BC-Monitor: Towards a Routinely Accessible Circulating Tumor DNA-Based Tool for Real-Time Monitoring Breast Cancer Progression and Treatment Effectiveness.

Circulating tumor DNA (ctDNA) is increasingly employed in the screening, follow-up, and monitoring of the continuously evolving tumor; however, most ctDNA assays validated for clinical use cannot maintain the right balance between sensitivity, coverage, sample requirements, time, and cost. Here, we report our BC-monitor, a simple, well-balanced ctDNA diagnostic approach using a gene panel significant in breast cancer and an optimized multiplex PCR-based NGS protocol capable of identifying allele variant frequencies below 1% in cell-free plasma DNA. We monitored a cohort of 45 breast cancer patients prospectively enrolled into our study receiving neoadjuvant chemotherapy or endocrine therapy or palliative therapy for metastatic diseases. Their tumor mutation status was examined in the archived tumor samples and plasma samples collected before and continuously during therapy. Traceable mutations of the used 38-plex NGS assay were found in approximately two-thirds of the patients. Importantly, we detected new pathogenic variants in follow-up plasma samples that were not detected in the primary tumor and baseline plasma samples. We proved that the BC-monitor can pre-indicate disease progression four-six months earlier than conventional methods. Our study highlights the need for well-designed ctDNA monitoring during treatment and follow-up, integrated into a real-time treatment assessment, which could provide information on the active tumor DNA released into the blood.

Loss of CDX2 gene expression is associated with DNA repair proteins and is a crucial member of the Wnt signaling pathway in liver metastasis of colorectal cancer.

Caudal type homeobox 2 (CDX2) has been well-established as a diagnostic marker for colorectal cancer (CRC); however, less is known about its regulation, particularly its potential interactions with the DNA repair proteins, adenomatous polyposis coli (APC) and ß-catenin, in a non-transcriptional manner. In the present study, the protein expression of CDX2 was analyzed, depending on the expression of the DNA repair proteins, mismatch repair (MMR), O6-methylguanine DNA methyltransferase (MGMT) and excision repair cross-complementing 1 (ERCC1), and its importance in Wnt signaling was also determined. A total of 101 liver metastases were punched into tissue microarray (TMA) blocks and serial sections were cut for immunohistochemistry. For each protein, an immunoreactive score was generated according to literature data and the scores were fitted to TMA. Subsequently, statistical analysis was performed to compare the levels of expression with each other and with clinical data. CDX2 loss of expression was observed in 38.5% of the CRC liver metastasis cases. A statistically significant association between CDX2 and each of the investigated MMRs was observed: MutL Homolog 1 (P<0.01), MutS protein Homolog (MSH) 2 (P<0.01), MSH6 (P<0.01), and postmeiotic segregation increased 2 (P=0.040). Furthermore, loss of MGMT and ERCC1 was also associated with CDX2 loss (P=0.039 and P<0.01, respectively). In addition, CDX2 and ERCC1 were inversely associated with metastatic tumor size (P=0.038 and P=0.027, respectively). Sustained CDX2 expression was associated with a higher expression of cytoplasmic/membranous ß-catenin and with nuclear APC expression (P=0.042 and P<0.01, respectively). In conclusion, CDX2 loss of expression was not a rare event in liver metastasis of CRC and the results suggested that CDX2 may be involved in mechanisms resulting in the loss of DNA repair protein expression, and in turn methylation; however, its exact function in this context remains to be elucidated.

Clinical benefit of fulvestrant monotherapy in the multimodal treatment of hormone receptor and HER2 positive advanced breast cancer: a case series.

Fulvestrant is a pure estrogen receptor (ER) antagonist approved for the treatment of metastatic ER positive breast cancer in postmenopausal women with disease progression following antiestrogen therapy. The clinical results of fulvestrant demonstrated encouraging activity in tumors in spite of HER2 positivity, but data about its use after progression on anti-HER2 agents are limited. Partial responses and durations of response of 12, 25, and 38 months in three cases with multiple metastases of ER positive and HER2 positive breast cancer were observed; all patients had been treated with 1-4 regimens of an anti-HER2 agent in combination with chemotherapy or an aromatase inhibitor before the initiation of fulvestrant. Fulvestrant is a valuable option with limited toxicity and durable response in metastatic HER2 and ER positive breast cancer after progression on anti-HER2 agents as well. Therapeutic benefit even in extensive skin metastases and (irradiated) brain metastases may be expected. Further investigations are warranted to establish where it fits into the multimodal management of ER and HER positive breast cancer.

Expression of ERCC1, RRM1, TUBB3 in correlation with apoptosis repressor ARC, DNA mismatch repair proteins and p53 in liver metastasis of colorectal cancer.

Liver metastasis in colorectal cancer is common and the primary treatment is chemotherapy. To date, there is no routinely used test in clinical practice to predict the effectiveness of conventional chemotherapy. Therefore, biomarkers with predictive value for conventional chemotherapy would be of considerable benefit in treatment planning. We analysed three proteins [excision repair cross-complementing 1 (ERCC1), ribonucleoside-diphosphate reductase 1 (RRM1) and class III ß-tubulin (TUBB3)] in colorectal cancer liver metastasis. We used tissue microarray slides with 101 liver metastasis samples, stained for ERCC1, RRM1 and TUBB3 and established scoring systems (fitted for tissue microarray) for each protein. In statistical analysis, we compared the expression of ERCC1, RRM1 and TUBB3 to mismatch proteins (MLH1, MSH2, MSH6 and PMS2), p53 and to apoptosis repressor protein (ARC). Statistically significant correlations were found between ERCC1, TUBB3 and MLH1, MSH2 and RRM1 and MSH2, MSH6. Noteworthy, our analysis revealed a strong significant correlation between cytoplasmic ARC expression and RRM1, TUBB3 (p=0.000 and p=0.001, respectively), implying an additional role of TUBB3 and RRM1 not only in therapy resistance, but also in the apoptotic machinery. Our data strengthens the importance of ERCC1, TUBB3 and RRM1 in the prediction of chemotherapy effectiveness and suggest new functional connections in DNA repair, microtubule network and apoptotic signaling (i.e. ARC protein). In conclusion, we showed the importance and need of predictive biomarkers in metastasized colorectal cancer and pointed out the relevance not only of single predictive markers but also of their interactions with other known and newly explored relations between different signaling pathways.

Cardiac Surveillance Findings During Adjuvant and Palliative Trastuzumab Therapy in Patients with Breast Cancer.

BACKGROUND/AIM: Trastuzumab therapy, the standard treatment for human epidermal growth factor receptor type-2 (HER2)-positive breast cancer, is associated with possible cardiotoxicity. We set out to retrospectively analyze the cardiac follow-up data of patients with breast cancer receiving trastuzumab treatment. PATIENTS AND METHODS: The study involved 47 and 31 patients receiving adjuvant or palliative chemotherapy plus trastuzumab, respectively. Cardiovascular system assessments including echocardiography were regularly performed. RESULTS: A significant heart abnormality was detected in 44.7% of the operable and 41.9% of metastatic cases. In the adjuvant setting, left ventricular ejection fraction changes occurred mostly during treatment and less frequently after its completion (40.4% vs. 19.4%), while in the palliative setting, 35.5% and 40% in the first and the second year of therapy. An asymptomatic atrial septum aneurysm was detected in 8.5% and 13% of the patients in the two groups. CONCLUSION: Trastuzumab-related cardiotoxicity is mostly manifested in an asymptomatic decrease in left ventricular ejection fraction; hypertension, a high body mass index and left-sided irradiation are its predictors.