Tilsotolimod Exploits the TLR9 Pathway to Promote Antigen Presentation and Type 1 IFN Signaling in Solid Tumors: A Multicenter International Phase I/II Trial (ILLUMINATE-101)

This article is featured in Highlights of This Issue, p. 3403

T cell–targeted cancer immune therapies such as checkpoint inhibitors (CPI) provide durable, systemic antitumor responses to some patients, but the effects rarely extend to less immunogenic tumors. Treatments that stimulate the innate immune system and reverse the immunosuppressive tumor microenvironment represent an emerging strategy to promote antitumor immunity either alone or in combination with CPIs for patients with immunologically cold tumors. Toll-like receptors (TLR) are expressed by an array of immune cells and enable a nonspecific immune response to pathogen-associated molecular patterns, biochemical patterns unique to pathogens (1). They trigger innate immunity by increasing the production of interferon and other cytokines and activating antigen-presenting cells, resulting in increased activation of effector T cells. TLR9 is located in endosomal compartments and is predominantly expressed by plasmacytoid dendritic cells (pDC) and B cells, elements of the innate and adaptive immune systems, respectively (2, 3). Direct, intratumoral stimulation of TLR9 with synthetic agonists promotes a local antitumor response via activation of the innate and ultimately the adaptive immune systems that can subsequently lead to systemic immunity. This limits systemic immune-related toxicities compared with the systemic administration of CPIs (4).

The TLR9 agonist IMO-2125 (tilsotolimod) is a synthetic phosphorothioate oligodeoxynucleotide consisting of two strands that are 3′–3′ linked. Intratumoral (i.t.) tilsotolimod demonstrated immune-mediated antitumor efficacy in murine syngeneic lymphoma and colon carcinoma models by an increase in CD3⁺ T lymphocytes within the tumor microenvironment and an upregulation of selected checkpoint genes including PD-1, PD-L1, CEACAM1, OX40, OX40L, CTLA-4, LAG3, and TIM3 (5). Additionally, in preclinical studies, tilsotolimod induced high levels of IFN-alpha from dendritic cells (DC) along with B-cell proliferation and differentiation and significantly augmented cytotoxic T-cell responses against tumor antigens leading to regression of injected and distant, noninjected tumors, suggestive of an abscopal effect (6–8). These findings are suggestive of both potential single-agent activity use and in combination with CPIs. In a phase I/II clinical study of i.t. tilsotolimod plus ipilimumab or pembrolizumab in patients with advanced melanoma who progressed on or after treatment with a PD-1 inhibitor, both combinations were generally well tolerated (9, 10). In this study, evaluable patients who received the 8 mg dose of tilsotolimod combined with ipilimumab (N = 49) achieved an overall response rate (ORR) of 22.4% and a disease control rate of 71%. Tumor reduction was observed in both the injected and noninjected lesions. Seven of the 11 responses lasted for ≥ 6 months, including an ongoing complete response of nearly 4 years. Tumor biopsies revealed a robust, early activation of a type I interferon response and DC activation in the injected lesion along with CD8⁺ T-cell proliferation in both injected and noninjected tumors (9). This led to a phase III randomized control trial in patients with advanced melanoma (ILLUMINATE-301; ref. 11).

Here, we present a phase I/II dose escalation trial of single-agent tilsotolimod in patients with refractory solid tumors to determine feasibility, safety, recommended phase II dose (RP2D), pharmacokinetics, clinical activity, immunologic activity, and biomarkers for immunologic assessment. This is one of the first clinical trials to study TLR9 agonists via i.t. injection in solid tumors other than advanced melanoma and to investigate the feasibility and safety of visceral i.t. injections. We hypothesized that treatment with tilsotolimod will modulate the tumor microenvironment in microsatellite stable solid tumors and lead to DC activation, increases in type I IFN, stimulate cytotoxic T-cell responses, upregulate checkpoint genes, and demonstrate an abscopal effect. Herein, we provide mature results from both the dose-escalation and expansion cohorts.

