Safety and Tolerability of SYNB1891 Injection Alone or in Combination With Atezolizumab in Adult Participants

This Phase 1, open-label, multicenter, 2-arm study was designed to evaluate SYNB1891 when administered either as monotherapy (Arm 1) or in combination with atezolizumab (Arm 2) in participants with advanced/metastatic solid tumors or lymphoma. The primary objective was to evaluate the safety and tolerability of study treatment, with a secondary objective of assessing preliminary tumor response to treatment and exploratory objectives of evaluating the pharmacokinetics/pharmacodynamics (PK/PD) of study treatment.

Arm 1 comprised intratumoral (IT) injections of SYNB1891 monotherapy to determine the single-agent maximum tolerated dose (MTD). The starting dose of SYNB1891 in the first cohort was 1 × 10\^6 live cells and was increased in approximately 3-fold increments in subsequent cohorts until MTD determination in accordance with the modified toxicity probability interval (mTPI) algorithm (Ji et al 2013). Dose escalation was to proceed until the target dose-limiting toxicity (DLT) range (approximately 30%) for SYNB1891 monotherapy was determined based on DLTs observed in Cycle 1. The dose selected as achieving the target DLT range was to be considered the MTD. DLTs were defined in the protocol as certain treatment-related Grade 3/4 laboratory values, sepsis, toxicity resulting in death, discontinuation of Cycle 1, or delay of Cycle 2.

Arm 2 comprised IT injections of SYNB1891 combined with standard dose atezolizumab (1200 mg intravenously \[IV\] every 3 weeks \[Q3W\]). Arm 2 dosing began after all participants in Arm 1 Cohort 4 completed their Cycle 1 DLT safety evaluation. The starting dose of IT SYNB1891 in the first cohort of Arm 2 was at the SYNB1891 Arm 1 Cohort 3 dose level. SYNB1891 dosing in the Arm 2 cohorts increased in approximately 3-fold increments in subsequent cohorts until recommended Phase 2 dose (RP2D) determination. SYNB1891 combination dosing in Arm 2 was always at least 1 dose level below the SYNB1891 monotherapy dose being evaluated in Arm 1, with combination doses not escalated above the SYNB1891 single-agent MTD established in Arm 1.

In both arms, after an initial 4 cycles (21 days per cycle) of study treatment, participants who did not have progressive disease may have received additional cycles of their assigned study treatment for up to 24 months (i.e., Cycles 5 to 35) after the initial dose.

The maximum time of study participation for a participant may have been up to 26 months, including the screening period (up to 28 days), treatment administration period (up to 24 months), and Safety Follow-up period (30 ± 5 days after the last dose).

Eligibility Criteria:
Inclusion Criteria:

1. Able and willing to voluntarily complete the informed consent process (participant or participant's representative).
2. Adults aged ≥ 18 years (on the day of signing informed consent) with histologically- or cytologically-confirmed stage III or IV advanced/metastatic solid tumor or lymphoma for which no therapeutic options were available to extend survival or for which the participant was not a candidate for standard-of-care therapy.
3. Eastern Cooperative Oncology Group performance status ≤ 1.
4. Life expectancy ≥ 3 months.
5. ≥ 1 injectable, measurable (≥ 10 mm in diameter, or ≥ 15 mm for nodal lesions), eligible lesion as defined by Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 (Eisenhauer et al 2009), immune-related RECIST (iRECIST) (Seymour et al 2017), and/or Lymphoma Response to Immunomodulatory Therapy Criteria (LYRIC) (Cheson et al 2016) and as assessed by the Investigator. Eligible lesions must not have been located in the thoracic cavity, spleen, pancreas, gastrointestinal tract (liver injection allowed), or cranium and must have been amenable to percutaneous injection and away from major blood vessels or neurological structures.
6. Able to provide biopsies for biomarker analysis from injected and (if available) noninjected lesions at baseline and other time points during the study.
7. Oxygen saturation \> 90% without the use of supplemental oxygen.
8. Adequate cardiac function, defined as follows:

