The 2021 San Antonio Breast Cancer Symposium (SABCS) included multiple scientific research posters focused on invasive lobular carcinoma (ILC), including one sharing the results of a survey on ILC imaging submitted by the Lobular Breast Cancer Alliance Inc. (LBCA). It should be noted that “posters” at research conferences are tools used to present new information, often the synopsis and key findings of a research study, in a graphic. At conferences there are typically many posters presented and a designated room for display. Online they are shared as PDFs.
LBCA is pleased to present lay summaries of many of the excellent ILC-focused research posters presented at SABCS in December 2021. All posters and abstracts from SABCS will be available this March in AACR’s Cancer Research supplement. LBCA thanks members of our Scientific Advisory Board, LBCA Board of Directors, and poster authors for creating these summaries.
Spotlight Session Posters
Each year the conference organizers select several posters to “spotlight.” At SABCS 2021, there was an entire “spotlight session” devoted to posters about ILC research. Posters with numbers beginning with “PD” below were spotlighted.
PD14-01 Comprehensive molecular characterization of patients with metastatic invasive lobular carcinoma (ILC): Using real-world data to describe this unique clinical entity
Population/Study Goal: There is some overlap between lobular and ductal cancers in cell appearance and behavior, making some mixed tumors difficult to characterize on pathology as one or the other. A hallmark of ILC is loss of E-Cadherin (CDH1) expression, a protein that when lost, allows cells to grow in specific ways. Thus, tumors with loss of CDH1 are characterized as ILC.
This study examined biopsy samples from metastatic ILC (mILC) and metastatic mixed lobular/ductal cancer to look for mutations in CDH1, as well as other genes that might identify specific mILC biology as opposed to mixed. The study was a retrospective analysis of 150 mILC samples and 51 “mixed” samples. In each sample the disease was “staged” using data closest to the date of biopsy collection.
Results:
- 65.3% of the mILC samples had CDH1 mutations.
- Median tumor mutational burden (i.e., overall number of mutations in the tumor) was significantly higher in the mILC CDH1-mutant samples.
- CDH1 mutant samples were more frequently ER+ subtype.
- Identified PIK3CA as a new gene that was enriched in CDH1 mutant mILC tumors.
- Mixed pathology patients with CDH1 mutations had lower CDH1 expression than those without mutations.
- Authors conclude that mILC tumors with CDH1 mutations behave differently than those without.
- mILC also differs from mixed pathology type at a gene transcription level, with lower CDH1 expression seen in mILC vs. mixed, regardless of CDH1 mutational status.
What does this mean for patients?
- May lead to better pathologic characterization of mixed samples as mILC based on CDH1 expression levels.
- PIK3CA may be investigated as a target for future therapies in CDH1 mutant mILC cases.
Lay summary by Tracy Cushing, MD
PD14-02 Unravelling spatial tumor organization and heterogeneity in lobular breast cancer using spatial transcriptomics
Recent studies showed that underlying heterogeneity found within the same tumor (intratumor heterogeneity, ITH) has an impact on patient outcome. Here, we aimed to better characterize this heterogeneity in ILC using spatial transcriptomics (ST) together with high-resolution morphological annotation of the relative histological slides. Spatial transcriptomics was performed on frozen tumor samples obtained from 15 patients with estrogen receptor-positive, HER2-negative ILC, of which four developed disease relapses.
For each sample, we annotated hematoxylin/eosin sections with a single cell resolution. Here we aimed at describing ITH as well as at identifying common or specific characteristics across different tumors. We performed clustering analysis in each sample, identifying 45 tumor clusters among the 15 samples, highlighting intratumor heterogeneity.
The different clusters were also characterized based on different biological processes. All samples showed at least a cluster enriched in estrogen signaling, while other tumor clusters were enriched for different pathways. Of note, samples from patients who experienced disease relapse were enriched for protein secretion, demonstrating inter-tumor heterogeneity, and characterized by more heterogeneity as described by a less organized spatial distribution of the clusters.
