INSIDER - Reflections during Breast Cancer Awareness Month

INSIDER is a new series that aims to highlight exciting developments from the world of cancer research and clinical trials. These posts are written by our research laboratory heads and the opinions expressed in these posts are those of the contributing author.

By Prof Robin Anderson, Head of the Translational Breast Cancer Program and Metastasis Research Laboratory

October is Breast Cancer Awareness month.  As we reflect on the unexpected events of 2020 that have forced us to make drastic, but hopefully temporary changes in lifestyle, it is a good time to reflect on the remarkable progress made over the past 30 years in the treatment of breast cancer, thanks to medical research.

The current five-year survival rate in Australia, based on data from 2012 to 2016, is 91% for women and 85% for men.

These five-year survival rates have increased from the 1987-1991 period when they were 75% for women and 76% for men. The increase in survival has been more dramatic for women, assisted by the increasing public awareness of symptoms, free mammography since 1994 and, importantly, medical research.  Breast cancer is a rare disease in men and is likely to be diagnosed at a more advanced stage.

These dramatic increases in survival rates are the result of many years of research in cancer institutes around the world, gaining new knowledge about the basic biology of breast cancer and seeking more effective therapies.  Breast cancer treatment varies depending on the type of cancer that develops.  The most common type of breast cancer typically responds well to endocrine therapy as the cancer needs the hormone estrogen to keep growing.  If deprived of estrogen, the cancer dies.  Endocrine therapy has been the mainstay of this type of breast cancer for the past three decades and results in close to 100% survival after 5 years. However, a few of these cancers can recur after many years and are more challenging to treat.  Medical researchers have now discovered new cell cycle inhibitors (specifically inhibiting CDKs 4 and 6) that dramatically prolong progression free survival when used in combination with endocrine agents.

Another type of breast cancer is called HER2 amplified breast cancer, so called because it is driven by a gene called Her2.  In the early 1990’s, patients with this type of cancer had a life expectancy of only 3-5 years.  The discovery of an antibody that specifically blocks Her2 function by medical researcher and medical oncologist Dennis Slamon revolutionised the treatment and now nearly all Her2 positive patients survive beyond 5 years.

The third major subtype, the so called triple negative breast cancer, is the most challenging type to treat and is the focus of researchers at the ONJCRI.  These patients are treated with standard chemotherapy to which most respond well, but it is not uncommon for the cancer to return.  There is very active research at ONJCRI and at many other cancer research institutes around the world seeking better therapies for this type of breast cancer.  Many new experimental treatments are emerging, but will need to be tested in clinical trials to see if they really can improve the outcome of triple negative breast cancer patients.


INSIDER – Using CRISPR to inactivate genes in tumour cells

INSIDER is a new series that aims to highlight exciting developments from the world of cancer research and clinical trials. These posts are written by our research laboratory heads and the opinions expressed in these posts are those of the contributing author.

By Prof Matthias Ernst, ONJCRI Director and Head of the Cancer and Inflammation Laboratory

Immunotherapy has without a doubt provided a quantum leap to the treatment of cancer not seen since the introduction of chemotherapy in the 1960’s. Our understanding on how we can re-engage a patient’s immune system to kill cancer cells is continuously being refined, since the ground-breaking work by the 2018 Nobel laureates Tasuku Honjo and James Allison and the many seminal contributions of immunologists, molecular biologists and oncologists before and after them. However, we still need to go a long way before we can deploy the exquisite power and specificity of the immune system to control a majority of cancer.

A recent study published in Nature by a team at the University of Toronto suggests that there is a potentially dazzling number of genes in cancer cells that enable them to evade being killed by the soldiers of the immune system, so called cytotoxic T lymphocytes (CTL). The team around Jason Moffat used a Nobel Prize awarded genome editing technique called CRISPR to inactivate each gene in tumour cells derived from genetically diverse models of breast, colon, kidney, and skin cancer to ask which gene prevents the killing of cancer cells when put together in a culture dish with CTLs.

