Research highlights on contributions of mitochondrial DNA microsatellite instability in solid cancers - an overview.

Head and neck squamous cell carcinomas (HNSCC), which refer to malignancies of the oral cavity, pharynx, and larynx, are the sixth most common cancer by incidence worldwide. Mitochondrial MSI in HNSCC has been studied by several teams. In the case of premalignant head and neck lesions, Ha et al . analysed the mtMSI in 93 patients and uncovered D310 sequence aberrations at a frequency of 36.6% in lesions [ 127 ]. Surprisingly, the author noted that the frequency of mtMSI increased according to the disease progression from benign hyperplasia (21.9%), mild dysplasia (33%), moderate dysplasia (36%), and severe dysplasia (50%), to carcinoma in situ (61.5%). Therefore, these aberrations might be used as markers of clonal progression premalignancy to cancer. Also, the authors examined mtMSI at 6 loci in patients that comprise metachronous and synchronous lesions and found that most cases displayed a clonal relationship between these lesions. In 2003 a study that involved the entire mtDNA mutation screening using temporal temperature gradient gel electrophoresis and followed by sequencing in oral cancers, Tan et al . reported that 44.4% of the cases (8/18) were found to have insertions or deletions in the D310 repeat [ 128 ]. In the subsequent year, Poetsch et al . sequenced 2 parts of the D-loop as well as MTND1 and MTND5 genes in 67 primary HNSCC from 56 patients [ 129 ]. They similarly demonstrated D310 repeat instability in 42% of the tumours. The nMSI at IGFIIR, hMSH3, hMSH6, and 5 other dinucleotide repeats were also tested on the same tumours for comparison. The frequency of nMSI (36% low nMSI and 13% high nMSI) determined in this series was less than mtMSI, and no correlation existed between them. In a large series of 109 HNSCC cases from the Laennec Hospital (Paris, France) was conducted by Lièvre et al . in 2006 [ 130 ]. This group revealed a rate of 17.4% (19/109) cases harboured alterations in the D310 repeat. In a more recent study, Shu et al . analysed 10 mtDNA microsatellite markers on 8 tongue squamous cell carcinomas, adjacent non-cancerous tissue, and peripheral blood samples from 8 Chinese patients [ 131 ]. The authors determined 25% (2/8) of mtMSI, and they were localized in the D310 region.

Despite impressive worldwide progress in the field of cancer diagnosis, prevention, and treatment strategies, cancer is still one of the leading causes of death globally. This is because cancer development and progression are driven by multiple molecular alterations that arise at the genomic and epigenomic levels [ 1 ]. This multi-step event has been termed the “hallmarks of cancer”, affecting both the nuclear and mitochondrial genome. Mitochondria have been acknowledged as extremely dynamic organelles where their biogenesis and bioenergetic functions are closely monitored and controlled by the nuclear genome [ 2 ]. Almost a century ago, a team led by Otto Warburg described the very first theoretical origin of cancer, i.e. that cancer formation might result from altered mitochondrial metabolism [ 3 ]. Since then, researchers have made remarkable progress in every aspect of the mitochondria in understanding how defective mitochondria can contribute to cancer development and progression.

