0455) were established by Crownbio

as previously described (24). Osimertinib resistant PDX model LUN210-4a was

established by GenenDesign after serial in vivo passages of LUN210 tumors under

long term Osimertinib pressure. Tumor-bearing mice were dosed once weekly with

SHR-A1307 or control antibodies via intraperitoneal or intravenous injections;

Erlotinib and Osimertinib were administered orally once daily; dosing schedule was

described in the results. Percentage of Tumor growth inhibition (TGI%) was calculated

by 100x (1- (average Vfinal – Vinitial of treatment group) / (average Vfinal – Vinitial of control

group)).

Pharmacokinetic and toxicity studies in Cynomolgus monkeys

All monkey studies were conducted in AAALAC (Association for Assessment and

Accreditation of Laboratory Animal Care) accredited research facilities, and protocols

were approved by Institutional animal care and use committee (IACUC). For




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Pharmacokinetic study (Jiangsu Tripod preclinical Research laboratories), 6 male

cynomolgus monkeys weighing from 3.91 to 5.56kg were assigned (3 animals each)

to receive a single dose of 5.0mg/kg of hR3 or hR3-YTE by intravenous injection.

Blood samples were collected before and after drug dosing as described in the results,

and serum concentrations of antibodies were detected by ELISA. Briefly, EGFR

antigens (sino Biological Inc.) were coated on the 96-well plates and washed with

blocking buffer three times. Samples were added and incubated at 37℃ for 1.5hrs,

and goat Anti-Human IgG Fc (HRP) (Abcam) and TMB were added. Signals were

detected by microplate reader at 450nm wavelength. In the repeat-dose, non-GLP

toxicity study (WestChina-Frontier Pharmatech), 3 cynomolgus monkeys (2 male and

1 female) were designed to receive 10mg/kg intravenous SHR-A1307 once weekly,

for up to 5 repeated doses, and a 4-week post-dosing recovery phase was planned for

extended observation. Vital signs, ECG and body weight were monitored throughout

the study. Blood samples were collected at various time points to measure drug

concentrations, hematology, clinical chemistry and coagulation parameters. Monkeys

were euthanized by exsanguination under anesthesia for gross necropsy and

histopathological examinations at the end of study. Pharmacokinetic and

Toxicokinetic parameters were calculated using a non-compartmental model analysis

of Phoenix WinNonlin 6.3 software.

Statistical Analysis

Data were analyzed by two-sided Student’s t test (to determine statistical significance

of mean values at one point in time) or Wilcoxon matched-pairs signed-rank test (to

compare statistical significance of tumor growth or plasma drug concentration over

time) using Microsoft office Excel or GraphPad Prism 6.0 software. Difference with

p<0.05 was considered statistically significant




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Results

Identification of a novel EGFR-targeting ADC SHR-A1307

Nimotuzumab (hR3), compared with other approved EGFR antibodies, has

demonstrated similar anti-tumor efficacy but significantly lower on-target, off-tumor

toxicities in patients (19). Mechanistic study suggested that the moderate EGFR

affinity of hR3 requires bivalent binding for stable attachment to cell surface, which

leads to selectively targeting of EGFR highly expressed tumors but sparing normal

tissues with low EGFR expression (20, 21, 25). In agreement with these reports, we

found that hR3 had ~20 fold weaker binding affinity compared with Cetuximab in

Biacore analysis (sup Fig1A), and ~5 fold weaker cell binding activity than

Panitumumab (sup Fig1B-C). Although both antibodies showed comparable growth

inhibition activities in HCC827 cells, hR3 had no impact on the proliferation of normal

mammary epithelial cells (MCF10A), in contrast to the remarkable MCF10A growth

inhibition caused by Panitumumab (sup Fig1D-E). Furthermore, when both antibodies

were coupled with potent microtubule inhibitor monomethyl auristatin F (MMAF)

(DAR=~2.0) (sup Fig1F), hR3 derived ADC demonstrated equally potent anti-cancer

activity but much lower MCF10A cytotoxicity (sup Fig1G-H), suggesting at

least >25-fold higher cancer vs. normal cell selectivity than Panitumumab.

Due to low tumor penetration efficiency of injected antibody (26), highly toxic

payloads were utilized by ADCs to achieve maximal anti-tumor efficacy. However,

serious toxicities may emerge if the toxins were deconjugated from ADCs and taken

up by normal tissues. Accumulated clinical evidence suggested that the majority of

ADC related toxicities were derived from systemically released free drugs (27), risking

their further clinical developments. To address this issue, we designed a series of

auristatin analogs with intention to reduce the cytotoxicity in free drug form while

maintaining their anti-cancer activity in ADC. Using an anti-Her2 based toxin

screening platform, five lead compounds were selected based on their high cancer

killing potency when conjugated to Pertuzumab (IC50 ranging from 0.026 to 0.33nM in

Her2 positive SK-BR-3 cells) (Table1). In addition, excellent selectivity in receptor




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negative HepG2 cells was also demonstrated (IC50>667nM in all tested ADCs), and

there were no liabilities in hERG (IC50>30uM) or CYP450 (IC50 in 1A2, 2C9, 2D6, 3A4

and 2C19 enzymes all >10uM). Free toxin counter screening assays demonstrated

that SHR151526, SHR151247 and SHR152852 had significantly reduced cytotoxicity

as free drugs in multiple cell lines (IC50 value range from 47.7~614.5nM), compared

with 1.28~10.6nM of MMAF, indicating >30 fold higher ADC vs. free toxin selectivity.

