Immune Profile Selection by FctDNA On Patients With Advance Non-Small Cell Lung Cancer Treated With Immunotherapy (IO)

Martin E. Gutierrez, M.D.

John Theurer Cancer Center

Executive Summary

Cancer, when advanced is a devastating disease that can reduce everyday hope and happiness and shorten life expectancy. Any of us having watched a loved-one suffer through cancer realizes that it is not just the disease but the side effects of traditional therapies like chemotherapy and radiation therapy that contribute to patients feeling so poorly during their care. Many patients rightfully ask is this therapy worth it knowing it may not cure them and the extension of life associated with administration of the traditional therapy may not be worth the suffering.

Immunotherapy has changed the paradigm of traditional cancer care and not only offers hope for better outcomes including more cures but is significantly less toxic than traditional therapy, so the “gain” is worth the “pain”. Our scientists and doctors are working towards a future that results in higher cure rates while avoiding traditional chemotherapy and possible radiation therapy. The program outlined below when completed can lead to a change in clinical care around the globe and offer patients greater hope, better outcomes and a significantly better quality of life in the process.

The program is intended to identify patients who will not need chemotherapy and need only immune therapy and compare this group to standard care.


Non- Small Cell Lung Cancer: In 2008, worldwide there were approximately 2.1 million patients diagnosed with non-small lung cancer with an estimated 1.7 million deaths [1]. In the United States, there are over 230,000 new cases of lung cancer and 130,000 deaths annually [2]. However, targeted therapies and immunotherapy have improved non-small cell lung cancer patients’ survival [3].

Non-small cell lung cancer (NSCLC) accounts for the majority of lung cancers (approximately 85 percent) with the remainder mostly small cell lung cancer (SCLC). Most patients present for diagnostic evaluation because of symptoms suspicious for lung cancer or an incidental finding on chest imaging.

Tumor immunology: An efficient and specific cytotoxic immune response against a tumor requires a complex, rapidly evolving interaction between various immune cell types in the adaptive and innate immune system. That includes CD8+ lymphocytes and Th1/Th2 subclasses of CD4+ T lymphocytes, traditionally referred to as cytotoxic T cells and helper T cells. T- Cell CD8+ and CD4+ lymphocytes initiate the distinction between self and non-self-antigens, through recognition at the "immune synapse." In addition, Natural Killer (NK) cells, as part of the innate immune system, do not require antigen presentation by the major histocompatibility complex (MHC) for cytotoxic activity. In fact, NK cells target cells with low MHC class 1 expression for destruction. Like T cells, NK cells express numerous inhibitory molecules as well, most notably various killer immunoglobulin-like receptor (KIR) subtypes [2]. Additional cell types, such as FoxP3+ CD25+ CD4+ T regulatory (Treg) and myeloid derived suppressor cells (MDSCs) largely inhibit cytotoxic T lymphocyte activity [3, 4]. Th17 cells, subsets of CD4+ T cells that secrete interleukin (IL)-17, are implicated in autoimmunity and cancer [5]. Macrophages differentiate into at least two different phenotypes: M1 macrophages, which release interferon (IFN) gamma and are responsible for phagocytosis, and M2 macrophages, which release cytokines such as IL-4, IL-10, transforming growth factor beta (TGF-beta), and dampen inflammatory responses and foster tolerance [4].

The most important co-stimulatory signal in naïve T cells is CD28, which binds to B7-1 and B7-2 (CD80/86) on the APC. This co-stimulatory process is tightly regulated by both "agonist" molecules (eg, GITR, OX40, ICOS) and inhibitory signals on the APC and T cells, often collectively referred to as "immune checkpoint" molecules. Examples of co-inhibitory or "immune checkpoint" molecules include cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), programmed cell death-1 (PD-1), TIM3, and LAG3. Chronic recognition of an antigen may lead to feedback inhibition of effector T cell function, resulting in a phenotype termed "exhaustion" [5].