Mitochondrial MSI has also previously been examined by some groups in several other solid cancers such as lung cancer, brain cancer, prostate cancer, bladder cancer, renal cancer, and gallbladder cancer. Fifty glioma paraffin-embedded tissues were evaluated by Kirches et al . using a combination of laser microdissection and PCR technique for D-loop poly-C tract alteration analysis [ 90 ]. The authors determined only 9% of mtMSI frequency in analysed specimens. Also, they conducted direct sequencing of the whole D-loop of 17 frozen glioblastoma samples and matched blood and revealed that 6 of these tumours harboured mtMSI at other repeats in the D-loop region (35.3%). More recently, we detected mtMSI in 20% of a series of 40 primary brain tumours. We observed a total of 8 different alterations within 2 loci, D310 and D16184 , of mtMSI markers [ 141 ]. Displacement-loop sequence alterations were examined by Chen et al . in 16 patients with prostate cancer [ 142 ]. Deletions or insertions appeared in 4 nucleotide positions; 50% of the patient samples showed a mutational hotspot for mtMSI at D310. In 2012, the study of Ashtiani et al . involved sequencing the complete mtDNA D-loop of 40 Iranian prostate cancer cases and benign prostatic hyperplasia patients [ 143 ]. They found the insertion of 1 or 2 nucleotide C at D310 was more frequent in both cancer and benign prostatic hyperplasia groups. Several groups of researchers have addressed mtMSI in lung cancer. The study conducted by Sanchez-Cespedes et al . discovered D310 mtMSI in 16% of 100 lung tumours [ 75 ], meanwhile Lee et al . determined in 13% of 31 tumours [ 99 ]. In 2003, Suzuki et al . amplified a 336-bp amplicon of the D-loop that contained D310 to detect polymeric C tract instability in 55 non-small cell lung cancers [ 144 ]. About 20% (11/55) of the tumours harboured these instabilities. Furthermore, the authors also determined that about 50% of D-loop alterations in 28 lung cancer cell lines occurred in this polymeric C tract. In a previous study, the mtMSI of 37 lung carcinoma and paired adjacent normal lung tissues from Chinese patients were analysed by Dai et al . using PCR-SSCP assay and sequence analysis [ 145 ]. It was reported that the percentage of mtMSI in lung carcinoma samples was 32.4% (12/37). Also, the lung carcinoma samples that harboured mtMSI were demonstrated to be correlated with a lower average of mtDNA content compared to cancer tissues without mtMSI. In 2015, an analysis of 48 patients with adenocarcinoma and 42 with squamous cell carcinoma of the lung from Dongsan Medical Centre, Korea uncovered mtMSI at a frequency of 20.8% and 19%, respectively [ 71 ]. The occurrence of D310 alterations in gallbladder cancers has been extensively studied by Tang et al . in 2004 [ 146 ]. Genetic analysis of archival gallbladder tissues from 123 tumours, 53 dysplasias, 90 histologically normal tissues adjacent to cancer, and 15 normal gallbladders were performed using PCR and sequencing assay for D310 alterations. This study reported a frequency of D310 alterations in the tumour, dysplastic lesions, and normal adjacent tissues of 38.2%, 57.1%, and 45.5%, respectively. Moreover, these D310 insertions/deletions were not only identified in cancer specimens, but the authors also found them in dysplastic and normal epithelial-derived from surrounding chronic cholecystitis (non-tumor) specimens. These outcomes proposed that there might be an early role of D310 alterations in the progressive series pathogenesis of gallbladder cancer. Wada et al . published the work regarding mtMSI in the urinary bladder and renal cancers in 2006 [ 147 ]. They examined genetic alterations in the mtDNA D-loop region as well as oxidative DNA damage by evaluating the level of 8-hydroxy-2’-deoxyguanosine (8-OHdG) in a panel of 31 bladder and 21 renal cancers in Japanese patients. Overall, 12.9% of bladder and 14.3% renal cancer cases had mtMSI, particularly at D310. Importantly, the 8-OHdG content was significantly increased in the cancerous renal tissues as compared with the paired non-cancerous tissues. Nevertheless, many tumour cases with mtMSI harboured lower 8-OHdG content than mtMSI-negative tumours. A 2021 recently published report by Alam et al . investigated the possible mtMSI in skin cancer, which comprised 57 patients with keratoacanthoma (KA) and 43 patients with squamous cell carcinoma (SCC) [ 148 ]. This report determined that mtMSI rates of 3.5% (2/57) of KA patients and 18.6% (8/43) of SCC patients, respectively. The authors also revealed that the mtMSI spot/location was found to be significant with tumour type, whereby the D514 alterations were detected in KA patients, and only SCC patients harboured mtMSI at D310.