This open-label, monotherapy, multicenter phase Ib dose evaluation study (NCT03052205) assessed the safety, pharmacokinetics (PK), preliminary clinical activity, immune effects, and recommended phase II dose (RP2D) of i.t. tilsotolimod monotherapy in multiple solid tumor types. The study was run in two parts, the dose evaluation/escalation portion, and the dose expansion portion. In the dose escalation phase, patients received i.t. tilsotolimod in doses escalating from 8 to 32 mg (8, 16, 23, and 32 mg) into a single lesion on day 1 of weeks 1, 2, and 3 (3-week cycle), and then on day 1 of week 1 of a 3-week cycle, for up to a maximum of 17 total cycles (Supplementary Fig. S1 and Supplementary Table S1). Cohorts of approximately eight patients were planned to be enrolled sequentially with dose escalation proceeding unless >2 patients experienced a DLT. An additional eight patients were planned to be enrolled at the RP2D. Patients with advanced melanoma were enrolled into an expansion cohort at the RP2D of 8 mg i.t. tilsotolimod previously established based on another study, ILLUMINATE-204 (NCT02644967), in combination with ipilimumab for metastatic melanoma refractory to a PD-1 inhibitor. The study was approved by institutional review boards at each participating center. The study was conducted according to the Declaration of Helsinki and Good Clinical Practices. Written informed consent was obtained prior to enrollment.

The primary objective of the dose escalation cohort was safety. Secondary objectives were to establish an RP2D, assess clinical activity of i.t. tilsotolimod monotherapy, and characterize its pharmacokinetics.

The primary objective of the melanoma dose expansion cohort was investigator-assessed ORR using RECIST v1.1. Secondary objectives were safety, other measures of clinical benefit, and pharmacokinetics.

Exploratory objectives were similar between the two cohorts and included characterizing biomarkers for immunologic assessment and assessing antidrug antibodies.

Eligible patients were adults with a histologically or cytologically confirmed diagnosis of metastatic solid tumors malignancies that are refractory to available therapies. Patients were required to have at least one lesion accessible for i.t. injection, an Eastern Cooperative Oncology Group performance status (ECOG PS) of ≤2, and adequate hematologic, renal, and hepatic function. Patients with a diagnosis for which a PD-(L)1/PD-1 inhibitor has been approved must have previously received treatment with one of these therapies. Key exclusion criteria included prior therapy with a TLR agonist, treatment with IFNα within the previous 6 months, active autoimmune disease requiring disease-modifying therapy, concurrent systemic steroid therapy (>10 mg/day of prednisone or equivalent), and active central nervous system metastases.

Tilsotolimod was administered as a series of i.t. injections into a single tumor selected for injection. The injected (primary) tumor for each patient was selected from pathologic draining lymph nodes, deep (visceral) metastases, and superficial or subcutaneous metastases. Deep injected tumors required real-time image-guided (US, CT) delivery utilizing interventional radiology techniques. In the event a full dose could no longer be practically administered into the injected tumor, another tumor could be selected for injection. In the case of complete remission, remaining injections were to be administered into the tumor bed, except in the case of visceral lesions where remaining injections were to be administered subcutaneously at a location based on the investigator's preference and discretion. Tilsotolimod was thoroughly distributed within the injected tumor while avoiding necrotic areas, using a fanning method to distribute the administered volume throughout the injected lesion. The total injected dose was to remain constant for each patient; however, injection volume could be adjusted depending on the size of tumor.

Clinical and laboratory safety assessments were conducted at baseline, weekly during cycle 1, then on day 1 of subsequent cycles. Adverse events were graded using the Common Terminology Criteria for Adverse Events (CTCAE) version 4.03. Imaging and efficacy assessments occurred every 6 weeks (dose-escalation cohort) or 9 weeks (melanoma expansion cohort) during the treatment period and then at least every 12 weeks during follow-up and included clinical examination and CT or magnetic resonance imaging of known sites of disease. Tumor response was assessed using RECIST v1.1 and immune-related Response Evaluation Criteria in Solid Tumors (irRECIST). Blood samples for plasma preparation and bioanalysis of tilsotolimod concentrations were collected prior to each injection during cycles 1–3, during the follow-up safety visit, and post-dose at intervals (30 minutes, 1, 2, 3, 4, and 24–48 hours) on day 1 of cycles 1 and 2.