   1. Left ventricular ejection fraction (LVEF) \> 50% by multi-gated acquisition (MUGA) scan or echocardiogram (ECHO) performed within 6 months prior to the first dose of study treatment provided the participant had not received any potential cardiotoxic agents in the intervening period. Symptoms relating to left ventricular dysfunction, cardiac arrhythmia, or cardiac ischemia must all have been Grade \< 1 per National Cancer Institute (NCI) Common Terminology Criteria for Adverse Events (CTCAE) version 5.0.
   2. QTc interval corrected for heart rate using Fridericia's formula (QTcF) \< 480 msec at screening.
9. Laboratory values within the following ranges:

   * Absolute neutrophil count ≥ 1500/µL
   * Lymphocyte count ≥ 500/µL
   * Platelets ≥ 100,000/µL without transfusion
   * Hemoglobin ≥ 9.0 g/dL (participants may have been transfused to meet this criterion)
   * Estimated glomerular filtration rate (eGFR) \> 50 mL/min/1.73 m\^2 per the Cockcroft-Gault formula
   * Total bilirubin ≤ 1.5 × upper limit of normal (ULN) OR direct bilirubin ≤ ULN for participants with total bilirubin levels \> 1.5 × ULN or ≤ 3 × ULN for participants with Gilbert's syndrome
   * Aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase ≤ 2.5 × ULN (AST and ALT ≤ 5 × ULN for participants with liver metastases and alkaline phosphatase ≤ 5 × ULN for participants with liver or bone metastases)
   * International normalized ratio (INR) or prothrombin time (PT) or activated partial thromboplastin time (aPTT) ≤ 1.5 × ULN unless participant was receiving anticoagulant therapy as long as PT or aPTT was within therapeutic range of intended use of anticoagulants
10. Agreed to use an acceptable method of contraception after informed consent, throughout the study, and for 5 months after the last dose of SYNB1891 or atezolizumab.

    Additional inclusion criteria that applied only to Arm 2 study participants were as follows:
11. Thyroid-stimulating hormone (TSH) within the normal reference range or the participant was receiving stable thyroid replacement therapy.
12. Participants must have received treatment with an anti-programmed cell death protein-1 (PD-1)/programmed death-ligand 1 (PD-L1) monoclonal antibody (mAb) administered either as monotherapy, or in combination with other checkpoint inhibitors (CPIs) or other therapies, if indicated (if CPI therapy was not indicated as a part of standard-of-care therapy, participants may have been CPI naïve).

Exclusion Criteria

1. Chemotherapy, radiation, or biological cancer therapy within 14 days prior to the first dose of study treatment, or failure of any adverse events (AEs) to recover to Baseline or NCI CTCAE version 5.0 Grade 1, except for any grade of alopecia caused by cancer therapeutics administered \> 28 days earlier.
2. Systemic immunostimulatory agents (including, but not limited to, interferon and interleukin-2) within 28 days or 5 drug elimination half-lives (whichever was longer) prior to initiation of study treatment. CPIs were not considered systemic immunostimulatory agents for this criterion and were subject to the washout period listed in exclusion criterion #1.
3. Allogeneic hematopoietic stem cell transplantation that required current use of immunosuppressors.
4. Receipt of a live vaccine within 90 days prior to the first dose of study treatment or anticipation of a need for such a vaccine during treatment.
5. Receipt of antibiotics within 7 days prior to the first dose of study treatment.
6. Participation in a study of an investigational agent and receipt of study therapy or use of an investigational device within 28 days prior to the first dose of study treatment.
7. Diagnosed immunodeficiency or current use of chronic systemic steroid therapy in excess of replacement doses (prednisone ≤ 10 mg/day or equivalent was acceptable) or any other form of immunosuppressive medication within 7 days prior to the first dose of study treatment.
8. For injection and biopsy of visceral (deep) lesions: participants must not have been on long-acting antiplatelet agents such as aspirin or clopidogrel or on therapeutic doses of anticoagulants, with the exception of participants receiving a preventative dose of low molecular weight heparin. In participants receiving preventative low molecular weight heparin, treatment must have been stopped 24 hours before the intratumoral injection and resumed again 24 hours after the injection (Marabelle et al 2018).
9. Previous or concurrent malignancy, with the exception of:

