Giuliana Restante

Detection of ALK and KRAS Mutations in Circulating Tumor DNA of Patients With Advanced ALK-Positive NSCLC With Disease Progression During Crizotinib Treatment

Background In patients with anaplastic lymphoma kinase (ALK)‐positive non–small‐cell lung cancer (NSCLC), disease progression occurs after a median of 9 to 10 months of crizotinib treatment. Several mechanisms of resistance have been identified and include ALK mutations and amplification or the activation of bypassing signaling pathways. Rebiopsy in NSCLC patients represents a critical issue and the analysis of circulating cell‐free DNA (cfDNA) has a promising role for the identification of resistance mechanisms. Patients and Methods Twenty patients with advanced ALK‐positive NSCLC were enrolled after disease progression during crizotinib treatment; cfDNA was analyzed using digital droplet polymerase chain reaction (BioRad, Hercules, CA) for ALK (p.L1196M, p.G1269A, and p.F1174L) and Kirsten rat sarcoma (KRAS) (codons 12 and 13) mutations. Results ALK secondary mutations (p.L1196M, p.G1269A, and p.F1174L) were identified in 5 patients; 1 patient had 2 ALK mutations (p.L1196M and p.G1269A). Overall, 10 patients presented KRAS mutations (7 p.G12D, 2 p.G12V, and 1 p.G12C mutations, respectively). In 3 patients KRAS mutations were associated with ALK mutations. cfDNA was monitored during the treatment with second‐generation ALK inhibitors and the amount of ALK as well as KRAS mutations decreased along with tumor regression. Conclusion ALK and KRAS mutations are associated with acquired resistance to crizotinib in ALK‐positive NSCLC. In particular, ALK acquired mutations can be detected in plasma and could represent a promising tumor marker for response monitoring. Micro‐Abstract We investigated mechanisms of acquired resistance in a group of anaplastic lymphoma kinase (ALK) translocated lung cancer patients whose disease progressed during crizotinib treatment using plasmatic circulating DNA extracted from blood samples. We identified 5 patients who presented ALK plasmatic mutations and 10 patients with Kirsten rat sarcoma (KRAS) gene mutations, both associated with crizotinib resistance. We showed that plasmatic decrease of mutation levels was associated with radiological response confirming that resistance ALK and KRAS mutations are detectable and their monitoring could serve as response parameters.

Pharmacogenetics and Metabolism from Science to Implementation in Clinical Practice: The Example of Dihydropyrimidine Dehydrogenase

BACKGROUND Fluoropyrimidines are widely used in the treatment of solid tumors and remain the backbone of many combination chemotherapy regimens. Despite their clinical benefit, they are associated with frequent gastrointestinal and hematological toxicities, which often lead to treatment discontinuation. Fluoropyrimidines undergo complex anabolic and catabolic biotransformation. Enzymes involved in this pathway include dihydropyrimidine dehydrogenase (DPD), which breaks down 5-FU and its prodrugs. Candidate gene approaches have demonstrated associations between 5-FU treatment outcomes and germline polymorphisms in DPD. The aim of this review is to report and discuss the latest results on fluoropyrimidine pharmacogenetics. METHODS Literature from PubMed databases and bibliography from retrieved publications have been analyzed according to terms such DPD, DPYD, fluoropyrimdines, polymorphisms, toxicity, pharmacogenetics. RESULTS To date, many sequence variations have been identified within DPYD gene, although the majority of these have no functional consequences on enzymatic activity. Nowadays, there is a general agreement on the clinical significance of the importance of DPD deficiency in patients who suffer from severe, life-threatening drug toxicity although preemptive testing is not applied to all patients. CONCLUSION Considering the published literature, clinicians are strongly encouraged to consider testing for DPD poor metabolizer variants as a rational pre-treatment screening for patients candidate to a fluoropyrimidine-based regimens, in order to prevent toxicities and personalise treatments.

