Silvestri Marco, Reduzzi Carolina, Feliciello Giancarlo, Vismara Marta, Schamberger Thomas, Köstler Cäcilia, Motta Rosita, Calza Stefano, Ferraris Cristina, Vingiani Andrea, Pruneri Giancarlo, Daidone Maria Grazia, Klein Christoph A, Polzer Bernhard, Cappelletti Vera
Biomarker Unit, Department of Applied Research and Technological Development, Fondazione IRCCS Istituto Nazionale dei Tumori di Milano, Via Giovanni Antonio Amadeo 42, 20133 Milano, Italy.
Division Personalized Tumor Therapy, Fraunhofer-Institute for Toxicology and Experimental Medicine, Biopark 1|Am Biopark 9, 93053 Regensburg, Germany.
Cancers (Basel). 2021 Mar 19;13(6):1409. doi: 10.3390/cancers13061409.
Circulating tumor microemboli (CTMs) are clusters of cancer cells detached from solid tumors, whose study can reveal mechanisms underlying metastatization. As they frequently comprise unknown fractions of leukocytes, the analysis of copy number alterations (CNAs) is challenging. To address this, we titrated known numbers of leukocytes into cancer cells (MDA-MB-453 and MDA-MB-36, displaying high and low DNA content, respectively) generating tumor fractions from 0-100%. After low-pass sequencing, ichorCNA was identified as the best algorithm to build a linear mixed regression model for tumor fraction (TF) prediction. We then isolated 53 CTMs from blood samples of six early-stage breast cancer patients and predicted the TF of all clusters. We found that all clusters harbor cancer cells between 8 and 48%. Furthermore, by comparing the identified CNAs of CTMs with their matched primary tumors, we noted that only 31-71% of aberrations were shared. Surprisingly, CTM-private alterations were abundant (30-63%), whereas primary tumor-private alterations were rare (4-12%). This either indicates that CTMs are disseminated from further progressed regions of the primary tumor or stem from cancer cells already colonizing distant sites. In both cases, CTM-private mutations may inform us about specific metastasis-associated functions of involved genes that should be explored in follow-up and mechanistic studies.
循环肿瘤微栓子（CTMs）是从实体瘤中脱离的癌细胞簇，对其研究可以揭示转移的潜在机制。由于它们经常包含未知比例的白细胞，因此分析拷贝数改变（CNA）具有挑战性。为了解决这个问题，我们将已知数量的白细胞滴定到癌细胞中（MDA-MB-453和MDA-MB-36，分别显示高和低DNA含量），产生0-100%的肿瘤比例。经过低通量测序，ichorCNA被确定为构建肿瘤比例（TF）预测线性混合回归模型的最佳算法。然后，我们从6例早期乳腺癌患者的血样中分离出53个CTM，并预测了所有簇的TF。我们发现所有簇中癌细胞的比例在8%至48%之间。此外，通过比较CTM中鉴定出的CNA与其匹配的原发性肿瘤，我们注意到只有31%-71%的畸变是共享的。令人惊讶的是，CTM特有的改变很丰富（30%-63%），而原发性肿瘤特有的改变很少（4%-12%）。这要么表明CTM是从原发性肿瘤进一步进展的区域播散而来，要么源于已经在远处定植的癌细胞。在这两种情况下，CTM特有的突变可能会让我们了解相关基因的特定转移相关功能，这些功能应在后续的机制研究中进行探索。