Worldwide, breast cancer is the most diagnosed cancer among women and one of the most significant causes of death among them. mtMSI frequency in breast cancer has ranged from 10.8% to 42.5% in various studies. The initial study on the occurrence of mtMSI in breast cancer was published in 2000 by Richard et al . [ 69 ]. Tumour and matched normal tissues of 40 patients were analysed for the D514 microsatellite site and 4 Mnl I sites located between the 16,108- and 16,420-bp positions of the D-loop region. They revealed a Mnl I site alterations frequency of 47.5% in the tumours and this rate was 216-fold greater than the spontaneous rate in the female germline. Furthermore, the mtMSI rate was shown to be 42.5%, which indicated a 16-fold higher rate of spontaneous mutations. Mitochondrial DNA genome D-loop region, particularly at D310, was observed to be a target area that is mostly found altered in breast cancer. In 2001, Parrella et al . performed direct sequencing analysis to investigate the frequency and distribution of mtDNA mutations in 18 primary breast cancers [ 117 ]. Eleven tumours were reported to harbour 12 somatic mutations at a mutational frequency of 61%, with 5 alterations in a homopolymeric C-stretch at D310. Moreover, a rapid PCR-based assay revealed 7 more D310 alterations in 46 additional cases of breast tumours, which resulted in a D310 alteration rate of 19% (12/64) in overall breast tumour cases. In 2003, a further study of D310 instability was carried out by Parrella et al . in 4 different tumour types, which included 20 cases of breast tumour [ 118 ]. The author later determined a 15% D310 abnormality percentage in breast tumour cases. In another study, Alazzouzi et al . carried out an extensive experiment to investigate the alternative patterns of microsatellite alterations in cancer cells by analysing mtMSI as well as nMSI in breast ductal adenocarcinomas [ 119 ]. They revealed results in which no MSI was identified in any of the dinucleotide and mononucleotide repeats of the nuclear genome in these tumours. Also, the frequency of mtMSI was determined to be 10.8% with the alterations that occurred in the transcription control region. None of the mitochondrial alterations was observed in the normal matched tissues, showing that this mtMSI appeared as sporadic somatic mutations in the tumour cells. In 2006, Wang’s group screened the stability status of 12 mtDNA microsatellite loci in a series of common female cancers and revealed that 29.4% had a positive instability in breast cancer [ 120 ]. Instability was discovered at 3 of 12 microsatellite markers, which were located in the D-loop. According to their data, all mtMSIs in breast cancer were reported at nucleotide positions D310, D514, and D16184 , with rates of 13.7%, 7.8%, and 15.7%, respectively. In a different study, Tseng et al . analysed somatic mutations in the D-loop region in 60 Taiwanese women patients by using direct DNA sequencing [ 121 ]. They noticed that 30% (18/60) of tumours had somatic mutations, with the majority of alterations (13/18 = 72.2%) being in npD310 poly-C tract. The genomic instability in both the nuclear and mtDNA of 64 sporadic breast cancer and paired adjacent non-cancerous breast tissues from Brazilian patients were analysed by Santos et al . using nuclear STR loci genotyping and direct sequencing of the HVI and HVII regions [ 122 ]. It was reported that 42.2% of cases had somatic mtDNA alterations, with the most prevalent base change being at region D310 poly-C (17/64 = 26.6%). Additionally, about half of cancer cases harboured nMSI, with the majority at D13S790 loci for the 13q31 region. The authors concluded that instabilities at the nuclear 13q31 locus and in mtDNA may play an important role in the development of breast cancer. There are also a few studies that report a negative finding of this mtMSI in breast cancer patients. In 2013, Tipirisetti et al . published the results of a study in which mtMSI was not uncovered in any of the 11 mtDNA microsatellite loci analysed in breast cancer patients except at D310, which indicated presumably mutational hotspots rather than mtMSI [ 123 ]. This finding is consistent with an earlier study by Tan et al ., who reported that no MSI were found in all breast tumours and their matched normal tissues [ 124 ].