Core biopsies of the injected (primary) tumor and another distant tumor (if available) were taken at screening, 24 hours after the first dose of i.t. tilsotolimod on day 1 of cycle 1, and approximately 6 weeks after the first dose (primary biopsy only). Core biopsies were optional in the melanoma expansion cohort.

Immune analyses included NanoString (NanoString Technologies) and/or flow cytometry of type I interferon (IFN) pathway activation, IFNγ levels, activation of dendritic cell subsets, and changes in T-cell status.

RNA was extracted from core needle biopsies preserved in RNA later using the Qiagen AllPrep Universal Kit (cat. #80224) according to the manufacturer's instructions. Purity and concentration were assessed using NanoDrop. Gene set scores were generated from the Pan-Cancer Immune Panel (NanoString Technologies) and analyzed using the nSolver Advanced Analysis Software.

The first part of the study was dose evaluation. Patients were treated in four dose escalation cohorts (8, 16, 23, and 32 mg). DLTs and treatment-emergent adverse event (TEAE) were graded based on CTCAE v4.03. For the dose evaluation cohort, results were summarized descriptively. Summary statistics for continuous variables included number of observations, mean, standard deviation (SD), median, minimum, and maximum. Categorical variables were summarized using frequency counts and percentages. Descriptive statistics were provided for all PK parameter values by dose and time. Data from all participating sites were pooled prior to analysis. Baseline was defined as the most recent, nonmissing value prior to the date and time of the first dose of the study drug. Study population analysis and safety analyses were based on the safety set. Summaries of AEs included only TEAEs, defined as AEs with a start date on or after the date of the first dose of study drug or existing AEs, which increased in CTCAE grade after the start of study treatment. All other safety analyses included only post-baseline measures. All efficacy analyses were based on the efficacy evaluable set. The best overall response was based on RECIST v1.1 criteria and response assessments of complete response (CR) and partial response (PR) required confirmation by imaging ≥4 weeks after the initial documentation of CR or PR. The primary analysis of PK was conducted on the PK population. A nonlinear power model was used to assess the dose proportionality of tilsotolimod on day 1 of cycle 1 for the four dose evaluation cohorts. The melanoma dose expansion portion used a Simon's Optimal Two-Stage design with a type I error rate of 0.05 and 80% statistical power to test the hypothesis that the response rate per RECIST v1.1 was ≥ 30%, which is clinically meaningful in this setting.

The authors confirm that the data supporting the findings of this study are available within the article and/or its supplementary materials.

The first patient was enrolled on May 15, 2017, and the last patient completed treatment on July 31, 2019. Fifty-four patients with refractory advanced solid tumors were enrolled, including 38 in the dose-evaluation portion and 16 patients with refractory melanoma in the expansion cohort. Deep, visceral lesions were injected with the aid of image guidance in 91% of patients (n = 49). All patients successfully received injected tilsotolimod doses to specified lesions.

Overall, patients in the dose evaluation portion of the study had a mean age of 56.3 years (range, 20–77 years). The majority of the patients were females (25, 65.8%). In the dose-evaluation cohort, the most common cancer types were pancreatic (n = 12) and colorectal cancer (n = 7). Most (n = 35, 92.1%) patients had stage IV disease, 23 (60.5%) had received prior targeted therapy, and 8 (21.1%) had received prior anti–PD-1 therapy (Table 1). Eight, 10, and 9 patients received the 16, 23, or 32 mg i.t. tilsotolimod dose, respectively, whereas 11 patients received the RP2D of 8 mg (Supplementary Table S1).

