Translational Research

EHW_4578.jpgCDK12 inactivation defines a new class of prostate cancer

We identified a new subtype of prostate cancer that occurs in about 7 percent of patients with advanced disease. The subtype is characterized by loss of the gene CDK12. It was found to be more common in metastatic prostate cancer compared with early stage tumors that had not spread. CDK12 mutant cases are associated with elevated neoantigen burden ensuing from fusion-induced chimeric open reading frames and increased tumor T cell infiltration/clonal expansion and thereby defines a distinct class of metastatic prostate cancers that may benefit from immune checkpoint immunotherapy.


Clinical genomics of metastatic cancer

Recently, we carried out a comprehensive molecular analysis of metastatic solid tumors of diverse lineage and biopsy site from 500 adult patients (MET500 cohort) by performing clinical-grade integrative whole exome (tumor/normal) and transcriptome sequencing. Sequencing matched tumor and normal samples from patients identified potentially pathogenic germline alterations as well as provided high resolution copy number landscapes. RNA sequencing analysis provided insights into the tumor lineage, functional gene fusions, transcriptional pathway activation, viral pathogen, and immune cell landscape. We found that the most prevalent genes somatically altered in metastatic cancer included TP53, CDKN2A, PTEN, PIK3CA, and RB1. Putative pathogenic germline variants were present in 12.2% of cases of which 75% were related to defects in DNA repair genes, most commonly occurring in BRCA1, BRCA2, CHEK2 and MUTYH. Tiering of the molecular alterations identified in metastatic cancers provided a rationale for clinical trial or registry study enrollment in 72% of cases and guideline based recommendations in 16% of cases. Our results demonstrate that integrative sequence analysis provides clinically relevant, multidimensional view of the complex molecular landscape and microenvironment of metastatic cancers. The manuscript presenting the analysis of the MET500 cohort has been published in Nature (2017 Aug 2. doi: 10.1038/nature23306).


Immunohistochemical Detection of HLRCC Markers

Dr. Rohit Mehra led a study on hereditary leiomyomatosis and renal cell carcinoma (HLRCC) that is caused by germline mutations in the FH gene, and is associated with increased incidence of leiomyomas and a potentially aggressive variant of renal cell carcinoma (HLRCC-associated RCC). Absence of fumarate hydratase (FH) expression as determined by immunohistochemistry has previously been used to diagnose HLRCC-associated RCC, but immunohistochemical staining of leiomyomas is not standard practice. Here, Dr. Mehra and colleagues performed immunohistochemistry (IHC) on whole sections from consecutive cutaneous leiomyomas from archived samples (96 samples from 87 patients) to evaluate for both FH and succinate dehydrogenase B expression, in addition to clinicopathologic data collection and review of all hematoxylin and eosin-stained slides for blinded morphologic evaluation of features reported to be seen in HLRCC-associated uterine leiomyomas. The overall sensitivity and specificity of negative FH expression in leiomyomas for detection of patients with HLRCC were 70.0% and 97.6%, respectively. Inclusion of cases classified as equivocal increased sensitivity to 75.0%. Succinate dehydrogenase B expression was retained in 95 specimens and equivocal in 1 specimen. None of the evaluated morphologic features showed any association with leiomyomas in HLRCC. Absent FH immunohistochemical expression in cutaneous leiomyomas is a sensitive and specific marker for detection of HLRCC, thus suggesting a role for prospective FH IHC in patients with these tumors to screen for HLRCC. The results of this study was published in Am J Surg Pathol (2017 Jun;41(6):801-809). 


Integrative Clinical Sequencing in the Management of Refractory or Relapsed Cancer in Youth

Through the pediatric arm of MI-ONCOSEQ (PEDS-ONCOSEQ), we published the results from the first 102 pediatric patients enrolled in our clinical sequencing study (JAMA, Vol. 314, No. 9, Sept. 1, 2015). A total of 91 pediatric patients with advanced or relapsed cancer underwent complete sequence analysis. Forty-two patients (46%) had actionable findings that changed their cancer management and individualized actions were taken in 23 patients based on actionable integrative clinical sequencing findings, including change in treatment for 14 patients and genetic counseling for future risk for 9 patients. Nine of the personalized clinical interventions resulted in ongoing partial clinical remission of 8 to 16 months or helped sustain complete clinical remission of 6 to 21 months. All 9 patients and families with actionable incidental genetic findings agreed to genetic counseling and screening. The study is the first to report on combined multiple genome sequencing approaches (tumor as well as normal DNA and tumor RNA) in real-time, in children and young adults with relapsed cancers.


Gene sequencing reveals mutations in the estrogen receptor

From the MI-ONCOSEQ cohort, we analyzed 9 ER-positive treated metastatic breast cancer patients; the samples were subjected to integrative sequencing including whole exome and transcriptome analysis that allows a mutational landscape of coding genes including point mutations, indels, amplifications, deletions, gene fusions/translocations, and outlier gene expression. The most remarkable observation in the mutational landscape of these treated ER positive patients was the finding of nonsynonymous mutations in the ligand binding domain (LBD) of ESR1 (n=4). The four index patients MO_1031, MO_1051, MO_1069, and MO_1129 had LBD mutations in amino acids L536Q, Y537S, D538G, and Y537S, respectively. All had been treated with anti-estrogens and estrogen deprivation therapies. A survey of The Cancer Genome Atlas (TCGA) identified 4 endometrial cancers with similar mutations of ESR1. The 5 novel LBD mutations of ESR1 identified here (L536Q, Y537S, Y537C, Y537N, and D538G) were shown to be constitutively active and continue to be responsive to anti-estrogen therapies in vitro. Taken together, these studies suggest that activating mutations of ESR1 are an important mechanism of acquired endocrine resistance in breast cancer therapy. Results of this study were published in the journal Nature Genetics (Nat Genet. 2013 Dec;45(12):1446-51).


