Supplementary Online Content 2

© 2018 American Medical Association. All rights reserved.

Supplementary Online Content 1

2

Ferris RL, Saba NF, Gitlitz BJ, et al. Effect of adding motolimod to standard combination 3 chemotherapy and cetuximab treatment of patients with squamous cell carcinoma of the head4 and neck: the Active8 randomized clinical trial. JAMA Oncol. Published online June 21, 2018.5 doi:10.1001/jamaoncol.2018.1888 6

7

eMethods. 8 eResults. 9 eFigure 1. Kaplan-Meier analysis of PFS by investigator-assessed irRECIST and independent central 10 assessment by RECIST 1.1. 11 eFigure 2. Treatment effect on PFS and OS. 12 eFigure 3. In vivo responses for selected analytes. 13 eFigure 4. Kaplan-Meier analysis of PFS and OS in motolimod patients with ISR vs placebo. 14 eFigure 5. Association between FcɣR IIa polymorphisms and overall survival. 15 eTable 1. Baseline demographics and clinical characteristics. 16 eTable 2. Prior treatments. 17 eTable 3. Dose modifications. 18 eTable 4. Response rates in ITT population and HPV+ oropharyngeal subgroup. 19 eTable 5. Treatment-emergent adverse events occurring in ≥ 25% of subjects. 20 eReferences. 21

22

This supplementary material has been provided by the authors to give readers additional information 23 about their work. 24

25


eMETHODS 26 27

Patients 28

Eligible patients were ≥18 years with histologically confirmed recurrent/metastatic (R/M) squamous cell 29

carcinoma of the oral cavity, oropharynx, hypopharynx, or larynx with measurable disease per Response 30

Evaluation Criteria in Solid Tumors (RECIST 1.1), Eastern Cooperative Oncology Group (ECOG)11 31

performance status of 0 or 1, and adequate bone marrow, renal, and hepatic function. No previous systemic 32

treatment for R/M disease was allowed nor was disease that was amenable to curative local therapy. 33

Major surgery or irradiation within 4 weeks of randomization was not allowed nor was prior systemic treatment 34

with anti-cancer therapy, unless administered for localized head and neck squamous cell carcinoma (HNSCC) 35

and completed more than 6 months before disease recurrence. Patients were ineligible if they had active 36

autoimmune or cardiovascular disease, history of another (or second) active primary malignancy, or active brain 37

metastases. Patients who had previous treatment with immunotherapy, had a prior serious infusion reaction to 38

cetuximab, had received oral or parenteral corticosteroids within 2 weeks, or an investigational agent within 4 39

weeks prior to study randomization were also excluded. 40

41

Study conduct 42

Interactive response technology (IXRS) generated the random allocation sequence, investigators enrolled 43

participants through IXRS, and IXRS assigned participants to interventions. Randomization (block size 4) was 44

stratified by prior systemic therapy (yes or no), Eastern Cooperative Oncology Group (ECOG) performance 45

status (0 versus 1), and platinum therapy type. Patients received a maximum of 6 chemotherapy cycles. On 46

Cycle 1 Day 1, cetuximab 400 mg/m2 was given as a 2-hour IV infusion, completed at least one hour prior to 47

chemotherapy; thereafter weekly cetuximab 250 mg/m2 was given as a 1-hour infusion. During the 48

chemotherapy portion, investigational product (motolimod or placebo; IP) was given SC at 3.0 mg/m2 on Days 49

8 and 15. After chemotherapy portion completion, patients received weekly cetuximab with motolimod or 50


placebo on Days 8 and 22 of each 28-day cycle. Treatment was administered until disease 51

progression/unacceptable toxicity. Dose reductions were permitted per protocol guidelines. Patients 52

experiencing severe infusion reactions with the first cetuximab dose were removed from study. Safety 53

monitoring included surveillance and recording of AEs per National Cancer Institute (NCI) Common 54

Terminology Criteria for Adverse Events (CTCAE), Version 4.012. The trial was conducted in accordance with 55

the Declaration of Helsinki and Good Clinical Practice guidelines. The protocol was approved by each center's 56

institutional review board (see study protocol in Supplementary Materials). Patients provided written informed 57

consent prior to enrollment. 58

Statistical Analyses 59

A sample size of 175 patients receiving ≥ 1 dose of motolimod or placebo was targeted, with event-driven 60

timing of the primary analysis. A required 134 progression-free survival PFS events would detect a hazard ratio 61

