Cytoreductive surgery with or without hyperthermic intraperitoneal chemotherapy in patients with advanced ovarian cancer (OVHIPEC-1): final survival analysis of a randomised, controlled, phase 3 trial.

BACKGROUND: The OVHIPEC-1 trial previously showed that the addition of hyperthermic intraperitoneal chemotherapy (HIPEC) to interval cytoreductive surgery resulted in improved progression-free and overall survival compared with cytoreductive surgery alone at 4·7 years of follow-up in patients with stage III epithelial ovarian cancer who were ineligible for primary cytoreduction. We report the final survival outcomes after 10 years of follow-up. METHODS: In this open-label, randomised, controlled, phase 3 trial, patients with primary epithelial stage III ovarian cancer were recruited at eight HIPEC centres in the Netherlands and Belgium. Patients were eligible if they were aged 18-76 years, had not progressed during at least three cycles of neoadjuvant carboplatin plus paclitaxel, had a WHO performance status score of 0-2, normal blood counts, and adequate renal function. Patients were randomly assigned (1:1) to undergo interval cytoreductive surgery without HIPEC (surgery group) or with HIPEC (100 mg/m2 cisplatin; surgery-plus-HIPEC group). Randomisation was done centrally by minimisation with a masked web-based allocation procedure at the time of surgery when residual disease smaller than 10 mm diameter was anticipated, and was stratified by institution, previous suboptimal cytoreductive surgery, and number of abdominal regions involved. The primary endpoint was progression-free survival and a secondary endpoint was overall survival, analysed in the intention-to-treat population (ie, all randomly assigned patients). This study is registered with ClinicalTrials.gov, NCT00426257, and is closed. FINDINGS: Between April 1, 2007, and April 30, 2016, 245 patients were enrolled and followed up for a median of 10·1 years (95% CI 8·4-12·9) in the surgery group (n=123) and 10·4 years (95% CI 9·5-13·3) in the surgery-plus-HIPEC group (n=122). Recurrence, progression, or death occurred in 114 (93%) patients in the surgery group (median progression-free survival 10·7 months [95% CI 9·6-12·0]) and 109 (89%) patients in the surgery-plus-HIPEC group (14·3 months [12·0-18·5]; hazard ratio [HR] 0·63 [95% CI 0·48-0·83], stratified log-rank p=0·0008). Death occurred in 108 (88%) patients in the surgery group (median overall survival 33·3 months [95% CI 29·0-39·1]) and 100 (82%) patients in the surgery-plus-HIPEC group (44·9 months [95% CI 38·6-55·1]; HR 0·70 [95% CI 0·53-0·92], stratified log-rank p=0·011). INTERPRETATION: These updated survival results confirm the long-term survival benefit of HIPEC in patients with primary stage III epithelial ovarian cancer undergoing interval cytoreductive surgery. FUNDING: Dutch Cancer Foundation (KWF Kankerbestrijding).

Pembrolizumab, radiotherapy, and an immunomodulatory five-drug cocktail in pretreated patients with persistent, recurrent, or metastatic cervical or endometrial carcinoma: Results of the phase II PRIMMO study.

A phase II study (PRIMMO) of patients with pretreated persistent/recurrent/metastatic cervical or endometrial cancer is presented. Patients received an immunomodulatory five-drug cocktail (IDC) consisting of low-dose cyclophosphamide, aspirin, lansoprazole, vitamin D, and curcumin starting 2 weeks before radioimmunotherapy. Pembrolizumab was administered three-weekly from day 15 onwards; one of the tumor lesions was irradiated (8Gyx3) on days 15, 17, and 19. The primary endpoint was the objective response rate per immune-related response criteria (irORR) at week 26 (a lower bound of the 90% confidence interval [CI] of > 10% was considered efficacious). The prespecified 43 patients (cervical, n = 18; endometrial, n = 25) were enrolled. The irORR was 11.1% (90% CI 2.0-31.0) in cervical cancer and 12.0% (90% CI 3.4-28.2) in endometrial cancer. Median duration of response was not reached in both cohorts. Median interval-censored progression-free survival was 4.1 weeks (95% CI 4.1-25.7) in cervical cancer and 3.6 weeks (95% CI 3.6-15.4) in endometrial cancer; median overall survival was 39.6 weeks (95% CI 15.0-67.0) and 37.4 weeks (95% CI 19.0-50.3), respectively. Grade ≥ 3 treatment-related adverse events were reported in 10 (55.6%) cervical cancer patients and 9 (36.0%) endometrial cancer patients. Health-related quality of life was generally stable over time. Responders had a significantly higher proportion of peripheral T cells when compared to nonresponders (p = 0.013). In conclusion, PRIMMO did not meet its primary objective in both cohorts; pembrolizumab, radiotherapy, and an IDC had modest but durable antitumor activity with acceptable but not negligible toxicity.Trial registration ClinicalTrials.gov (identifier NCT03192059) and EudraCT Registry (number 2016-001569-97).