   1. Adequately treated basal or squamous cell carcinoma, in situ carcinoma of the cervix, or ductal carcinoma in situ of the breast;
   2. Localized prostate cancer definitively treated with surgery or radiation or stable on hormone therapy;
   3. Other cancer from which the participant had been disease free for at least 2 years.
10. Clinically active central nervous system metastases and/or carcinomatous meningitis (treated brain metastases were permitted if radiologically stable for ≥ 28 days prior to the first dose of study treatment).
11. Allergy to antibiotics that precluded treatment for infection with Escherichia coli Nissle 1917.
12. Hepatitis B or C infection(s) unless screening tests indicated a negative viral load, and/or human immunodeficiency virus infection unless screening tests indicated a negative or below the limit of quantitation viral load.
13. Known active tuberculosis.
14. Grade 3 or higher infection according to NCI CTCAE within 28 days prior to initiation of study treatment, including, but not limited to, hospitalization for complications of infection, bacteremia, or severe pneumonia.
15. Failure to fully recover from the effects of major surgery. Surgeries that required general anesthesia must have been completed \> 14 days before the first dose of study drug. Surgery requiring regional/epidural anesthesia must have been completed \> 72 hours before the first dose of study treatment and participants should have recovered.
16. Heart failure of New York Heart Association Class 3 or greater, restrictive cardiomyopathy, or unstable angina or recent myocardial infarction within 3 months prior to the first dose of study treatment.
17. Any other medical condition that might have confounded the results of the study, interfered with the participant's participation, or was not in the best interest of the participant, in the opinion of the treating Investigator.
18. Pregnant or breastfeeding or anticipated conception or fathering of children within the projected duration of the study and for 5 months after the last dose of SYNB1891 or atezolizumab.

    Additional exclusion criteria that applied only to Arm 2 study participants were as follows:
19. History of immune-mediated AEs on prior PD-1/PD-L1 therapies requiring discontinuation or immunosuppressor therapy, excluding endocrinopathies.
20. Active or history of underlying autoimmune disease or immune deficiency, including, but not limited to, myasthenia gravis, myositis, autoimmune hepatitis, systemic lupus erythematosus, rheumatoid arthritis, inflammatory bowel disease, antiphospholipid antibody syndrome, Wegener granulomatosis, Sjögren syndrome, Guillain-Barré syndrome, or multiple sclerosis, with the following exceptions:

    1. Participants with a history of autoimmune-related hypothyroidism who were on thyroid replacement hormone were eligible for the study.
    2. Participants with controlled Type 1 diabetes mellitus who were on an insulin regimen were eligible for the study.
    3. Participants with eczema, psoriasis, lichen simplex chronicus, or vitiligo with dermatologic manifestations only (e.g., participants with psoriatic arthritis are excluded) were eligible for the study provided all of following conditions were met:

       * Rash covered \< 10% of body surface area;
       * Disease was well controlled at baseline and required only low potency topical corticosteroids;
       * No occurrence of acute exacerbations of the underlying condition requiring psoralen plus ultraviolet A radiation, methotrexate, retinoids, biologic agents, oral calcineurin inhibitors, or high potency or oral corticosteroids within the previous 12 months.
21. History of a Grade ≥ 3 allergic anaphylactic reaction following treatment with a PD-1 mAb or to chimeric, human, or humanized antibodies or fusion proteins.
22. History of idiopathic pulmonary fibrosis, organizing pneumonia (e.g., bronchiolitis obliterans), drug-induced pneumonitis requiring treatment, or idiopathic pneumonitis, or evidence of active pneumonitis.
23. Known hypersensitivity to Chinese hamster ovary cell products or any component of the atezolizumab formulation.