(HR) of 0.67 with 85% power at a 1-sided alpha of 0.10. This HR represents a 50% PFS improvement, 62

translating to an increase in median PFS from 6 months (control arm) to 9 months (combination arm). An 63

estimated 175 patients accrued over 18 months (plus a 9-month follow-up) were required to provide the 64

necessary number of PFS events. Primary analysis was planned at 134 PFS events or earlier, pending sufficient 65

number of PFS events for at least an 80% power to detect a HR of 0.67 at a 1-sided alpha or 0.10. Overall 66

survival (OS) analysis was planned for when all randomized patients had been followed for at least 12 months 67

after final enrollment. One hundred twelve OS events were estimated to accrue during this time, providing 68

72.8% power at a 1-sided alpha of 0.10 to detect a HR of 0.70. This HR represents a 30% reduction in the risk 69

of death, and a 42.96% increase in the median OS. With an 11-month median OS assumed in the control arm, 70

median OS in the combination arm would be 15.7 months. 71

Primary endpoint analyses were performed using the intent-to-treat (ITT) population (defined as all patients 72

randomized on the study, regardless of eligibility or treatment received) using a log rank test stratified by 73

randomization strata. Immune-related PFS (irPFS) and OS were summarized by treatment arm using Kaplan-74

Meier methods, and the HR between the 2 treatment arms, and the associated 1-sided 90% confidence interval, 75


were calculated using a Cox proportional hazards regression model. PFS per RECIST 1.1 (PFS1.1) and 76

investigator-determined irPFS were assessed using similar methodology. 77

A pre-specified analysis was conducted in the patient subset with oropharyngeal disease to compare irPFS and 78

OS in human papilloma virus (HPV)+ vs HPV- patients. The number of HPV+ patients for enrollment was not 79

pre-specified, but was a natural prevalence of patients with HPV+ disease. Six patients with oropharyngeal 80

disease and unknown HPV status were excluded. Similar methods were used as for primary analyses of irPFS 81

and OS. Analyses were conducted with using SAS® version 9.4 or higher. 82

Exploratory Endpoints and Analyses 83

Exploratory endpoints included associations between clinical outcome and: pharmacodynamic (PD) response to 84

motolimod (plasma immune cytokines and chemokines); single nucleotide polymorphisms (SNP) in the gene 85

encoding toll-like receptor 8 (TLR8); polymorphisms in FcɣR IIIa and FcɣR IIa; and immune score—the 86

density and location of tumor-infiltrating lymphocytes (TIL) including CD3+, CD8+, CD45RO+, PD-L1+, 87

CD68+, and FOXP3+ cells—assessed on fresh or archived pretreatment tumor specimens (Supplementary 88

Methods). Motolimod population pharmacokinetics (PK) and subgroups of interest including patients 89

experiencing an injection site reaction (ISR) due to motolimod injection were evaluated. 90

Disease response and progression were evaluated using magnetic resonance imaging (MRI) or computed 91

tomography (CT) scans at baseline, Weeks 12 and 18, and then every 8 weeks thereafter until centrally 92

confirmed progressive disease or placement on non-protocol therapy. 93

Safety was evaluated from first dose of study treatment until post-treatment visit. All motolimod-related AEs 94

were followed until resolution/resolution with sequelae. 95

Whole blood for genotyping (TLR8 SNP, FcɣR IIIa, FcɣR IIa) and plasma samples were obtained before and 96

after IP dosing for PK and PD assessments. 97

HPV Testing 98


HPV status was collected for patients with oropharyngeal carcinoma only. If HPV status was not available in 99

the subject’s medical history, testing for HPV status by p16 immunohistochemistry, in situ hybridization, or 100

polymerase chain reaction was conducted at the time of screening. 101

Tumor Specimen Collection 102

A fresh or archived pretreatment tumor specimen was collected whenever possible; specimens were obtained 103

for 88% of patients. 104

Evaluation of Pharmacodynamic Responses 105

The HumanMAP® panel (Myriad RBM; Austin, TX), used to assess the in vivo immunopharmacologic 106

response to motolimod, is a qualitative and quantitative, multiplexed immunoassay comprising a panel of more 107

than 100 protein biomarkers, including those indicative of TLR8 activation. Blood samples are drawn into 108

standard lavender-top collection tubes prior to, and approximately 8 hours after, subcutaneous dosing with 109

motolimod or placebo. Tubes are centrifuged immediately after collection, and the supernatant is analyzed via 110

the HumanMAP assay. Plasma samples were collected on cycle 1 day 8 and cycle 3 day 8. 111