Evaluation of a new online cognitive assessment tool in breast cancer survivors with cognitive impairment: a prospective cohort study.

INTRODUCTION: Currently cancer-related cognitive impairment (CRCI) is mainly assessed by means of questionnaires, which is very laborious for the patients and the supervising physician. We evaluated a new online cognitive assessment tool, the MyCognition Quotient (MyCQ, Cambridge) in breast cancer survivors with CRCI, and compared the results with a psychometric test measuring cognitive complaints, depression, and anxiety. MATERIALS AND METHODS: In this prospective study, 46 adult patients between 18 and 70 years old with a diagnosis of BC were studied, all complaining of disturbing cognitive impairment. They participated in a physical cognitive rehabilitation program. The patients had an online cognitive assessment (MyCQ Med by MyCognition) every 4 weeks on their home computer. In addition patients were assessed in the outpatient clinic by the principal investigator at baseline, after 3 and 6 months using the following validated neuro-psychological surveys: the Hospital Anxiety and Depression Scale (HADS), Beck Cognitive Insight Scale (BCIS), and Cognitive Failure Questionnaire (CFQ). MyCQ scores were correlated with the results of these surveys. RESULTS: Only weak correlations could be found between overall MyCQ or the MyCQ subtests with the psychometric tests (between - 0.43 and 0.458) at baseline and when combining data at time point 0, 3, and 6 months. Linear mixed models showed there was a significant association between Latency Choice Reaction Time and CFQ (continuous; p = 0.026). An AUC of 0.640 and a cut-off of 481.5 ms in Latency Choice Reaction Time were found to distinguish patients with CFQ below 44 to patients with CFQ above 44 (sensitivity 0.63 and specificity 0.73). In Latency Coding an AUC of 0.788 and a cut-off of 1316 ms were found to distinguish non-depressive patients from patients likely to present with depressive symptoms (sensitivity 0.75 and specificity 0.76). CONCLUSION: MyCQ cannot replace the various psychometric tests. However, abnormal Latency in cognitive tests, Choice Reaction Time and Coding, seems promising to be used as a screening tool to detect specific aspects of abnormal cognitive functioning in patients with cognitive complaints and depressive symptoms.

The potential and controversy of targeting STAT family members in cancer.

The Signal Transducer and Activator of Transcription (STAT) family of proteins consists of transcription factors that play a complex and essential role in the regulation of physiologic cell processes, such as proliferation, differentiation, apoptosis and angiogenesis, and serves to organize the epigenetic landscape of immune cells. To date, seven STAT genes have been identified in the human genome; STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b and STAT6. They all account for diverse effects in response to extracellular signaling proteins, mainly by altering gene transcription in the effector cells. Members of the STAT family have been implicated in human cancer development, progression, metastasis, survival and resistance to treatment. Particularly STAT3 and STAT5 are of interest in cancer biology. They are currently considered as oncogenes, but their signaling is embedded into a complex and delicate balance between different (counteracting) transcription factors, and thus, in some contexts they can have a tumor suppressive role. Assessing STAT signaling mutations as well as screening for aberrant STAT pathway activation may have a role to predict sensitivity to immunotherapy and targeted STAT inhibition. In the present comprehensive review of the literature, we discuss in-depth the role of each STAT family member in cancer, assemble cutting-edge information on the use of these molecules as potential biomarkers and targets for treatment, and address why their clinical implementation is controversy.

Targeting the PD-1 Axis with Pembrolizumab for Recurrent or Metastatic Cancer of the Uterine Cervix: A Brief Update.

Even though cervical cancer is partly preventable, it still poses a great public health problem throughout the world. Current therapies have vastly improved the clinical outcomes of cervical cancer patients, but progress in new systemic treatment modalities has been slow in the last years. Especially for patients with advanced disease this is discouraging, as their prognosis remains very poor. The pathogen-induced nature, the considerable mutational load, the involvement of genes regulating the immune response, and the high grade of immune infiltration, suggest that immunotherapy might be a promising strategy to treat cervical cancer. In this literature review, we focus on the use of PD-1 blocking therapy in cervical cancer, pembrolizumab in particular, as it is the only approved immunotherapy for this disease. We discuss why it has great clinical potential, how it opens doors for personalized treatment in cervical cancer, and which trials are aiming to expand its clinical use.

Acceptability of quality indicators for the management of endometrial, cervical and ovarian cancer: results of an online survey.

