Blood Draw Predicts Breast Cancer Relapse

Circulating tumor DNA (ctDNA) is a promising biomarker for noninvasive assessment of cancer burden, but existing ctDNA detection methods have insufficient sensitivity or patient coverage for broad clinical applicability. Here Newman et al., (2014) introduce cancer personalized profiling by deep sequencing (CAPP-Seq), an economical and ultrasensitive method for quantifying ctDNA. They implemented CAPP-Seq for non–small-cell lung cancer (NSCLC) with a design covering multiple classes of somatic alterations that identified mutations in >95% of tumors. They detected ctDNA in 100% of patients with stage II–IV NSCLC and in 50% of patients with stage I, with 96% specificity for mutant allele fractions down to ~0.02%. Levels of ctDNA were highly correlated with tumor volume and distinguished between residual disease and treatment-related imaging changes, and measurement of ctDNA levels allowed for earlier response assessment than radiographic approaches. Finally, we evaluated biopsy-free tumor screening and genotyping with CAPP-Seq. They envision that CAPP-Seq could be routinely applied clinically to detect and monitor diverse malignancies, thus facilitating personalized cancer therapy. Blood contains two types of cancer-derived materials that are susceptible to detailed molecular analysis: intact circulating tumor cells (CTC) and cell-free circulating tumor DNA (ctDNA). The former are shed from primary or metastatic tumor deposits, and although they are rare, they are thought to be enriched for metastatic precursors (Hater & Velculescu, 2014). Initially detected in an 1869 autopsy within the blood of a patient with widespread breast cancer (Ashworth, 1869), CTCs are now isolated with increasingly sophisticated technologies (Maheswaran & Haber, 2010; Pantel & Alix-Panabieres, 2010; Yu et al., 2011). However, the advantage of applying multiple DNA, RNA, and protein-based assays to study whole tumor cells in the circulation (so-called liquid biopsies) is currently restricted by the need for complex cellular isolation platforms. Recent advances in technologies to analyze circulating tumor cells and circulating tumor DNA are setting the stage for real-time, noninvasive monitoring of cancer and providing novel insights into cancer evolution, invasion, and metastasis. Source: Newman AM, Bratman SV, To J, et al. Nature Medicine. 20, 548–554 (2014) doi:10.1038/nm.3519 References: Ashworth TR. A case of cancer in which cells similar to those in the tumors were seen in the blood after death. Aust Med J 1869;14:146–9. Hater DA, Velculescu VE. Blood-Based Analyses of Cancer: Circulating Tumor Cells and Circulating Tumor DNA. Cancer Discovery June 2014 4; 650 doi: 10.1158/2159-8290.CD-13-1014 Maheswaran S, Haber DA. Circulating tumor cells: a window into cancer biology and metastasis. Curr Opin Genet Dev 2010;20:96–9. Pantel K, Alix-Panabieres C. Circulating tumour cells in cancer patients: challenges and perspectives. Trends Mol Med 2010;16:398–406. Pantel K, Alix-Panabieres M, Stott S, Toner M, Maheswaran S, Haber DA. Circulating tumor cells: approaches to isolation and characterization. J Cell Biol 2011;192:373–82. The management of metastatic breast cancer needs improvement. As clinical evaluation is not very accurate in determining the progression of disease, the analysis of circulating tumor DNA (ctDNA) has evolved to a promising noninvasive marker of disease evolution. Indeed, ctDNA was reported to represent a highly sensitive biomarker of metastatic cancer disease directly reflecting tumor burden and dynamics. However, at present little is known about the dynamic range of ctDNA in patients with metastatic breast cancer. In this study, 74 plasma DNA samples from 58 patients with metastasized breast cancer were analyzed with a microfluidic device to determine the plasma DNA size distribution and copy number changes in the plasma were identified by whole-genome sequencing (plasma-Seq). Furthermore, in an index patient we conducted whole-genome, exome, or targeted deep sequencing of the primary tumor, metastases, and circulating tumor cells (CTCs). Deep sequencing was done to accurately determine the allele fraction (AFs) of mutated DNA fragments. Although all patients had metastatic disease, plasma analyses demonstrated highly variable AFs of mutant fragments. analyzed an index patient with more than 100,000 CTCs in detail. Heidary et al., (2014) first conducted whole-genome, exome, or targeted deep sequencing of four different regions from the primary tumor and three metastatic lymph node regions, which enabled us to establish the phylogenetic relationships of these lesions, which were consistent with a genetically homogeneous cancer. Subsequent analyses of 551 CTCs confirmed the genetically homogeneous cancer in three serial blood analyses. However, the AFs of ctDNA were only 2% to 3% in each analysis, neither reflecting the tumor burden nor the dynamics of this progressive disease. These results together with high-resolution plasma DNA fragment sizing suggested that differences in phagocytosis and DNA degradation mechanisms likely explain the variable occurrence of mutated DNA fragments in the blood of patients with cancer. The dynamic range of ctDNA varies