Patients in the melanoma expansion cohort study had a mean age of 61.7 years (range, 18–86 years) and nine of the 16 patients (56.3%) were male. All patients had stage IV disease and had received prior checkpoint inhibitor therapy: 13 (81.3%) anti–PD-1, 10 (62.5%) anti–CTLA-4, and 5 (31.25%) as a combination. Most patients (n = 10, 62.5%) had elevated LDH, and 4 (25%) had BRAF mutation-positive melanoma.

No treatment-related DLTs were observed. All 54 patients had at least one TEAE (Table 2). Treatment-related adverse events (TRAE) were mostly ≤ grade 2 with 8 (14.8%) patients having grade 3. The most common TRAEs were pyrexia, fatigue, chills, nausea, and vomiting. Fatigue was the only frequent TRAE to increase with the dose. Only one patient required dose reduction due to AEs, and there were no discontinuations due to AEs. There were no AEs related to the technical aspects of it delivery of the agent nor during the acquisition of core biopsy material. There were no AEs resulting in death in both the dose evaluation and dose-expansion cohorts. There were no DLTs observed in the four dose-escalation cohorts and responses were seen across the different doses levels and hence the recommended phase II (RP2D) was determined to be 8 mg.

Thirty-five patients were evaluable for clinical response assessment with a median follow-up of 2.6 months. Twelve (34.3%) patients achieved a best overall response of stable disease (SD; Table 3). Tumor reduction was observed in both the injected lesion and distant, noninjected lesion; five patients demonstrated a reduction in the longest diameter of either the injected or noninjected lesions (Fig. 1A). Noninjected lesions were all visceral lesions and included hepatic, pulmonary, or other intrabdominal masses. The median duration of SD was 5.1 months (minimum 1.5, maximum 12.6), with one patient ongoing for more than 1 year (Fig. 1B and C). SD was observed in at least two patients at each dose level and in several tumor types, including three of seven patients with soft-tissue sarcoma and four of seven patients with colorectal cancer. There was no correlation between a best overall response of SD and baseline characteristics or any treatment-related AE. Three patients (two colorectal cancer and one sarcoma) achieved SD for more than 10 months after progressing on all lines of standard-of-care therapy (Fig. 1B). The two colorectal cancer patients were in the 23 mg cohort and had stable disease for >24 weeks and both developed PD at the end of cycles 15 and 17. The sarcoma patient had uterine leiomyosarcoma and was in the 32 mg cohort and had SD for > 24 weeks and during active follow-up visits continued to have SD.

All 16 patients were evaluable for clinical response assessment with a median follow-up of 6.4 months. Similar to the dose evaluation cohort, tumor reduction in either the injected or noninjected lesions occurred in five patients (31.3%; Fig. 1A). In this monotherapy expansion cohort in heavily pretreated population with advanced melanoma, SD was the best overall response (n = 3, 18.8%; Fig. 1A). Patients had a median progression-free survival of 2.2 months (95% CI, 1.9–4.0) and median overall survival (OS) of 8.5 months (95% CI, 4.5–not evaluable: NE).

Due to the intratumoral route of tilsotolimod administration, variability in drug plasma concentration is expected and influenced by multiple factors. Tilsotolimod was quickly absorbed from the injection site into the systemic circulation with a T_max of approximately 0.5 hours and a half-life of < 2 hours. There was no quantifiable tilsotolimod 1 week after dosing. Drug exposure was dose proportional for AUC_inf and C_max. The PK profiles were similar between the cohorts.

Similar to prior preclinical data of i.t. tilsotolimod, a rapid, robust activation of the type I IFN pathway was consistently observed in biopsies obtained 24 hours. after tilsotolimod i.t. injection compared with baseline levels (Fig. 2). Specifically, several IFN-signaling genes were upregulated including genes that encode transcription factors (STAT1, STAT2, IRF7), antiviral proteins (Cig5, OAS3, MX1), cell death/survival factors (MG1P3, IFI27), and signaling regulators (IFIT4, Ly6E).

