Molecular profiling of ctDNA in pancreatic cancer: Opportunities and challenges for clinical application

Pancreatic ductal adenocarcinoma (PDAC) is predicted to become the second leading cause of cancer-related mortality within the next decade, with limited effective treatment options and a dismal long-term prognosis for patients. Genomic profiling has not yet manifested clinical benefits for diagnosis, treatment or prognosis in PDAC, due to the lack of available tissues for sequencing and the confounding effects of low tumour cellularity in many biopsy specimens. Increasing focus is now turning to the use of minimally invasive liquid biopsies to enhance the characterisation of actionable PDAC tumour genomes. Circulating tumour DNA (ctDNA) is the most comprehensively studied liquid biopsy analyte in blood and can provide insight into the molecular profile and biological characteristics of individual PDAC tumours, in real-time and in advance of traditional imaging modalities. This can pave the way for identification of new therapeutic targets, novel risk variants and markers of tumour response, to supplement diagnostic screening and provide enhanced scrutiny in treatment stratification. In the roadmap towards the application of precision medicine for clinical management in PDAC, ctDNA analyses may serve a leading role in streamlining candidate biomarkers for clinical integration. In this review, we highlight recent developments in the use of ctDNA-based liquid biopsies for PDAC and provide new insights into the technical, analytical and biological challenges that must be overcome for this potential to be realised.     

The use of personalized biomarkers and liquid biopsies to monitor treatment response and disease recurrence in locally advanced rectal cancer after neoadjuvant chemoradiation

Neoadjuvant chemoradiotherapy (nCRT) followed by surgery is the mainstay treatment for locally advanced rectal cancer. Variable degrees of tumor regression are observed after nCRT and alternative treatment strategies, including close surveillance without immediate surgery, have been investigated to spare patients with complete tumor regression from potentially adverse outcomes of radical surgery. However, clinical and radiological assessment of response does not allow accurate identification of patients with complete response. In addition, surveillance for recurrence is similarly important for these patients, as early detection of recurrence allows salvage resections and adjuvant interventions. We report the use of liquid biopsies and personalized biomarkers for monitoring treatment response to nCRT and detecting residual disease and recurrence in patients with rectal cancer. We sequenced the whole-genome of four rectal tumors to identify patient-specific chromosomal rearrangements that were used to monitor circulating tumor DNA (ctDNA) in liquid biopsies collected at diagnosis and during nCRT and follow-up. We compared ctDNA levels to clinical, radiological and pathological response to nCRT. Our results indicate that personalized biomarkers and liquid biopsies may not be sensitive for the detection of microscopic residual disease. However, it can be efficiently used to monitor treatment response to nCRT and detect disease recurrence, preceding increases in CEA levels and radiological diagnosis. Similar good results were observed when assessing tumor response to systemic therapy and disease progression. Our study supports the use of personalized biomarkers and liquid biopsies to tailor the management of rectal cancer patients, however, replication in a larger cohort is necessary to introduce this strategy into clinical practice.     

Toward liquid biopsies in cancer treatment: application of circulating tumor DNA

Circulating tumor DNA (ctDNA) refers to the fraction of cell‐free DNA in a patient's blood originating from tumor cells. Increased knowledge about tumor genomics, improvements in targeted therapies, and accompanying advances in DNA‐sequencing technologies have increased the interest in using ctDNA as a minimally invasive tool in cancer diagnostics and treatment. Especially, early tumor detection including identification of minimal residual disease and stratification of adjuvant therapy are promising approaches. Also, ctDNA showed to be reliable in treatment monitoring and can be used to assess therapy resistance due to the broad variety of tumor subclones captured in ctDNA. Therefore, using ctDNA in the clinical setting has the potential to improve therapeutic outcomes. In the present review, we summarize the status of ctDNA in oncology with focus of being an alternative to tissue biopsies in early detection and treatment monitoring.     