Pharmacodynamic Response Modeling 112

A pre-specified set of analytes that are known to be induced in vivo by motolimod based on previously 113

completed non-clinical and clinical trials were included in the primary analysis model, and include: IL-1ra, IL-114

6, IL-8, IL-10, MMP-9, G-CSF, MCP-1, MIP1-β, and GH. For each dose of motolimod (cycle 1 day 8 and cycle 115

3 day 8), the mean log-fold change across the set of analytes was calculated as the difference between the pre- 116

and post-dose analyte levels for each subject, and modeled quantitatively and with the use of categories based 117

on median response (e.g., low vs high, tertiles). 118

119

Both OS and PFS by irRECIST outcomes were modeled separately. When applicable, correction for multiple 120

testing was performed using the approach of Benjamini and Hochberg (1995). 121


Additional analyses were performed using penalized logistic regression via the Elastic Net as implemented in 122

the glmnet R package to explore optimal multivariate models that could predict irPFS or OS. 123

For the comparison of treatment groups, a two sample t-test was used to compare responses between motolimod 124

and placebo recipients. 125

Pharmacokinetic (PK) Analysis 126

Limited plasma samples for population pharmacokinetics were collected during the investigational product dose 127

on cycle 1 day 8 (predose, 45 minutes ± 15 minutes post-dose, 8 hours ± 15 minutes post-dose). For each 128

patient, the parameters measured included maximum serum concentration (Cmax; ng/mL), last observed plasma 129

concentration (Clast; ng/mL), and area under the curve 0 to 8 hours (AUC(0-8); ng.hr/mL). 130

Immune Score 131

Tissue was analyzed by immunohistochemistry (IHC) with multiple immune markers. The density (positive 132

cells/HPF) and location (tumor core vs invasive margin) of the following markers were assessed: CD3, CD8, 133

CD45RO, PD-L1, CD68, and FOXP3. PD-L1 membrane expression was evaluated using a mouse anti-human 134

PD-L1 antibody (clone 9A11)1 and scored at pre-specified expression level of ≥1% to be deemed “positive” in a 135

minimum of 100 evaluable tumor cells. The percentage of tumor cells expressing PD-L1 could not be 136

determined for 71 or 195 subjects, including 17 subjects in the HPV+ subgroup. 137

A Cox proportional hazard model was used to test for association between all reported immune score variables 138

and irPFS and OS. Immune score variables were modeled on the square root scale to decrease the influence of 139

large outliers. This analysis was repeated dichotomizing the variables as low/high around the median across 140

subjects. The correlation between irPFS and OS and PD-L1 expression was further assessed by evaluating the 141

percentage of intratumoral PD-L1 positive cells as a continuous variable, as well as discretize as ≤5% versus 142

>5%.143

144

145


eRESULTS 146

Exploratory Analyses 147

Aside from the ISR analysis reported, no association was found between outcome and AEs. Additionally, 148

outcomes of patients receiving carboplatin or cisplatin were analyzed, but no differences in PFS or OS were 149

observed (data not shown). 150

Interestingly, post-hoc analyses revealed that ISR+ motolimod-treated subjects had significantly improved OS 151

and PFS vs. placebo controls. Although these data arise from exploratory analyses and several explanations may 152

account for this result, one hypothesis is that ISR represents a proxy for immune activation by motolimod 153

within the tumor microenvironment 154

Exploratory Correlative Studies 155

Previous nonclinical and clinical data with motolimod were used to identify the panel of cytokines and 156

chemokines expected to be induced by motolimod in vivo3,5,7,13. At 8 hours post-dose, near the early cytokine 157

mediator response peak, levels of responsive cytokines in patients treated with motolimod were significantly 158

increased. As expected, at 8 hours post-dose, plasma levels of these mediators in the placebo group were 159

comparable to pre-dose levels. In motolimod-treated patients, mean changes in the responsive plasma mediators 160