When looking across all tumour models, the researchers identified nearly 200 genes, whose deletion altered the sensitivity or the resistance of cancer cells to CTL-mediated killing.

The team was able to validate their experimental approach because they identified genes already known to be mutated in patients who stopped responding to immunotherapy, confirming that their experimental approach worked. However, most surprisingly many of the genes they found were never before linked to evasion of cancer cell detection and killing by CTL such as cancer cell autophagy, a conserved pathway activated by distressed cells. Indeed, induction of autophagy in cancer cells made them resistant to cell CTL killing induced by the cytokines IFNγ and TNF.

The findings of this paper powerfully illustrate how the combination of large-scale screening efforts, together with already existing extensive cancer genome information, allows us to identify a plethora of potential new targets to further improve immunotherapy. The challenge will be to filter through the many candidates targets to strategically invest in those that are ultimately providing the broadest benefit to cancer patients.

Source:  NATURE, Vol 586, pages120–126 (2020); KA Lawson et al Functional genomic landscape of cancer-intrinsic evasion of killing by T cells"


Evaluating the efficacy and mechanism of action of tyrosine kinase inhibitors in triple negative breast cancer

Evaluating the efficacy and mechanism of action of tyrosine kinase inhibitors in triple negative breast cancer

2021 Honours opportunity

Supervisors: Dr Normand Pouliot and Dr Delphine Denoyer

Laboratory: Matrix Microenvironment and Metastasis

Subjects prerequisites: MED3LAB or MED3PRJ

 

HER2+ve and triple negative breast cancer (TNBC) are aggressive subtypes of breast cancer that have a high propensity to spread (metastasise) to the brain. Antibody-based therapies for breast cancer effectively control systemic disease and extend life in some patients but are largely ineffective against brain metastases due to the poor permeability of antibodies across the blood-brain barrier and/or acquired resistance that almost inevitably develops. Small molecule tyrosine kinase inhibitors (TKIs) provide a promising alternative to antibody due to their high potency, small size and greater brain permeability. We have evaluated the efficacy of multiple TKIs in pre-clinical mouse models of HER2+ve breast cancer brain metastasis and identified neratinib as a potent inhibitor with a unique mechanism of action called ferroptosis, a process distinct from apoptosis. We are currently testing new combination therapies to overcome resistance to neratinib in vitro and in vivo.

This project seeks to extend these findings to TNBC by comparing the efficacy of various TKIs alone or in combination with cell adhesion receptor inhibitors in vitro. Whether neratinib inhibits TNBC growth through ferroptosis or other cell death pathways will be investigated in standard in vitro assays. The efficacy of neratinib mono and combination therapies will be evaluated in mouse models of TNBC metastasis. The project will make use of a variety of techniques including standard cell culture, proliferation assays, metabolic assays, western blotting, flow cytometry, immunohistochemistry, qPCR and animal models.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au


Limitrin: a new cell adhesion receptor regulating breast cancer metastasis

Limitrin: a new cell adhesion receptor regulating breast cancer metastasis

2021 Honours opportunity

Supervisors: Dr Normand Pouliot and Dr Delphine Denoyer

Laboratory: Matrix Microenvironment and Metastasis

Subjects prerequisites: MED3LAB or MED3PRJ

 

Adhesion of cancer cells to the surrounding matrix and endothelium is critical for successful metastasis. This project seeks to characterise the expression and function of a novel cell adhesion receptor called limitrin/DICAM, in breast cancer metastasis. Limitrin has been shown to mediate cell-cell adhesion in normal epithelial and endothelial cells but its role in cancer remains unknown. We have found that the expression of limitrin is increased in breast cancer cell lines and tumours that spread (metastasise) to the brain. Preliminary results in various breast cancer models indicate that limitrin may promote cancer cell adhesion to, and migration through, the endothelium thereby facilitating colonisation of the brain.