BACKGROUND: Measuring quality indicators (QI's) is a tool to improve the quality of care. The aim of this study was to evaluate the acceptability of 36 QI's, defined after a literature search for the management of endometrial, cervical and ovarian cancer. Relevant specialists in the field of interest were surveyed. METHODS: To quantify the opinions of these specialists, an online survey was sent out via mailing to members of gynaecological or oncological societies. The relevance of each QI was questioned on a scale from one to five (1 = irrelevant, 2 = less relevant, 3 = no opinion/neutral, 4 = relevant, 5 = very relevant). If a QI received a score of 4 or 5 in 65% or more of the answers, we state that the respondents consider this QI to be sufficiently relevant to use in daily practice. RESULTS: The survey was visited 238 times and resulted in 53 complete responses (29 Belgian, 24 other European countries). The majority of the specialists were gynaecologists (45%). Five of the 36 QI's (13,9%) did not reach the cut-off of 65%: referral to a tertiary center, preoperative staging of endometrial cancer by MRI, preoperative staging of cervical cancer by positron-emission tomography, incorporation of intracavitary brachytherapy in the treatment of cervical cancer, reporting ASA and WHO score for each patient. After removing the 5 QI's that were not considered as relevant by the specialists and 3 additional 3 QI's that we were considered to be superfluous, we obtained an optimized QI list. CONCLUSION: As QI's gain importance in gynecological oncology, their use can only be of value if they are universally interpreted in the same manner. We propose an optimized list of 28 QI's for the management of endometrial, cervical and ovarian cancer which responders of our survey found relevant. Further validation is needed to finalize and define a set of QI's that can be used in future studies, audits and benchmarking.

Triple-negative breast cancer-Role of immunology: A systemic review.

Recently, the complex role of immune therapy has been the target of increased attention in breast cancer, particularly in triple-negative breast cancer (TNBC). Although TNBC is sensitive to chemotherapy, the recurrence and mortality rates are worse compared with the other breast cancer types. In addition, TNBC still lacks targeted treatment options. With the improved understanding of the immune system in TNBC, it is expected that new predictive and prognostic markers will be identified, and innovative treatment modalities will be developed. The aim of this review was to provide an overview of the effector cells in the TNBC's microenvironment and to highlight a novel approach to treat this kind of cancer. A computer-based literature research was carried out using PubMed, American Society of Clinical Oncology Annual Meeting (ASCO) and San Antonio Breast Cancer Symposium (SABCS). To date, studies have shown that tumor-infiltrating lymphocytes (TILs) and tumor-associated macrophages (TAMs) play a very important role in the TNBC's microenvironment. Tumor-infiltrating lymphocytes can even be considered as biomarkers to predict chemotherapy response in TNBC. Furthermore, TNBC was shown to have immune active subtypes, and therefore, the use of immunotherapy may be an attractive treatment approach. In this respect, several randomized studies have been designed or are currently ongoing to explore the combination of chemotherapy with immunotherapy in TNBC. Combination of chemo- and immunotherapy is likely to be beneficial in a subgroup of patients with TNBC.

The Non-Bone-Related Role of RANK/RANKL Signaling in Cancer.

RANK ligand (RANKL) is a member of the tumor necrosis factor alpha superfamily of cytokines. It is the only known ligand binding to a membrane receptor named receptor activator of nuclear factor-kappa B (RANK), thereby triggering recruitment of TNF receptor-associated factor (TRAF) adaptor proteins and activation of downstream pathways. RANK/RANKL signaling is controlled by a decoy receptor, osteoprotegerin (OPG), but also has additional more complex levels of regulation. It is crucial for the differentiation of bone-resorbing osteoclasts and is deregulated in disease processes such as osteoporosis and cancer bone metastasis. Cells expressing RANK and RANKL are commonly found in the tumor environment. In many tumor types, the RANK/RANKL pathway is overexpressed, and this is in most cases correlated with poor prognosis. RANK signaling plays an important role in the innate and adaptive immune response, generates regulatory T (Treg) cells, and increases the production of cytokines. It is also involved in chemo resistance in vitro. Recent evidence suggests that RANKL blockade improves the efficacy of anti-CTLA-4 antibodies against solid tumors and experimental metastasis. Therefore, there is increasing interest to use RANKL inhibition as an immunomodulatory strategy in an attempt to make immune-resistant tumor responsive to immune therapy.

PRIMMO study protocol: a phase II study combining PD-1 blockade, radiation and immunomodulation to tackle cervical and uterine cancer.