The prevalence of mtMSI in gastric cancers is approximately 10-63%. An initial report of mtMSIs in gastric cancer was demonstrated by Habano et al . in 2000 [ 68 ]. In their study, 10 out of 62 (16.1%) Japanese gastric carcinoma cases had the mtMSI phenotype in the D-loop region. They believed that the mtMSI phenotype exhibited in some gastric carcinoma cases might play a significant role in tumourigenesis. In the following year, a study performed by Máximo et al . identified mtMSI in 18.8% of 32 gastric carcinomas [ 89 ]. In the same year, Sanchez-Cespedes et al . reported a higher frequency of mtMSI (62.5%) in primary gastric tumour patients [ 75 ]. This observation was further supported by a report from Han et al . [ 109 ]. They analysed 22 patients with gastric tumours in China and discovered that 59.1% of tumours and 40.9% of the adjacent normal tissues carried the D310 or D16184 instabilities. In the year 2003 Ling et al . observed a 36.7% (11/30) frequency of mtMSI in gastric cancer [ 110 ]. They also demonstrated that mtMSI-positive gastric cancer had significantly higher interleukin-8 activity levels than mtMSI-negative gastric cancer. One year later, the same group extended the study by examining 68 cases of advanced gastric cancer, 40 cases of chronic gastritis, 30 cases of intestinal metaplasia, and 20 cases of dysplasia. They found that a progressive accumulation of mtMSI occurred in the histological progression from chronic gastritis to cancer through intestinal metaplasia and dysplasia (12.5−38.2%) [ 111 ]. Interestingly, this study also observed a statistically significant association between mtMSI with intestinal-type and distal location of gastric cancer. In 2005, 3 groups of researchers from Taiwan, Japan, and China examined mtMSI in gastric cancer. Lee et al . reported the frequency of mtMSI to be 32.3% in 31 gastric cancers with alterations in the mononucleotide and dinucleotide repeat of D310 [ 99 ]. Kose et al . published their report on using microsatellite assay investigating 96 Japanese patients and identified an mtMSI rate of 15% at the D310 region [ 98 ]. Zhoa et al . analysed the D-loop region mutation in 20 Chinese specimens and found sequence mutations in 35% of the specimens including 4 microsatellite instabilities in the mutations [ 112 ]. In 2006, Kose’s team expanded the microsatellite instabilities study by examining 74 gastrointestinal stromal tumours for mtMSI at D310 as well as nMSI at BAT26 [ 113 ]. The authors noted that the MSI percentage at D310 was 16%, higher than that at BAT26 (5%), despite the lack of statistical significance between them in gastrointestinal stromal tumours. The first endeavour to explore a possible connection between the RUNX3 promoter methylation, nMSI, and mtMSI in gastric cancer was attempted by Gargano et al . and was published in 2007 [ 114 ]. One hundred cases of gastric cancer were selected for the nMSI and mtMSI analysis using the National Cancer Institute guidelines (the Bethesda-recommended panels of microsatellite markers) and D310, respectively. According to their work, mtMSI was identified with a frequency of 11% while high-level nMSI was detected in 13% of cases. This study also highlighted that mtMSI was significantly associated with tumour-node-metastasis staging. Furthermore, an essential correlation between MSI-H status, mtMSI, and RUNX3 methylation was demonstrated, proposing a potential connection between these events in gastric carcinogenesis. In a series of 49 gastric cancer patients from the general surgery division at Dongsan Medical Centre, Jeong et al . reported a mtMSI rate of 10.2% [ 115 ]. Moreover, this study also showed that all mtMSI were significantly observed in intestinal-type cancer. Interestingly, the author also noted a similar frequency of mtMSI occurring in gastric dysplasia as a precancerous lesion (12.5%, 3/24), suggesting that mtMSI might contribute to the early stages in the progression of gastric carcinogenesis. In the year 2016, Ling et al . explored the association of mtMSI with Helicobacter pylori (H. pylori) infection and the level of interleukin-8 (IL-8) in gastric cancer [ 116 ]. They published the results of their analysis regarding mtMSI in 122 samples of gastric carcinomas and found a mtMSI rate of 23%. According to their study, an important significant association was revealed between mtMSI, H. pylori infection, and increased IL-8 level. The authors concluded that mtMSI might play a critical role in different mechanisms of gastric cancer development. In a recent study, Wang et al . used a sequencing assay to detect mtMSI in the whole mtDNA D-loop in 69 gastrointestinal cancers in Chinese patients, of whom 18 patients had gastric cancer [ 97 ]. They reported a rate of mtMSI in the D16184 , D514, and D310 regions of 11.7%, 37.2%, and 40.3%, respectively.