These results were further confirmed using hR3-ADCs in EGFR positive cells (Table1).

SHR152852 also demonstrated a 4-fold higher IC50 value and 20% lower maximum

inhibition than MMAF in primary human hepatocytes (sup Fig2A), suggesting a

potentially lower risk of liver toxicity as free toxin. Pharmacokinetics of SHR152852

was comparable to MMAF in rat (sup Fig2B). To analyze the plasma stability, hR3

conjugated with SHR151329, SHR152852 or MMAF at equal DAR (2.2~2.4) were

incubated in human plasma for 7 days, and all 3 ADCs exhibited good stability with

low level of deconjugated free auristatin analogs and non-detectable MC-auristatin

analog levels (sup Fig2C). Furthermore, these ADCs were tested in HCC827

xenograft model for in vivo efficacy comparison, and SHR152852 demonstrated

significantly better anti-tumor efficacy than SHR151329 when conjugated to hR3

(p<0.01), and comparable efficacy with MMAF (sup Fig2D). Given the largely

improved free drug safety profile and high anti-cancer potency, SHR152852 was

selected for later synthesis of EGFR-ADC.

hR3 antibody was reported to have relatively short half-life in human (t1/2~47.5h)

compared with other FDA approved EGFR antibodies (~112hr for Cetuximab and

~180hr for Panitumumab) and required weekly dosing to maintain sufficient drug

exposure, which was inconvenient and cost inefficient in clinical practice (28-30).

Moreover, it is very important for ADCs to have sustained exposure and long half-life

to reduce the risk of toxin accumulation caused by frequent dosing (27). To address

this question, we introduced the triple mutation M252Y/S254T/T256E (YTE) on Fc

portion of hR3, which was reported to increase neonatal Fc receptor (FcRn) binding at

acidic endosomes (pH<6.0) and facilitate FcRn mediated IgG recycling (31).

Consistent with previous reports, hR3-YTE exhibited at least 3-fold stronger binding to




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human and cyno FcRn compared with hR3 at pH=6.0, but no such increase at pH=7.4

in Biacore analysis (sup Fig3A). To confirm the effects in vivo, pharmacokinetics was

investigated in cynomolgus monkeys. The Cmax was very similar for hR3 and hR3-YTE

after a single iv. administration (5mg/kg), but hR3 levels decreased more rapidly than

hR3-YTE (sup Fig3B). Plasma half-life and area under the curve (AUC) were almost

doubled in hR3-YTE group (p=0.0001). In addition, binding and internalization

capabilities were not changed in hR3-YTE (sup Fig3C-D), suggesting a significantly

improved PK profile after Fc YTE engineering without affecting EGFR binding

properties.

To generate EGFR-targeting ADC SHR-A1307, hR3-YTE was coupled with

SHR152852 via non-cleavable L2-MC- linker with average DAR of 2 (range 0~6)

(Fig1A and sup Fig4A). The purity of SHR-A1307 was >97% analyzed by

size-exclusion chromatography (sup Fig4B), and free SHR152852 was 0.724ng/mL

(<0.0007%) and MC-SHR152852 was 3.28ng/mL (<0.003%) in SHR-A1307 solution

(10mg/mL). To further evaluate the in vivo stability, 3mg/kg SHR-A1307 was injected

(iv.) in to SD rats for pharmacokinetic analysis. Plasma concentrations of total

antibody and ADC were comparable over time, and free SHR152852 and

MC-SHR152852 were both below detection limits, indicating a good in vivo stability.

SHR-A1307 was produced in large scale and subject to further in vitro and in vivo

characterizations.

In vitro cytotoxicity of SHR-A1307 in drug resistant tumor cells harboring EGFR

or downstream gene alterations

To explore the therapeutic potential of EGFR-ADC, we examined SHR-A1307 in a

panel of cancer cell lines with different oncogenic mutations and drug resistance

patterns. In NSCLC cells with EGFR activating mutations, SHR-A1307 was very

potent in cancer cell killing, with IC50 of 0.57nM and 1.9nM in HCC827 and H1975

cells respectively, which were about 5~15 fold more potent than Erlotinib

(IC50=8.95nM) or Osimertinib (IC50=11.98nM) (Fig.1B). To evaluate if SHR-A1307

was effective in cancer cells harboring Cys797Ser (C797S) mutation, a recently




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identified resistance mechanism to Osimertinib, we constructed in-cis EGFR C797S

cell models via lenti-viral delivered EGFR Del19/T790M/C797S (DTC) or

L858R/T790M/C797S (LTC) triple mutation genes, and examined their cell viability

after treatment with different anti-EGFR agents. Consistent with previous reports,

