Monitoring of breast cancer progression via aptamer-based detection of circulating tumor cells in clinical blood samples

Introduction: Breast cancer (BC) diagnostics lack noninvasive methods and procedures for screening and monitoring disease dynamics. Admitted CellSearch® is used for fluid biopsy and capture of circulating tumor cells of only epithelial origin. Here we describe an RNA aptamer (MDA231) for detecting BC cells in clinical samples, including blood. The MDA231 aptamer was originally selected against triple-negative breast cancer cell line MDA-MB-231 using cell-SELEX.Methods: The aptamer structure in solution was predicted using mFold program and molecular dynamic simulations. The affinity and specificity of the evolved aptamers were evaluated by flow cytometry and laser scanning microscopy on clinical tissues from breast cancer patients. CTCs were isolated form the patients’ blood using the developed method of aptamer-based magnetic separation. Breast cancer origin of CTCs was confirmed by cytological, RT-qPCR and Immunocytochemical analyses.Results: MDA231 can specifically recognize breast cancer cells in surgically resected tissues from patients with different molecular subtypes: triple-negative, Luminal A, and Luminal B, but not in benign tumors, lung cancer, glial tumor and healthy epithelial from lungs and breast. This RNA aptamer can identify cancer cells in complex cellular environments, including tumor biopsies (e.g., tumor tissues vs. margins) and clinical blood samples (e.g., circulating tumor cells). Breast cancer origin of the aptamer-based magnetically separated CTCs has been proved by immunocytochemistry and mammaglobin mRNA expression.Discussion: We suggest a simple, minimally-invasive breast cancer diagnostic method based on non-epithelial MDA231 aptamer-specific magnetic isolation of circulating tumor cells. Isolated cells are intact and can be utilized for molecular diagnostics purposes

Age determination of sediment core LV76-21

Eight samples of monospecific planktic foraminifera of Neogloboquadrina pachyderma (sin.) with a weight of 4–6 mg were picked from the >125‐μm size fraction and measured in the Beta Analytic Radiocarbon Dating Laboratory. Conventional accelerator mass spectrometer (AMS) 14C ages were calibrated to calendar ages using Calib 8.20 with the Marine20 calibration curve (Heaton et al., 2020) and the ΔR of 350 ± 40 years according to the reservoir age of the far NW Pacific equal to 900 years (Max et al., 2012). The resultant AMS 14C data along with other time points were used for age model construction

Results of analysis of sediment core LV76-21 from Detroit Seamount

The sediment core LV76-21 was recovered from northwest of the Detroit Seamount (51°34’N, 167°15.7’E; water depth 2,769 m) using a gravity corer on the R/V “Akademik M.A. Lavrentyev” during joint Russian–Chinese expeditions in 2016. The core sediments were predominantly gray–olive gray terrigenous silty clay–clayey silt with rare sand. The sediment was coarser in the intervals of 144–167 and 434–456 cm. The upper 17 cm of the sediment was represented by an oxidized brown–gray-brown ooze. In the intervals 0–38 cm, 76–167 cm, 356–364 cm, 407–434 cm, 456–475 cm, and 519–546 cm, the sediments were represented by grayish-olive–olive weakly diatomaceous oozes. The sediments in the intervals of 0–144 cm and 519–546 cm were enriched with foraminifera shells, and sediments were weakly enriched with foraminifera at the intervals of 407–434 cm, 456–482 cm, and 546–557 cm. The lithological description shows three visible ash layers at depths of 364–366 cm, 397–404 cm, and 467–468.5 cm, with thin ash lenses at a depth of 477–479 cm

