Publications by authors named "Suzanne Kamel-Reid"

165 Publications

Optimal duration of imatinib treatment/deep molecular response for treatment-free remission after imatinib discontinuation from a Canadian tyrosine kinase inhibitor discontinuation trial.

Br J Haematol 2021 May 20;193(4):779-791. Epub 2021 Apr 20.

Department of Oncology, McMaster University, Hamilton, Canada.

Although total duration of tyrosine kinase inhibitor (TKI) therapy and of molecular response at 4 log reduction or deeper (MR4) correlates with treatment-free remission (TFR) success after TKI discontinuation, the optimal cut-off values of the duration remain unresolved. Thus, 131 patients were enrolled into the Canadian TKI discontinuation study. The molecular relapse-free survival (mRFS) was defined from imatinib discontinuation till molecular recurrence, that is, major molecular response (MMR) loss and/or MR4 loss. We evaluated mRFS at 12 months after imatinib discontinuation, analyzed it according to the imatinib treatment duration and MR4 duration, and calculated P value, positive (PPV) and negative predictive value (NPV) in the yearly cut-off period of time. The shortest cut-off was sought that met the joint criteria of a P value ≤ 0·05, PPV ≥ 60% and NPV ≥ 60%. We propose six years as the shortest imatinib duration cut-off with a P value 0·01, PPV 68% and NPV 62%: The patients treated with imatinib duration ≥ 6 years showed a superior mRFS rate (61·8%) compared to those with less treatment (36·0%). Also, 4·5 years MR4 duration as the shortest cut-off with a P value 0·003, PPV 63% and NPV 61%: those with MR4 duration ≥ 4·5 years showed a higher mRFS rate (64·2%) than those with a shorter MR4 duration (41·9%).
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http://dx.doi.org/10.1111/bjh.17447DOI Listing
May 2021

The Prevent Ovarian Cancer Program (POCP): Identification of women at risk for ovarian cancer using complementary recruitment approaches.

Gynecol Oncol 2021 Apr 13. Epub 2021 Apr 13.

Gynecologic Oncology, The University Health Network, Toronto, Canada; Department of Obstetrics and Gynecology, University of Toronto, Toronto, Canada. Electronic address:

Background: Up to 20% of high-grade serous ovarian carcinomas (HGSOC) are hereditary; however, historical uptake of genetic testing is low. We used a unique combination of approaches to identify women in Ontario, Canada, with a first-degree relative (FDR) who died from HGSOC without prior genetic testing, and offer them multi-gene panel testing.

Methods: From May 2015-Sept 2019, genetic counseling and testing was provided to eligible participants. Two recruitment strategies were employed, including self-identification in response to an outreach campaign and direct targeting of FDRs of deceased HGSOC patients treated at our institution. The rate of pathogenic variants (PV) in established/potential ovarian cancer risk genes and the benefits/challenges of each approach were assessed.

Results: A total of 564 women enrolled in response to our outreach campaign (n = 473) or direct recruitment (n = 91). Mean age at consent was 52 years and 96% did not meet provincial testing criteria. Genetic results were provided to 528 individuals from 458 families. The rate of PVs in ovarian cancer risk genes was highest when FDRs were diagnosed with HGSOC <60 years (9.4% vs. 3.9% ≥ 60y, p = 0.0160). Participants in the outreach vs. direct recruitment cohort had a similar rate of PVs; however, uptake of genetic testing (97% vs. 89%; p = 0.0036) and study completion (95% vs. 87%; p = 0.0062) rates were higher in the former. Eleven participants with pathogenic variants have completed risk-reducing gynecologic surgery, with one stage I HGSOC and two breast cancers identified.

Conclusion: Overall PV rates in this large cohort were lower than expected; however, we provide evidence that genetic testing criteria in Ontario should include individuals with a deceased FDR diagnosed with HGSOC <60 years of age.
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http://dx.doi.org/10.1016/j.ygyno.2021.04.011DOI Listing
April 2021

Assessing the Diagnostic Yield of Targeted Next-Generation Sequencing for Melanoma and Gastrointestinal Tumors.

J Mol Diagn 2020 04 6;22(4):467-475. Epub 2020 Feb 6.