Osimertinib lost its activity in both HCC827-DTC and H1975-LTC cells (IC50=1856nM

and 7914nM respectively). In contrast, SHR-A1307 demonstrated potent killing

activities in these cells (IC50=0.17 and 6.5nM respectively), indicates potential to

overcome EGFR C797S mediated drug resistance. In CRC and HNSCC cells

harboring EGFR downstream mutations in RAS/MAPK or PI3K pathways,

SHR-A1307 also exhibited effective cell killing with IC50 values of 8.1nM and 17.2nM

in Lovo (KRASG13D) and Detroit562 (PIK3CAH1047R) respectively, whereas Cetuximab

or Panitumumab had only minimal cytotoxic effects. Moreover, control IgG1-ADC had

no cytotoxic effects in any of the tested cells, suggesting a target-mediated cell killing

mechanism of SHR-A1307. To correlate in vitro potency with EGFR receptor density

on the cell surface, a panel of cancer cell lines with various EGFR receptor levels was

selected based on previously published results, and EGFR expressions were

confirmed by FACS in selected cell lines (Table2, sup Fig5) (32-35). Cell surface

EGFR levels in general correlated well with in vitro potency of SHR-A1307, with IC50

values ranging from 0.17~17.2nM in cells with high to moderate EGFR expression,

while in EGFR low or negative cells, SHR-A1307 had negligible effect. Together,

these results suggested that SHR-A1307 can be very effective against EGFR positive

cancer cells that are resistant to current EGFR targeted therapies in vitro, regardless

of the mutation status of EGFR or downstream mediators.

Efficacy of SHR-A1307 in anti-EGFR therapy resistant xenograft models

To assess the anti-tumor activity of SHR-A1307 in vivo, we performed studies in cell

line derived xenograft models harboring various representative activating or drug

resistant mutations. In HCC827 (EGFRDel19) xenograft model, and SHR-A1307 at

1.2mg/kg and 4mg/kg resulted in significant tumor reductions (TGI%=78.8% and

110.9% respectively), including 100% tumor remissions at 4mg/kg group (8/9 CRs




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and 1/9 PR) (Fig1C and F). To further investigate the vivo efficacy of EGFR-ADC

against drug-resistant C797S triple mutant, SHR-A1307 was evaluated in

HCC827-DTC xenograft model compared with high dose Osimertinib (50mg/kg), and

the mixture of hR3-YTE and SHR152852 at a 1:2 molar ratio (Ab+toxin Mix). As

expected, Osimertinib only induced transient tumor stasis at initial treatment phase,

and tumors quickly progressed after day10 even in the presence of high drug

exposure (Fig1D). In contrast, treatment with 4mg/kg SHR-A1307 induced significant

tumor regressions in 7 out of 8 mice, including 5 CRs, and the response continued

after treatment cessation for additional 41 days (Fig1D). As control, Ab+toxin Mix only

had minor impact on tumor growth, suggesting that sufficient anti-tumor activity of

cytotoxic payloads required antibody mediated delivery into targeted tumor cells.

There were no overt toxicities or body weight loss in mice except for those treated

with high dose Osimertinib (sup Fig6C).

To investigate the therapeutic potential of SHR-A1307 in KRAS mutated CRCs,

we tested it in Lovo xenograft model with KRASG13D mutation, which was reported to

be intrinsically resistant to high dose Cetuximab in vivo (32). Consistent with previous

studies, 40mg/kg hR3-YTE antibody had no effect on tumor growth. However,

1.2mg/kg and 4mg//kg SHR-A1307 caused dose dependent tumor growth inhibition,

including 4/10 PRs at 4mg/kg treated group. In addition, near maximal anti-tumor

efficacy was observed at 12mg/kg and 40mg/kg doses, and all mice in these 2 groups

had tumor regressions (sup Fig6A). No signs of adverse effects were observed in all

treated mice even at the highest dose of 40mg/kg (sup Fig6D-E).

To better predict clinical response of SHR-A1307 in cancer patients who are

resistant or refractory to current anti-EGFR therapies, PDX models representing

different resistance mechanisms were utilized for efficacy evaluation. Tumor EGFR

expression was analyzed by IHC based scoring system, ranging from 0 to 3 points in

EGFR positivity, and gene amplification and mutations were detected using RNAseq

(sup Table1). In an Erlotinib resistant model LU1429, SHR-A1307 caused remarkable

tumor regression at 3mg/kg and tumor stasis at 1mg/kg with TGI% of 119% (p=0.005)

and 70% (p=0.038) respectively. 50 mg/kg Erlotinib showed only marginal antitumor




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efficacy, and unconjugated antibody and low dose of SHR-A1307 (0.3mg/kg) were

undistinguishable from IgG1 control (Fig2A). In another EGFR positive NSCLC PDX

model LUN210-4a with acquired resistance to Osimertinib (mutation not defined),

treatment with 10mg/kg Osimertinib had no impact on tumor growth, but twice weekly

administration of 3mg/kg SHR-A1307 led to significant tumor growth inhibition

(TGI=111%, p=0.001) and partial tumor regressions in 4/5 of animals (Fig2B). In

NSCLC PDX model LU2503, high EGFR levels along with MET amplification were

detected in tumors, which were insensitive to high dose Erlotinib or Cetuximab

treatment, suggesting a RTK bypass mechanism of EGFR inhibitor resistance (36).

