Discussion
Detection of nuclear expression of MLH1, MSH2, PMS2 and MSH6 proteins by immunohistochemistry (IHC) is the gold standard for identifying MMR status, with nuclear loss of at least one of these markers within tumour cells considered dMMR.143–145 Notably, dMMR is increasingly being detected indirectly by somatic MMR gene alterations on NGS, and small studies have demonstrated concordance with IHC MMR protein loss.146 147 To determine MSI status, earlier studies employed PCR while more contemporary studies used NGS. The established Bethesda PCR panel consists of five mononucleotides (BAT25, BAT26, D2S123, D5S346 and D17S250), of which at least two must be altered to diagnose MSI-H.148 149 Although developed for use in colorectal carcinoma, the panel has been used in other solid tumours including UC to detect MSI status.47 87 95 When only one locus was altered, which defined microsatellite instability-low (MSI-L), the clinical and biological implications were equivocal. Additional mononucleotide repeats in the panel were suggested to increase sensitivity.149 NGS studies including whole exome sequencing and targeted sequencing assays can determine MSI status by sequencing around microsatellite regions with comparison between tumour and normal tissue.13 150–153 In large cohorts, validation with PCR and IHC showed 99.1%–99.4% concordance.154 155 NGS-based methods are more sensitive as they are capable of assessing hundreds to thousands of loci, compared with a limited panel with PCR, which reduces the likelihood of detection of MSI-L tumours.153 This overcomes the possible subjectivity that arises from using PCR by providing quantitative results. However, since the majority of MSI is caused by epigenetic changes, detecting these by NGS requires more complex analysis and interpretation.
Notably, the pooled weighted prevalences of MSI-H and somatic dMMR gene alterations were higher in localised BC (stage M0) compared with metastatic BC, and in UTUC, the pooled weighted prevalence of MSI-H was also higher in localised disease. The higher prevalence of these characteristics in localised disease has also been observed in colorectal cancer156 157 and may be due to higher neoantigen load of MSI-H/dMMR tumours stimulating an antitumour immune response; hence reducing the likelihood of metastases.157 Interestingly, there was also a difference in the pattern of MMR protein loss in UTUC compared with BC. In UTUC, MSH2 and MSH6 were the most frequent MMR proteins and/or genes lost or altered (both somatic and germline), whereas the trend was not as clear in BC. This contrasts with dMMR colorectal cancer, in which 90% of cases are due to loss of MLH1 and MSH2 genes.158 159
We found a higher prevalence of germline MMR gene alterations in patients with UTUC or a positive family history; hence, germline testing should be discussed in these situations. In colorectal cancer, in which the prevalence of Lynch syndrome is also relatively low, around 5%,160 universal germline testing is recommended.161 The increasingly routine performance of NGS, especially in metastatic UC, will identify more patients with either MSI-H or somatic MMR gene alterations, which will likely result in more germline testing being performed. Identification of a germline MMR gene alteration will have significant implications for patients and their families. A prior study showed that the standardised incidence ratio of developing BC was 8.2-fold and 16.2-fold higher in males and females, respectively, harbouring MMR germline variants compared with the general population.162 The risk was highest for MSH2 germline carriers. Moreover, the risk of developing subsequent urothelial cancers was >100 fold higher in MMR germline carriers. Another study showed that in patients harbouring pathogenic Lynch syndrome-associated variants, the risk of developing urinary tract tumours by age 75 was 24.9% for MSH2 carriers, 11% for MSH6 carriers and 8% for MLH1 carriers. In comparison, risks of developing colorectal cancer were 43%, 15% and 45.8%, for MSH2, MSH6 and MLH1 carriers, respectively.163 For individuals with germline MMR gene alterations, the National Comprehensive Cancer Network guidelines do not recommend routine surveillance for UC due to lack of clear supporting evidence, although it is recommended for individuals with a family history of UC. The considerably higher risk of developing BC, especially among MSH2 carriers, could inform surveillance guidelines. For colorectal cancer, the detection of germline MMR gene alterations has clear surveillance implications, including earlier and more frequent colonoscopies.161