at Cycle 1 were comparable to Cycle 3, indicating that repeat motolimod dosing did not result in desensitization 161

or augmentation of the TLR8 response. Univariate and multivariate statistical analyses did not reveal any 162

relationship between the PD markers at 8 hours and either AEs or clinical outcome, including in patient 163

subgroups (data not shown). 164

TLR8 single nucleotide polymorphism (SNP) did not predict outcomes in irPFS or OS. Furthermore, FcɣR IIIa 165

polymorphisms were not prognostic and did not predict irPFS outcomes. However, prespecified analyses 166

revealed a significant treatment effect of FcɣR IIa 15 for OS. 167

Immune score in this cohort was not prognostic or predictive of irPFS or OS in the ITT population. 168


Based on a limited sampling schedule, PK analyses were consistent with results from other trials with 169

motolimod, with expected interpatient variability. 170

Pharmacogenomics 171

TLR8 SNPs 172

TLR8 SNPs (rs3764880) were reported as A only (A/A), G only (G/G), or both (G/A, A/G). Based on the 173

hypothesis that carriers of the G allele (GG and GA/AG) would activate NFκB more effectively than those 174

without2, the primary analysis grouped patients with AA and GA/AG genotypes to determine if subjects with 175

these haplotypes had improved irPFS or OS compared to those of GG patients following treatment with 176

motolimod. 177

Specimens of whole blood for genomic DNA were collected from patients who provided appropriate informed 178

consent at the time of study entry. These specimens were genotyped for TLR8 SNPs. A total of 156 specimens 179

were analyzed and resulted. Of these, 109 subjects (69.9%) were TLR8-A/A, 38 subjects (24.4%) were TLR8-180

G/G, and 9 subjects (5.8%) were TLR8-A/G. 181

Fc-Gamma Receptor (FcɣR) Polymorphisms 182

Specimens of whole blood for genomic DNA were collected from patients who provided appropriate informed 183

consent at the time of study entry. These specimens were genotyped for FcɣR polymorphisms. A total of Total 184

of 158 specimens were analyzed and resulted for both FcɣR IIIa (rs396991) and FcɣR IIa (rs1801274). For 185

FcɣR IIIa, 68 subjects (43.0%) are FF, 16 subjects (10.1%) are VV, and 74 subjects (46.8%) are VF. For FcɣR 186

IIa, 36 subjects (22.8%) are HH, 43 subjects (27.2%) are RR, and 79 subjects (50.0%) are HR. 187

A Cox proportional hazard model adjusting for stratification (randomization) variables was used to test for 188

significant association between TLR8 SNPs and/or FcɣR polymorphisms and irPFS or OS. 189

TRANSLATIONAL CORRELATIVE RESULTS 190

Pharmacodynamic Data 191


In plasma analytes that were significantly increased following dosing with motolimod, there were no significant 192

statistical associations with clinical outcomes (irPFS or OS). 193

Immune Score 194

No immune score variable was prognostic nor predictive for irPFS or OS in the ITT population, and none was 195

not prognostic or predictive for irPFS in the HPV+ subgroup. When PD-L1 expression was discretized by 196

percent positive cells (≤5 versus >5), OS is significantly improved in HPV+ subjects with a low percentage of 197

cells expressing PD-L1 (1-sided p=0.073). When the effect of treatment with motolimod is included, OS is 198

significantly improved based on PD-L1 expression (1-sided p=0.038) and treatment with motolimod (1-sided 199

p=0.046), suggesting that HPV+ subjects benefited from motolimod independently of their PD-L1 score. Given 200

the small number of subjects, and in noting that this effect was not observed when PD-L1 was assessed as a 201

continuous variable, these data should be interpreted with caution. 202

TLR8 SNPs 203

TLR8 SNPs did not predict response in irPFS or OS. 204

Fc-Gamma Receptor (FcγR) Polymorphisms 205

Polymorphisms in FcɣR IIIa were not prognostic and did not predict irPFS or OS outcomes (eFigure 5). 206