Immune profiling of gene families revealed that IFNγ, IFN downstream, and inflammatory chemokines were the most heavily upregulated gene sets in 24-hour biopsies and then returned to near baseline levels 6 weeks after dosing (Fig. 3A). Analysis of paired biopsies before and 24 hours after tilsotolimod administration also found that the IFNα gene-expression signature was substantially upregulated in many tumor types (Fig. 3B). Importantly, all 24-hourbiopsies had a highly significant increase in IFNα gene-expression signature (P < 10E−5).

To examine the potential functional effects of elevated IFN signaling, the maturation of type 1 myeloid dendritic cells (mDC1) was assessed by flow cytometry in fresh biopsy samples. Upregulated HLA-DR expression indicative of mDC1 activation was observed in one out of three available samples [number of CD1c-positive cells (mDC1) increased by 4.4-fold], whereas mDC2 maturation was observed in two of three samples [numbers of CD141-positive cells (mDC2) increased by 2.3-fold and 3.3-fold; Fig. 3C].

The capacity for robust antigen presentation was also elevated in 24-hours post-tilsotolimod biopsies. TAP1 and TAP2, transporters responsible for MHC peptide loading, were upregulated by 2.4-fold and 2.2-fold, respectively, along with IFN-induced protein with tetratricopeptide repeats 2 (IFIT2) by 6.3-fold (Fig. 2).

Genes for checkpoints and costimulatory proteins were also modulated 24 hours after tilsotolimod injection. PD-1, PD-L2, TIM3, and LAG3 were all significantly increased, though CTLA-4 expression was not significantly changed. Additionally, the expression of costimulatory proteins CD80 and CD86 was increased (Supplementary Table S2). Overall, tilsotolimod had a substantial impact on the innate immune activation profile of solid tumors.

We have demonstrated feasibility, safety, and tolerability as well as an immunomodulatory effect of a synthetic CpG oligonucleotide injected i.t. in non–MSI-H refractory solid tumors (12). Treatment with tilsotolimod revealed potential preliminary evidence of clinical activity across multiple solid tumors including those traditionally unresponsive to immunotherapy. Of the 45 patients who were evaluable for response, 33.3% had stable disease including some who had rapidly progressed on prior treatment, such as two colorectal cancer patients and one sarcoma patient who achieved SD for more than 10 months.

This was a dose-escalation trial; however, no DLTs were observed and no patient discontinued treatment due to TRAEs. The most common TRAEs (pyrexia, fatigue, nausea, and chills) were similar to side effects previously reported with other CpG agonists (13, 14). As no DLTs were observed and efficacy was seen across different doses, the recommended phase II (RP2D) was determined to be 8 mg. This trial also demonstrated a unique finding of a decrease in size of not only injected but also noninjected measurable lesions, suggesting an abscopal effect. This was seen previously in a clinical trial investigating the combination of TLR9 agonist plus radiation in low-grade non-Hodgkin lymphoma; however, we report this finding as monotherapy in solid tumors which suggests CpG agonist as the cause and not only radiation (14). Immune monitoring analysis showed robust activation of the type I interferon pathway in injected lesions demonstrated by increased IRF7, IFIT1, and IFIT2 gene expression and early increases in type I interferon signaling. Subsequent analysis also demonstrated an increase in dendritic cell activation, upregulation of MHC class II, and upregulation of interferon-alpha signaling, suggesting improved antigen presentation. This was observed across multiple tumor types, and changes were consistent with those observed in a previous phase I/II clinical trial of patients with metastatic melanoma (9). Moreover, immune profiling gene expression from baseline revealed an approximately 2-fold increase in the expression of immune-checkpoint gene mRNA, including LAG-3, PD-1, and PD-L2 (12). These findings provided, in part, the rationale for exploring the potential for tilsotolimod to complement CPIs in advanced solid tumors, which is being investigated in patients with colorectal cancer in combination with nivolumab and ipilimumab (ILLUMINATE-206) and a phase III trial (ILLUMINATE-301) studying the combination of tilsotolimod plus ipilimumab versus ipilimumab in patients with advanced melanoma refractory to PD-1. However, it is important to note that the ILLUMINTAE-301 failed to meet its primary endpoint of ORR. We look forward to reviewing the final publication to evaluate the details of efficacy analysis including OS and correlative biomarkers. It is important to note that the ILLUMINTAE-301 enrolled patients with malignant melanoma only and not patients with advanced solid tumors as is the case in our current trial.