The findings of our study highlight the presence of shared biological processes across ILCs, as well as the role of heterogeneity and specific pathways potentially associated to the risk of recurrence. Indeed, these tumor clusters must be further characterized to identify therapeutic biomarkers and tumor features which may not be evident from common sequencing analyses. Further validation of our findings is warranted and currently ongoing in a larger cohort.
Lay summary by Christos Sotiriou, MD, PhD
PD14-05 Portraying tumor evolution of lobular breast cancer through phylogenetic analysis
ILC of the breast is the most common special type of breast cancer. Despite therapeutic advancements, a substantial number of patients with treated early-stage ILC will still relapse as late as 20 years after the initial tumor diagnosis, thus making follow-up a challenging task.
The main objective of this study was to characterize the underlying genomic events that best portray the trajectory of cancer cells from the primary tumor to metastasis and to paint the landscape of tumor evolution in ILC. Herein, by sequencing the genome of 38 metastatic ILCs, analyzing their genomic aberrations (abnormalities), and representing tumor evolution as phylogenetic trees, we identified different patterns of tumor dissemination, with both early and late metastatic seeding patterns being possible mechanisms.
Interestingly, we observed that in a small subset of patients, the metastasis harbored less genomic aberrations than the primary tumor, indicating that possibly a decelerated tumor progression is in place.
Lastly, we observed that, in approximately 60% of the patients, genomic aberrations were acquired early during tumor evolution, highlighting the importance of early detection through cancer screening. The findings of this study provided insight of how ILCs evolve and could potentially influence the clinical management of ILC.
Lay summary by Christos Sotiriou, MD, PhD
PD14-06 Does chemotherapy benefit patients with HR+/HER2 – invasive lobular breast cancer?
Study Goal: This study was a retrospective analysis of a large database of patients with ILC conducted to evaluate the benefits of chemotherapy.
Population/Study: The study looked at hormone receptor + (ER+) HER 2 negative (Her2 –) ILC patients in the National Cancer Database. There were 17,987 patients in the database from the years 2010-2016. They were classified into four groups depending on the treatment they had: surgery only, endocrine therapy (ET) only, chemotherapy only (CT), and combined endocrine/chemotherapy (CET). As grouped, 60% received ET, 26% received CET, 2% had CT, and 12% had surgery only. The overall survival was evaluated for each group at 5 years.
Results: The 5-year overall survival was significantly better for patients on ET or CET (90%) vs. CT alone or surgery alone (80%).
- A subgroup analysis accounting for age, stage, histology, radiation therapy, and Oncotype DX score showed similar results: 5-year overall survival was similar for ET and CET groups after adjusting for these variables. Of note, patients with both low (<26) and high (>26) Oncotype DX scores had similar 5-year outcomes whether they had ET or CET.
What does this mean for patients?
- Chemotherapy did not improve overall survival at 5 years in this large sample of ER+ Her2- ILC patients regardless of stage, lymph node involvement, or Oncotype DX score.
- This might decrease the number of ILC patients undergoing systemic chemotherapy.
- Future research might examine which subset of ILC patients may benefit most from chemotherapy: as of now, we do not know.
Lay summary by Tracy Cushing, MD
PD14-07 Bromodomain and Extra-terminal motif (BET) inhibitors are a rational therapeutic choice for the treatment of invasive lobular carcinoma
Study Goal: Big picture: to look for targeted therapies to treat ILC.
Focused goal: To study whether extra-terminal motif (BET) inhibitors can prevent oncogenesis/cell proliferation in cancer by preventing the interaction between bromodomains (BRDs) and the molecules they interact with to drive cell growth. Previous research from this lab showed some ILC lines were BET-inhibitor sensitive, and others were not. Targeting a growth factor (FGFR1) in combination with a BET inhibitor was effective for the ILC models studied here. High expression of BRD-3, one of the BET proteins, is associated with a poor prognosis in ILC.