Advanced Molecular Diagnostics Laboratory, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario; Division of Genome Diagnostics, Department of Clinical Laboratory Genetics, Laboratory Medicine Program, University Health Network, Toronto, Ontario; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada. Electronic address:

A common rationale in molecular diagnostic laboratories is that implementation of next-generation sequencing (NGS) enables simultaneous multigene testing, allowing increased information benefit compared with non-NGS assays. However, minimal published data exist to support this justification. The current study compared clinical diagnostic yield of TruSight Tumor 26 Sequencing Panel (TST26) in melanoma, colorectal (CRC), and gastrointestinal stromal (GIST) tumors with non-NGS assays. A total of 1041 formalin-fixed, paraffin-embedded tumors, of melanoma, CRC, and GIST, were profiled. NGS results were compared with non-NGS single-gene or single-variant assays with respect to variant output and diagnostic yield. A total of 79% melanoma and 94% CRC tumors were variant positive by panel testing. TST26 panel improved serine/threonine-protein kinase B-raf (BRAF) variant detection in melanoma compared with single-variant BRAF Val600Glu/Lys (V600E/K) routine tests by 24% and detected variants in genes other than BRAF, NRAS, and KIT, which could impact patient management in 20% additional cases. NGS enhanced diagnostic yield in CRC by 36% when compared with routine single-gene assays. In contrast, no added benefit of NGS-based testing for GIST tumors was observed. TST26 panel either missed or inaccurately called complex insertion/deletion variants in KIT exon 11, which were accurately identified by non-NGS methods. Findings of this study demonstrate the differential impact of cancer site and variant type on diagnostic test information yield from NGS assays.
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http://dx.doi.org/10.1016/j.jmoldx.2019.12.008DOI Listing
April 2020

A 4-gene signature from histologically normal surgical margins predicts local recurrence in patients with oral carcinoma: clinical validation.

Sci Rep 2020 02 3;10(1):1713. Epub 2020 Feb 3.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Prognostic biomarkers for recurrence of Oral Squamous Cell Carcinoma (OSCC) are urgently needed. We aimed to independently validate a 4-gene expression signature (MMP1, COL4A1, P4HA2, THBS2) predictive of OSCC recurrence risk. Gene expression was measured using Nanostring nCounter in 245 histologically normal surgical resection margins from 62 patients. Association between risk scores for individual patients and recurrence was assessed by Kaplan-Meier analysis. Signature performance was quantified by concordance index (CI), hazard ratio (HR) and the area under receiver operating characteristics (AUC). Risk scores for recurrence were significantly higher than recurrence-free patients (p = 9.58e-7, Welch's t-test). A solid performance of the 4-gene signature was determined: CI = 0.64, HR = 3.38 (p = 1.4E-4; log-rank test), AUC = 0.71. We showed that three margins per patient are sufficient to preserve predictive performance (CI = 0.65; HR = 2.92; p = 2.94e-3; AUC = 0.71). Association between the predicted risk scores and recurrence was assessed and showed HR = 2.44 (p = 9.6E-3; log-rank test, N = 62). Signature performance analysis was repeated using an optimized threshold (70 percentile of risks), resulting in HR = 3.38 (p = 1.4E-4; log-rank test, N = 62). The 4-gene signature was validated as predictive of recurrence risk in an independent cohort of patients with resected OSCC and histologically negative margins, and is potentially applicable for clinical decision making on adjuvant treatment and disease monitoring.
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http://dx.doi.org/10.1038/s41598-020-58688-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6997450PMC
February 2020

Phase II Trial of Cabozantinib in Recurrent/Metastatic Endometrial Cancer: A Study of the Princess Margaret, Chicago, and California Consortia (NCI9322/PHL86).

Clin Cancer Res 2020 06 28;26(11):2477-2486. Epub 2020 Jan 28.

Princess Margaret Cancer Centre, Toronto, Ontario, Canada.

Purpose: The relevance of the MET/hepatocyte growth factor pathway in endometrial cancer tumor biology supports the clinical evaluation of cabozantinib in this disease.

Patients And Methods: PHL86/NCI#9322 (NCT01935934) is a single arm study that evaluated cabozantinib (60 mg once daily) in women with endometrial cancer with progression after chemotherapy. Coprimary endpoints were response rate and 12-week progression-free-survival (PFS). Patients with uncommon histology endometrial cancer (eg, carcinosarcoma and clear cell) were enrolled in a parallel exploratory cohort.