In our trial, most of the patients were heavily pretreated and had aggressive malignancies with guarded prognosis such as pancreatic adenocarcinoma, soft-tissue sarcoma, and osteosarcoma. These data with single-agent intratumoral tilsotolimod administration are promising and support further investigation of use in combination with other immunotherapeutic agents such as CPIs. Although it is important to note the negative phase III trial (ILLUMINTAE-301) in patients with malignant melanoma and the median OS of 8.5 months in the melanoma expansion cohort in our current trial indicate no role as single agent and questionable efficacy when combined with CPI in this disease. In patients with colorectal cancer, we look forward to reviewing the results from the ILLUMINATE-206 trial.

In conclusion, tilsotolimod shows clinical activation of the innate and adaptive immune response via rapidly increasing dendritic cell activation, upregulation of MHC class II, and IFN-alpha (antigen presentation), and induces a 2-fold increase in the expression of immune checkpoint genes including inducing an “abscopal effect” in refractory non–MSI-H solid tumors. These findings provide a rationale for ongoing trials in solid tumors combined with CPI; however, this will need to be revisited after evaluating upcoming data from the ILLUMINATE-301 phase III trial.

H. Babiker reports grants from Idera during the conduct of the study as well as other support from Myovant, Idera, Bayer, CARIS, Guardant360, Celgene, SirTex, and Coherus Biosciences and grants from Novocure outside the submitted work. E. Borazanci reports grants from Idera during the conduct of the study as well as consulting work for Vivacitas, Nanology, TD2, and BioNTech. V. Subbiah reports grants from Idera Pharma, Eli Lilly/Loxo Oncology, Blueprint Medicines Corporation, Turning Point Therapeutics, Boston Pharmaceutical, and Helsinn Pharmaceuticals and an advisory board/consultant position with Eli Lilly/Loxo Oncology during the conduct of the study as well as grants from Roche/Genentech, Bayer, GlaxoSmithKline, Nanocarrier, Vegenics, Celgene, Northwest Biotherapeutics, Berghealth, Incyte, Fujifilm, D3, Pfizer, Multivir, Amgen, AbbVie, Alfa-sigma, Agensys, Boston Biomedical, Idera Pharma, Inhibrx, Exelixis, Blueprint Medicines, Altum, Dragonfly Therapeutics, Takeda, National Comprehensive Cancer Network, NCI-CTEP, University of Texas MD Anderson Cancer Center, Turning Point Therapeutics, Boston Pharmaceuticals, Novartis, Pharmamar, and Medimmune; advisory board/consultant positions with Helsinn, Incyte, QED Pharma, Daiichi-Sankyo, Signant Health, Novartis, Relay Therapeutics, Roche, and Medimmune; travel funds from Pharmamar, Incyte, ASCO, and ESMO; and other support from Medscape outside the submitted work. A. Algazi reports other support from Idera during the conduct of the study as well as personal fees and nonfinancial support from Oncosec Medical and Sensei, personal fees from Actigal, and other support from Valitor and multiple biotech companies outside the submitted work. M. Lotem reports grants from Dr. Miriam and Sheldon G. Adelson Medical Research Foundation, other support from Merck and BMS, and personal fees from Oncohost and Novartis outside the submitted work. C. Maurice-Dror reports other support from Idera Pharmaceuticals during the conduct of the study as well as personal fees from Biomica, MSD, BMS, Medison, and Pfizer outside the submitted work. S. Rahimian reports other support from Idera Pharmaceuticals outside the submitted work. H. Minderman reports other support from Idera Pharmaceuticals outside the submitted work. C. Haymaker reports grants from Idera Pharmaceuticals during the conduct of the study as well as grants from Iovance, Dragonfly, BTG plc, and Sanofi; other support from Briacell; and personal fees from Nanobiotix outside the submitted work; in addition, C. Haymaker has a patent for TLR9 modulators for treating cancer pending. C. Bernatchez reports grants from Idera Pharmaceuticals during the conduct of the study; in addition, C. Bernatchez has a patent for biomarkers associated with clinical response to tilsotolimod + ipilimumab in PD-1 refractory in metastatic melanoma patients pending. S. Chunduru reports other support from Idera during the conduct of the study as well as other support from Idera Pharmaceuticals outside the submitted work; in addition, S. Chunduru has a patent for TLR9 modulators for treating cancer pending to Idera Pharmaceuticals. A. Diab reports grants and personal fees from Idera during the conduct of the study. I. Puzanov reports personal fees from Iovance, Amgen, Nektar, Merck, and Oncorus outside the submitted work. No disclosures were reported by the other authors.