Population/Study:
- Cells from 126 ILC patients and 1161 IDC patients were studied for their expression of several BRD proteins, and the master regulator proteins (MTPs) that influence them.
- Two ILC cell lines known to be endocrine therapy resistant were evaluated for their sensitivity to 9 different BET inhibitors; the 5 most potent were selected.
- The ILC lines were then exposed to the 5 most potent BET inhibitors to see which genes it most down- and up-regulated in the exposed cells.
- Researchers wanted to know which MTPs would be affected by knocking out BRD-3: they identified JQ1.
- Researchers ultimately tested JQ1 inhibition in vivo in mice in an endocrine-resistant cell line, in combination with fulvestrant, or in combination with tamoxifen, compared with fulvestrant or tamoxifen alone.
Results:
- In cells exposed to BET inhibitors, genes associated with negative regulation of growth and apoptosis (cell death) increased – meaning the process that went awry in cancer growth seems to be counteracted by this inhibition.
- They identified a panel of potential BRD-3 regulated MTPs for other scientists to study as targets of new therapies.
- The JQ1 inhibition in mice resulted in decreased tumor burden in combination with tamoxifen than tamoxifen alone in this ER- cell line.
What does this mean for patients?
This study explored possible genetic targets for therapies in ILC, especially promising for estrogen receptor (ER) negative patients in whom therapy can be limited. BET-inhibitors will be a future direction of therapeutic research for ILC.
Lay summary by Tracy Cushing, MD
PD14-09 APOBEC signature, clinical characteristics, and outcome in hormone receptor-positive (HR+) HER2-negative (HER2-) breast cancer patients in real-world data
Hormone-receptor positive (HR+) breast cancer represents the most common type of breast cancer, encompassing approximately 70% of diagnoses. The growth of HR+ tumors is driven by estrogen, a hormone naturally produced by the body. The use of drugs that block estrogen production or the estrogen receptor have been transformative in the treatment of HR+ disease with significant survival and quality of life benefits. However, a subset of patients will fail to have a long-term benefit from therapy and will have their disease return quickly.
Previous research has identified a subset of tumors that have acquired a high number of mutations across the tumor genome. These mutations can come from a number of sources including tobacco smoke, ultraviolet radiation, and defects in repair processes. In breast cancer, high mutation counts are associated with APOBEC mutational processes. APOBEC is a family of proteins that protect against viral infections in part through mutation of viral nucleotides. Typically, these mutations are specific to viruses, however, in a subset of breast and other cancers, a large number of APOBEC-associated mutations can be found across the DNA of the tumor cells.
Clinical experience from physicians at the Mayo and Duke clinics suggested that patients with high numbers of APOBEC-associated mutations (APOBEC+) did worse on the therapies standardly given to HR+ patients. To confirm their experience, we explored a clinical dataset of HR+ breast cancer patients treated at approximately 800 sites of care in the Flatiron Health Network. Examining this large dataset, we found that tumors with APOBEC-associated mutations (APOBEC+) were common (approximately 8% of patients) and that ILCs were more often APOBEC+. APOBEC+ patients progressed more quickly on standard of care relative to APOBEC- patients. This confirmed the clinical experience at Mayo and Duke. These findings suggest that the population of APOBEC+ patients need additional therapy options.
In 2020, the FDA approved the use of pembrolizumab, a drug that activates a patient’s immune system to attack the tumor, for use in patients with a high number of tumor mutations. Use of this drug was approved in all advanced solid tumors; however, limited information is available describing how well this drug works in HR+ breast cancer patients. We therefore looked at the experience of APOBEC+ patients on these immune-activating drugs. Several patients stayed on therapy for a long time, including one patient with 3+ years on therapy, suggesting benefit from these drugs.
This study suggests that APOBEC+ patients are at high risk of progression on standard HR+ therapies, but there are possible options to treat patient that have progressed, including with immune-activating drugs.