We found that dMMR and MSI-H metastatic UC is responsive to ICI, with included studies reporting ORRs of 50%–90%,35 36 39 101 deep responses35 36 39 101 121 126 164 and long durations of response121 123 126 132 (online supplemental tables S3 and S4). These data show that dMMR and MSI-H may be predictive biomarkers for response to ICI in UC, which has already been prospectively demonstrated in solid tumours.20 Indeed, pembrolizumab received accelerated FDA approval for use in any metastatic solid tumours with dMMR or MSI-H based on clinical activity demonstrated in the phase 2 Keynote-158 trial.20 23 The response of dMMR/MSI-H metastatic UC to chemotherapy is less encouraging, with high rates of primary progression101 122 132 133 and short PFS.101 The high rates and long durations of response to ICI and the lower likelihood of disease control with chemotherapy shown in this systematic review raise the question of whether these patients should receive ICI monotherapy upfront for metastatic disease. This question remains even with the standard of care for untreated metastatic UC recently changing to enfortumab vedotin (EV) plus pembrolizumab as it is unknown whether this subset of patients will do just as well receiving the agents sequentially instead of concurrently, which will limit or delay treatment related toxicity.165 166 For localised disease, there is conflicting evidence. Some studies suggest superior DFS compared with pMMR/MSI-L/MSS patients and high rates of local control67 77; other studies suggest no difference in outcomes.96 98 Some studies suggest a propensity for dMMR/MSI-H patients to have multifocal, bilateral and/or both bladder and upper tract disease, either synchronously or metachronously.67 77 A novel approach for the treatment of localised dMMR/MSI-H tumours which has seen impressive results in rectal24 and colon cancer25 26 is definitive treatment with ICI, known as immunoablation. In the phase 2 dMMR rectal cancer trial, there was a 100% clinical CR rate among the first 14 participants with stage II and stage III disease who were treated with the anti-PD-1 antibody dostarlimab, and no recurrences at a median follow-up of 6.8 months. None of the patients had required chemotherapy, radiotherapy or surgery up to the latest data cut-off; however, longer follow-up is needed to confirm the success of this approach.24 167 In the phase 2 NICHE-2 trial which enrolled 112 patients with locally advanced dMMR colon cancer, patients were given two cycles of ICI (first ipilimumab/nivolumab, then nivolumab) before proceeding to surgery. Remarkably, 95% of patients attained a major pathological response, defined as <10% residual viable tumour, and 67% of patients attained a pathologic CR.26 A systematic review assessing neoadjuvant ICI in 423 patients with dMMR/MSI-H localised colorectal cancer reported CR (pathological CR plus clinical CR) of 72% and a complete resection (R0) rate of 99.3%.168 These results suggest a need to develop similar trials in dMMR/MSI-H muscle-invasive or locally advanced UC, as these could revolutionise the treatment landscape for these patients. This approach may allow patients to avoid the quality-of-life impacts of radical surgery or trimodality therapy and potentially be effective for multifocal localised disease without the need for multiple procedures.
Survival in patients with dMMR/MSI-H may be favourable. In one study, all 10 patients with advanced or metastatic disease survived beyond 15.5 months after ICI39; in another study, 77% of 26 patients lived beyond 2 years.101 Univariate analyses have demonstrated superior OS in patients with dMMR/MSI-H compared with patients without these characteristics84 95 113; however, other studies show no difference in survival.96 109 Of note, most of these studies are small and retrospective, which increases the risk of bias. Comparatively, in localised and advanced colorectal cancer, MSI-H has been shown to be favourably prognostic for survival.169–171 In UC, it is likely that OS will depend on treatment for advanced disease. Newer studies with more patients receiving ICI may demonstrate a greater degree of improvement in survival compared with pMMR/MSI-L/MSS patients.
Study limitations
There was heterogeneity in MSI detection techniques among included studies due to their significant evolution over the last two decades. Consequently, the MSI-H prevalence data that were meta-analysed were likely not completely standardised. The focus of this systematic review was on dMMR and MSI-H in UC; however, our search results also yielded studies focusing on germline and somatic gene alterations, which we included due to their relevance to the topic. As this study was not primarily designed to search for genetic and genomic studies, our search may not have comprehensively included these studies. The response rates of dMMR/MSI-H patients to ICI and chemotherapy were pooled from separate studies; hence, they are only crude estimates. Lastly, the survival of patients with dMMR or MSI-H UC and the prognostic value of these biomarkers could not be estimated or meta-analysed due to a lack of uniformity in the data.