Critical focus on mechanisms of resistance and toxicity of m-TOR inhibitors in pancreatic neuroendocrine tumors

Pancreatic neuroendocrine tumors (pNETs) are rare neoplasms representing less than 2% of all pancreatic malignancies. The PI3K-AKT-mTOR pathway is often deregulated in pNETs and seems to play a key role in tumorigenesis. Everolimus, an inhibitor of the mTOR pathway, has demonstrated efficacy in the treatment of pNETs. Nevertheless de novo or acquired drug resistance is responsible for disease progression and represents a major obstacle to overcome by clinicians. Blocking the PI3K/AKT/mTOR pathway may cover the supposed main mechanisms of resistance to everolimus. Therefore, BEZ-235, a potent oral dual PI3K/mTOR inhibitor was investigated in clinical trials. Globally more than 250 patients with different types of solid tumors were treated. Two studies were conducted in pNETs with BEZ-235 as single agent. The former was a phase 2 trial conducted in pNETs resistant to everolimus while the latter a randomized trial comparing everolimus and BEZ-235. Unfortunately, both the studies disappointed the expectations and were prematurely halted mainly due to severe toxicity. On this basis we reviewed m-TOR inhibitors in pNETs, focusing on their mechanisms of resistance and toxicity.

Concise Review: Chronic Myeloid Leukemia: Stem Cell Niche and Response to Pharmacologic Treatment

Nowadays, more than 90% of patients affected by chronic myeloid leukemia (CML) survive with a good quality of life, thanks to the clinical efficacy of tyrosine kinase inhibitors (TKIs). Nevertheless, point mutations of the ABL1 pocket occurring during treatment may reduce binding of TKIs, being responsible of about 20% of cases of resistance among CML patients. In addition, the presence of leukemic stem cells (LSCs) represents the most important event in leukemia progression related to TKI resistance. LSCs express stem cell markers, including active efflux pumps and genetic and epigenetic alterations together with deregulated cell signaling pathways involved in self‐renewal, such as Wnt/β‐catenin, Notch, and Hedgehog. Moreover, the interaction with the bone marrow microenvironment, also known as hematopoietic niche, may influence the phenotype of surrounding cells, which evade mechanisms controlling cell proliferation and are less sensitive or frankly resistant to TKIs. This Review focuses on the role of LSCs and stem cell niche in relation to response to pharmacological treatments. A literature search from PubMed database was performed until April 30, 2017, and it has been analyzed according to keywords such as chronic myeloid leukemia, stem cell, leukemic stem cells, hematopoietic niche, tyrosine kinase inhibitors, and drug resistance. Stem Cells Translational Medicine 2018;7:305–314

Patients with NSCLC may display a low ratio of p.T790M vs. activating EGFR mutations in plasma at disease progression: implications for personalised treatment

Introduction NSCLC harboring activating mutations of EGFR is highly sensitive to first-line EGFR-tyrosine kinase inhibitors (TKIs), but drug resistance depending on the EGFR mutation p.T790M will occur in about 50-60% of patients. Detailed information on the amount of p.T790M plasmatic level associated with resistance to EGFR-TKIs and guidance to treatment with p.T790M-effective TKI depending on these levels, is lacking. Methods This study enrolled p.T790M-positive patients (n=49) affected by EGFR-mutated NSCLC at progression to first-line EGFR-TKIs and, in selected cases (n=5), after second-line treatment with osimertinib. Cell-free circulating tumor DNA (cftDNA) was extracted from plasma and the quantitative analysis of EGFR ex19del, p.L858R and p.T790M was performed by digital droplet PCR. Results The mean amount of mutated alleles at progression to first-line EGFR-TKIs was 108,492 copies/ml for ex19del, 97,336 copies/ml for p.L858R, but only 8,754 copies/ml for p.T790M. There was no significant correlation between progression-free survival and the ratio of p.T790M over EGFR activating mutations. The analysis of cftDNA in 5 patients treated with osimertinib revealed a marked decrease of all EGFR mutant alleles. Conclusions The amount of p.T790M in plasma can be much lower than activating EGFR mutations. Despite this finding, osimertinib is effective in p.T790M-positive patients. These results indicate that clones driving resistance to EGFR-TKIs represent a minority among cells bearing activating EGFR-mutations. In addition, the identification of a threshold level of p.T790M is not a strict requirement for the selection of patients to be treated with osimertinib, since treatment showed a decrease in all EGFR mutated cells.

