Neoadjuvant Treatment of Locally-advanced Breast Cancer Patients With Ribociclib and Letrozole

Study Overview
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Brief Summary
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Patients with locally advanced (stage III) breast cancer (LABC) are characterized by a significantly worse prognosis compared to patients with primarily operable breast cancer. While neoadjuvant chemotherapy has been the first choice in this situation for several decades, recent evidence suggests that some patients may experience an extraordinary effect from neoadjuvant endocrine treatments involving aromatase inhibitors as monotherapy or in modern drug combinations.Selected LABC patients admitted for treatment will be offered combination therapy including letrozole and ribociclib. The overall goal of the project is to improve understanding of tumor responses and resistance in patients suffering from ER-positive/HER-2 negative locally advanced breast cancer, focusing on the role of the immune system including the gut microbiome.

Detailed Description
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Neoadjuvant endocrine therapy (NET) offers a good treatment option to reduce the size of large (inoperable) primary breast tumors and/or advanced axillary lymph node metastasis prior to definite surgery. For selected patients (postmenopausal women with ER-positive breast cancer), primary endocrine therapy has been shown to be as effective as standard neoadjuvant chemotherapy. It is the general opinion that aromatase inhibitors of the third-generation (letrozole, anastrozole, exemestane) are the preferable drugs for neoadjuvant endocrine therapy in ER-positive, postmenopausal breast cancer patients. Anastrozole, letrozole and exemestane have been shown to suppress total body aromatisation as well as plasma and tissue estrogen levels by > 90% in vivo. All three drugs are currently used as standard care in the neoadjuvant, adjuvant and metastatic setting worldwide, including in Norway. In 2015, a new class of anti-cancer drugs was introduced known as cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors. Following pivotal trials showing dramatic effects when combined with aromatase inhibitors or steroidal antiestrogens like fulvestrant, these compounds are now well established in distinct combinations during therapy for metastatic breast cancer. All in all, the addition of a CDK4/6 inhibitor to standard anti-hormone therapy for breast cancer doubled the time to disease progression and caused significant improvements in overall survival. Based on these findings, CDK4/6 inhibitors have also been tested in the neoadjuvant setting in clinical trials with very promising results. However, the combination of an aromatase inhibitor and a CDK4/6-inhibitor is not currently approved as standard neoadjuvant treatment for patients with locally advanced breast cancer (LABC) in Norway. Thus, the study described here will make this highly promising drug combination available for all patients who participate in this study. At the same time, the protocol will allow investigators to optimize the selection of patients benefitting from this treatment, highlight relevant biomarkers for personalized medicine and treatment, evaluate predictive markers and study the basic biology underlying treatment effects and resistance. In recent years researchers have noted a growing body of evidence concerning the involvement of the immune system in the onset and prognosis of breast cancer both locally (in the tumor or connected tissues) and through the involvement of the immune system as a whole. Interestingly, the gastrointestinal (gut) microbiota seems to play an important role in determining whether the immune system is able to fight against several cancer types. Pre-clinical studies have shown that the microbes in the gut may influence the repertoire and activity of immune cells, such as T cells, potentially priming the immune system for cancer cell recognition and destruction. This hypothesis is supported by the finding that treatment responses to immune checkpoint inhibitors (ICIs) in a variety of cancer types is dependent on the distinct gut microbiome of an individual patient as the efficacy of the ICIs relies on the presence of cancer-recognizing T cells. In addition, recent findings indicate that an emerging group of small molecules / targeting cancer therapies have immune altering properties. The MAPK kinase (MEK) inhibitors exemplify this by contributing to anti-tumor immunity through increasing the levels of cytotoxic T lymphocytes in preclinical model systems. Similar findings have recently been reported for both CDK4/6 inhibitors and PI3K-inhibitors. Thus, CDK4/6 inhibitors are believed to increase the antigen presentation of cancer cells, increase the activity of tumor infiltration by CD45+ cells and effector T cell activation, as well as decreasing Treg cell proliferation. While these mechanisms have been well described in in vitro and animal models, they need to be studied in humans as soon as possible to understand them and allow investigators to take advantage of these novel aspects in clinical decision making. These findings may also pave the way for microbiome and immune-related data to be used as potential biomarkers for patient selection and response evaluations during therapy. In addition to the clinical effects of being the standard care for breast cancer patients suffering from locally advanced breast cancer, neoadjuvant therapy is widely used to study the endocrinology of breast cancer in general and is recognized as one of the best model systems to predict responses in other clinical settings (early breast cancer or metastatic breast cancer). Tumor biopsies obtained before initiation of treatment and following six months of therapy will allow correlations to the individual type of clinical response (partial responses vs complete responses etc.). The research will focus on intratumoral mechanisms of adaption through tumor characterization using single cell technology which will allow investigators to follow both the different cancer clones as well as the evolution of different immune cells during therapy. Investigators will use novel approaches to evaluate liquid biopsies (measurement of cytokines levels, metabolites and cell free DNA-fragments etc.) during letrozole and ribociclib therapy using state of the art laboratory methods available at the host hospital and in the laboratories of the listed collaborators. Thus, all in all, the planned study will be able to contribute to the basic understanding of this very potent new drug combination that may be used in large groups of breast cancer patients in the future, including locally advanced breast cancer, hopefully reducing the use of traditional chemotherapy in this setting.