Notably, treatment with 3mg/kg SHR-A1307 resulted in strong inhibition of tumor

growth with 92% TGI (p=0.0005) (Fig2C), suggesting a promising strategy of using

EGFR-ADC to overcome RTK bypass instead of EGFR/MET dual blocking (37).

Furthermore, the anti-tumor efficacy of SHR-A1307 was analyzed in additional PDX

models (LU3075 and LU2512), representing several difficult to treat mutations (sup

Table1). Although tumors didn’t regress, SHR-A1307 caused a trend of tumor growth

inhibition in both models (Fig2D-E). The weaker potency of EGFR-ADC in these two

models was possibly due to the relatively low expression of EGFR receptors or some

intrinsic resistance to anti-mitotic agents in these tumors. In an EGFR negative CRC

PDX model CR0455, no tumor growth inhibition was noted even at highest level of

SHR-A1307 dose (40mg/kg QW) (Fig2F), suggesting that high tumor EGFR levels are

required for anti-tumor efficacy of SHR-A1307. Together, SHR-A1307 exhibited much

better anti-tumor efficacy than EGFR antibodies or TKIs in multiple tumor models, and

may provide a therapeutic opportunity to treat EGFR positive tumors that are resistant

or refractory to current anti-EGFR therapies due to inherent or acquired mechanisms.

Preclinical toxicology study of SHR-A1307 in Cynomolgus monkeys

Results from mice xenograft studies have demonstrated anti-tumor efficacy of

SHA-1307 in the absence of overt toxicities after repeated drug administration (up to

40mg/kg), indicating an excellent small animal safety profile. But since hR3 does not

cross react with murine EGFR (20), mouse is not a suitable species for on-target




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toxicity evaluation. Cynomolgus monkey is a more human relevant species as

monkey EGFR has high amino acid homology with human, and hR3 antibody

cross-binds monkey and human EGFR with similar binding affinity (38). Therefore,

cynomolgus monkey was selected to assess preclinical safety in a non-GLP

toxicology study. 10mg/kg of SHR-A1307 was administered via weekly intravenous

infusion, and high exposure was achieved in toxicokinetics analysis after first dosing

(Fig3A), which were about 3-fold higher than hR3 exposure in human under 200mg

clinical dose (Cmax=68.55+10.345 µg/ml, AUC0-inf=4521.9+1181.6 µg*h/ml) (28).

SHR-A1307 was generally well tolerated in monkeys. Mild to moderate hair loss,

desquamation and scratches on the forearms were observed, in accordance with

previously reported on-target toxicity of EGFR inhibition in epithelial cells. One

monkey had nose bleeding after 4th dosing, and was recovered after medical

treatments. Liver enzymes were normal after first administration of SHR-A1307, but

significant elevation of AST and ALP levels were detected after 4th dosing, including 3

and 1 monkeys exhibited >3x Upper Limit of Normal AST and ALP levels respectively

(Fig3B). Hematological parameters (neutrophil, lymphocyte, platelet counts)

fluctuated near normal range of variations during drug treatment. One case of

thrombocytopenia was observed and considered related to the nose bleeding in this

monkey. Most of the toxicities emerged after 4th and 5th dosing of 10mg/kg

SHR-A1307, consistent with the observation of increased exposure levels of

SHR-A1307 (nearly doubled Cmax and 4-fold AUC levels after 5th dosing vs. day0) due

to slow clearance and drug accumulation (Fig3A). Necropsy revealed mild changes in

appearances of internal organs, as decreased thymus, GI track inflammation, dark

red spots in lung and heart. Overall, SHR-A1307 exhibited well tolerable safety profile

under high drug exposure and therapeutic index was adequate for further

development.