BACKGROUND: Immunotherapeutic approaches have revolutionized oncological practice but are less evaluated in gynecological malignancies. PD-1/PD-L1 blockade in gynecological cancers showed objective responses in 13-17% of patients. This could be due to immunosuppressive effects exerted by gynecological tumors on the microenvironment and an altered tumor vasculature. In other malignancies, combining checkpoint blockade with radiation delivers benefit that is believed to be due to the abscopal effect. Addition of immune modulation agents has also shown to enhance immune checkpoint blockade efficacy. Therefore we designed a regimen consisting of PD-1 blockade combined with radiation, and different immune/environmental-targeting compounds: repurposed drugs, metronomic chemotherapy and a food supplement. We hypothesize that these will synergistically modulate the tumor microenvironment and induce and sustain an anti-tumor immune response, resulting in tumor regression. METHODS: PRIMMO is a multi-center, open-label, non-randomized, 3-cohort phase 2 study with safety run-in in patients with recurrent/refractory cervical carcinoma, endometrial carcinoma or uterine sarcoma. Treatment consists of daily intake of vitamin D, lansoprazole, aspirin, cyclophosphamide and curcumin, starting 2 weeks before the first pembrolizumab dose. Pembrolizumab is administered 3-weekly for a total of 6 cycles. Radiation (3 × 8 Gy) is given on days 1, 3 and 5 of the first pembrolizumab dose. The safety run-in consists of 6 patients. In total, 18 and 25 evaluable patients for cervical and endometrial carcinoma respectively are foreseen to enroll. No sample size is determined for uterine sarcoma due to its rarity. The primary objective is objective response rate at week 26 according to immune-related response criteria. Secondary objectives include safety, objective response rate at week 26 according to RECIST v1.1, best overall response, progression-free survival, overall survival and quality of life. Exploratory, translational research aims to evaluate immune biomarkers, extracellular vesicles, cell death biomarkers and the gut microbiome. DISCUSSION: In this study, a combination of PD-1 blockade, radiation and immune/environmental-targeting compounds is tested, aiming to tackle the tumor microenvironment and induce anti-tumor immunity. Translational research is performed to discover biomarkers related to the mode of action of the combination. TRIAL REGISTRATION: EU Clinical Trials Register: EudraCT 2016-001569-97 , registered on 19-6-2017. Clinicaltrials.gov: NCT03192059 , registered on 19-6-2017.

RANK-RANKL Signaling in Cancer of the Uterine Cervix: A Review.

RANK ligand (RANKL) is a member of the tumor necrosis factor alpha superfamily of cytokines. It is the only known ligand binding to a membrane receptor named receptor activator of nuclear factor-kappa B (RANK), thereby triggering recruitment of tumor necrosis factor (TNF) receptor associated factor (TRAF) adaptor proteins and activation of downstream pathways. RANK/RANKL signaling is controlled by a decoy receptor called osteoprotegerin (OPG), but also has additional more complex levels of regulation. The existing literature on RANK/RANKL signaling in cervical cancer was reviewed, particularly focusing on the effects on the microenvironment. RANKL and RANK are frequently co-expressed in cervical cancer cells lines and in carcinoma of the uterine cervix. RANKL and OPG expression strongly increases during cervical cancer progression. RANKL is directly secreted by cervical cancer cells, which may be a mechanism they use to create an immune suppressive environment. RANKL induces expression of multiple activating cytokines by dendritic cells. High RANK mRNA levels and high immunohistochemical OPG expression are significantly correlated with high clinical stage, tumor grade, presence of lymph node metastases, and poor overall survival. Inhibition of RANKL signaling has a direct effect on tumor cell proliferation and behavior, but also alters the microenvironment. Abundant circumstantial evidence suggests that RANKL inhibition may (partially) reverse an immunosuppressive status. The use of denosumab, a monoclonal antibody directed to RANKL, as an immunomodulatory strategy is an attractive concept which should be further explored in combination with immune therapy in patients with cervical cancer.

A dynamic clinical pathway for the treatment of patients with early breast cancer is a tool for better cancer care: implementation and prospective analysis between 2002-2010.