Thyroid cancer is the most common neoplasm of the endocrine system, and its incidence has been steadily rising worldwide. Several studies have been carried out to explore the occurrence of mtMSI in thyroid cancer. The first manuscript to be reported concerning mtMSI in thyroid cancer was published in 2002 by Lohrer et al . [ 132 ]. Two series of tumour tissues from 126 radiation-associated and 40 sporadic thyroid tumour patients were analysed for the instability of the hypervariable region II of the mtDNA. The authors revealed an extremely low frequency of D310 poly-C tract in 2 out of 126 radiation-associated tumours from Belarus and 6 out of 40 tumours from Munich. Nevertheless, the frequencies were found to be associated with the patient’s age. A lower percentage of the mtMSI in thyroid cancer has also been reported by Tong et al . [ 133 ]. This study involved the analysis of D310 region alterations using PCR-based methods in 72 thyroid cancers, which included papillary, medullary, anaplastic, follicular, and insular thyroid carcinoma. The overall rate of D310 instability was shown to be 6.9%. In 2005, a study of 66 thyroid tumours comprising benign adenomas, follicular, and papillary carcinomas from a Portuguese series by Máximo et al . further revealed a high rate of mtMSI [ 93 ]. These mtMSIs were not limited to cancer cases only but were also identified in benign lesions. The authors detected 48.5% (32/66) of thyroid patients contained somatic instability in D-loop microsatellite regions with the most occurring at D310 (30.3%, 20/66), followed by D514 (22.7%, 15/66), and D568 mononucleotide repetitive (4.6%, 3/66). In a panel of 101 patients with malignant thyroid carcinomas or non-tumour nodular goitres (benign thyroid adenomas) from the Wenzhou area, Ding et al . discovered a mtMSI rate of 10.4% (8/77) in malignant thyroid carcinomas and 10.5% (2/19) in nodular goitres [ 134 ]. This study indicates that mtDNA alterations could play a pivotal role during the initial stage of thyroid tumorigenesis. In 2019, Bircan et al . observed a higher frequency of D310 somatic mutations in the hot thyroid nodules group (22.2%) than in cold thyroid nodules (10.5%) and papillary thyroid carcinoma samples (10.4%) [ 135 ]. Also, a higher percentage of the mtMSI was found in D16184 (20.8%) and D514 (25%) of papillary thyroid carcinoma. They concluded that D310 instability might be utilized as a diagnostic clonal expansion biomarker for premalignant thyroid tumour cells and considered D514 as a prognostic biomarker for benign to malign transformation in thyroid tumours [ 135 ].