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Discussions

EGFR alterations are among the best studied oncogenes in a variety of solid tumors,

and EGFR targeted therapeutics have resulted in marked anti-tumor responses and

significant survival benefit in cancer patients with advanced diseases (2). However,

not all patients responded effectively to EGFR inhibitors, as in NSCLC patients with

EGFR insensitizing mutations and CRC patients with RAS family mutations, no

clinical benefit was found due to primary resistance (6, 39). Moreover, despite the

initial clinical efficacy in some patients, acquired resistance invariably develops and

disease relapses typically occur within 1~2 years (12). To overcome resistance to

current anti-EGFR therapies, many novel compounds targeting resistance mutations

(MET, HER2, PIK3CA) or downstream mediators (MEK, AKT, mTOR) were in clinical

development, as single agent or in combinations. Notably, although Erlotinib plus

anti-MET antibody Onartuzumab showed significant improvements in both PFS and

OS in MET-positive patients in a retrospective phase 2 analysis, phase 3 trial of the

combination therapy was disappointing, failing to show improved outcomes in the

same patient population (40). In addition, MEK inhibitors were tested for the treatment

of refractory RAS-driven NSCLC or CRC patients with limited success due to

insufficient signal blockade as single agent or increased toxicities as combination (41,

42).

An alternative approach is to re-invigorate anti-EGFR antibodies by coupling

them with highly potent cytotoxic payloads. Herein, we described SHR-A1307 as an

example of novel EGFR targeting ADC that is potent, tumor selective and well

tolerable in mice and monkeys. SHR-A1307 has multiple features enabling it to

effectively kill tumor cells while minimizing toxicities, including 1) Intermediate EGFR

affinity of hR3 to selectively bind EGFR expressing cancer cells while sparing normal

cells; 2) non-cleavable linker to increase ADC stability and decrease toxin

deconjugation in circulation; 3) free drug cytotoxicity attenuated payload SHR152852

to further decrease the risk of systemic toxicity; 4) PK improvement by Fc domain

engineering to reduce dosing frequency and potentially decrease toxin accumulation.




17

Moreover, the ability of SHR-A1307 to efficiently kill cancer cells that were

non-responsive to current EGFR inhibitors provides an attractive therapeutic

opportunity to overcome drug resistance in patients with EGFR overexpression

tumors. Particularly, SHR-A1307 had demonstrated low nanomolar in vitro cytotoxic

activities in a broad spectrum of cancer cells harboring different drug resistance

mutations. This finding implies that potent cancer killing effect of anti-mitotic payload

SHR152852 is independent of EGFR or downstream mutations. Remarkably, in

engineered C797S mutated (DTC and LTC) NSCLC cells that are resistant to

available EGFR TKIs, SHR-A1307 demonstrated >1000 fold higher potency than

Osimertinib. This anti-tumor activity was further confirmed in HCC827-DTC mice

xenograft model, and sustained tumor regressions were observed after only 3 weekly

injections of SHR-A1307. Lovo and Detroit562 cells represent patients

refractory/relapsed from EGFR mAb treatments due to RAS or PIK3CA mutations

respectively (11, 32). Proliferation of both cells was significantly inhibited by

SHR-A1307 in vitro, and dose dependent tumor growth response was observed in

SHR-A1307 treated Lovo xenograft tumors. More importantly, we found that the in

vitro and in vivo anti-tumor activities of SHR-A1307 were closely associated with

surface EGFR levels, suggesting tumor cell associated EGFR expression could be a

useful biomarker to predict clinical response in patients. These findings were further

substantiated using PDX tumor models. SHR-A1307 demonstrated potent anti-tumor

activity in multiple PDX tumors regardless of their gene mutation or drug resistance

status, and a trend of correlation between EGFR IHC intensity scores and in vivo

tumor responses was also established.

The safety profiles of SHR-A1307 were assessed in small and large animals. In

xenograft studies, we found no overt toxicities in mice repeatedly dosed with up to

40mg/kg SHR-A1307, which was more than 10~20 fold of in vivo efficacious doses.

Although SHR-A1307 lacks cross reactivity with murine EGFR and on-target toxicities

were not evaluable, the excellent safety profiles in mice indicated low risk of

non-specific ADC uptake and negligible free toxin related toxicities, which were

usually considered as important contributing factors to ‘antigen-independent' adverse




18

effects (43). To evaluate the tolerability of SHR-A1307 in a more human-related

species, toxicology study was performed in cynomolgus monkeys. SHR-A1307 was

generally well tolerable in monkeys, with low incidence of serious adverse effects.

EGFR target-related toxicities were observed, but mostly well manageable and low

grade in severity. Due to slow clearance and drug accumulation, the exposure level of

SHR-A1307 in monkeys significantly increased after repeated weekly dosing, and late

onset of thrombocytopenia and liver function abnormalities were identified after 4th or

5th dosing. One monkey had severe platelet counts drop, manifested as nose bleeding

and mucosal hemorrhage. Since severe thrombocytopenia was uncommon in human

patients treated with hR3 or other EGFR inhibitors (3-5, 19, 21), this AE is not likely an

EGFR on-target effect. Rather, thrombocytopenia induced by ADCs is thought to be

related to megakaryocyte progenitor inhibition by tubulin acting agents (27), and

mechanistic study of T-DM1 suggested that FcγRIIa expressed on megakaryocyte

progenitors may contribute to target-independent ADC uptake and impairment of

megakaryocyte differentiation (44). As SHR-A1307 is an IgG1 based ADC like T-DM1,

it was possible that at high plasma concentration, SHR-A1307 may trigger transient

FcγRIIa mediated internalization to impair platelet production. Other common ADC

related hematological adverse effects, like lymphopenia or neutropenia, were not

observed in 10mg/kg SHR-A1307 treated monkeys, suggesting a low risk of bone

marrow suppression thanks in part to the attenuated free drug cytotixicity of payload

SHR152852 (27). Elevated liver enzymes were also observed as common AEs after

SHR-A1307 administration. It has been proposed that mannose receptor expressed

on hepatic sinusoidal cells can mediate target-independent uptake of ADC into the

liver (45), which could be a major contributing factor to SHR-A1307 caused hepatic

toxicity.