substantially in patients with metastatic breast cancer. This has important implications for the use of ctDNA as a predictive and prognostic biomarker. Clinical response evaluation of patients with cancer is often not accurate. Therefore, the use of ctDNA has been proposed as a biomarker for monitoring tumor burden and treatment response. Indeed, several studies have suggested that ctDNA analysis is an effective indicator of tumor load, allowing more accurate monitoring of tumor dynamics (Diehl et al., 2008; Leary et al., 2010; McBride et al., 2010). Circulating tumor DNA was successfully detected in 29 of the 30 women (97%) in whom somatic genomic alterations were identified; CA 15-3 and circulating tumor cells were detected in 21 of 27 women (78%) and 26 of 30 women (87%), respectively. Circulating tumor DNA levels showed a greater dynamic range, and greater correlation with changes in tumor burden, than did CA 15-3 or circulating tumor cells. Among the measures tested, circulating tumor DNA provided the earliest measure of treatment response in 10 of 19 women (53%). In breast cancer, a recent study has provided evidence that ctDNA levels had a greater dynamic range and greater correlation with changes in tumor burden than CA 15-3 or CTCs (Dawson et al., 2013). Source Heidary M, Auer M, Ulz P, et al. The dynamic range of circulating tumor DNA in metastatic breast cancer. Breast Cancer Research 2014, 16:421 http://breast-cancer-research.com/content/16/4/421 References Dawson SJ, Tsui DW, Murtaza M, et al. Analysis of circulating tumor DNA to monitor metastatic breast cancer. N Engl J Med 2013, 368:1199–1209. Diehl F, Schmidt K, Choti MA, et al. Circulating mutant DNA to assess tumor dynamics. Nat Med 2008, 14:985–990. Leary RJ, Kinde I, Diehl F, et al. Development of personalized tumor biomarkers using massively parallel sequencing. Sci Transl Med 2010, 2:20ra14. McBride DJ, Orpana AK, Sotiriou C, et al. Use of cancer-specific genomic rearrangements to quantify disease burden in plasma from patients with solid tumors. Gene Chromosome Canc 2010, 49:1062–1069. Tracking tumor DNA in the blood of early breast cancer patients after surgery can detect relapse 7.9 months earlier than conventional imaging, according to the results of a study published in Science Translational Medicine. Using a non-invasive circulating tumor DNA (ctDNA) analysis, Isaac Garcia-Murillas, PhD, of the Institute of Cancer Research in London, and colleagues tracked breast tumor–specific mutations in 55 patients who had undergone surgery and chemotherapy as a potentially curative treatment. The results of the prospective study suggest that patients at risk for relapse may be identified earlier and given more aggressive treatment to prevent metastasis. Of the 15 patients who relapsed on study, the presence of ctDNA predicted the relapse of 12 patients. Among the patients who did not relapse, 96% had no detectable ctDNA in either the post-surgery sample (24 of 25; P = .0038) or during temporal tracking of tumor mutations (27 of 28; P < .0001). One patient, with triple-negative disease, had detectable ctDNA after surgery but did not relapse on study. All metastatic tumors were detectable by ctDNA except for three patients who had recurrence in the brain. The study also showed that ctDNA analysis could identify the genetic events that define minimal residual disease among breast cancer patients. This genetic analysis of minimal residual disease also predicted the genetics of the relapsed tumor better than sequencing analysis of the primary breast tumor. The study authors concluded that tracking of ctDNA linked to cancer relapse could facilitate the tailoring of adjuvant therapies based on the mutations captured in patients’ blood samples. The utility of ctDNA analysis in late-stage cancer patients has already been demonstrated in previous research studies, although the technique is not yet ready to be used in the clinical setting. The current study, however, is among the first to demonstrate the ability to capture ctDNA from blood samples of earlier stage cancer patients. Tumor DNA circulates in the blood in minute amounts, and whether these molecules can be consistently detected using current technologies has been an open question. Garcia-Murillas and his colleagues first sequenced each patient’s primary tumor, identifying tumor-specific somatic mutations to track in the ctDNA following surgery. ctDNA detected at baseline, prior to any therapy, was not associated with early relapse. ctDNA detected at 2 to 4 weeks after surgery was indicative of early relapse—those who had detectable ctDNA (19%; 7 of 37 patients) had a median disease-free survival (DFS) of 6.5 months; median DFS among patients with no detectable ctDNA was not reached. “In addition to confirming the feasibility of ctDNA detection in non-metastatic breast cancer, Garcia-Murillas et al have extended these findings in a prospective study, demonstrating that longitudinal monitoring of ctDNA is more reliable than single baseline measurements in predicting recurrence in women treated with neoadjuvant chemotherapy for localized breast cancer,” stated Tilak Sundaresan, MD, and Daniel Haber, MD, PhD, of the Massachusetts General Hospital Cancer Center in Boston.