Results: A total of 102 patients were accrued. Among 36 endometrioid histology patients, response rate was 14%, 12-week PFS rate was 67%, and median PFS was 4.8 months. In serous cohort of 34 patients, response rate was 12%, 12-week PFS was 56%, and median PFS was 4.0 months. In a separate cohort of 32 patients with uncommon histology endometrial cancer (including carcinosarcoma), response rate was 6% and 12-week PFS was 47%. Six patients were on treatment for >12 months, including two for >30 months. Common cabozantinib-related toxicities (>30% patients) included hypertension, fatigue, diarrhea, nausea, and hand-foot syndrome. Gastrointestinal fistula/perforation occurred in four of 70 (6%) patients with serous/endometrioid cancer and five of 32 (16%) patients in exploratory cohort. We observed increased frequency of responses with somatic mutation [four partial responses (PRs) in 10 patients, median PFS 7.6 months] and concurrent and mutations (three PRs in 12 patients, median PFS 5.9 months).

Conclusions: Cabozantinib has activity in serous and endometrioid histology endometrial cancer. These results support further evaluation in genomically characterized patient cohorts.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-2576DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7269808PMC
June 2020

Latency and interval therapy affect the evolution in metastatic colorectal cancer.

Sci Rep 2020 01 17;10(1):581. Epub 2020 Jan 17.

Princess Margaret Cancer Centre, Toronto, Ontario, Canada.

While comparison of primary tumor and metastases has highlighted genomic heterogeneity in colorectal cancer (CRC), previous studies have focused on a single metastatic site or limited genomic testing. Combining data from whole exome and ultra-deep targeted sequencing, we explored possible evolutionary trajectories beyond the status of these mutations, particularly among patient-matched metastatic tumors. Our findings confirm the persistence of known clinically-relevant mutations (e.g., those of RAS family of oncogenes) in CRC primary and metastases, yet reveal that latency and interval systemic therapy affect the course of evolutionary events within metastatic lesions. Specifically, our analysis of patient-matched primary and multiple metastatic lesions, developed over time, showed a similar genetic composition for liver metastatic tumors, which were 21-months apart. This genetic makeup was different from those identified in lung metastases developed before manifestation of the second liver metastasis. These results underscore the role of latency in the evolutionary path of metastatic CRC and may have implications for future treatment options.
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http://dx.doi.org/10.1038/s41598-020-57476-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6969060PMC
January 2020

Measurable residual disease monitoring provides insufficient lead-time to prevent morphologic relapse in the majority of patients with core-binding factor acute myeloid leukemia.

Haematologica 2021 01 1;106(1):56-63. Epub 2021 Jan 1.

Princess Margaret Cancer Centre, University Health Network, Toronto, Canada.

Core-binding factor acute myeloid leukemia is characterized by t(8;21) or inv(16) and the fusion proteins RUNX1-RUNX1T1 and CBFB-MYH11. International guidelines recommend monitoring for measurable residual disease every 3 months for 2 years after treatment. However, it is unknown if serial molecular monitoring can predict and prevent morphologic relapse. We conducted a retrospective single-center study of 114 patients in complete remission who underwent molecular monitoring with RT-qPCR of RUNX1-RUNX1T1 or CBFB-MYH11 transcripts every 3 months. Morphologic relapse was defined as re-emergence of >5% blasts and molecular relapse as ≥1 log increase in transcript level between 2 samples. Over a median follow-up time of 3.7 years (range 0.2-14.3), remission persisted in 71 (62.3%) patients but 43 (37.7%) developed molecular or morphologic relapse. Patients who achieved <3 log reduction in RUNX1-RUNX1T1 or CBFB-MYH11 transcripts at end of chemotherapy had a significantly higher risk of relapse compared to patients who achieved ≥3 log reduction (61.1% vs. 33.7%, p=0.004). The majority of relapses (74.4%, n=32) were not predicted by molecular monitoring and occurred rapidly with <100 days from molecular to morphologic relapse. Molecular monitoring enabled the detection of impending relapse and permitted pre-emptive intervention prior to morphologic relapse in only 11 (25.6%) patients. The current practice of molecular monitoring every 3 months provided insufficient lead-time to identify molecular relapses and prevent morphologic relapse in the majority of patients with core-binding factor acute myeloid leukemia treated at our institution. Further research is necessary to determine the optimal monitoring strategies for these patients.
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http://dx.doi.org/10.3324/haematol.2019.235721DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7776265PMC
January 2021

HPV-independent Vulvar Squamous Cell Carcinoma is Associated With Significantly Worse Prognosis Compared With HPV-associated Tumors.

