The breast tumour microenvironment: A target for therapy

BCAC committee member, Louise Malone, attended the San Antonio Breast Cancer Symposium in December 2017 and gives us an update on the latest cutting-edge research into new targeted immunotherapy treatments.

As cancer researchers better understand the complex interchanges between tumour cells and immune cells and the microenvironment in which they operate, new targets for therapy are emerging. 

Tumour microenvironments are complex. As well as cancer cells, they include ‘stroma’ composed of endothelial cells, immune cells, fibroblasts and adipocytes, all held in a framework of structural and regulatory proteins (the extracellular matrix). There are many different kinds of immune cells in a tumour, some working to help the body fight the cancer and some subverted by the cancer to help the tumour grow. (See the diagram at the end of this article for an overview of different types of immune cells.)

Immune cells found in tumours include TILs or tumour-infiltrating lymphocytes, which are comprised of a variety of T cells. These can be cytotoxic (T cells which kill other cells, e.g. CD8+), or helper T cells, which are needed to kick other immune cells into action (e.g. CD4+ helper), or regulatory T cells (Tregs), which put the brakes on the immune response and stop it from attacking healthy cells (as in autoimmune disorders). However, cancer Tregs can block anti-tumour responses and promote cancer growth.

TAMs or tumour-associated macrophages also behave differently in the tumour microenvironment from how normal macrophages would. Instead of stimulating inflammation and a normal immune response, TAMs produce high levels of cytokines, growth factors and proteases, which stimulate angiogenesis (blood supply to the tumour) and enhance tumour cell migration and invasion. Cancer-associated fibroblasts (CAFs) have been shown to play many roles in assisting a cancer to grow and spread. Cancer-associated adipocytes (CAAs) also seem to help cancer cells but are not yet well understood.

Tumours can also secrete growth factors and cytokines that direct the bone marrow to produce a set of special immune cells known as myeloid derived suppressor cells (MDSCs) which then infiltrate the tumour. MDSCs assist the cancer by inhibiting cytotoxic T cells and natural killer cells, accelerating angiogenesis, tumour cell invasion and metastasis.

Cells in the tumour communicate via biochemicals (soluble mediators) and receptor-ligand interactions, whereby molecules on the surfaces of cells lock onto each other in a kind of handshake between cells. ‘Immune checkpoints’ are receptor proteins on the surface of T cells, which allow them to tell if other cells are friends or foes, and therefore whether or not to attack. PD-1 (programmed cell death protein 1) is a well-known immune checkpoint found on the surface of T cells. If a T cell encounters another cell with the interlocking PD-L1 protein then it will not attack. Unfortunately, in many cancers PD-L1 is produced not only by normal cells but also tumour cells, allowing the cancer to evade T cell attack. The good news is that immune checkpoint inhibitors (monoclonal antibodies designed to block the PD-1/PD-L1 interaction) can overcome this and boost the immune response to cancer cells.

Therapies targeting the PD-1/PD-L1 interaction have seen great success in cancers such as melanoma and non-small cell lung cancer. However, clinical trials with breast cancer patients and checkpoint inhibitors of PD-1 (pembrolizumab) and PD-L1 (avelumab and atezolizumab) have shown only modest success in shrinking tumours and mixed results with regard to the expected relationship between the amounts of PD-1 expressed by the cancer and the effectiveness of the inhibitors. To date, there is no FDA-approved immunotherapy drug for breast cancer.

Why is this, and what can be done about it? These questions were addressed at SABCS 2017 in two sessions where the speakers looked for answers “beyond the PD-1 axis”.

Dr Robert Vonderheide, from the University of Pennsylvania, noted that in human clinical studies, only 5 to 18% of subjects with triple negative breast cancer (TNBC) that were treated with PD-1 or PD-L1 monoclonal antibodies responded with tumour shrinkage or disappearance, and that response to CTLA-4 (another checkpoint inhibitor) in breast cancer was rare, especially in estrogen- progesterone-positive breast cancer. Dr Vonderheide explained that the genomic and transcriptomic profiles of breast cancers, and their RNA sequence signatures, differ from other types of cancer, suggesting that ‘cutting and pasting’ immune therapies that work in one cancer to another will not always be successful. He posed the question: can we convert ‘cold’ breast cancer tumours that do not respond to immunotherapy into ‘hot’ tumours that do?

This might be accomplished by ‘vaccinating’ the tumour to induce it to produce more T cells. In a clinical trial with metastatic breast cancer patients, CAR-T cells (chimeric antigen receptors on T cells) were taken from tumours, engineered in the lab to produce an antibody to a cell surface protein (c-Met) and injected back into the tumour. Examination of the tumours after subsequent removal showed greater necrosis of tumour tissue and furthermore, engineered CAR-T cells could be found in peripheral blood suggesting that the procedure had made the tumours ‘hot’ for immune response. A new trial is now enrolling patients to test the effects of intravenous c-Met CAR-T cells.

Radiation could also act as a ‘vaccine’ on tumours as it produces inflammation which can stimulate the immune response and thus response to immune therapy. Dr Vonderheide described a case where a woman with progressive and refractory metastatic estrogen-positive breast cancer was treated with radiotherapy targeted specifically to one lung tumour and then given immune checkpoint inhibitors against CTLA-4 and PD-L1. The targeted tumour had shrunk at eight weeks, as had other tumours in her lungs and liver, suggesting that even a localised treatment of one tumour had generated a systemic boost in her immune response such that the therapies could produce positive results in other tumours.