Lay summary by Ethan Sokol, PhD
Additional Posters focusing on ILC
P1-02-09: Results of a worldwide survey on the currently used histopathological diagnostic criteria for invasive lobular breast cancer (ILC)
ILC is a specific breast cancer type because of its typical growth and appearance. In contrast to most other breast cancers, ILC cells have inactivated a protein called E-cadherin. E-cadherin can be regarded as the ‘glue’ that keeps cells together. Loss of E-cadherin results in ILC, a tumor type in which breast cancer cells typically do not adhere to each other and are able to survive on their own.
These features yield the specific growth patterns of ILC when studied under a microscope (histology). The presence of E-cadherin (expression) can simultaneously be judged using antibodies in a technique called immunohistochemistry (IHC). Together, examining histology and IHC for E-cadherin expression can be used to optimally diagnose ILC.
The current study presents data on a world-wide survey from December 2020 until July 2021 to chart how lobular breast cancer is diagnosed, and which histological ILC types are registered. Also, it probes if the diagnosis for ILC is based on in E-cadherin expression and which antibody tools are used for this purpose. Input from 147 participants from 34 countries were collated and analyzed. The largest contributors were Europe (63%), Asia (18%) and the Americas (15%).
Results
Approximately half of all participants use E-cadherin IHC to support ILC diagnosis, mostly and only in case of doubt. The use of E-cadherin IHC to aid diagnosis of lobular carcinoma in situ is rare (3%).
Interestingly, there is quite a lot of diversity in the type of E-cadherin antibodies. The main antibody clone within this survey is NCH-18 (38%), followed by Clone 36 (15%) and EP700Y (14%). Several (8) other clones are used by a minority of different participants. An inventory of the diagnosed subtypes was made whereby the most prevalent were classic, pleomorphic, solid, alveolar and histiocytic.
Conclusions and future directions
E-cadherin IHC is used in approximately half of the cases, whereby the type of antibodies and procedures followed still differ substantially. This non-uniform way of diagnosing lobular breast cancer possibly results in different diagnostic outcomes. The E-cadherin antibodies used should be better scrutinized for specificity and reproducibility, and a uniform Global protocol for ILC diagnosis should be developed and implemented. This is essential to design, perform, and improve intervention trials tailored specifically for ILC.
Lay summary by Patrick W.B. Derksen, PhD
P1-05-03 Integrating spatial transcriptomics and high-resolution morphological annotation to investigate tumor heterogeneity and PAM50 molecular subtyping in lobular breast cancer
The goal of this research is to investigate the underlying heterogeneity of ILC, both within the tumor of the same patient (intra-tumor, ITH) as well as between tumors from different patients (inter-tumor), in terms of microenvironment composition, molecular subtypes (PAM50 classification) and proliferation.
By combining the cutting-edge technology of spatial transcriptomics (ST), which allows us to explore gene-expression profiles at a resolution of 1-10 cells retaining geographic information, and the accurate morphological annotation of the ST slides, primary tumor samples from 15 patients with estrogen receptor-positive, HER2-negative ILC were analyzed. The histological slides were annotated at the cell level into different histomorphological categories (e.g., tumor cell, stroma cell, fat tissue, etc.). Our results highlighted a higher proportion of fat tissue together with higher co-localization with tumor cells in samples from the four patients who experienced disease relapse). Higher spatial variability of proliferation was also associated with poor prognosis. Moreover, the proportion of stroma and fat tissue affected the PAM50 molecular classification of the tumors. The findings of this study highlight the molecular heterogeneity of ILC at an unprecedented level and help to understand the role of the microenvironment in disease relapse.
Lay summary by Christos Sotiriou, MD, PhD
P1-08-13 Comparison of gene expression profiling results and clinical outcomes among patients with pleomorphic ILC and classic ILC
The goal of this project was to determine how Oncotype Dx testing is used for patients with ILC, with special attention for how use of the test and results differ between patients with pleomorphic ILC (pILC) and classical ILC (cILC). The records for 691 patients with ILC treated at Cleveland Clinic were reviewed. Out of these records, approximately 15% of patients had pILC. The patients with pILC were more likely to be HER2 positive (12% for pILC compared to 7% with cILC) and were more likely to have larger breast tumors but less lymph node involvement.