BACKGROUND: Due to increasing the complexity of breast cancer treatment it is of paramount importance to develop structured care in order to avoid a chaotic and non-consistent management of patients. Clinical pathways, a result of the adaptation of the documents used in industrial quality management namely the Standard Operating Procedures, can be used to improve efficiency and quality of care. They also aim to re-centre the focus on the patient's overall journey, rather than the contribution of each specialty or caring function independently. METHODS: The effect of the implementation and prospective systematic evaluation of a clinical care pathway for the management of patients with early breast cancer in a single breast unit is evaluated over a long time interval (between 2002 and 2010). Annual analysis of predefined clinical outcome measures, service indicators, team indicators, process indicators and financial indicators was performed. Pathway quality control meetings were organized at least once a year. Systematic feedback was given to the team members, and if necessary the pathway was adapted according to evidence based literature data and in house pathway related data in order to improve quality. RESULTS: The annual number of patients included in the pathway (289 vs. 390, P <0.01), proportion of patients with Tis-T1 tumors (42% vs. 58%, P <0.01), negative lymph nodes (44% vs. 58%, P <0.01) and no metastases at diagnosis (91.5% vs. 95.9%) has risen significantly between 2002 and 2010. Evolution of mandatory quality indicators defined by EUSOMA shows a significant improvement of quality of cancer care. Particularly, the proportion of patients having anti-hormonal therapy (84.8% vs. 97.4%, P = 0.002) and adjuvant chemotherapy according to the guidelines (72% vs. 95.6%, P = 0.028) increased dramatically. Patient satisfaction improved significantly (P <0.05). Progression free 4-year survival was significantly higher for all patients, for T1 tumors only and for T2-T4 tumors only, treated between 2006 to 2008 compared to between 1999 to 2002 and 2003 to 2005 (P = 0.006, P = 0.05, P = 0.06, respectively). Overall 4-year survival of the entire population treated between 2006 and 2008 was significantly better (P = 0.05). CONCLUSIONS: Although the patient characteristics changed over the years due to better screening, this clinical pathway and regular audit of quality indicators for the treatment of patients with operable breast cancer proved to be important tools to improve the quality of care, patient satisfaction and outcome.

The immune microenvironment characterisation and dynamics in hormone receptor-positive breast cancer before and after neoadjuvant endocrine therapy.

BACKGROUND: Oestrogen receptor positive (ER+)/HER-2 negative breast cancer (BC) is considered to be an immunologically cold tumour compared to triple negative breast cancer. Therefore, the tumour microenvironment (TME) of ER+/HER-2 negative BC is understudied. The aim of this project is to investigate the TME and the immune response during neoadjuvant endocrine therapy (NET) and to correlate this with the treatment response in a real life setting. METHODS: Expression of immune checkpoint receptors and immune cells was examined immunohistochemically, pre- and post-NET in a cohort of 56 ER+/HER-2 negative BC patients. They were treated with tamoxifen (n = 16), an aromatase inhibitor (n = 40) or a combination of an aromatase inhibitor with a PI3K inhibitor (n = 11) for a median duration of 6 months (range 1-32 months). Immunohistochemical staining with monoclonal antibodies for PDL-1, PD-1, TIM-3, LAG-3, CTLA-4, CD4, CD68 and FOXP3 were performed. All staining procedures were done according to validated protocols, and scoring was done by a pathologist specialized in breast cancer. Positivity was defined as staining >1% on TILs. Response to NET was evaluated according to tumour size change on imaging and Ki-67 change. RESULTS: The median age was 61.02 (37-90) years. Diameter of tumour size decreased with a mean of 8.1 mm (-16 mm to 45 mm) (p < 0.001) during NET and the value of Ki-67 value decreased with a median of 9 after NET (p < 0.001). An increase in PD-L1 expression after NET showed a trend towards significant (p = 0.088) and CD-4+ T cells significantly increased after NET (p = 0.03). A good response to NET defined as a decrease in tumour size and/or decrease of Ki-67 was found to be associated with a longer duration of NET, a change of CD4+ T-cells and a higher number of CD68+ tumour-associated macrophages before the start of NET. CONCLUSION: The immune microenvironment plays an important role in ER+/HER-2 negative BC. NET influences the composition and functional state of the infiltrating immune cells. Furthermore, changes in the immune microenvironment are also associated with treatment response.

Humoral and Cellular Immune Responses against SARS-CoV-2 after Third Dose BNT162b2 following Double-Dose Vaccination with BNT162b2 versus ChAdOx1 in Patients with Cancer.

PURPOSE: Patients with cancer display reduced humoral responses after double-dose COVID-19 vaccination, whereas their cellular response is more comparable with that in healthy individuals. Recent studies demonstrated that a third vaccination dose boosts these immune responses, both in healthy people and patients with cancer. Because of the availability of many different COVID-19 vaccines, many people have been boosted with a different vaccine from the one used for double-dose vaccination. Data on such alternative vaccination schedules are scarce. This prospective study compares a third dose of BNT162b2 after double-dose BNT162b2 (homologous) versus ChAdOx1 (heterologous) vaccination in patients with cancer. EXPERIMENTAL DESIGN: A total of 442 subjects (315 patients and 127 healthy) received a third dose of BNT162b2 (230 homologous vs. 212 heterologous). Vaccine-induced adverse events (AE) were captured up to 7 days after vaccination. Humoral immunity was assessed by SARS-CoV-2 anti-S1 IgG antibody levels and SARS-CoV-2 50% neutralization titers (NT50) against Wuhan and BA.1 Omicron strains. Cellular immunity was examined by analyzing CD4+ and CD8+ T-cell responses against SARS-CoV-2-specific S1 and S2 peptides. RESULTS: Local AEs were more common after heterologous boosting. SARS-CoV-2 anti-S1 IgG antibody levels did not differ significantly between homologous and heterologous boosted subjects [GMT 1,755.90 BAU/mL (95% CI, 1,276.95-2,414.48) vs. 1,495.82 BAU/mL (95% CI, 1,131.48-1,977.46)]. However, homologous-boosted subjects show significantly higher NT50 values against BA.1 Omicron. Subjects receiving heterologous boosting demonstrated increased spike-specific CD8+ T cells, including higher IFNγ and TNFα levels. CONCLUSIONS: In patients with cancer who received double-dose ChAdOx1, a third heterologous dose of BNT162b2 was able to close the gap in antibody response.