Official Title
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Presurgical Treatment With Ribociclib and Letrozole in Patients With Locally Advanced Breast Cancer: the NEOLETRIB Study.

Conditions
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Breast Cancer, HER2-negative Breast Cancer, ER-positive Breast Cancer, Locally Advanced Breast Cancer, Luminal A Breast Cancer, Luminal B Breast Cancer

Intervention / Treatment
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* Drug: Letrozole 2.5mg oral tablet; Ribociclib 600mg oral tablet
* Drug: Goserelin


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Females ≥ 18 years old at the time of the ICF signature histologically confirmed locally advanced breast carcinoma, defined as either large T2 (>3cm in diameter) or T3-T4, and/or N2-3 primary breast cancer ER-positive (defined by ER-pos. in ≥ 50% of cancer cells) and HER-2 negative, luminal A/B breast cancer postmenopausal status (natural status or induced by treatment with the LHRH-analogue goserelin 3.6mg implant s.c. given every 4 weeks); definition of natural post-menopausal status: age above 55 years or age above 50 years and at least 2 years of amenorrhea in addition to LH-, FSH-, and plasma estradiol levels in the post-menopausal range. Patient has adequate bone marrow and organ function as defined by the following laboratory values (as assessed by central laboratory eligibility): Absolute neutrophil count ≥ 1.0x109/L; platelets ≥ 100 x 109/L; Hemoglobin ≥ 9.0g/dL; INR≤1.5 (unless the patient is receiving anticoagulants and the INR is within the therapeutic range of intended use for that anticoagulant within 7 days prior to the first dose of study drug); Estimated glomerular filtration rate (eGFR) ≥ 30mL/min/1.73m2 according to the Modification of Diet in Renal Disease (MDRD) formula; total bilirubin < ULN except for patients with Gilbert's syndrome who may only be included if the total bilirubin is ≤ 3.0 x ULN or direct bilirubin ≤ 1.5 x ULN; aspartate transaminase (AST) < 2.5 x ULN; Alanine transaminase (ALT) < 2.5 x ULN; patient must have the following laboratory values within normal limits or corrected to within normal limits with supplements before the first dose of study medication: potassium, magnesium, total calcium (corrected for serum albumin). Standard 12-lead ECG values defined as the mean of the triplicate ECGs [QTcF interval at screening < 450 msec (QT interval using Fridericia's correction), mean resting heart rate 50-90 bpm (determined from the ECG)] Performance status: Eastern Cooperative Oncology Group (ECOG) score 0-1 Ability and willingness to comply with study visits, treatment, testing and to comply with the protocol. Exclusion Criteria: Any prior treatment for primary invasive breast cancer Patient with a known hypersensitivity to any of the excipients of ribociclib or letrozole Patient with known hypersensitivity to peanuts or soya-products Any evidence of distant metastasis Triple-negative breast cancer HER-2 positive disease, suitable for neoadjuvant therapy with trastuzumab, pertuzumab and taxanes, etc. Other conditions rendering patients in need of other treatment options with immediate effect like chemotherapy Concomitant medications that are known strong inducers of CYP3A4/5 Clinically significant, uncontrolled heart disease and/or cardiac repolarization abnormality, including any of the following: History of documented myocardial infarction (MI), angina pectoris, symptomatic pericarditis, or coronary artery bypass graft (CABG) within 6 months prior to study entry; documented cardiomyopathy; Left Ventricular Ejection Fraction (LVEF) < 50% as determined by Multiple Gated acquisition (MUGA) scan or echocardiogram (ECHO); Long QT syndrome or family history of idiopathic sudden death or congenital long QT syndrome, or any of the following: Risk factors for Torsades de Pointe (TdP) including uncorrected hypocalcemia, hypokalemia or hypomagnesemia, history of cardiac failure, or history of clinically significant/symptomatic bradycardia, Concomitant medication(s) with a known risk to prolong the QT interval and/or known to cause Torsades de Pointe that cannot be discontinued or replaced by safe alternative medication (e.g., within 5 half-lives or 7 days prior to starting study drug) or Inability to determine the QTcF interval; Clinically significant cardiac arrhythmias (e.g., ventricular tachycardia), complete left bundle branch block, high-grade AV block (e.g., bifascicular block, Mobitz type II and third degree AV block); Systolic Blood Pressure (SBP) >160 or <90 mmHg Patient is currently receiving or has received systemic corticosteroids ≤ 2 weeks prior to starting study drug, or who have not fully recovered from side effects of such treatment. Note: The following uses of corticosteroids are permitted: a short duration (<5 days) of systemic corticosteroids; any duration of topical applications (e.g. for rash), inhaled sprays (e.g., for obstructive airways diseases), eye drops or local injections (e.g., intraarticular). Pregnant or breast-feeding (lactating) women or women who plan to become pregnant or breast-feed during the trial Women of child-bearing potential defined as all women physiologically capable of becoming pregnant, unless they are using highly effective methods of contraception during the study treatment and for 21 days after stopping the treatment. Highly effective contraception methods include: Total abstinence (when this is in line with the preferred and usual lifestyle of the patient. Periodic abstinence (e.g., calendar, ovulation, symptothermal, post-ovulation methods) and withdrawal are not acceptable methods of contraception; Female sterilization (have had surgical bilateral oophorectomy with or without hysterectomy), total hysterectomy or tubal ligation at least 6 weeks before taking study treatment. In case of oophorectomy alone, only when the reproductive status of the woman has been confirmed by follow up hormone level assessment; Male partner sterilization (at least 6 months prior to screening). For female patients on the study, the vasectomized male partner should be the sole partner for that patient and the success of the vasectomy must be medically confirmed as per local practice; Placement of an intrauterine device (IUD); Note: Use of oral (estrogen and progesterone), transdermal, injected, implanted, hormone containing intrauterine systems (IUS) or any other hormonal methods of contraception is not allowed in this study Autoimmune disorders or significant allergies (i.e. rheumatoid arthritis, asthma, psoriasis, etc.) Known HIV infection, Hepatitis B or C infection (testing not mandatory) History of autoimmune celiac, inflammatory bowel disease, or other chronic GI disease Recent use (within past month) of more than 3 days of antibiotics use Current use of probiotic supplements Taking proton pump inhibitors, steroids, other non-steroidal antiinflammatory drugs such as ibuprofen or acetyl salicylic acid Past bariatric surgery