Tumor evasion of immune surveillance: Cancer immunoediting proceeds in three phases [6]: The elimination phase consists of innate and adaptive immune responses to specific tumor-associated antigens. The equilibrium phase is a balance between immune-mediated destruction by the adaptive immune system and persistence of rare malignant clones and Immunologic escape describes the phase where malignant clones have acquired the ability to evade the adaptive immune system.

Some of the proposed mechanisms for escape from immune surveillance [7] include:
Loss or alteration of specific antigens or antigenic machinery [8, 9]; tumor induced immune-tolerant microenvironment by manipulation of cytokines (increased secretion of IL-6, IL-10, and TGF-beta; consumption of IL-2) that encourage infiltration of Treg cells, myeloid derived suppressor cells (MDSCs), and other cell types that inhibit cytotoxic T cell function [10, 11,12]; and lastly tumors can upregulate the expression of immune checkpoint molecules such as programmed cell death ligand 1 (PD-L1) that promote peripheral T cell exhaustion [13].


The treatment of patients with non-small cell lung cancer treatment will be determined based on the absence / presence of a gene driving mutation and the absence / presence of a high level of programmed cell death ligand 1 (PD-L1) expression. We are interested in the largest populations of NSCLC, approximately 70%, that have a positive expression of PDL-1. We would like to identify the population of patients who will require chemotherapy in addition to a checkpoint inhibitor in this group of patients vs. the subset of patients who may require a chemotherapy free - immunotherapy base only treatment. We recognize that tumor PD-L1 expression as a biomarker has limitations. PD-L1 and PD-1 expression are dynamic markers that change in relation to local cytokines and other factors, and the thresholds that separate "positive" and "negative" PD-L1 expression remain under debate [14-17]. PDL-1 expression associates with increased likelihood of response to checkpoint inhibitors / it neither guarantees response in those with high tumor PDL1 expression nor eliminates the possibility of response in those tumors that lack PDL1 expression. This limitation makes the identification of patients who will benefit on immunotherapy only treatment rather difficult.

We proposed a randomized clinical trial in which the primary endpoint will determine the PFS of subjects treated with IO therapy only vs the subjects that may need the addition of chemotherapy to checkpoint inhibition. We propose to use fctDNA as a biomarker of response to initial IO treatment; then subjects will be randomized to IO therapy or IO / Chemotherapy treatment pending fctDNA changes after two cycles of IO therapy only. Secondary and exploratory endpoints will focus on define the immune profile of subjects in both groups.

Study design: Eligible subjects will be required to have positive PDL-1 expression. Subjects will be stratified by the level of PDL-1 expression > or < less than 50% expression.

Subjects will have a pre-treatment evaluation of fraction of tumor circular in DNA (fctDNA), tumor methylation status, and the immune profile of the tumor by multiplex immunohistochemistry, t-cell mRNA and microbiome.

Subjects will start treatment with IPI/Nivo if PDL-1 expression 1%-49% or Pembrolizumab if PDL-1 expression >50%. At six weeks of treatment, in the absence of clear progression by irRECIST, subjects will have an evaluation fctDNA. If fctDNA reveals < 50% reduction, subjects will be randomized 1:1 to either continue the immunotherapy alone or to escalate treatment and add chemotherapy for a total of four cycles. fctDNA will be evaluated at six weeks of treatment and at the time of progression. CT staging w/u will be done every 6 weeks until disease progression.

disease progression chart

Primary Endpoint

  • Progression Free Survival

Secondary Endpoint

  • Overall survival (OS) and duration of response (DOR)
  • Response rate


  • Tumor methylation status
  • Tumor immune profile multiplex immunohistochemistry
  • Tumor DNA/RNA NGS
  • T-cell mRNA
  • Microbiome


We expect to need 104 randomized subjects to establish a sadistic difference between subjects treated with immunotherapy alone vs immunotherapy and chemotherapy.

At JTCC we evaluated approximately 140 stage IV subjects. We predict that 50% of the subjects will be eligible to be randomized, therefore we estimate that it will take 2 years to enroll and completed the study.


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