Expression profiling of cancerous and normal breast tissues identifies microRNAs that are differentially expressed in serum from patients with (metastatic) breast cancer and healthy volunteers.

INTRODUCTION: MicroRNAs (miRNAs) are a group of small noncoding RNAs involved in the regulation of gene expression. As such, they regulate a large number of cellular pathways, and deregulation or altered expression of miRNAs is associated with tumorigenesis. In the current study, we evaluated the feasibility and clinical utility of circulating miRNAs as biomarkers for the detection and staging of breast cancer. METHODS: miRNAs were extracted from a set of 84 tissue samples from patients with breast cancer and eight normal tissue samples obtained after breast-reductive surgery. After reverse transcription and preamplification, 768 miRNAs were profiled by using the TaqMan low-density arrays. After data normalization, unsupervised hierarchical cluster analysis (UHCA) was used to investigate global differences in miRNA expression between cancerous and normal samples. With fold-change analysis, the most discriminating miRNAs between both tissue types were selected, and their expression was analyzed on serum samples from 20 healthy volunteers and 75 patients with breast cancer, including 16 patients with untreated metastatic breast cancer. miRNAs were extracted from 200 µl of serum, reverse transcribed, and analyzed in duplicate by using polymerase chain reaction (qRT-PCR). RESULTS: UHCA showed major differences in miRNA expression between tissue samples from patients with breast cancer and tissue samples from breast-reductive surgery (P < 0.0001). Generally, miRNA expression in cancerous samples tends to be repressed when compared with miRNA expression in healthy controls (P = 0.0685). The four most discriminating miRNAs by fold-change (miR-215, miR-299-5p, miR-411, and miR-452) were selected for further analysis on serum samples. All miRNAs at least tended to be differentially expressed between serum samples from patients with cancer and serum samples from healthy controls (miR-215, P = 0.094; miR-299-5P, P = 0.019; miR-411, P = 0.002; and miR-452, P = 0.092). For all these miRNAs, except for miR-452, the greatest difference in expression was observed between serum samples from healthy volunteers and serum samples from untreated patients with metastatic breast cancer. CONCLUSIONS: Our study provides a basis for the establishment of miRNAs as biomarkers for the detection and eventually staging of breast cancer through blood-borne testing. We identified and tested a set of putative biomarkers of breast cancer and demonstrated that altered levels of these miRNAs in serum from patients with breast cancer are particularly associated with the presence of metastatic disease.

Improved Characteristics of RANKL Immuno-PET Imaging Using Radiolabeled Antibody Fab Fragments.

PURPOSE: RANKL expression in the tumor microenvironment has been identified as a biomarker of immune suppression, negating the effect of some cancer immunotherapies. Previously we had developed a radiotracer based on the FDA-approved RANKL-specific antibody denosumab, [89Zr]Zr-DFO-denosumab, enabling successful immuno-PET imaging. Radiolabeled denosumab, however, showed long blood circulation and delayed tumor uptake, potentially limiting its applications. Here we aimed to develop a smaller radiolabeled denosumab fragment, [64Cu]Cu-NOTA-denos-Fab, that would ideally show faster tumor accumulation and better diffusion into the tumor for the visualization of RANKL. EXPERIMENTAL DESIGN: Fab fragments were prepared from denosumab using papain and conjugated to a NOTA chelator for radiolabeling with 64Cu. The bioconjugates were characterized in vitro using SDS-PAGE analysis, and the binding affinity was assessed using a radiotracer cell binding assay. Small animal PET imaging evaluated tumor targeting and biodistribution in transduced RANKL-ME-180 xenografts. RESULTS: The radiolabeling yield of [64Cu]Cu-NOTA-denos-Fab was 58 ± 9.2%, with a specific activity of 0.79 ± 0.11 MBq/µg (n = 3). A radiotracer binding assay proved specific targeting of RANKL in vitro. PET imaging showed fast blood clearance and high tumor accumulation as early as 1 h p.i. (2.14 ± 0.21% ID/mL), which peaked at 5 h p.i. (2.72 ± 0.61% ID/mL). In contrast, [64Cu]Cu-NOTA-denosumab reached its highest tumor uptake at 24 h p.i. (6.88 ± 1.12% ID/mL). [64Cu]Cu-NOTA-denos-Fab specifically targeted human RANKL in transduced ME-180 xenografts compared with the blocking group and negative ME-180 xenograft model. Histological analysis confirmed RANKL expression in RANKL-ME-180 xenografts. CONCLUSIONS: Here, we report on a novel RANKL PET imaging agent, [64Cu]Cu-NOTA-denos-Fab, that allows for fast tumor imaging with improved imaging contrast when compared with its antibody counterpart, showing promise as a potential PET RANKL imaging tool for future clinical applications.