Globally, colorectal cancer has been ranked in third place among the commonest malignancies, and based on the World Health Organization GLOBOCAN database there are over 1.84 million predicted with new cases and around 880,792 deaths in 2018 ( https://gco.iarc.fr/today/ ). Mitochondrial MSI is commonly studied in colorectal cancer, and about 8–69% of patients with colorectal cancer have mtMSI. The first study on the involvement of MSI in the mitochondrial genome of colorectal carcinomas was reported by Habano et al ., who screened 9 mtMSI sites in 45 sporadic colorectal carcinomas and revealed a mtMSI frequency of 44% (20/45) [ 47 ]. They also identified 3 cases with frameshift mutations that corresponded to polyadenosine (A)8 or polycytidine (C)6 tract in the mitochondrial NADH dehydrogenase ( ND ) genes ( ND1 and ND5 ), which were predicted to result in truncated of complex I proteins. One year later, the same group extended the study by analysing the entire mtDNA amplification scanning in the same series of 45 carcinomas [ 96 ]. They found ND1 or N5 gene alterations in 15.6% (7/45) of cases, which included 3 frameshift mutations in the mononucleotide repeat sequences, 2 missense mutations, and one small 15-bp deletion. Six of 7 cases with mutations also presented mtMSI of the (C)n sequence in the D-loop region [ 96 ]. Studies by Venderbosch et al . [ 30 ] and Wang et al . [ 97 ] have shown a high frequency of mtMSI, at 54.4% and 82.4%, respectively. However, in contrast to these, in a study by Kose et al . only 8.3% (11/133) of mtMSI was identified in a large panel of 138 colorectal cancer samples [ 98 ]. The frequency of mtMSI in colorectal cancers also varies from community to community. Mitochondrial MSI frequency is 36%, 8.3%, and 43.2% in Taiwanese [ 99 ], Japanese, and Vietnamese patients [ 100 ], respectively. Lim et al . also reported a high frequency of mtMSI 46.3% (25/54) in Korean colorectal cancer, which mainly occurred in D310 (35%) and D16184 poly C (19%) [ 101 ]. In another study, Lee et al . in 2015 [ 102 ] reported a 30% frequency of mtMSI in the Korean group of colorectal cancer, which was similar to that found by Tsai et al . [ 103 ]. Also recognized as a mutational hotspot in some tumour types, mtMSI in the D310 site is commonly found in several types of colorectal carcinomas. In the year 2001, Sanchez-Cespedes et al . were the first to uncover D310 as a mutational hotspot in several kinds of primary tumours [ 75 ]. A study by Sanchez-Cespedes et al . that analysed 25 colorectal tumours showed that 28% of tumours harboured D310 mtMSI. In 2005, Lièvre et al . conducted an experiment to investigate the presence of D-loop mutations in a series of 365 colorectal cancer patients and found that 36.2% of the cases carried alterations in the D310 microsatellite sequence [ 104 ]. They also demonstrated that the prevalence of D310 alterations significantly increased with the number of cytosines in the sequence, in which C 7 TC 6 was 9% (16/178), C 8 TC 6 was 49.6% (56/113), and C 9 TC 6 was 73.3% (11/15), respectively. While, in the same year, D310 alterations together with repeats in the ND1 and ND5 genes were examined by Guleng et al . in a group of 95 colorectal cancer patients and 95 control subjects [ 70 ]. The authors detected that 34% of the carcinomas had D310 alterations, while there were only 2% in the control group. In addition, they failed to detect any mtMSI in the ND1 and ND5 genes and concluded that D310 instability was not correlated with nMSI in their cases [ 70 ]. In another study, also in the same year, Lee et al . analysed the nucleotide sequence of the D-loop as well as mtDNA content in a series of solid tumours that comprised of 25 colorectal cancers [ 99 ]. Impressively, the results indicated that 40% (10/25) of the cases studied had mutations in the D-loop with a 90% (9/10) rate of altered D310 microsatellite sequence. They proposed that mtDNA instability, in conjunction with the diminish in the mtDNA content, plays a crucial role in human carcinogenesis [ 99 ]. In 2006, Kim et al . used PCR-SSCP and DNA sequencing analysis to examine 48 micro-dissected cancer epithelia and adjacent stroma of sporadic colorectal cancer [ 105 ]. They identified that 27.1% (13/48) of cancer epithelia and 10.4% (5/48) of stroma contained D-loop mtMSI. Three years later, Pinheiro et al . aimed to analyse D310 as well as ND1 and ND5 for MSI in a series of 38 rectal carcinomas in comparison with 25 sigmoid carcinomas [ 106 ]. They revealed a D310 MSI frequency of 34.3% and 37.5% of rectal and sigmoid carcinomas, respectively, while ND5 alterations were observed in 5.3% and 