The assessment of PK results suggested that dosage optimization of SHR-A1307

could be applied to further improve therapeutic index. We noticed high peak and

trough plasma concentrations in SHR-A1307 dosed monkeys, which were much

higher than the efficacious concentration required for cancer killing. Dose de-titration

was practicable to reduce peak concentration and minimize Cmax related toxicity, while




19

maintaining sufficient plasma levels required for efficacy. Furthermore, weekly

administration of SHR-A1307 resulted in accumulation of the drug, due to slow

clearance and prolonged t1/2 of 4~5 days after Fc YTE engineering, which caused late

onset of toxicities (after 4 or 5 cycles). Adjusting the dosing frequency from weekly to

biweekly administration may reduce the drug accumulation and keep plasma drug

exposure under toxicity threshold.

In conclusion, SHR-A1307 has demonstrated robust and sustained anti-tumor

efficacy in multiple cell lines and xenograft tumor models that were resistant to current

anti-EGFR therapies. Toxicology studies in monkeys suggested a well acceptable

safety profile and sufficient therapeutic index to support future clinical testing in

human.




20

Acknowledgments

The design, conduct and financial support for the study was provided by Shanghai

Hengrui Pharmaceutical Co., Ltd (a subsidiary of Jiangsu Hengrui Medicine Co., Ltd.).

The authors thank contributions by the SHR-A1307 project team, including but not

limited to Xiangdong Qu, Xiaoyan Zhu, Limin Zhang, Juan Luo, Jingwen Zeng, Zairan

Chen, Yanbin Guan, Changyong Yang, Zhendong Xue, Yuchang Mao and Penghui

Sun.




21

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24

Table1. Activities of MMAF and its derivatives in human cells as free toxin or ADC

Cytotoxicity

IC50 nM

Structure

Her2-ADC a

Cytotoxicity

EGFR-ADC a

Cytotoxicity

Free toxin Cytotoxicity ADC / Free

toxin activity

ratio

SK-BR-3

(Her2+)

HepG2

(Her2-)

HCC827

(EGFR+)

HepG2

(EGFR-)

SK-BR-3 HCC827 HepG2

MMAF

0.16 >667 1.09 >667 1.28 4.47 10.6 8~16

SHR151329

0.026 >667 0.35 >667 1.11 1.40 8.60 8~85

SHR151522

0.15 NDb ND ND 1.06 ND 4.12 ~14

SHR151526

0.097 >667 0.45 >667 81.6 370.1 174.7 1645~1682

SHR151247

0.33 ND 1.0 ND 81.7 595.5 614.5 495~1191

SHR152852

0.031 >667 0.26 >667 47.7 214.7 283.7 1652~3077

a Toxin payload conjugated to either Her2 antibody (pertuzumab) or EGFR antibody (hR3);

DAR=1.9~2.4

b ND: not determined;




25

Table2. EGFR expression and in vitro cytotoxicity of SHR-A1307 in a panel of solid

tumor cell lines with different EGFR and downstream gene alterations

Cell line Cancer

type Gene alterations

Resistance to

EGFR targeted

therapies

SHR-A1307

Cytotoxicity

IC50 nM

EGFR expression (refs 32-35)

Reported EGFR

expression

Reported EGFR

Number / cell

DiFi CRC wtEGFR amp Sensitive 9.9 + 3.7 High ~1.8x106

HCC827 NSCLC EGFR Del19 Sensitive 0.57 + 0.09 High

HCC827-DTC NSCLC EGFR

Del19/T790M/C797S 3rd gen TKI 0.17 + 0.09

H1975 NSCLC EGFR L858R/T790M 1st gen TKI 1.9 + 0.56 High

H1975-LTC NSCLC EGFR

L858R/T790M/C797S 3rd gen TKI 6.5 + 0.8

Detroit 562 HNSCC PIK3CA H1047R EGFR mAb 17.2 + 1.0 Moderate

Lovo CRC KRAS G13D EGFR mAb 8.1 + 2.6 Moderate 1.28x105

SW480 CRC KRAS G12V EGFR mAb >220 Moderate ~9x104

HCT116 CRC KRAS G13D EGFR mAb >220 Low

HT-29 CRC B-RAF V600E EGFR mAb >133 Low 9000

COLO-205 CRC B-RAF V600E EGFR mAb No inhibition* Low

SW620 CRC KRAS G12V EGFR mAb No inhibition Negative 0

*No significant growth inhibition @ highest concentration tested (167nM ~ 667nM)