Int J Gynecol Pathol 2020 Jul;39(4):391-399

Department of Pathology and Laboratory Medicine, Henry Ford Health System, Detroit, Michigan (G.A.) Departments of Radiation Oncology (M.L.Y., J.C., M.M.) Obstetrics and Gynecology, Division of Gynecologic Oncology (S.F.) Medical Oncology (H.M.) Laboratory Medicine and Pathobiology (S.K.R., I.W., D.G., B.A.C.) Biostatistics (M.P.), Princess Margaret Cancer Centre/University Health Network, and University of Toronto, Toronto, ON, Canada.

Vulvar squamous cell carcinomas (VSCC) represent the most common carcinoma of the female external genitalia, with increasing incidence. Although high-risk human papillomavirus (HPV) infection has long been implicated in the majority of cervical and anal squamous cell carcinomas, there is uncertainty about its prevalence and prognostic impact in VSCC. In this study, we conducted a retrospective integrated morphologic and multimodal HPV analysis of a cohort of 114 VSCC cases treated at the Princess Margaret Cancer Centre/University Health Network, Toronto, Canada between 2000 and 2010. VSCC histology was reviewed. We analyzed the cohort for HPV using polymerase chain reaction based method, and tissue microarray DNA and RNA in situ hybridization (ISH), and p16 immunohistochemistry. Among the 114 cases (age 70±16 yr), 36.7% of cases were classified as having histomorphology of HPV infection. HPV was detected in 31.9% (polymerase chain reaction), 14.0% (DNA ISH), and 27.3% (RNA ISH) of cases. p16 immunohistochemistry was positive in 37.8% of cases. On univariate analysis, HPV morphology (P=0.009), p16+ (P=0.00013), DNA ISH+ (P=0.021), and RNA ISH+ (P=0.00061) were associated with better 5-yr progression-free survival. DNA ISH+ (P=0.049) was associated with better 5-yr overall survival. On multivariate analysis, HPV morphology (P=0.033), p16+ (P=0.01), and RNA ISH+ (P=0.035) were associated with better 5-yr progression-free survival. In conclusion, a subset of VSCC is associated with HPV, which correlates with better outcome. Relatively inexpensive tests such as histomorphologic evaluation, p16 immunohistochemistry, and HPV RNA ISH can be used to predict outcome in VSCC. Therefore, routine reporting of HPV status in VSCC is recommended.
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http://dx.doi.org/10.1097/PGP.0000000000000620DOI Listing
July 2020

Performance Comparison of Different Analytic Methods in Proficiency Testing for Mutations in the , , and Genes: A Study of the College of American Pathologists Molecular Oncology Committee.

Arch Pathol Lab Med 2019 10 10;143(10):1203-1211. Epub 2019 Apr 10.

From the Office of the Director, The Joint Pathology Center, Silver Spring, Maryland (Dr Moncur); the Department of Pathology, St Joseph Mercy Hospital, Ypsilanti, Michigan (Dr Bartley); the Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha (Dr Bridge); the Department of Pathology, University Health Network, University of Toronto, Toronto, Ontario, Canada (Dr Kamel-Reid); the Department of Pathology, The University of Texas MD Anderson Cancer Center, Houston (Dr Lazar); the Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts (Drs Lindeman and Kim); Biostatistics (Mr Long) and Proficiency Testing (Ms Vasalos), College of American Pathologists, Northfield, Illinois; the Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill (Dr Merker); the Department of Pathology and Cell Biology, Columbia University Medical Center, New York, New York (Dr Rai); the Department of Pathology, Yale University School of Medicine, New Haven, Connecticut (Dr Rimm); and the Department of Pathology and Laboratory Medicine, Strong Memorial Hospital, University of Rochester Medical Center and Pathology and Laboratory Medicine, Molecular Diagnostic Laboratory, Rochester, New York (Dr Rothberg). Dr Moncur is employed by the US Army. The identification of specific products or scientific instrumentation is considered an integral part of the scientific endeavor and does not constitute endorsement or implied endorsement on the part of the authors, the Department of Defense, or any component agency. The views expressed in this article are those of the authors and do not reflect the official policy of the Department of Army/Navy/Air Force, Department of Defense, or US government. The other authors have no relevant financial interest in the products or companies described in this article.