8% of rectal and sigmoid carcinomas, respectively. Moreover, only rectal carcinoma cases were reported to harbour ND1 alterations at an even lower frequency, with a MSI frequency of 2.6%. The prevalence of D310 alterations in colorectal adenomas and liver metastases of colorectal cancers were reported in 27.4% (17/62) and 33.3% (5/15), respectively, in a study performed by Legras et al . [ 107 ]. Their investigations were continued with a case-control study by assessing the possible role of D310 polymorphism in colorectal adenoma risk. The authors proved that germline D310 polymorphism might not be a risk factor for colorectal adenomas. Nevertheless, they suggested that D310 mutation could be considered as a potential biomarker for the early detection of colorectal cancer. Several studies have evaluated the correlation of mtMSI with clinicopathological parameters of colorectal cancer. Lee et al . found mtMSI in 30% of 100 colorectal cancers and later demonstrated that patients with mtMSI had shorter overall survival [ 102 ]. In a 2017 study carried out by Kleist et al . detected mtMSI in 37.5%, and this was the first to show that D-loop MSI was significantly associated with TP53 wild-type status of primary colorectal cancer [ 78 ]. Additionally, D-loop MSI was not associated with gender, clinical stage, tumour location, survival or KRAS, NRAS, BRAF, or PIK3CA mutations. Several studies have investigated the prognostic value of mtMSI in colorectal cancer; however, their results were considered controversial. In a previous study, Chang et al . reported that mtMSI had no prognostic effect in colorectal cancer patients [ 108 ]. In contrast, a study by Tsai et al . revealed that Dukes’ stage C colorectal cancer patients with mtMSI had better disease-free and overall survival [ 103 ]. They concluded that mtMSI provides a good prognosis in Dukes’ C colorectal cancer. In a study by Lim et al ., it was shown that there was a higher frequency of mtMSI in larger size tumours and more advanced tumor node metastasis stage, indicating mtMSI as a risk factor for unfavourable outcome [ 101 ].

Over the past decades, the nDNA has been the focus of studies in the search for cancer susceptibility genes; nevertheless, accumulating evidence now converges to demonstrate that mtDNA may also play an essential role in the genetic aetiology and pathophysiology of cancer. The present review provided a brief overview of several aspects of the mitochondrial genome and then summarized evidence of previous studies concerning the role of mtMSI in most solid cancers. The study of mtDNA alterations in the context of cancer progression has been recognized for centuries, and there are a vast number of outstanding questions and opportunities in the area that require further investigation. According to cumulative evidence generated from previous studies, this review indicates that mtMSI could contribute to increased cancer risk. However, it is still an ongoing concern whether these instabilities are merely reflecting secondary epiphenomena of the underlying cancer process or comprise a true causative connection. Future larger prospective studies should be conducted to further elucidate the role of mtMSI in solid cancers.

Worldwide, oesophageal cancer is the sixth leading cause of cancer mortality. Each year ~572,034 new oesophageal cancer cases are diagnosed and > 508,000 patients die of oesophageal cancer. Similarly to hepatocellular cancer, less attention and research has been focused on the mtMSI in patients with oesophageal cancer. In 2004, Kumimoto et al . sequenced the HVR-I and HVR-II regions of mtDNA from 38 primary oesophageal tumours and matched normal tissues for the somatic mutation rate [ 76 ]. They discovered 14 somatic mutations in 13 patients, and a 28.9% (11/38) mtMSI rate was noted at region D310 poly-C stretch. In another study that involved a series of 82 Japanese cancer cases, Kose et al . reported a prevalence of mtMSI in the D310 region of 14% [ 98 ]. A previous study involving oesophageal squamous cell carcinomas among Taiwanese patients conducted by Lin et al . determined a high rate (77.8%, 56/72) of somatic D310 alterations [ 125 ]. The author also claimed that these variants were significantly correlated with a poorer prognosis. Two years later, the same group extended their research by examining 66 thoracic oesophageal squamous cell carcinomas for mtMSI and nMSI using microsatellite analysis for markers D310 and D17S960 ( TP53 ), respectively [ 126 ]. They observed a higher frequency of 66.7% of samples with the D310 alterations. Moreover, only 6.1% of samples harboured nMSI at D17S960.