Is DCIS Cancer?

  As many as 60,000 American women each year are told they have a very early stage of breast cancer — Stage 0, as it is commonly known — a possible precursor to what could be a deadly tumor. And almost every one of the women has either a lumpectomy or a mastectomy, and often a double mastectomy, removing a healthy breast as well. Yet it now appears that treatment may make no difference in their outcomes. Patients with this condition had close to the same likelihood of dying of breast cancer as women in the general population, and the few who died did so despite treatment, not for lack of it, researchers reported Thursday in JAMA Oncology. continue reading

Gambogenic acid

Cancer: Breast
Action: Overcomes MDR, Adriamycin
Adriamycin (ADR) is beneficial for the treatment of breast cancer. However, its wide application often leads to drug resistance in clinic practice, which results in treatment failure. Gambogenic acid (GNA), a polyprenylated xanthone isolated from the traditional medicine gamboge, has been reported to effectively inhibit the survival and proliferation of cancer cells.
An MTT assay was used to evaluate the inhibitory effect of the drugs on the growth of MCF-7 and MCF-7/ADR cell lines. The effects of drugs on apoptosis were detected using Annexin-V APC/7-AAD double staining. The expression of apoptosis-related proteins and the proteins in the PTEN/PI3K/AKT pathway were evaluated by Western blot analysis.
In the MCF-7/ADR cell lines, the IC50 (half maximal inhibitory concentration) of the group that received combined treatment with GNA and ADR was significantly lower than that in the ADR group, and this value decreased with an increasing concentration of GNA. In parallel, GNA treatment increased the chemosensitivity of breast cancer cells to ADR. T
The study by He et al., (2015) has indicated a potential role for GNA to increase the chemosensitivity of breast cancer cells to ADR. This modulatory role was mediated by suppression of the PTEN/PI3K/AKT pathway that led to apoptosis in MCF-7/ADR cells. This work suggests that GNA may be used as a regulatory agent for treating ADR resistance in breast cancer patients.
Source
He Y, Ding J, Lin Y, et al. Gambogenic acid alters chemosensitivity of breast cancer cells to Adriamycin. BMC Complementary and Alternative Medicine 2015, 15:181 doi:10.1186/s12906-015-0710-8