Results are expressed as mean + SD

1st gen TKI: erlotinib, gefitinib

3rd

gen TKI: Osimertinib

EGFR mAb: cetuximab, panitumumab




26

Figure legends

Figure 1. In vitro and in vivo activities of SHR-A1307 in cancer cells harboring

different EGFR mutations. (A) Chemical structure of SHR-A1307; (B) In vitro

activities of SHR-A1307 and anti-EGFR therapies in multiple cancer cell lines with

different mutations; IgG1-ADC control is chimeric human IgG1 anti-HIV antibody C25

conjugated with the same linker drug as SHR-A1307 (DAR~2.2); (C) HCC827

xenograft model with EGFR Del19 activating mutation (9 mice in each group); (D)

Osimertinib resistant HCC827-DTC tumor model represents patients with acquired

C797S mutation after progression on Osimertinib (8 mice in each group). (E) EGFR

antibody resistant Lovo tumor model represents patients resistant to Cetuximab or

Panitumumab due to downstream RAS/RAF mutations (10 mice in each group).

Tumor growth curve after treatment with small molecule TKI (Osimertinib), hR3-YTE

mAb, Ab+toxin Mix, or SHR-A1307 showing the mean tumor volumes (+ SEM); (F)

Waterfall plots of tumor size change from baseline showing individual tumor

responses in mice. QW: once weekly; Complete remission (CR) and partial remission

(PR) of tumors are defined as tumor size change after treatment achieved 100%

and >30% reduction respectively compared with initial tumor size before treatment.

Statistical significance: p<0.05 (*), p<0.01(**), p<0.001(***), ns. non-significant;

Figure 2. Anti-tumor efficacy of SHR-A1307 in multiple drug resistant PDX

models with different EGFR expression levels. (A) Erlotinib resistant NSCLC

model LU1429 treated with 0.3, 1 and 3mg/kg SHR-A1307, 50mg/kg Erlotinib, 3mg/kg

hR3-YTE and IgG1 controls; B, Osimertinib resistant NSCLC model LUN210-4a

treated with 3mg/kg SHR-A1307 and 10mg/kg Osimertinib; NSCLC models LU2503

(C) and LU3075 (D) and LU2512 (E) were treated with 3mg/kg SHR-A1307; (F) CRC

model CR0455 was treated with high dose (40mg/kg) SHR-A1307; Gene alternations

in each PDX model were summarized in Supplementary table 1; Representative

pictures of EGFR IHC staining in PDX tumors were shown. IHC intensity was

analyzed by pathologists using 4-point scoring system: 0 (negative); 1 (weak staining




27

on membrane and <10% positive area); 2 (medium staining on membrane and 10~50%

positive area); 3 (strong staining on membrane and >50% positive area).

Figure 3. Toxicology study of SHR-A1307 in Cynomolgus monkey

A, Monkey plasma drug concentrations and toxicokinetic parameters after first and

last (5th) drug dosing; Data was shown as Mean values + SEM; B, Clinical chemistry

and hematology parameters with significant changes after dosing; Grey areas denote

the normal range of laboratory values.




HCC827-DTC(EGFR Del19/T790M/C797S)

-2 0 2 4

0

50

100

Log[nM]

Inhi

bitio

in%

hR3-YTE

SHR-A1307

OsimertinibErlotinib

IgG1-ADC

HCC827(EGFR Del19)

-2 0 2 4-20

0

20

40

60

80

100

Log[nM]

Gro

wth

Inhi

bitio

in%

hR3-YTESHR-A1307OsimertinibErlotinibIgG1-ADC

A.

Lovo(KRAS G13D)

-2 0 2 4-20

0

20

40

60

80

Log[nM]

Gro

wth

Inhi

bitio

in% IgG1-ADC

hR3-YTE

SHR-A1307

Panitumumab

H1975(EGFR L858R/T790M)

-2 0 2 4-20

0

20

40

60

80

Log[nM]

Gro

wth

Inhi

bitio

in%

hR3-YTE

SHR-A1307

OsimertinibErlotinib

IgG1-ADC

B.

SHR-A1307

Detroit562(PIK3CA H1047R)

-2 0 2 4-20

0

20

40

60

Log[nM]

Gro

wth

Inhi

bitio

in%

hR3-YTESHR-A1307CetuximabIgG1-ADC

H1975-LTC(EGFR L858R/T790M/C797S)

-2 0 2 4-20

0

20

40

60

Log[nM]

Gro

wth

Inhi

bitio

in%

hR3-YTESHR-A1307OsimertinibIgG1-ADC

Lovo Xenograft

Day

Tum

or v

olum

e m

m3

0 2 7 9 13 17 200

500

1000

1500

2000

2500Vehicle (PBS)

SHR-A1307 4mg/kg QWx3

hR3-YTE 40mg/kg QWx3

SHR-A1307 1.2mg/kg QWx3

*

*

HCC827-DTC Xenograft

1 4 7 10 14 17 21 24 28 31 35 39 42 46 49 52 560

1000

2000

3000

Day

Tum

or v

olum

e m

m3

Vehicle (PBS)Osimertinib 50mg/kg QDx21Ab+toxin Mix 4mg/kg QWx3SHR-A1307 4mg/kg QWx3

***

ns.