H. Babiker: Conceptualization, data curation, formal analysis, investigation, methodology, writing–original draft, writing–review and editing. E. Borazanci: Investigation, methodology, writing–review and editing. V. Subbiah: Formal analysis, investigation, methodology, writing–review and editing. S. Agarwala: Investigation, methodology, writing–review and editing. A. Algazi: Formal analysis, investigation, methodology, writing–review and editing. J. Schachter: Investigation, methodology, writing–review and editing. M. Lotem: Investigation, methodology, writing–review and editing. C. Maurice-Dror: Investigation, methodology, writing–review and editing. D. Hendler: Investigation, methodology, writing–review and editing. S. Rahimian: Conceptualization, resources, data curation, formal analysis, supervision, funding acquisition, validation, investigation, methodology, project administration, writing–review and editing. H. Minderman: Formal analysis, investigation, methodology, writing–review and editing. C. Haymaker: Formal analysis, investigation, methodology, writing–review and editing. D. Mahadevan: Investigation, methodology, writing–review and editing. C. Bernatchez: Formal analysis, investigation, writing–review and editing. R. Murthy: Formal analysis, investigation, methodology, writing–original draft. R. Hultsch: Investigation, methodology, writing–review and editing. N. Kaplan: Investigation, visualization, methodology, writing–review and editing. G. Woodhead: Investigation, visualization, writing–review and editing. C. Hennemeyer: Investigation, visualization, writing–review and editing. S. Chunduru: Resources, data curation, software, formal analysis, funding acquisition, validation, visualization, methodology, project administration, writing–review and editing. P.M. Anderson: Investigation, visualization, methodology, writing–review and editing. A. Diab: Investigation, methodology, writing–review and editing. I. Puzanov: Supervision, investigation, methodology, writing–review and editing.

H. Babiker worked previously at the University of Arizona Cancer Center and currently is employed by the Mayo Clinic. We thank Andy Johnson (Idera) and James Mancuso (MDACC) for assistance in writing and editing. V. Subbiah is an Andrew Sabin Family Foundation Fellow at The University of Texas MD Anderson Cancer Center. V. Subbiah acknowledges the support of The Jacquelyn A. Brady Fund. V. Subbiah is supported by NIH grant R01CA242845. MD Anderson Cancer Center Department of Investigational Cancer Therapeutics is supported by the Cancer Prevention and Research Institute of Texas (RP1100584), the Sheikh Khalifa Bin Zayed Al Nahyan Institute for Personalized Cancer Therapy (1U01 CA180964), NCATS Grant UL1 TR000371 (Center for Clinical and Translational Sciences), and the MD Anderson Cancer Center Support Grant (P30 CA016672). This work was supported by the Roswell Comprehensive Cancer Center and NCI grants P30CA016056 and 1R50CA211108 (H. Babiker) involving the use of Roswell Park Comprehensive Cancer Center's Flow and Image Cytometry. The clinical trial was supported by Idera Pharmaceuticals.

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