Cancer: Lung
Action: Autophagy
Gambogenic acid (GNA) is one of the active compounds of Gamboge, a traditional medicine that was used as a drastic purgative, emetic, or vermifuge for treating tapeworm. Recently, increasing evidence has indicated that GNA exerts promising anti-tumor effects. In the present paper, Mei et al., (2014) found that GNA could induce the formation of vacuoles, which was linked with autophagy in A549 and HeLa cells. Further studies revealed that GNA triggers the initiation of autophagy based on the results of MDC staining, AO staining, accumulation of LC3 II, activation of Beclin 1 and phosphorylation of P70S6K. Similar results were obtained using a xenograft model. Their findings show, for the first time, that GNA can cause aberrant autophagy to induce cell death and may suggest the potential application of GNA as a tool or viable drug in anticancer therapies.
Source
Mei W, Dong C, Hui C, Bin L, Fenggen Y, Jingjing S, et al. (2014) Gambogenic Acid Kills Lung Cancer Cells through Aberrant Autophagy. PLoS ONE 9(1): e83604. doi:10.1371/journal.pone.0083604

Cancer: Triple Negative Breast
Action: Decrease bcl-2, inhibited cell proliferation, apoptosis
Zhou et al., (2013) used nude mouse models to detect the effect of gambogenic acid on breast tumors, analyzing expression of apoptosis-related proteins in vivo by Western blotting. Effects on cell proliferation, apoptosis and apoptosis-related proteins in MDA-MB-231 cells were detected by MTT, flow cytometry and Western blotting. Inhibitors of caspase-3,-8,-9 were also used to detect effects on caspase family members.
They found that gambogenic acid suppressed breast tumor growth in vivo, in association with increased expression of Fas and cleaved caspase-3,-8,-9 and bax, as well as decrease in the anti-apoptotic protein bcl-2. Gambogenic acid inhibited cell proliferation and induced cell apoptosis in a concentration-dependent manner.
Gambogenic acid suppressed breast cancer MDA-MB-231 cell growth by mediating apoptosis through death receptor and mitochondrial pathways in vivo and in vitro.
Source:
Zhou J, Luo YH, Wang JR, Lu BB, Wang KM, Tian Y. Gambogenic acid induction of apoptosis in a breast cancer cell line. Asian Pac J Cancer Prev. 2013;14(12):7601-5.

Potential Role of Ginseng in the Treatment of Colorectal Cancer

Colorectal cancer remains one of the most prevalent cancer and a leading cause of cancer related death in the US. Many currently used chemotherapeutic agents are derived from botanicals. Identifying herbal sources, including those from ginseng family, to develop better anti-cancer therapies remains an essential step in advancing the treatment of the cancer. In this article, potential roles of ginseng herbs, especially American ginseng and notoginseng, in colorectal cancer therapeutics are presented. The major pharmacologically active constituents of ginsengs are ginsenosides, which can be mainly classified as protopanaxadiol and protopanaxatriol groups. Structure-activity relationship between their chemical structures and pharmacological activities are discussed.

In addition, various steaming temperature and time treatment of the ginseng herbs can change ginsenoside profiles, and enhance their anti-cancer activities. This heat treatment process may increase the role of ginseng in treating colorectal cancer.

Source
Wang C-Z, Yuan C-S. Am. J. Chin. Med. 36, 1019 (2008). DOI: 10.1142/S0192415X08006545