Ages Eligible for Study
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Minimum Age: 18 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Treatment
Intervention Model: Single Group Assignment
Interventional Model Description: Multicentre, single-arm, open-label, neoadjuvant study model
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: Ribociclib and Letrozole Arm<br>Patients entered into this study will be given letrozole (FemarTM) in combination with ribociclib (KisqaliTM) for at least 6 months. Premenopausal women will also receive treatment with goserelin 3.6 mg s.c. every 4 weeks. | Drug: Letrozole 2.5mg oral tablet; Ribociclib 600mg oral tablet<br>* Patients will be given letrozole 2.5mg and ribociclib 600mg daily, per oral for a period of 21 days followed by 7 days of letrozole only.<br>* Other names: Femara; Kisqali;Drug: Goserelin<br>* Premenopausal women will be given goserelin 3.6 mg (subcutaneous) every 4 weeks in concert with their Letrozole and Ribociclib treatment.<br>* Other names: Zoladex;|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| To study the change in levels of direct and indirect immunologic biomarkers of targeted cancer therapy with letrozole and ribociclib given in combination for patients with locally-advanced, ER-positive, HER-2 negative, luminal A7B breast cancer | In order to accurately profile changes in the composition of the tumor over time and identify potentially-related biomarkers of response within the tumor, single cell RNA panels will be performed at three points during the treatment period. Samples of tumor tissue will be obtained, and the cells dissociated at the single cell level. These dissociated cells will then be subjected to RNA profiling via scRNA-seq. Genes expressed at a rate higher than the threshhold (generally 50 reads per kb per million reads) will be examined using t-Distributed Stochastic Neighbor Embedding, (tSNE) clustering, generally shown as a scatterplot, to allow the characterisation of phenotypes (clones). Using this technique, it will be evident which tumour phenotypes are eradicated by the treatment combination and which are not. This examination will also help to monitor patient response. | Baseline, Day 21 and at time of surgery (Day 180) | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Measurement of changes in the tumor through DNA profiling throughout the treatment cycle with letrozole and ribociclib | In order to examine the potential genetic changes within the tumor during treatment, samples of tumor tissue will be taken and examined using standard DNA profiling. Baseline samples will be compared to those at day 21 and at the time of surgery (6 months). This outcome will allow identification of overall tumour composition and changes to over the course of the therapy. | Baseline, Day 21 and at time of surgery (Day 180) | 
| Confirmation of the breast cancer subtype | To determine the subtypes of the tumor, a PROSIGNA-test will be undertaken on all samples to confirm their subtype. | Baseline | 
| Changes in neoantigens and single T-cell receptor function after treatment with letrozole and ribociclib | In order to examine the effects of letrozole and ribociclib on the presence of immune cells within the tumor throughout treatment, samples of tumor tissue will be extracted and cells will be dissociated at the single-cell level using enzymatic and mechanical means. Measurement of tumor neoantigens and testing of the ability of the T-cell receptors to recognise these neoantigens will assist in the understanding of the rules and molecular events underlying immune-mediated tumor destruction. Further, responders and non-responders will be compared to identify whether they belong to a new yet undiscovered subtype. | Baseline, Day 21 and at time of surgery (Day 180) | 