The project will test the hypothesis that suppressing the expression of limitrin will prevent or delay the outgrowth of brain metastases in clinically relevant mouse models of breast cancer brain metastasis. The project will involve knocking out limitrin in mammary carcinoma lines using CRISPR/Cas9 technology and validation of knockout by flow cytometry, RT-PCR, immunofluorescence and immunohistochemistry. The effect of limitrin suppression on cellular functions will be assessed in in vitro proliferation, adhesion, migration, invasion and trans-endothelial migration assays. The relationship between limitrin and metastasis will be addressed in vivo using preclinical models of breast cancer brain metastasis.

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au


Understanding metastatic breast cancer by single cell transcriptomics

Understanding metastatic breast cancer by single cell transcriptomics

2021 Honours opportunity

Supervisor: Dr Bhupinder Pal

Laboratory: Cancer Single Cell Genomics

Subjects prerequisites:MED3LAB, MED3PRJ and/or GEN3LAB

 

Metastatic breast cancer refers to a disease that has spread beyond the breast and regional lymph nodes to distant sites, with variable locations and volumes of organ involvement. Targeting metastatic breast cancer cells is proving difficult due to the molecular heterogeneity that arise from different factors including the cell of origin, somatic mutations in breast cancer susceptibility genes, and genetic alterations including mutations, deletions, fusions or amplifications of key genes. Cancer cells that escape and seed at new sites usually undergo molecular changes in their genetic and epigenetic landscape. Furthermore, the tumour microenvironment can also influence cancer cell survival, treatment outcome and cancer metastasis.

The technological advancements in the field of single cell biology has allowed the study of molecular heterogeneity in mixed cell populations and shed light on cell lineage relationships. Our work has revealed variable heterogeneity in breast cancer cells and identified novel cell clusters in breast tissue.

The primary objective of this project is to gain novel molecular insights into aggressive and drug resistant breast cancers. We will use high-throughput single cell transcriptomic/genomic analysis approach to study clinical samples from breast cancer patients undergoing treatment. Expected outcome: 1) Understand the role of tumour cell heterogeneity in promoting cancer disease. 2) identify specific immune/stromal gene signatures that can predict metastatic resurgence. 3) discover biomarkers associated with metastatic progression. Novel biomarkers will be evaluated utilising breast cancer patient derived xenografts and organoid assays.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au


Studying the function of gamma delta T cells in colon and stomach cancer

Studying the function of gamma delta T cells in colon and stomach cancer

2021 Honours opportunity

Supervisor: Dr Lisa Mielke

Laboratory: Mucosal Immunity and Cancer

Subjects prerequisites:MED3LAB, MED3PRJ and/or GEN3LAB

 

Modulation of the immune system has revolutionised the treatment of some cancer types, such as melanoma, non-small cell lung cancer and renal cancer. Despite its huge success, immunotherapy with drugs blocking immune inhibitory receptors PD-1 and CTLA4, fails in gastric and colon cancer. Immunotherapy in cancer patients also results in severe side effects in a significant proportion of patients due to systemic activation of the immune system, causing autoimmune-like syndromes.

The Mucosal Immunology and Cancer Laboratory focuses on identifying new immune targets that can be explored to develop novel therapeutics to treat stomach and colon-specific cancers, while avoiding side effects. We use mouse models and patient samples to understand the organ-specific functions of intraepithelial lymphocytes (IELs), that consists of heterogeneous populations of T cells that are distinct in their frequency and function across different organs of the gastrointestinal tract (GI). Our preliminary studies have shown that one population, known as gamma delta T cells, plays a protective role in colon cancer. In this project we will use single-cell RNA sequencing, flow cytometry and multiplexed fluorescent immunohistochemistry technologies to understand the function of gamma delta T cells in mouse models of colon and stomach cancer and patient samples.