HCC827 Xenograft

Day

Tum

or v

olum

e m

m3

1 4 7 11 14 18 21 25 28 32 340

500

1000

1500

2000Vehicle (PBS)hR3-YTE 4mg/kg QWx3

SHR-A1307 4mg/kg QWx3SHR-A1307 1.2mg/kg QWx3

**

***

Tum

or V

olum

e %

cha

nge

-100

-50

0

50

100

500

1000

1500

VehiclehR3-YTE Ab SHR-A1307 1.2mg/kg

SHR-A1307 4mg/kgOsimeritinib

Ab+toxin mix

HCC827 EGFR Del19 HCC827-DTC EGFR C797S Lovo KRAS G13D

C. D.

E. F.

O

NO O

NH

O

OF

O

ON

N

N

O

NO

O

O

SHNAb

Figure 1.

SHR152852Maleimidocaproyl linker (MC)

ThioetherLinker (L2)




A. B.

C. D.

E.

0 5 1 0 1 5 2 0 2 5

0

5 0 0

1 0 0 0

1 5 0 0

Ig G 1 c o n t ro l 3 m g /k g Q W x 3

h R 3 -Y T E 3 m g /k g Q W x 3

S H R -A 1 3 0 7 0 .3 m g /k g Q W x 3

S H R -A 1 3 0 7 1 m g /k g Q W x 3

S H R -A 1 3 0 7 3 m g /k g Q W x 3

E r lo t in ib 5 0 m g /k g Q D x 1 8

D a y s

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

L U 1 4 2 9

*

**

n s .

0 5 10 15 20 250

200

400

600

800

1000

1200

Vehicle

SHR-A1307 3mg/kg BIWx3

Osimertinib 10mg/kg QDx21

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

LUN210-4a

Days

***

ns.

0 5 1 0 1 5 2 0 2 5

0

1 0 0 0

2 0 0 0

3 0 0 0

V e h ic le

S H R -A 1 3 0 7 3 m g /k g Q W x 4

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

L U 2 5 0 3

D a y s

***

0 5 10 15 20 250

500

1000

1500

2000

2500

3000Vehicle

SHR-A1307 3mg/kg QW x3

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

LU2512

Days

ns.

0 5 10 15 20 25 300

500

1000

1500

Vehicle

SHR-A1307 3mg/kg QW x3

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

Days

LU3075

ns.

0 5 10 15 20 250

500

1000

1500

2000

Tu

mo

r V

olu

me

(m

m3

)

me

an

S

EM

CR0455

Days

Vehicle QW x3

SHR-A1307 40 mg/kg QW x3

ns.

F.

IHC score: 2.5

IHC score: 3.0

IHC score: 2.0

IHC score: 2.0

IHC score: 1.5 IHC score: 0

Figure 2.




Time (hrs)

Plas

ma

Leve

ls (u

g/m

l)

0 50 100 150 20010

100

1000

after 5th dosingafter 1st dosing

TK(mean + SD)

10mg/kg(1st dosing)

10mg/kg (5th dosing)

Cmax (μg/mL) 266.9±15.4 452.8±8.0

AUC（0-t ) (μg·h/mL) 5838±3571.3 22979.5±3243.5

AUC（0-inf) (μg·h/mL) 10030.3±7496.3 29736.2±7488.1

t1/2 (h) 120.8±25.9 79.9±21.8

Cl (mL/h/kg) 0.3±0.2 0.35±0.08

A.

B.

IU /

L

0 3 7 17 22 24 280

100

200

300

400

Time (day)

AST1M0011M0021F001

IU /

L

0 3 7 17 22 24 280

50

100

150

Time (day)

ALT

1F001

1M0011M002

IU /

L

0 3 7 17 22 24 280

500

1000

1500

2000

Time (day)

ALP

1F001

1M0011M002

109 /

L

0 3 7 17 22 24 280

5

10

15

20

Time (day)

Lymphocyte

1M0021F001

1M001

109 /

L

0 3 7 17 22 280

200

400

600

800

1000

Time (day)

Platelet

1F001

1M0011M002

109 /

L

0 3 7 17 22 24 280

5

10

15

20

Time (day)

Neutrophil1M0011M0021F001

Figure 3.




Published OnlineFirst April 8, 2019.Mol Cancer Ther Kaijie He, Jianyan Xu, Jindong Liang, et al. refractory to anti-EGFR therapiesSHR-A1307, for the treatment of solid tumors resistant or Discovery of a novel EGFR targeting antibody-drug conjugate,

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