Other research aimed at improving the efficacy of PD-1 immune checkpoint inhibition in breast cancer was reported by Dr Dmitri Gabrilovich, from the Wistar Institute in Philadelphia, and Dr Cynthia Zahnow from Johns Hopkins University, Baltimore.

Dr Gabrilovich has found that combining PD-1 inhibition with inhibition of CSFR1 (colony stimulating factor receptor 1) and inhibition of the chemokine receptor CXCR2 significantly reduces tumour size in mouse models.

Dr Zahnow has been looking at opportunities for using epigenetic therapies in cancer. Epigenetics describes processes which are not driven directly by our genes but which regulate the actions of our genes in profound and lasting ways. Examples include DNA methylation, histone modifications and the actions of transcription factors. In cancer cells, abnormal methylation occurs and proteins such as histone deacetylases (HDAC) modify the functioning of DNA. This suggests that DNA methyltransferase inhibitors (e.g. AZA) and HDAC inhibitors (e.g. givinostat or entinostat) could be useful in combination as cancer therapies and might assist the action of PD-1 inhibitors. Dr Zahnow showed that mice with ovarian cancer given AZA, entinostat and anti-PD-1 had better survival than those given only AZA and anti-PD-1 or those given AZA, givinostat and anti-PD-1. She has also seen some responses to entinostat and AZA in mice with breast cancer, suggesting that this line of research could yield useful new combinations of therapies.

Researchers Dr Jeff Rathmell (Vanderbilt University) and Dr David DeNardo (Washington University School of Medicine, St Louis) are looking for completely new approaches to immune targeting in breast cancer by learning more about the basic biology of tumours.

By examining the metabolism of T cells during an immune response and how this may be altered in the tumour microenvironment, Dr Rathmell is hoping to gain insights that could lead to completely new types of therapy.  He has found that the tumour microenvironment is very heterogeneous and that nutrients such as glucose and lactate can be low in some parts and high in others. Different types of T cells have different metabolic requirements and T cells in tumours have multiple defects, including impairments in glucose uptake and mitochondria. These defects contribute to the inadequate functioning of T cells inside tumours. Experiments with cell culture models have shown that some of these defects could be overcome by providing particular biochemicals to allow the immune cells’ metabolic pathways to run normally again. This research is still at a very basic stage, but could lead to new therapies in the future.

Dr DeNardo is also looking deeply into the cellular biology of the immune response, but his focus is on dendritic cells. These are produced in the bone marrow and their function is to pick up antigens from foreign or cancer cells anywhere in the body and present them to T cells in the lymph nodes. This activates these cytotoxic T cells to travel to the tumour and kill the cancer cells. Dr DeNardo explained that, as part of their counterattack, tumours produce factors which travel to the bone marrow and direct over-production of cells which suppress the immune response and allow tumour growth. They do this at the expense of the production of the dendritic cells. Bone marrow samples from patients with localised solid 2cm breast tumours have abnormally low numbers of dendritic cells and their pre-cursors. Furthermore, cancer patients with low numbers of dendritic cells in the bone marrow do not respond as well to chemotherapy as those with higher numbers.

GCSF (granulocyte colony-stimulating factor) may be one of the factors at play here. Dr DeNardo has shown that GCSF levels are high in the blood of mice (and humans) with breast cancer, that GCSF can drive down differentiation of bone marrow myeloid progenitors into dendritic cells in cell culture. The development of dendritic cells also relies on a transcription factor called IRF8 and GCSF inhibits IRF8; if GCSF is neutralised then dendritic cell deficiency is reversed. Pancreatic cancer patients with high levels of IRF8 have better survival than those with low IRF8.

Using all this information, Dr DeNardo’s team are looking for ways to overcome dendritic cell deficiency in the hope that this will make tumours more responsive to immunotherapy. He presented some very new results showing that animals with tumours given a three-way combination of a PD-1 inhibitor, an anti-GCSF compound, and another compound (Flt3L) which controls dendritic cell development, had better survival than those given only Flt3L or Flt3L and anti-GCSF. 

Finally, Dr Robert Vonderheide raised the tantalising prospect of using immunotherapy for prevention of cancer in healthy people, just as childhood vaccines protect against infections. This would require the discovery of a ‘universal tumour antigen’ and Dr Vonderheide suggested that the telomerase reverse transcriptase, hTERT, could be a candidate. hTERT is expressed in nearly all human cancer, but expression is restricted in normal cells, and it has a critical functional role in oncogenesis (so a therapy targeting it would not be easily overcome by the cancer). In clinical trials using hTERT peptides in patients with metastatic breast cancer, there was a robust immune response without toxicity and this response correlated with overall survival. This is a promising early start, but much more research is needed before this approach could be translated into a treatment.

In conclusion these presentations showed that, in spite of some initial disappointment that breast cancer does not seem to respond as readily as some other cancers to current immunotherapies, there are many exciting avenues being explored that could lead to real breakthroughs in breast cancer treatment.

Diagram 1: Immune cells produced from stem cells in bone marrow.

Presentations cited:

DeNardo, D. G. Do tumours overcome immune surveillance by blocking dendritic cell development? SABCS ES6-2

Gabrilovich, D. I. Myeloid-derived suppressor cells: many facets of regulation of tumour progression. SABCS BS1-2

Rathmell, J. C. How tumour metabolism contributes to the immunosuppressive microenvironment. SABCS ES6-1

Vonderheide, R. H. Opportunities for immune therapy and prevention. SABCS ES6-3

Zahnow, C. A. Epigenetic therapy activates type 1 interferon signalling to reduce immunosuppression and tumour burden. SABCS BS1-1

14 May 2018

 

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