Identification of targetable FGFR gene fusions in diverse cancers.

In four index MI-ONCOSEQ cases, we identified gene rearrangements of FGFR2, including patients with cholangiocarcinoma, breast cancer, and prostate cancer. We then extended the screening of FGFR rearrangements across multiple tumor cohorts and identified additional FGFR fusions with intact kinase domains in lung squamous cell cancer, bladder cancer, thyroid cancer, oral cancer, glioblastoma, and head and neck squamous cell cancer. All FGFR fusion partners tested exhibited oligomerization capability, suggesting a shared mode of kinase activation. Overexpression of FGFR fusion proteins induced cell proliferation. Two bladder cancer cell lines that harbor FGFR3 fusion proteins exhibited enhanced susceptibility to pharmacologic inhibition in vitro and in vivo. Because of the combinatorial possibilities of FGFR family fusion to a variety of oligomerization partners, clinical sequencing approaches that incorporate transcriptome analysis for gene fusions have the potential to identify rare, targetable FGFR fusions across diverse cancer types. (Cancer Discov. 2013 Jun;3(6):636-647).


MI-ONCOSEQ identifies rare cancer mutation

In the latest study led by Dr. Dan Robinson and published in the journal Nature Genetics, researchers utilized an integrative sequencing approach to identified a novel gene fusion (the shuffling and subsequent joining together of two separate genes in the genome causes “gene fusions” that can be an important cancer causing mechanism) as the driving mutation in solitary fibrous tumor. In this rare cancer for which there is no effective standard therapy the gene fusion, NAB2-STAT6, was found in 100% of the tumors examined. This important discovery identifies a specific target toward which novel drugs could be directed (Nat Genet. 2013Feb;45(2):180-5).


Mutational analysis of CRPC

We, along with colleagues from UM, Compedia Bioscience and Yale, undertook the task of analyzing the global mutational landscape of metastatic castrate-resistant prostate cancer (CRPC) by sequencing the exomes of pre-treated and treatment naïve metastatic CRPC patients. Results of this study revealed that the overall mutational rates in both groups were low, demonstrating a monoclonal origin of metastatic CRPC. We found that a deletion/mutation in the CHD1 gene (in 8% of cases) was associated with ETS gene fusion-negative status and recurrent mutations were found in several genes such as MLL2 (8.6 % of cases) that are involved in chromatin/histone modification. Novel recurrent mutations were also identified in FOXA1 gene (3.4% cases). Additionally, proteins that interact with the Androgen Receptor such as ERG gene fusion product, FOXA1, MLL2, UTX, and ASXL1 were discovered to be mutated in CRPC. The range of genetic aberrations identified here provides new insights into the evolution of resistance mechanisms in CRPC. (Nature. 2012 Jul 12;487(7406):239-43.). 


New urine test for prostate cancer

Our group along with external collaborators developed a new urine test that can aid in the early detection of and inform treatment decisions about prostate cancer. The test supplements the current prostate specific antigen (PSA) screen and could help some men delay or avoid a needle biopsy while identifying men at highest risk for clinically significant prostate cancer. The new test is designed to detect the gene fusion, TMPRSS2:ERG. Studies in prostate tissues show that the gene fusion almost always indicates cancer. But because the gene fusion is present only in about 50% of prostate cancer patients, we also included another marker identified earlier, PCA3. The combination was more predictive of cancer than either marker alone. Our goal is to incorporate this test in the clinical diagnosis of prostate cancer for early identification of metastatic disease that would require aggressive therapeutic intervention. We are working with Gen-Probe Inc., which has licensed the technology, to develop the assay for clinical use and we hope to offer it to U-M patients in the near future. We currently offer the PCA3 screening alone as follow-up to elevated PSA. (Sci Transl Med. 2011 Aug 3;3(94):94ra72).


MI-ONCOSEQ: Clinical sequencing in cancer

Complementing our ongoing efforts in the discovery of novel molecular sub-types of cancer, we have initiated a new project, Michigan Oncology Sequencing Center (MI-ONCOSEQ), to exploit the rapid advances in high throughput DNA sequencing technologies to realize the goals of “personalized medicine” for the treatment of cancer. We have begun to sequence patient genomes and transcriptomes to identify “actionable” driving mutations. We have completed a pilot study where 20 patients with advanced or refractory cancer who were eligible for clinical trials were enrolled. For each patient, we performed whole-genome sequencing of the tumor, targeted whole-exome sequencing of tumor and normal DNA, and transcriptome sequencing (RNA-Seq) of the tumor to identify potentially informative mutations in a clinically relevant time frame of 3 to 4 weeks. With this approach, we detected several classes of cancer mutations including structural rearrangements, copy number alterations, point mutations, and gene expression alterations. A multidisciplinary Sequencing Tumor Board (STB) deliberated on the clinical interpretation of the sequencing results obtained. The results of this pilot study was published in Science Translational Medicine (2011 Nov 30;3(111):111ra121).