Size fraction >150 µm determination of sediment core LV76-21

Sediment for IRD determination was sampled from 1 cm-thick slices with steps of 2–4 cm through the core. Samples with a dry weight of approximately 10 g were wet sieved using a 150 μm and 63 μm sieves. Coarse fraction >150 μm was decarbonized using 10% HCl, washed using distilled water, and dried. The ratio of weight percentage for the decarbonized coarse fraction (150–2,000 µm) relative to the weight of the dried bulk sediment were used as IRD. This is consistent with the commonly used size fraction > 150 μm for delimiting IRD (Hemming, 2004). Data in the intervals of the visible tephra layers were excluded from the IRD record because this material was delivered into sediment through the atmosphere, rather than by ice and, therefore, is not IRD. Need to understand that the nearby Kamchatka peninsula is area of active volcanism. Therefore, the determined IRD values (wt % of decarbonated fraction 150-2,000 µm) may include as terrigenous particles as volcanic particles, which were earlier fallout of the land during explosive volcanic eruptions, captured by sea ice and icebergs from the coast and land and then transferred into core location by water currents

Magnetic susceptibility of sediment core LV76-21

The sediment magnetic susceptibility (MS) was measured onboard using a Satis Geo KM-7 kappameter at a resolution of 1 cm. Within 2 h after coring, the sediment core was split, and one half of core was used for the MS measurements. Values of the sediment MS are determined by concentration of terrigenous ferromagnetic material and of volcanogenic material as well

Stable oxygen isotope record of Uvigerina auberiana of sediment core LV76-21

Sediments for the δ18O of planktic and benthic foraminifera analysis from core LV76-21 were sampled from 1 cm-thick slices with step of 2–5 cm along the core length. Foraminifera shells were picked from the sediment fractions of 150-250 µm in the samples where they were available. The δ18O of planktic and benthic foraminifera were measured at Tongji University (China) and at the Far Eastern Geological Institute (Russia). Isotope measurements from Tongji University were provided using a Finnigan‐MAT 252 mass spectrometer. The results were validated against a Chinese national carbonate standard (GBW04405) and NBS‐19. The standard deviation is 0.07‰ for δ18O. The isotope measurements from the Far Eastern Geological Institute were analyzed using a Finnigan-MAT 253 mass spectrometer with modification of Velivetskaya et al. (2009) without preliminary roasting. The standard deviation for the δ18O values is ± 0.05‰. All the values are reported on the Pee Dee Belemnite (PDB) scale

Size fraction >63 µm determination of sediment core LV76-21

Sediment for IRD determination was sampled from 1 cm-thick slices with steps of 2–4 cm through the core. Samples with a dry weight of approximately 10 g were wet sieved using a 150 μm and 63 μm sieves. Coarse fraction >150 μm was decarbonized using 10% HCl, washed using distilled water, and dried. The ratio of weight percentage for the decarbonized coarse fraction (150–2,000 µm) relative to the weight of the dried bulk sediment were used as IRD. This is consistent with the commonly used size fraction > 150 μm for delimiting IRD (Hemming, 2004). Data in the intervals of the visible tephra layers were excluded from the IRD record because this material was delivered into sediment through the atmosphere, rather than by ice and, therefore, is not IRD. Need to understand that the nearby Kamchatka peninsula is area of active volcanism. Therefore, the determined IRD values (wt % of decarbonated fraction 150-2,000 µm) may include as terrigenous particles as volcanic particles, which were earlier fallout of the land during explosive volcanic eruptions, captured by sea ice and icebergs from the coast and land and then transferred into core location by water currents

Carbon concentration of sediment core LV76-21

The total carbon and inorganic carbon (CaCO3) contents were measured in sediment slices of 1 cm thickness with 2 cm resolution using an AN-7529 coulometer (Gorbarenko et al., 1998). The total organic carbon (TOC) content in the sediment was calculated using the difference between the total carbon and inorganic carbon contents

Benthic foraminifera abundances of sediment core LV76-21

Benthic foraminifera were studied in the core LV76-21 sediments sampled in 1 cm-thick slices. All the samples were dried, weighted, and wet sieved using a 63 μm sieve. The benthic foraminifera shells were picked and counted from the 63–250 μm sediment fraction. In total, 222 samples were analyzed at an average resolution of 2.5 cm (range 1–5 cm). Almost all the samples had enough microfossils for representative evaluation, except for intervals 365 cm, 398–408 cm, 435–440 cm, 444–465 cm, and 490–510 cm, where foraminifera were rare or absent. The total benthic foraminifera abundance (BFA) expressed as the number of shells per 1 g of dry sediment (shells x g^−1)