Concise Review: Resistance to Tyrosine Kinase Inhibitors in Non‐Small Cell Lung Cancer: The Role of Cancer Stem Cells

Among the potential mechanisms involved in resistance to tyrosine kinase inhibitors (TKIs) in non‐small cell lung cancer, the manifestation of stem‐like properties in cancer cells seems to have a crucial role. Alterations involved in the development of TKI resistance may be acquired in a very early phase of tumorigenesis, supporting the hypothesis that these aberrations may be present in cancer stem cells (CSCs). In this regard, the characterization of tumor subclones in the initial phase and the identification of the CSCs may be helpful in planning a specific treatment to target selected biomarkers, suppress tumor growth, and prevent drug resistance. The aim of this review is to elucidate the role of CSCs in the development of resistance to TKIs and its implication for the management of patients. Stem Cells 2018;36:633–640

Pharmacogenetics of androgen signaling in prostate cancer: Focus on castration resistance and predictive biomarkers of response to treatment

Tumor heterogeneity strongly affects the molecular mechanisms driving resistance to hormonal therapies in castration-resistant prostate cancer. Since the current use of available treatments can be optimized on the basis of the molecular profile of tumor, the present review focuses on genetic biomarkers in prostate cancer and their application to a personalized treatment.

PD-L1 mRNA expression in plasma-derived exosomes is associated with response to anti-PD-1 antibodies in melanoma and NSCLC.

This corrects the article DOI: 10.1038/bjc.2017.85

Unusual gastrointestinal and cutaneous toxicities by bleomycin, etoposide, and cisplatin: a case report with pharmacogenetic analysis to personalize treatment

The standard treatment of testicular germ cell tumors is based on the combination of bleomycin, etoposide, and cisplatin (PEB). However, this treatment may be associated with severe adverse reactions, such as hematological and non-hematological toxicities. Here, we report a case of a patient suffering from severe PEB-related toxicities, to whom pharmacogenetic analyses were performed, comprising a panel of genes involved in PEB metabolism. The analysis revealed the presence of a complex pattern of polymorphisms in GSTP1, UGT1A1 (TA)6/7, UGT1A7, and ABCB1. The present case shows that a pharmacogenetic approach can help in the management of adverse drug reactions in order to predict, prevent, and personalize treatments.

KRAS mutations and its implications for acquired resistance to treatment in NSCLC

Rationale KRAS is the most common and, simultaneously, the most ambiguous oncogene implicated in human cancer. Despite KRAS mutations were identified in Non Small Cell Lung Cancers (NSCLCs) more than 20 years ago, selective and specific inhibitors aimed at directly abrogating KRAS activity are not yet available. Nevertheless, many therapeutic approaches have been developed potentially useful to treat NSCLC patients mutated for KRAS and refractory to both standard chemotherapy and targeted therapies. The focus of this review will be to provide an overview of the network related to the intricate molecular KRAS pathways, stressing on preclinical and clinical studies that investigate the predictive value of KRAS mutations in NSCLC patients. Materials and Methods A bibliographic search of the Medline database was conducted for articles published in English, with the keywords KRAS, KRAS mutations in non-small cell lung cancer, KRAS and tumorigenesis, KRAS and TKIs, KRAS and chemotherapy, KRAS and monoclonal antibody, KRAS and immunotherapy, KRAS and drugs, KRAS and drug resistance.