| Determination of the histopathological sub-type and status of the tumour | To ensure the correct patients are identified for participation in this study, and to record overall baseline information, samples of tumor tissue will be examined via standard histopathological investigations including subtyping, grading, ER-status, PGR-status, HER-2 status and level of Ki67-expression will be undertaken by a qualified pathologist. After the baseline, examination will confirm that biopsies consist of tumor tissue. | Baseline, Day 21 and at time of surgery (Day 180) | 
| Determination of the early and late mechanisms of adaptation and/or resistance to letrozole in combination with ribociclib | Using single-cell RNA-seq (scRNA-seq), thousands of individual cells will be profiled to build a cellular atlas of whole tumor biomarkers for targeted cancer treatments. scRNA-seq allows the characterisation of phenotypes (clones) to assist in determination of the phenotypes eradicated by the treatment, and to find biomarkers of response. Further knowledge of the phenotypes of the cells most affected by the therapy may assist in the identification of further drugs targeting any cells that remain or thrive. | Baseline and at time of surgery (Day 180) | 
| Change in PEPI status from baseline in response to targeted neoadjuvant therapy with letrozole and ribociclib | Preoperative endocrine prognostic index (PEPI) status will be evaluated across the treatment period using the procedure outlined in Ellis et al, 2008. | Baseline, Day 21, Day 90, Day 180 | 
| Change in Ki67 from baseline in response to targeted neoadjuvant therapy with letrozole and ribociclib | Levels of Ki67 will be evaluated across the study period using the XXXXX protocol. Complete Cell Cycle Arrest (CCCA) will be recorded as a Ki67 of less than 2.7%. | Baseline, Day 21, Day 90, Day 180 | 
| Change in PROSIGNA Risk of Recurrence (ROR) score from baseline in response to targeted neoadjuvant therapy with letrozole and ribociclib |  | Baseline, Day 21, Day 90, Day 180 | 
| Changes in the composition of the gut microbiota | A DNA-based approach to determination of fecal microbial composition will be performed. Stool samples will be collected at baseline, and during follow ups at day 21, day 90 and at time of surgery (day 180), and thereafter annually in years 1-5 (further if patient relapses). DNA purification from fecal samples will be performed using PSP Spin Stool DNA Plus Kit (Stratec Molecular GMBH). Next generation amplicon sequencing targeting the V4 region of the 16S rRNA gene (DNA) will be applied to detect the members of the fecal microbiota. | Baseline, 21 and 90, at time of surgery (Day 180), and annually in years 1-5 (extension if patient relapses) | 
| Identification of anti-tumor effects via changes in levels of circulating serum cytokines in liquid biopsies | Serum cytokines will be measured using 54-plex cytokine panel, and analyzed with Luminex xMAP 200. The assay includes a series of known concentrations giving standard curves. Samples will be analyzed in duplicate. | Baseline, 21 and 90, and at time of surgery (Day 180). | 
| Identification of anti-tumor effects via changes in levels of circulating serum metabolites in liquid biopsies | Circulating metabolites will be identified using high resolution magic angle spinning mass spectrometry as described in Bathen et al (2013). | Baseline, 21 and 90, and at time of surgery (Day 180). | 
| Identification of anti-tumor effects via circulating free tumor DNA (cftDNA) in liquid biopsies | Circulating free tumor DNA will be collected and identified using standard DNA extraction and analysis techniques. | Baseline, 21 and 90, and at time of surgery (Day 180). | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Breast Cancer, Letrozole, Ribociclib, Combination Therapy