Techniques employed will include:

  • Tissue harvest from mice
  • Tissue digestion and extraction of immune cells from mouse GI tract
  • Immunohistochemistry of mouse and human samples
  • Microscopy
  • Flow cytometry
  • Mouse handling.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au


Studying the molecular characteristics of metastatic clones

Studying the molecular characteristics of metastatic clones

2021 Honours opportunity

Supervisors: Dr Delphine Merino and Dr Jean Berthelet

Laboratory: Tumour Progression and Heterogeneity

Subjects prerequisites: MED3LAB or MED3PRJ

Breast cancer is a highly heterogeneous disease. Many studies have shown that patient tumours are composed of a large number of cells or group of cells called ‘clones’ that are genetically diverse. This high level of heterogeneity represents a major obstacle for cancer therapy, some of these cells may have a survival advantage to colonise vital organs and resistant standard treatments, leading to disease recurrence.

To colonise distant organs, tumour cells must intravasate into blood vessels as circulating tumour cells, extravasate and survive in their new micro-environment. One of our previous studies showed that only a minor proportion of cells present in the primary tumour will survive and form lethal metastases. Breast cancers are known to preferentially colonise lungs, bones, lymph nodes, liver, brain and ovaries, and how the microenvironments of these different organs impact on clonal selection remains unclear.

Our laboratory aims at understanding the biologic properties of metastatic clones. We would like to understand how they differ between different metastatic sites and resist current treatments. Therefore, we are using patient samples, cell lines and a new technology called ‘cellular barcoding’ to label cancer cells from patient samples and follow their behaviour. Using sequencing at bulk or single cell level we are planning to identify the pathways involved in metastatic spread and drug resistance, to ultimately propose better treatments for patients with breast cancer.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au

 


Role of the EHF transcription factor in breast cancer

Role of the EHF transcription factor in breast cancer

2021 Honours opportunity

Supervisors: Prof John Mariadason and Dr Ian Luk

Laboratory: Oncogenic Transcription

Subjects prerequisites:MED3LAB, MED3PRJ and/or GEN3LAB

EHF is a transcription factor that is highly expressed in the breast epithelium. We recently generated an Ehf knockout mouse and found that female mice are unable to feed their pups.

Examination of the mammary glands of Ehf KO mice revealed a pronounced defect in the development of this tissue during pregnancy. Expression of EHF is also downregulated in human breast cancers, particularly the triple negative subtype, where tumours with low EHF expression have a poorer outcome. Triple negative breast cancers (TNBC’s) comprise ~20% of all breast cancers and have limited treatment options. There is therefore an urgent need to identify the driver genes which give rise of this subtype so that new treatments can be developed.

 

Project outline

The goal of this Honours project is to determine the role of EHF in the growth, survival, migration and chemotherapy response in triple negative breast cancer cells. We will achieve this as follows:

  1. Determine the level of EHF mRNA and protein expression in 20 breast cancer cell lines,including 5 TNBC cell lines.
  2. Determine the effect of EHF re-expression on cell proliferation, survival, migration and response to chemotherapy in TNBC cells.
  3. Determine the effect of EHF knockdown in TNBC cells which express EHF on cell proliferation, survival, migration and response to chemotherapy.
  4. Identify the target genes of EHF in TNBC cells following EHF knockdown or overexpression.

The student undertaking this project will learn the fundamental concepts of cancer biology and use a variety of techniques including working with cell line models of breast cancer, western blotting, immunohistochemistry, transfections and assessing response to drug treatment.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au

 


Understanding the molecular mechanisms by which apoptosis and/or autophagy are regulated

Understanding the molecular mechanisms by which apoptosis and/or autophagy are regulated

2021 Honours opportunity

Supervisors: Dr Erinna Lee and A/Prof Doug Fairlie

Laboratory: Cell Death and Survival

Subjects prerequisites: MED3LAB, MED3PRJ and/or GEN3LAB


Background
Cells possess distinct pathways that promote their survival or death. These pathways are tightly regulated but when this regulation goes awry, diseases such as cancer ensue. Our lab is interested in two main pathways that control cell death and cell survival. These are apoptosis and autophagy respectively.