In 1998, Habano et al . [ 47 ], pioneers in the research of mtMSI in cancer, mainly in colorectal cancer, demonstrated that more attention must be directed toward the role of the mitochondrial genome in carcinogenesis. Since then, many studies have been conducted, determining the link between mtMSI and various types of cancer ( Table 1 ). Summary of studies in which mitochondrial microsatellite instability was examined in some types of solid cancers AII – astrocytoma WHO grade II, AIII – astrocytoma WHO grade III, ADC – adenocarcinoma, BC – bladder cancer, BTCC – bladder transitional cell carcinoma, CC – colon cancer, CTNs – cold thyroid nodules, D310 – (poly-C)n, between np 303-315, D514 – (CA)n, between np 514-523, D568 – (poly-C) n, between np 568-573, D16184 – (poly-C)n, between np 16184-1619, GBM – glioblastomas, HTNs – hot thyroid nodules, KA – keratoacanthoma, mtMSI – mitochondrial microsatellite instability, PTCs – papillary thyroid carcinomas, RC – rectal carcinoma, RLC – renal cancer, rTT – radiation-associated thyroid tumour, SC – sigmoid carcinoma, SCC – squamous cell carcinoma, sTT – sporadic thyroid tumour

Gynaecological cancer is a major health burden globally for women, and approximately 70,500 women are diagnosed with this cancer each year. The 3 main types of gynaecological cancer are cervical, endometrial, and ovarian. The frequency of mtMSI in gynaecological cancer ranges approximately between 20% and 50%. The mtMSI was first described in ovarian cancer samples (20%, 3/15) by Liu et al . in 2001 [ 91 ]. Two years later, a follow-up study of mtMSI was conducted by the authors on 50 samples of primary endometrial carcinomas, revealing a higher rate of mtMSI (50%), particularly detected in 3 different regions (D310, D514, and D16184 ) of the D-loop region and one in the 12S RNA gene [ 73 ]. In the year 2005, Wang et al . investigated the contribution of mtMSI in a series of gynaecological cancers [ 80 ]. In this large study that involved 262 pairs of female cancers and matched normal tissues, mtMSI was analysed for 12 microsatellite regions in primary cervical, endometrial, ovarian, and breast carcinomas. Most of the mtMSI belonged to D-loop microsatellite loci (95.6%), particularly in D310, D514, and D16184 . A year later in a follow-up study, Wang et al . assessed mtMSI in another 257 samples of common female cancers, included cervical, endometrial, ovarian, and breast cancers [ 120 ]. In agreement with their previous findings, the authors reported the greatest frequency of mtMSI in 48.4% of endometrial cancers followed by 29.4% of breast cancers, 25.4% of cervical cancers, and 21.9% of ovarian cancers. A 2009 study of 24 cervical carcinomas by Chen and Zhan demonstrated that 9 patients harboured 30 alterations in the D-loop region; 8 (27%) of these changes were mtMSI (8 mtMSI in 6 patients, 25%) [ 138 ]. In another study of 40 cervical cancer specimens, Zhao et al . reported that 21 mutations were detected in 12 patients, where there were 4 mtMSI in the mutations [ 139 ]. Mitochondrial MSI has also been studied in endometriosis patients by Govatati et al ., who claimed that less frequently mtMSI occurred in endometriosis cases [ 140 ].

Hepatocellular carcinoma (HCC) is believed to be the most common primary cancer of the liver and represents the third most common cause of cancer death and the sixth in incidence worldwide. So far, there have been very few studies carried out on the link of mtMSI in hepatocellular carcinogenesis. One of the earliest studies was published in 2002 by Nomoto et al . reported a D310 poly-C tract deletion/insertion alterations frequency of 42.1% (8/19) in Japanese HCC cases [ 136 ]. In 2004, and in an effort to report the frequency of mtMSI and nMSI in 52 HCC patients, a study was reported from China by Fang et al . [ 137 ]. A rate of 21.2% (11/52) and 5.8% (3/52) of HCC harboured mtMSI and nMSI, respectively. The authors stated that no association was found between both MSIs in the cases of HCC. In a further study that involved a series of solid tumours from Taiwanese patients, Lee et al . published an 18.5% (10/54) mtMSI frequency in HCC cases [ 99 ].