Twenty-five percent of patients with pILC had Oncotype Dx testing sent compared to 33% of patients with cILC. Low risk Oncotype Dx scores were seen in 8% of pILC patients who had testing and 18% of patients with cILC. The rate of intermediate Oncotype Dx recurrence score was similar for both groups (approximately 72%). High risk Oncotype Dx recurrence scores were found for 20% of pILC and 9% of cILC.
More patients with pILC received chemotherapy followed by anti-estrogen treatment compared to cILC (62% for pILC compared to 37% for cILC). There was no statistical difference in the breast cancer recurrence risk or overall survival rates for patients with pILC compared to those with cILC. This challenges the notion that pleomorphic ILC has worse prognosis compared to classic ILC. It is important to note that chemotherapy may be important for pILC to reach these equal long-term outcomes. The higher rate of HER-positivity for pILC is also likely an important factor in these results. There are plans to re-analyze the results separating out the HER2-positive and HER2-negative patients to see the observed results are different.
Lay summary by Megan Kruse, MD
P1-22-07 Surgical outcomes of lobular carcinoma in situ diagnosed on core biopsy in Dutch women between 2011 and 2020
Study Goal: This study was a retrospective cohort study looking at the upgrade from lobular carcinoma in situ (LCIS) to ILC in patients with classic, pleomorphic, and mixed types of lobular cancer.
Population/Study: Retrospective cohort study of Dutch women from 2011-2020. Researchers wanted to evaluate which pathologic subtypes were upgraded to invasive, in particular whether pleomorphic ILC subtypes were more aggressive than classic.
Results: There were 1,185 biopsy samples evaluated: 1,024 classic LCIS, 130 pleomorphic LCIS, 31 non-classic/mixed LCIS. Thirty-four percent of classic LCIS was upgraded to ILC, 31.5% for pleomorphic, and 38.8% for non-classic/mixed – these were not statistically significant findings.
What does this mean for patients? The similarity in advancement from LCIS to ILC appears to be the same across subtypes, which can be reassuring for patients with pleomorphic or mixed subtypes. It appears all LCIS upgrades to ILC in around 32% of patients.
Lay summary by Tracy Cushing, MD
P2-02-02 Differences in levels of circulating tumor cells (CTC) and disseminated tumor cells (DTC) in early-stage lobular versus ductal breast cancer
Previous studies have shown that in the metastatic setting, patients with ILC have more circulating tumor cells (CTC’s) than those with IDC. As blood-based markers (so-called “liquid biopsies”) become a more commonly used part of breast cancer care, it is important to understand whether the interpretation of these assays changes in different histologic types of breast cancer.
We therefore evaluated data from a study of patients with stage I-III breast cancer to determine whether or not patients with ILC also had more CTCs in this non-metastatic setting. Indeed, we found that regardless of stage, patients with ILC did have higher levels of CTC’s.
Whether this translates into different outcomes or different interpretations of study results by histologic subtype is not yet clear. But these findings support the need for subtype specific analyses in liquid biopsy.
Lay summary by Rita Mukhtar, MD
P3-01-25 Assessing the impact of preoperative breast MRI on time to treatment for invasive lobular carcinoma
The goal of this project was to determine if doing MRI before surgery for patients with invasive ILC would delay the time to surgery in a meaningful way. Prior studies have shown that delays in time to treatment for breast cancer (either time to pre-surgical chemotherapy or surgery itself) can cause worse outcomes for patients where the goal of treatment is cancer cure.
While breast MRI can have an important impact on surgery planning for patients with lobular breast cancer, these benefits may not be important if time to cancer treatment is delayed because of the time it takes to obtain the MRI and follow-up on any new concerns detected on the MRI.