Immuno-PET Molecular Imaging of RANKL in Cancer.

PURPOSE: The involvement of RANK/RANKL signaling in the tumor microenvironment (TME) in driving response or resistance to immunotherapy has only very recently been recognized. Current quantification methods of RANKL expression suffer from issues such as sensitivity, variability, and uncertainty on the spatial heterogeneity within the TME, resulting in conflicting reports on its reliability and limited use in clinical practice. Non-invasive molecular imaging using immuno-PET is a promising approach combining superior targeting specificity of monoclonal antibodies (mAb) and spatial, temporal and functional information of PET. Here, we evaluated radiolabeled anti-RANKL mAbs as a non-invasive biomarker of RANKL expression in the TME. EXPERIMENTAL DESIGN: Anti-human RANKL mAbs (AMG161 and AMG162) were radiolabeled with 89Zr using the bifunctional chelator DFO in high yield, purity and with intact binding affinity. After assessing the biodistribution in healthy CD-1 nude mice, [89Zr]Zr-DFO-AMG162 was selected for further evaluation in ME-180 (RANKL-transduced), UM-SCC-22B (RANKL-positive) and HCT-116 (RANKL-negative) human cancer xenografts to assess the feasibility of in vivo immuno-PET imaging of RANKL. RESULTS: [89Zr]Zr-DFO-AMG162 was selected as the most promising tracer for further validation based on biodistribution experiments. We demonstrated specific accumulation of [89Zr]Zr-DFO-AMG162 in RANKL transduced ME-180 xenografts. In UM-SCC-22B xenograft models expressing physiological RANKL levels, [89Zr]Zr-DFO-AMG162 imaging detected significantly higher signal compared to control [89Zr]Zr-DFO-IgG2 and to RANKL negative HCT-116 xenografts. There was good visual agreement with tumor autoradiography and immunohistochemistry on adjacent slides, confirming these findings. CONCLUSIONS: [89Zr]Zr-DFO-AMG162 can detect heterogeneous RANKL expression in the TME of human cancer xenografts, supporting further translation of RANKL immuno-PET to evaluate tumor RANKL distribution in patients.

The Evolution and Prognostic Role of Tumour-Infiltrating Lymphocytes and Peripheral Blood-Based Biomarkers in Inflammatory Breast Cancer Patients Treated with Neoadjuvant Chemotherapy.

INTRODUCTION: Inflammatory breast cancer (IBC) is a rare but aggressive form of breast cancer (BC) in which the (prognostic) role of stromal tumour-infiltrating lymphocytes (sTIL) and the peripheral circulating immune cells in patients with residual disease (RD) after neo-adjuvant chemotherapy (NACT) is not clearly established. METHODOLOGY: To describe the evolution of sTIL and some peripheral inflammation markers (Neutrophil-to-lymphocyte ratio, Platelet-to-lymphocyte ratio and Lymphocyte-to-monocyte ratio) after NACT in IBC, we retrospectively collected clinicopathological variables for 125 stage III IBC patients. sTILs were scored by three different researchers on an H&E slide of the mastectomy specimen. A cohort of subtype-matched non-IBC breast cancer patients (nIBC) treated with NACT was included for comparison. RESULTS: There was no significant difference in the pre- and posttreatment sTIL scores between IBC and nIBC and in both groups the number of sTIL was significantly lower after NACT. However, the IBC phenotype did correlate with a stronger decrease of sTIL after NACT (OR: 0.25, 95% CI: 0.073-0.76, p = 0.018). The change in the peripheral immune markers was not significantly different between IBC and nIBC. After NACT, 75 patients had residual disease. In this group, a high number of sTIL before NACT (HR: 0.23, 95% CI: 0.05-1.02, p = 0.05) was prognostic for a longer OS, while a low number of sTIL after NACT (HR: 0.33, 95% CI: 0.11-0.98, p = 0.046) and a low residual cancer cellularity (HR: 0.20, 95% CI: 0.08-0.52, p < 0.001) was associated with a longer DFS. CONCLUSIONS: IBC is associated with a significantly stronger decrease of sTIL after NACT compared to nIBC. Furthermore, a high number of sTIL after NACT was associated with a worse prognosis in IBC.