Melanoma and chronic inflammatory patterns

Inflammasome: activation mechanisms Inflammation is a rapid biologic response of the immune system in vascular tissues, directed to eliminate stimuli capable of causing damage and begin the process of repair. The macromolecular complexes known as “inflammasomes” are formed by a receptor, either NOD (NLR) or ALR, the receptor absent in melanoma 2 (AIM2). In addition, the inflammasome is formed by the speck-like protein associated to apoptosis (ASC) and procaspase-1, that may be activated by variations in the ionic and intracellular and extracellular ATP concentrations; and the loss of stabilization of the fagolisosomme by internalization of insoluble crystals and redox mechanisms. As a result, there is activation of the molecular platform and the processing of inflammatory prointerleukins to their active forms. There are two modalities of activation of the inflammasome: canonical and non-canonical, both capable of generating effector responses. Recent data associate NLRP 3, IL-1? and IL-18 in the pathogenesis of a variety of diseases, including atherosclerosis, type II diabetes, hyperhomocysteinemia, gout, malaria and hypertension. The inflammasome cascade is emerging as a new chemotherapeutic target in these diseases. In this review we shall discuss the mechanisms of activation and regulation of the inflammasome that stimulate, modulate and resolve inflammation. Source Suárez R, Buelvas N. Invest Clin. 2015 Mar;56(1):74-99. C-reactive protein as a marker of melanoma progression. Two independent sets of plasma samples from a total of 1,144 patients with melanoma (587 initial and 557 confirmatory) were available for CRP determination. Kaplan-Meier method and Cox regression were used to evaluate the relationship between CRP and clinical outcome. Among 115 patients who underwent sequential blood draws, we evaluated the relationship between change in disease status and change in CRP using nonparametric tests. Elevated CRP level was associated with poorer OS and MSS in the initial, confirmatory, and combined data sets (combined data set: OS hazard ratio, 1.44 per unit increase of logarithmic CRP; 95% CI, 1.30 to 1.59; P < .001; MSS hazard ratio, 1.51 per unit increase of logarithmic CRP; 95% CI, 1.36 to 1.68; P < .001). These findings persisted after multivariable adjustment. As compared with CRP < 10 mg/L, CRP ? 10 mg/L conferred poorer OS in patients with any-stage, stage I/II, or stage III/IV disease and poorer disease-free survival in those with stage I/II disease. In patients who underwent sequential evaluation of CRP, an association was identified between an increase in CRP and melanoma disease progression. CONCLUSION: CRP is an independent prognostic marker in patients with melanoma. CRP measurement should be considered for incorporation into prospective studies of outcome in patients with melanoma and clinical trials of systemic therapies for those with melanoma. Source Fang S, Wang Y, Sui D, et al. J Clin Oncol. 2015 Apr 20;33(12):1389-96. doi: 10.1200/JCO.2014.58.0209. Serum amyloid A as a prognostic marker in melanoma identified by proteomic profiling. Currently known prognostic serum biomarkers of melanoma are powerful in metastatic disease, but weak in early-stage patients. This study was aimed to identify new prognostic biomarkers of melanoma by serum mass spectrometry (MS) proteomic profiling, and to validate candidates compared with established markers. Two independent sets of serum samples from 596 melanoma patients were investigated. The first set (stage I = 102; stage IV = 95) was analyzed by matrix assisted laser desorption and ionization time of flight (MALDI TOF) MS for biomarkers differentiating between stage I and IV. In the second set (stage I = 98; stage II = 91; stage III = 87; stage IV = 103), the serum concentrations of the candidate marker serum amyloid A (SAA) and the known biomarkers S100B, lactate dehydrogenase, and C reactive protein (CRP) were measured using immunoassays. MALDI TOF MS revealed a peak at m/z 11.680 differentiating between stage I and IV, which could be identified as SAA. High peak intensities at m/z 11.680 correlated with poor survival. In univariate analysis, SAA was a strong prognostic marker in stage I to III (P = .043) and stage IV (P = .000083) patients. Combination of SAA and CRP increased the prognostic impact to P = .011 in early-stage (I to III) patients. Multivariate analysis revealed sex, stage, tumor load, S100B, SAA, and CRP as independent prognostic factors, with an interaction between SAA and CRP. In stage I to III patients, SAA combined with CRP was superior to S100B in predicting patients’ progression-free and overall survival. CONCLUSION: SAA combined with CRP might be used as prognostic serological biomarkers in early-stage melanoma patients, helping to discriminate low-risk patients from high-risk patients needing adjuvant treatment. Source Findeisen P, Zapatka M, Peccerella T, et al. J Clin Oncol. 2009 May 1;27(13):2199-208. doi: 10.1200/JCO.2008.18.0554.