This study involved looking back at records from patients with ILC treated at Cleveland Clinic from 2004-2017. The rates of MRI use were documented as well as time from diagnosis to surgery. Patients were sorted into four groups as follows: those who did not have MRI, those who had MRI and no new concerns found, those who had MRI with new concern found but not biopsied, and those who had MRI with new concern found and new concern was biopsied.
There were 691 patients included in this study and 65% had pre-surgery breast MRI. Most common time from diagnosis to surgery was 40 days for the entire study group. Patients who did not have breast MRI had most common time to surgery of 34 days while those who had MRI had most common time to surgery of 41 days.
For those who had MRI and a new area of concern was found, time to treatment was 42 days for those who had an additional biopsy while time to treatment was 36 days for those who did not have an additional biopsy. Average follow-up of these patients was 6 years. At 5 years of follow-up, survival and cancer recurrence rates were no different for patients who had breast MRI and those who did not.
These results suggest that having a pre-surgery breast MRI does not delay treatment in a negative way. Longer-term follow-up will be important for this study as we know that recurrences for patients with ILC can occur late so 10-year follow-up will be important.
Lay summary by Megan Kruse, MD
P5-06-11 Secreted frizzled related protein 1: From lobular involution to breast osteoimmunological disorder
Population/Study Goal: This study is based on previous research that identified secreted frizzled related protein 1 (SFRP1) as a protein that is downregulated in carcinogenesis (less SFRP1 = greater cell growth). SFRP1 is also expressed highly in the microcalcifications of non-tumor breast tissue and in bone resorption. Authors hypothesize that loss of SFRP1 function is responsible for microcalcification accumulation and early breast carcinogenesis.
Results:
- Down-regulation of SFRP1 in a non-tumoral atypical ductal hyperplasia cell line induced an increase in cell’s ability to proliferate (multiply) and migrate (move).
- Down-regulation of SFRP1 in this line also induced increase in estrogen receptor expression, suggesting a role for SFRP1 in carcinogenesis.
What does this mean for patients?
The results indicate a role for SFRP1 in the acquisition of a malignant phenotype (cell behavior becoming malignant) – when it has lower/decreased expression, cells can more easily multiply and migrate. What causes SFRP1 to be down-regulated in vivo/real patients remains unknown.
Lay summary by Tracy Cushing, MD
P5-12-03 WCRC-25: A novel luminal ER negative invasive lobular carcinoma cell line model
About 1 in 8 women in the United States will be diagnosed with breast cancer over their lifetime. Breast cancer can be categorized by the molecular and histologic presentation of the tumor where the major histologic subtypes observed in patients are invasive ductal carcinoma (IDC; approximately 80%) and invasive lobular carcinoma (ILC; ~10-15%). ILC tumors are typically seen as growing in a single file discohesive manner with their hallmark loss of E-cadherin. The lower prevalence of ILC results in this subtype being understudied in the field but in the last few years more attention has been given to this type of breast cancer to cater to precision medicine. Although the incidence is lower, it should be noted that ILC still affects a large number of women each year.
Clinically, patients with ILC are treated in a similar manner as those with IDC and much is still unknown regarding the unique biology and therapeutic vulnerabilities of ILC. Evidently, numerous unique features of ILC are continuously being reported, addressing the need to treat ILC as a unique disease.
To that end, more models that faithfully recapitulate the disease are needed for research to better study and characterize ILC. Here we report the successful establishment and characterization of a novel ILC cell line from a pleural aspirate (fluid extracted from the lung) of a patient with metastatic breast cancer, which represents some of the many characteristics seen in other ILC cell lines and in patients. This cell line replicated therapeutic vulnerabilities that were predicted from RNA sequencing and can offer a great value towards better understanding of ILC.
The comprehensive analysis reported here is only a start of future studies that can be done when coupled with other models and innovative ideas to advance our understanding of ILC and its therapeutic novelties.