A liquid biopsy for head and neck cancers

Head and neck cancer patients often present with advanced metastatic disease resulting in a poor 5-year survival. Therefore, there is a need for non-invasive diagnostic tools that could complement conventional imaging to inform clinicians of patient outcomes and treatment responses. A liquid biopsy addresses this unmet clinical need; a simple peripheral blood draw could provide information about the disseminated disease in terms of circulating tumor cells and circulating tumor DNA. Moreover, detectable tumor DNA in the saliva of head and neck cancer patients could signify the early signs of the disease and present an opportunity for clinical intervention. This review provides an overview of the current literature with regard to the feasibility of such a test in the head and neck cancer field and highlights the need for such a test.     

Plasma Circulating Tumor DNA and Clonal Hematopoiesis in Metastatic Renal Cell Carcinoma

BackgroundThere is a lack of molecularly-informed biomarkers for patients with metastatic renal cell carcinoma (RCC). Plasma cell-free DNA (cfDNA) sequencing is a minimally-invasive alternative to tissue for profiling the genome in other cancers but relevance in metastatic RCC remains unclear.Materials and MethodsWhole blood was collected from 55 patients with metastatic RCC. Plasma cfDNA and leukocyte DNA were subjected to targeted sequencing across 981 cancer genes. Matched tumor tissue from 14 patients was analyzed.ResultsThirty-three percent of patients had evidence for RCC-derived circulating tumor DNA (ctDNA), significantly lower than patients with metastatic prostate or bladder cancer analyzed using the same approach. Among ctDNA-positive patients, ctDNA fraction averaged only 3.9% and showed no strong association with clinical variables. In these patients, the most commonly mutated genes were VHL, BAP1, and PBRM1, and matched tissue concordance was 77%. Evidence of somatic expansions unrelated to RCC, such as clonal hematopoiesis of indeterminate potential, were detected in 43% of patients. Pathogenic germline mutations in DNA repair genes were detected in 11% of patients. CtDNA-positive patients had shorter overall survival and progression-free survival on first-line therapy. Patients with evidence of clonal hematopoiesis of indeterminate potential had an intermediate prognosis compared with ctDNA-positive and -negative patients.ConclusionsCfDNA sequencing enables straightforward characterization of the somatic RCC genome in a minority of patients with metastatic RCC. Owing to low ctDNA abundance, and the presence of non-RCC derived somatic clones in circulation, cfDNA sequencing may not be a simple pan-patient alternative to tissue biopsy in metastatic RCC.     

Intercalation of imidazoacridinones to DNA and its relevance to cytotoxic and antitumor activity

Imidazoacridinones (IA) are a class of antitumor agents which includes C-1311, an interesting drug in clinical trials. This study investigated the mechanism of IA binding to DNA for a series of 13 analogs that differ in their cytotoxic potency. Using C-1311 as a model compound, crystallographic, spectroscopic and biochemical techniques were employed to characterize drug-DNA interactions. X-ray crystallographic analysis revealed a planar structure of imidazoacridinone core that is capable of intercalative DNA binding. Accordingly, C-1311 binding to DNA followed 'classical' pattern observed for intercalation, as proved by the DNA topoisomerase I-unwinding experiments, with relatively weak binding affinity (K(i)=1.2 x 10(5)M(-1)), and the binding site size of 2.4 bp. Other IA also bound to DNA with the binding affinity in the range of 10(5)M(-1) and binding site size of 2-3 bp, suggesting a prevalence of the intercalative mechanism, similar to C-1311. Considerable DNA binding affinity was displayed by all the highly cytotoxic derivatives. However, none of the analyzed drug-DNA binding parameters was significantly correlated with IA biological activities such as cell growth, DNA and RNA synthesis inhibition, or tumor growth inhibition, which suggests that the IA ability to non-covalently bind to DNA is not crucial for their biological activity. These results show that the ability to intercalate into DNA is a prominent attribute of IA, although factors other than intercalative binding seem to be required for the biological activities of IA drugs.