Context.—: The performance of laboratory testing has recently come under increased scrutiny as part of important and ongoing debates on regulation and reimbursement. To address this critical issue, this study compares the performance of assay methods, using either commercial kits or assays designed and implemented by single laboratories ("home brews"), including next-generation sequencing methods, on proficiency testing provided by the College of American Pathologists Molecular Oncology Committee.

Objective.—: To compare the performance of different assay methods on College of American Pathologists proficiency testing for variant analysis of 3 common oncology analytes: , , and .

Design.—: There were 6897 total responses across 35 different proficiency testing samples interrogating 13 different variants as well as wild-type sequences for , , and . Performance was analyzed by test method, kit manufacturer, variants tested, and preanalytic and postanalytic practices.

Results.—: Of 26 reported commercial kits, 23 achieved greater than 95% accuracy. Laboratory-developed tests with no kit specified demonstrated 96.8% or greater accuracy across all 3 analytes (1123 [96.8%] acceptable of 1160 total responses for ; 848 [97.5%] acceptable of 870 total responses for ; 942 [97.0%] acceptable of 971 total responses for ). Next-generation sequencing platforms (summed across all analytes and 2 platforms) demonstrated 99.4% accuracy for these analytes (165 [99.4%] acceptable of 166 total next-generation sequencing responses). Slight differences in performance were noted among select commercial assays, dependent upon the particular design and specificity of the assay. Wide differences were noted in the lower limits of neoplastic cellularity laboratories accepted for testing.

Conclusions.—: These data demonstrate the high degree of accuracy and comparable performance across all laboratories, regardless of methodology. However, care must be taken in understanding the diagnostic specificity and reported analytic sensitivity of individual methods.
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http://dx.doi.org/10.5858/arpa.2018-0396-CPDOI Listing
October 2019

Effect of Coexisting KRAS and TP53 Mutations in Patients Treated With Chemotherapy for Non-small-cell Lung Cancer.

Clin Lung Cancer 2019 05 19;20(3):e338-e345. Epub 2018 Dec 19.

Division of Medical Oncology and Hematology, Princess Margaret Cancer Centre, University Health Network and Department of Medicine, University of Toronto, Toronto, ON, Canada.

Background: KRAS and TP53 are common mutations in non-small-cell lung cancer (NSCLC). The Lung Adjuvant Cisplatin Evaluation Biological Program group found adjuvant chemotherapy to be deleterious in patients with coexisting KRAS/TP53 mutations.

Patients And Methods: To validate these results, patients with NSCLC tested for KRAS and TP53 mutations and receiving chemotherapy for any stage NSCLC were selected. Mutation status was analyzed using next generation sequencing (Illumina) or multiplex recurrent mutation detection (MassARRAY, Agena Biosciences) assays, and was correlated with clinical and demographic data. Disease-free (DFS) or progression-free survival (PFS) was the main endpoint, and overall survival (OS) was the secondary endpoint.

Results: Among 218 patients, 28 had coexisting KRAS/TP53 mutations, 77 TP53, 37 KRAS, 76 had neither KRAS nor TP53 mutation (WT/WT). There was no DFS/PFS difference for the KRAS/TP53 group versus all others among 99 patients who received adjuvant chemotherapy (hazard ratio [HR], 1.22; 95% confidence interval [CI], 0.61-2.44; P = .57), 27 stage III patients who received chemo-radiation (HR, 0.87; 95% CI, 0.32-2.38; P = .8), and 63 patients who received palliative chemotherapy (HR, 0.68; 95% CI, 0.31-1.48; P = .33). OS was longer in the WT/WT group compared with any other group (KRAS: HR, 1.87; 95% CI, 1.02-3.43; P = .043; TP53: HR, 2.17; 95% CI, 1.3-3.61; P = .0028; KRAS/TP53: HR, 2.06; 95% CI, 1.09-3.88; P = .026). No OS difference was seen for KRAS/TP53 compared with the other groups (HR, 1.26; 95% CI, 0.75-2.13; P = .38).