Similarly, polymorphisms in FcɣR IIa were not prognostic and did not predict irPFS. However, there was a 207

significant interaction between the FcɣR IIa HH genotype and treatment (two-sided p=0.00498), whereby 208

motolimod-treated HH subjects had better overall survival. This effect was reversed in the placebo group. A log 209

rank test restricted to HH patients showed a significant motolimod treatment effect (two-sided p=0.00593). Of 210

note, the patient numbers in this analysis are small, with 36 subjects (18 placebo, 18 motolimod) having the HH 211

genotype and 122 subjects (56 placebo, 66 motolimod) having the RR+ RH genotype. Therefore, the 212

significance of this result should be interpreted with caution. However, while this finding needs further 213

investigation, it does offer support to the proposed mechanism of action of motolimod through enhancement of 214

antibody-dependent cellular cytotoxicity (ADCC).215


eFigure 1. Kaplan-Meier analysis of PFS by investigator-assessed irRECIST and independent central 216 assessment by RECIST 1.1. 217 218

219

220


eFigure 2: Treatment effect on PFS and OS 221

222 223


eFigure 3: In vivo responses for selected analytes. Pre-specified cytokines were measured for each dose of 224 motolimod/placebo (cycle 1 day 8 and cycle 3 day 8) and plotted as box plots on a log scale. Medians with 225 quartiles are graphed across the set of analytes. 226

227 228 229

230

G‐CSF

5

GH IL‐1ra IL‐6 IL‐8

Placebo Motolimod

IL‐10 MIP‐1 MMP9 MCP‐1

Cycle

1

Log

Cycle

3

0

0

5


eFigure 4. Kaplan-Meier analysis of PFS and OS in motolimod patients with ISR vs placebo. 231

232

233 ISR=injection site reaction. 234 235 236

237


eFigure 5. Association between FcɣR IIa Polymorphisms and Overall Survival 238

239

240

241

242

(n = 56)

(n = 66)

(n = 18)

(n = 18)


eTable 1 Baseline Demographics and Clinical Characteristics

EXTREME + Placebo

N = 95 (%)

EXTREME + Motolimod N = 100 (%)

Total

N = 195 (%) Median age, years (range) 60.0 (23, 78) 58.0 (23, 81) 58.0 (23, 81) Gender

Male 81 (85.3) 85 (85.0) 166 (85.1) Female 14 (14.7) 15 (15.0) 29 (14.9) Race

White 78 (82.1) 81 (81.0) 159 (81.5) Black or African American 12 (12.6) 15 (15.0) 27 (13.8) Asian 2 (2.1) 1 (1.0) 3 (1.5) Other 3 (3.2) 3 (3.0) 6 (3.1)

EthnicityHispanic or Latino 7 (7.4) 5 (5.0) 12 (6.2) Not Hispanic or Latino 87 (91.6) 95 (95.0) 182 (93.3) Unknown 1 (1.1) 0 (0.0) 1 (0.5)

Smoking or tobacco use Current or former 61 (64.2) 75 (75.0) 136 (69.7)

Never 34 (35.8) 25 (25.0) 59 (30.3) ECOG performance statusa

0 37 (38.9) 38 (38.0) 75 (38.5) 1 58 (61.1) 62 (62.0) 120 (61.5)

Prior systemic therapy for SCCHN (curative setting)b

Yes 62 (65.3) 65 (65.0) 127 (65.1) No 33 (34.7) 35 (35.0) 68 (34.9)

Prior treatment Surgery 53 (55.8) 56 (56.0) 109 (55.9) Radiotherapy 81 (85.3) 79 (79.0) 160 (82.1) Chemotherapy 55 (57.9) 63 (63.0) 118 (60.5) Chemotherapy + radiotherapy 40 (42.1) 49 (49.0) 89 (45.6) Other systemic therapyc 22 (23.2) 10 (10.0) 32 (16.4) Platinum therapy received at randomization

Cisplatin 18 (18.9) 21 (21.0) 39 (20.0) Carboplatin 77 (81.1) 79 (79.0) 156 (80.0)


HPV status at screeningd

Positive 28 (65.1) 24 (60.0) 52 (62.7) Negative 12 (27.9) 13 (32.5) 25 (30.1) Missing 3 (7.0) 3 (7.5) 6 (7.2)

Location of primary cancer Oral cavity 26 (27.4) 27 (27.0) 53 (27.2) Oropharynx 43 (45.3) 40 (40.0) 83 (42.6) Hypopharynx 5 (5.3) 4 (4.0) 9 (4.6) Glottic larynx 11 (11.6) 8 (8.0) 19 (9.7)