Apoptosis is a form of cell death required for the removal of damaged or unwanted cells. We are primarily interested in the mechanisms by which the BCL-2 family of proteins regulate this process and how they can be targeted with drugs for disease treatment. Autophagy is one mechanism of cell survival and is an evolutionarily conserved process of cellular self-cannibalism. Damaged or unnecessary cellular components are targeted to the lysosome for their removal once encapsulated in autophagic vesicles. A large focus of our work is on the autophagy inducer Beclin1 and how it contributes to cellular homeostasis and disease.

Project Description
Broadly, the Honours project(s) will be aimed at understanding the molecular mechanisms by which apoptosis and/or autophagy are regulated by our proteins of interest under normal physiological conditions and how this can go awry to give rise to disease. We are also interested in exploring the therapeutic value of molecules that modulate these pathways and developing new ways in which we can target these pathways for therapeutic benefit in cancer treatment.

Upon completion of the Honours year we expect our student to be equipped with fundamental laboratory techniques in cell biology, protein biochemistry and drug screening. Skills that will be essential for a future career as a lab scientist.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au

 


Reshaping the immunosuppressive tumour microenvironment of brain cancer

Reshaping the immunosuppressive tumour microenvironment of brain cancer

2021 Honours opportunity

Supervisors: Prof Matthias Ernst and Dr Ashleigh Poh

Laboratory: Cancer and Inflammation

Subjects prerequisites: MED3LAB, MED3PRJ and/or GEN3LAB

Glioblastoma (GBM) is the most common and fatal type of primary malignant brain cancer. A hallmark of GBM is an immunologically “cold” tumour microenvironment characterised by an abundance of immunosuppressive myeloid cells, which limit anti-tumour immune responses. Thus, readily translatable strategies that can reprogram the immunosuppressive tumour microenvironment and promote the recruitment and activation of cytotoxic immune cells represent a major advancement for the treatment of GBM.

Elevated expression of the myeloid-specific kinase Haematopoietic Cell Kinase (HCK) is observed in most solid cancers and correlates with poor patient survival. We have previously shown that constitutive HCK activation promotes tumour growth and progression by facilitating the polarisation of myeloid cells towards an immunosuppressive endotype (Poh et. A; Cancer Cell 2017, Cancer Immunology Research 2020). Accordingly, genetic ablation or pharmacologic inhibition of HCK impairs colon and gastric tumour growth by reducing the abundance of immunosuppressive myeloid cells.

Given the role of HCK signaling in myeloid-driven immunosuppression, and the contribution of these cells in GBM, we hypothesise that targeting HCK will present new opportunities for the treatment of brain cancer.

The overall aims of this project are to:

  • Dissect the immunological mechanisms by which constitutive HCK activation or loss of HCK signaling influences GBM development and progression
  • Validate the therapeutic efficacy of novel HCK-specific small molecule inhibitors in preclinical models of GBM.

Students will work in state-of-the-art research laboratories of the School of Cancer Medicine /Olivia Newton-John Cancer Research Institute, which is embedded in the ONJ Cancer Centre. Students will gain experience in multidisciplinary techniques in tumour biology, immunology, molecular biology, and pre-clinical mouse models.

 

How do I apply?
Please complete and return your Ranked Entry Project Selection Form (available in the 2021 projects bookletby 5pm Friday 6 November 2020 to I.Poon@latrobe.edu.au

How do I find out more?
Discover more about La Trobe University’s School of Molecular Sciences/LIMS Honours opportunities
Contact the LTU Honours Coordinator, Dr Ivan Poon
Phone: 03 9479 6488
E-mail: I.Poon@latrobe.edu.au