Blood Cytokine Analysis Suggests That SARS-CoV-2 Infection Results in a Sustained Tumour Promoting Environment in Cancer Patients.

Cytokines, chemokines, and (angiogenic) growth factors (CCGs) have been shown to play an intricate role in the progression of both solid and haematological malignancies. Recent studies have shown that SARS-CoV-2 infection leads to a worse outcome in cancer patients, especially in haematological malignancy patients. Here, we investigated how SARS-CoV-2 infection impacts the already altered CCG levels in solid or haematological malignancies, specifically, whether there is a protective effect or rather a potentially higher risk for major COVID-19 complications in cancer patients due to elevated CCGs linked to cancer progression. Serially analysing immune responses with 55 CCGs in cancer patients under active treatment with or without SARS-CoV-2 infection, we first showed that cancer patients without SARS-CoV-2 infection (n = 54) demonstrate elevated levels of 35 CCGs compared to the non-cancer, non-infected control group of health care workers (n = 42). Of the 35 CCGs, 19 were common to both the solid and haematological malignancy groups and comprised previously described cytokines such as IL-6, TNF-α, IL-1Ra, IL-17A, and VEGF, but also several less well described cytokines/chemokines such as Fractalkine, Tie-2, and T cell chemokine CTACK. Importantly, we show here that 7 CCGs are significantly altered in SARS-CoV-2 exposed cancer patients (n = 52). Of these, TNF-α, IFN-ß, TSLP, and sVCAM-1, identified to be elevated in haematological cancers, are also known tumour-promoting factors. Longitudinal analysis conducted over 3 months showed persistence of several tumour-promoting CCGs in SARS-CoV-2 exposed cancer patients. These data demonstrate a need for increased vigilance for haematological malignancy patients as a part of long COVID follow-up.

Anthracyclines Strike Back: Rediscovering Non-Pegylated Liposomal Doxorubicin in Current Therapeutic Scenarios of Breast Cancer.

Anthracyclines are among the most active chemotherapies (CT) in breast cancer (BC). However, cardiotoxicity is a risk and peculiar side effect that has been limiting their use in clinical practice, especially after the introduction of taxanes. Non-pegylated liposomal doxorubicin (NPLD) has been developed to optimize the toxicity profile induced by anthracyclines, while maintaining its unquestionable therapeutic index, thanks to its delivering characteristics that increase its diffusion in tumor tissues and reduce it in normal tissues. This feature allows NPLD to be safely administered beyond the standard doxorubicin maximum cumulative dose of 450-480 mg/m2. Following three pivotal first-line phase III trials in HER2-negative metastatic BC (MBC), this drug was finally approved in combination with cyclophosphamide in this specific setting. Given the increasing complexity of the therapeutic scenario of HER2-negative MBC, we have carefully revised the most updated literature on the topic and dissected the potential role of NPLD in the evolving therapeutic algorithms.

A video-game based cognitive training for breast cancer survivors with cognitive impairment: A prospective randomized pilot trial.

INTRODUCTION: We investigated whether a web-based cognitive training video game is an effective approach to improve cognitive decline in combination with our standard of care for rehabilitation of breast cancer (BC) patients. MATERIALS AND METHODS: Self-selected BC patients between 18 and 71 years old complaining of disturbing cognitive impairment were studied. The patients received access to a web-based internet video game and online cognitive assessments (Aquasnap, Cambridge, MyCQ™). The early intervention group (n = 23) had a training program of 6 months of at least three times a week for a minimum of 60 min of game playing per week at home in addition to standard of care rehabilitation. The delayed intervention (n = 23) received standard of care for three months, followed by three months of similar MyCQ training. Outcome measures were the MyCQ (sub)scores and Activity of Daily Life (ADL), mood, subjective cognition and functional cognitive status measured by classic neuropsychological tests. RESULTS: At baseline the means for CFQ (a measure of self-reported cognitive failure), anxiety, PSQI and self-reflectiveness were beyond normal range in both groups. CFQ improved significantly better in the intervention group (p = 0.029). Combining the evolution over time in the entire population a significant improvement was seen for overall MyCQ score, level of fear, physical and emotional role limitation, and health change (all p < 0.05), but self-reflectivess deteriorated (p < 0.05)). Significant differences in the various MyCQ subtests over time were: improved speed in choice reaction time, visual memory recognition, N back 1 and 2, coding, trail making test B, improved accuracy of N back 1 and 2 (all p < 0.05). CONCLUSION: A program of cognitive training improves cognitive functioning over time. "Aquasnap" has a beneficial effect on the perception of subjective cognitive functioning (CFQ) but the exact role of video gaming in this process remains uncertain.