Supraglottic larynx 9 (9.5) 14 (14.0) 23 (11.8) Primary unknown/other 1 (1.1) 7 (7.0) 8 (4.1)

Median time from diagnosis to randomization, (mos)

16.6 17.5 17.2

NOTE: Data are given as No. (%) unless otherwise noted a At screening b Must have been of curative-intent. c See eTable 2 for details on other systemic therapy. d Subjects with oropharyngeal carcinoma only

eTable 2. Prior Treatments

SoC + Placebo

N = 95 (%) SoC + Motolimod

N = 100 (%) Total

N = 195 (%)

Prior surgery No 44.2 44.0 44.1

Yes 55.8 56.0 55.9

Prior radiotherapy No 14.7 21.0 17.9

Yes, setting 85.3 79.0 82.1

Palliative 4.2 2.0 3.1

Adjuvant 33.6 33.0 33.3

Primary 53.7 49.0 51.3

Prior chemotherapy No 42.1 37.0 39.5

Yes 57.9 63.0 60.5

Adjuvant 6.3 6.0 6.2

Neoadjuvant 3.2 2.0 2.6

Concomitant with radiotherapy 42.1 49.0 45.6

Other 4.2 5.0 4.6

Other – radiosensitizing chemotherapya 4.2 6.0 5.1


Page 18 of 22

SoC + Placebo N = 95 (%)

SoC + Motolimod N = 100 (%)

Total N = 195 (%)

Other prior systemic therapy No 76.8 90.0 83.6

Yes 23.2 10.0 16.4

Prior cetuximab 21.1 10.0 15.4

Adjuvant 2.1 0 1.0

Neoadjuvant 1.1 0 0.5

Concomitant 17.9 10.0 13.8

Other prior systemic therapy excluding cetuximab 6.3 0 3.1

Adjuvant 1.1 0 0.5

Neoadjuvant 0 0 0

Concomitant 4.2 0 2.1

Other 1.1 0 0.5 a Radiosensitizing chemotherapy was not considered as prior systemic therapy. Note: For each therapy, multiple subjects were counted for more than one reason, therefore the count for Yes was not be the same as the sum as each of the reasons for that therapy.


Page 19 of 22

eTable 3. Dose Modifications

SOC + Placebo

N = 86 (%)

SOC + Motolimod

N = 89 (%)

Median duration of treatment (weeks) Cetuximab 25.3 26.0 Carboplatin 18.0 18.0 Cisplatin 16.9 14.1 Fluorouracil (5-FU) 18.0 19.0 Investigational product – motolimod or

Placebo 23.0 24.0

Patients with at least 1 dose delay

CetuximabSubjects with dose delay 48 (55.8) 49 (55.1) Subjects with dose reduction 3 (3.5) 14 (15.7)

Platinum therapy (carboplatin or cisplatin) Subjects with dose delay 55 (64.0) 49 (55.1) Subjects with dose reduction 36 (41.9) 38 (42.7)

Fluorouracil (5-FU) Subjects with dose delay 52 (60.5) 51 (57.3) Subjects with dose reduction 44 (51.2) 38 (42.7)

Investigational product – motolimod or placebo Subjects with dose delay 46 (52.5) 44 (49.4) Subjects with dose reduction 2 (2.3) 14 (15.7)


Page 20 of 22

eTable 4 Response rates in ITT population and HPV+ oropharyngeal subgroup ITT Population Subgroup

Independent Central Assessment by

irRECIST

Investigator Assessment

by irRECIST

Independent Central Assessment by

RECIST

HPV+ Oropharyngeal Patients

SOC + Placebo (n=95)

SOC + Motolimo

d (n=100)


SOC + Motolimo

d (n=100)


SOC + Motolimo

d (n=100)


SOC + Motolimo

d (n=24)

