Mitochondrial DNA mutations in breast cancer tissue and in matched nipple aspirate fluid

Abstract

Unlike nuclear (n)DNA, of which there is one paired copy per cell, there are many copies of mitochondrial (mt)DNA per cell, making PCR amplification of mtDNA easier in samples of limited cellularity. The aims of this study were to (i) determine the mutation patterns of breast cancers through a comprehensive screen of mtDNA mutations, and (ii) assess if mutations in the cancers are also detectable in breast nipple aspirate fluid (NAF), a physiologic fluid which contains shed ductal epithelial cells. Fifteen breast cancers, matched benign tissues and NAF were collected. Nine overlapping primer sets were used to sequence the entire mitochondrial genome from tissue samples. For NAF samples, we focused on the 19 nucleotide positions (np) where mutations were found in a 3701 bp region (np 15331 to 2463), which includes the displacement (D)-loop, a mtDNA mutation hot spot. Fourteen of the fifteen (93%) cancer samples had > or =1 somatic mtDNA mutation for a total of 45 at 35 np (9 np reported previously, 26 new). Nine of fifteen tumors had > or =2 mutations. The D-loop contained 17 of 45 (38%) and non-D-loop (coding) regions contained 28 (62%) mutations. Of the 28 mutations in the coding loci, 11 led to an amino acid change. The frequency of mtDNA mutations was higher in the D-loop region (1.5 versus 0.18% of loci). 155 polymorphisms were identified (98 reported previously, 57 new). Sixteen of forty-five (36%) mutations were located at polymorphism sites. Four of nineteen mtDNA mutations in 10 cancers located between np 15331 and 2463 were found in matched NAF (two of eleven mutations in the D-loop and two of eight in non-D-loop regions). No mutations were found in five matched NAF samples from women whose cancers lacked a mutation in the same region. In conclusion, mtDNA mutations in breast cancer occur both within and outside of the D-loop, though the mutation rate in the D-loop is over 7-fold higher than in coding areas. We identified 26 new mutation loci (25 in regions sequenced by others, one in an area not). The high frequency of mtDNA mutations at polymorphic loci requires further investigation. Specific mtDNA mutations can be detected in a subset of NAF samples from women with breast cancer.

Quantitative analysis of mitochondrial DNA 4977-bp deletion in sporadic breast cancer and benign breast diseases

“…The development of cancer involves the accumulation of various genetic alterations, which are present both in the mitochondrial and nuclear genomes (1). Human mitochondrial DNA (mtDNA) is a circular molecule consisting of 16,571 bp encoding 2 rRNAs, 22 tRNAs, and 13 polypeptides (2,3).…”

Rapid screening of the entire mitochondrial DNA for low-level heteroplasmic mutations

“…Recent studies demonstrated that mtDNA mutations accumulate with increasing age (Jaksch et al, 2001;Murdock et al, 2000), that the level of heteroplasmy is associated with the age of onset and progression of chronic conditions, such as diabetic complications (Iwase et al, 2001) and that the decline in mitochondrial function can be due to Mitochondrion 5 (2005) [282][283][284][285][286][287][288][289][290][291][292][293][294][295][296] www.elsevier.com/locate/mito the accumulation of mutations in mtDNA (Singh, 2004). Somatic homoplasmic and heteroplasmic changes have also been reported in a wide variety of cancers suggesting an influence on carcinogenesis (Singh, 2004;Yang et al, 2004;Zhu et al, 2005). Denaturing high performance liquid chromatography (DHPLC), a method which separates heteroduplex and homoduplex DNA molecules on a stationary phase under partially denaturating conditions has already been shown to successfully detect unknown alterations in nuclear encoded genes (O'Donovan et al, 1998;Torrente et al, 2004;Xiao and Oefner, 2001) and the mtDNA (Conley et al, 2003;Danielson et al, 2003;LaBerge et al, 2003;Liu et al, 2002;van Den Bosch et al, 2000).…”

Mitochondrial DNA mutations in renal cell carcinomas revealed no general impact on energy metabolism

“…Recently, a high incidence of specific mtDNA alterations has been reported for gastric (Maximo et al, 2001;Wu et al, 2005), prostate (Jeronimo et al, 2001;Petros et al, 2005), pancreatic (Jones et al, 2001), skin (Girald-Rosa et al, 2005), colorectal (Polyak et al, 1998;Hibi et al, 2001a;Lievre et al, 2005), urinary bladder (Fliss et al, 2000), thyroid (Yeh et al, 2000), oesophageal (Hibi et al, 2001b;Kumimoto et al, 2004), liver (Nishikawa et al, 2001), breast (Richard et al, 2000;Tan et al, 2002;Zhu et al, 2005), uterine cancers (Pejovic et al, 2004) as well as chromophobe renal cell carcinoma (Nagy et al, 2002). Of all mtDNA mutations reported in cancer tissues, only a few are known to be of pathological relevance as shown for patients with disorders of the mitochondrial energy metabolism.…”