Objective response rate*

32 (33.7) 38 (38.0) 32 (33.7) 40 (40.0) 32 (33.7) 38 (38.0) 11 (39.3) 7 (29.2)

p-value** P=0.536 P=0.350 P=0.536 P=0.249 Best overall response

CR 5 (5.3) 2 (2.0) 4 (4.2) 3 (3.0) 5 (5.3) 2 (2.0) 2 (7.1) 0 (0.0) PR 27 (28.4) 36 (36.0) 28 (29.5) 37 (37.0) 27 (28.4) 36 (36.0) 9 (32.1) 7 (29.2) SD 23 (24.2) 22 (22.0) 42 (44.2) 33 (33.0) 23 (24.2) 20 (20.0) 8 (28.6) 9 (37.5) NN 15 (15.8) 12 (12.0) 6 (6.3) 10 (10.0) 15 (15.8) 12 (12.0) 4 (14.3) 2 (8.3) PD 8 (8.4) 9 (9.0) - - 8 (8.4) 11 (11.0) 1 (3.6) 2 (8.3) NE 16 (16.8) 18 (18.0) 15 (15.8) 17 (17.0) 16 (16.8) 18 (18.0) 4 (14.3) 4 (16.7) ND 1 (1.1) 1 (1.0) - - 1 (1.1) 1 (1.0) 0 (0.0) 0 (0.0)

Abbreviations: ITT, intent to treat; HPV, human papillomavirus; irRECIST, immune-related Response Evaluation Criteria In Solid Tumors; RECIST, Response Evaluation Criteria In Solid Tumors; SOC, stand of care (5FU + cetuximab + cisplatin or carboplatin); CR, complete response; PR, partial response; SD, stable disease; NN, non-CR/non-PD; PD, progressive disease; NE, not evaluable; ND, no measurable disease at baseline. *Objective response rate is defined as the percentage of subjects who achieve best overall response of CR or PR**p-values are from Cochran-Mantel-Haenszel test controlling for randomization stratification factors and comparing tumor responserates between the treatment groups


Page 21 of 22

eTable 5 Treatment-emergent Adverse Events Occurring in 25% of Subjects

Adverse Event EXTREME + Placebo N = 86 (%)

EXTREME + Motolimod N = 86 (%)

Total N = 175 (%)

All Grades Grade 3 All Grades Grade 3 All Grades Grade 3 Neutropenia 45 (52.3) 31 (36.0) 50 (56.2) 35 (39.3) 95 (54.3) 66 (37.7) Thrombocytopenia 43 (50.0) 14 (16.3) 50 (56.2) 14 (15.7) 93 (53.1) 28 (16.0)

Anaemia 39 (45.3) 13 (15.1) 53 (59.6) 22 (24.7) 92 (52.6) 35 (20.0)

Fatigue 39 (45.3) 10 (11.6) 38 (42.7) 10 (11.2) 77 (44.0) 20 (11.4)

Stomatitis 42 (48.8) 13 (15.1) 34 (38.2) 7 (7.9) 76 (43.4) 20 (11.4)

Dermatitis acneiform 31 (36.0) 5 (5.8) 43 (48.3) 7 (7.9) 74 (42.3) 12 (6.9)

Leukopenia 29 (33.7) 15 (17.4) 38 (42.7) 17 (19.1) 67 (38.3) 32 (18.3)

Nausea 29 (33.7) 2 (2.3) 36 (40.4) 4 (4.5) 65 (37.1) 6 (3.4)

Hypomagnesaemia 31 (36.0) 2 (2.3) 26 (29.2) 0 (0.0) 57 (32.6) 2 (1.1)

Vomiting 24 (27.9) 4 (4.7) 32 (36.0) 5 (5.6) 56 (32.0) 9 (5.1)

Diarrhoea 26 (30.2) 3 (3.5) 28 (31.5) 5 (5.6) 54 (30.9) 8 (4.6)

Hypokalaemia 24 (27.9) 9 (10.5) 29 (32.6) 4 (4.5) 53 (30.3) 13 (7.4)

Constipation 23 (26.7) 0 (0.0) 30 (33.7) 0 (0.0) 53 (30.3) 0 (0.0) Pyrexia 10 (11.6) 0 (0.0) 38 (42.7) 2 (2.2) 48 (27.4) 2 (1.)

Rash 23 (26.7) 1 (1.2) 17 (19.1) 2 (2.2) 40 (22.9) 3 (1.7)

Chills 5 (5.8) 0 (0.0) 33 (37.1) 1 (1.1) 38 (21.7) 1 (0.6)

Injection site reactions 0 (0.0) 0 (0.0) 35 (39.3) 2 (2.2) 35 (20.0) 2 (1.1)


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Supplementary Online Content 2