Conclusions: There was no significant difference in DFS/PFS between the 4 groups. However, OS was longer for patients with TP53 and KRAS wild-type NSCLC who received chemotherapy for any stage compared with patients with KRAS, TP53 mutation, or double mutant tumors.
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http://dx.doi.org/10.1016/j.cllc.2018.12.009DOI Listing
May 2019

Somatic Tumor Variant Filtration Strategies to Optimize Tumor-Only Molecular Profiling Using Targeted Next-Generation Sequencing Panels.

J Mol Diagn 2019 03 19;21(2):261-273. Epub 2018 Dec 19.

Advanced Molecular Diagnostics Laboratory, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada; Department of Clinical Laboratory Genetics, Laboratory Medicine Program, University Health Network, Toronto, Ontario, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada. Electronic address:

A common approach in clinical diagnostic laboratories to variant assessment from tumor molecular profiling is sequencing of genomic DNA extracted from both tumor (somatic) and normal (germline) tissue, with subsequent variant comparison to identify true somatic variants with potential impact on patient treatment or prognosis. However, challenges exist in paired tumor-normal testing, including increased cost of dual sample testing and identification of germline cancer predisposing variants. Alternatively, somatic variants can be identified by in silico tumor-only variant filtration precluding the need for matched normal testing. The barrier to tumor-only variant filtration is defining a reliable approach, with high sensitivity and specificity to identify somatic variants. In this study, we used retrospective data sets from paired tumor-normal samples tested on small (48 gene) and large (555 gene) targeted next-generation sequencing panels, to model algorithms for tumor-only variants classification. The optimal algorithm required an ordinal filtering approach using information from variant population databases (1000 Genomes Phase 3, ESP6500, ExAC), clinical mutation databases (ClinVar), and information on recurring clinically relevant somatic variants. Overall the tumor-only variant filtration strategy described in this study can define clinically relevant somatic variants from tumor-only analysis with sensitivity of 97% to 99% and specificity of 87% to 94%, and with significant potential utility for clinical laboratories implementing tumor-only molecular profiling.
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http://dx.doi.org/10.1016/j.jmoldx.2018.09.008DOI Listing
March 2019

Heterogenous loss of mismatch repair (MMR) protein expression: a challenge for immunohistochemical interpretation and microsatellite instability (MSI) evaluation.

J Pathol Clin Res 2019 04 19;5(2):115-129. Epub 2018 Dec 19.

Department of Anatomical Pathology, Laboratory Medicine Program, University Health Network and University of Toronto, Toronto, Canada.

Immunohistochemistry (IHC) for mismatch repair (MMR) proteins is used to identify MMR status: being diffusely positive (intact/retained nuclear staining) or showing loss of nuclear tumour staining (MMR protein deficient). Four colonic adenocarcinomas and a gastric adenocarcinoma with associated dysplasia that displayed heterogenous IHC staining patterns in at least one of the four MMR proteins were characterised by next-generation sequencing (NGS). In order to examine a potential molecular mechanism for these staining patterns, the respective areas were macrodissected, analysed for microsatellite instability (MSI) and investigated by NGS and multiplex ligation-dependent probe amplification (MLPA) analysis of MLH1, MSH2, MSH6 and PMS2 genes, including MLH1 methylation analysis. One colonic adenocarcinoma showed heterogenous MSH6 IHC staining and molecular analysis demonstrated increasing allelic burden of two MSH6 frameshift variants (c.3261delC and c.3261dupC) in areas with MSH6 protein loss compared to areas where MSH6 was retained. Two colonic adenocarcinomas with heterogenous MLH1 staining showed no differences in sequence variants. In one of these cases, however, MLH1 was hypermethylated in the area of MLH1 loss. Another colon carcinoma with heterogenous PMS2 staining (but with retained MSH6) showed both MSH6 c.3261dupC and 3260_3261dupCC where PMS2 protein was lost and only c.3261dupC where PMS2 was retained. The gastric carcinoma showed complete loss of MSH6 in dysplastic foci, while the underlying invasive carcinoma showed retention of MSH6. Both these areas, however, were MSI-high and showed the same MSH6 variant: c.3261delC. The gastric dysplasia additionally showed MSH6 c.3261dupC. In four of the five cases where MMR protein was lost, these areas were MSI-high. Heterogenous MMR IHC (focal and/or zonal within the same tumour or between invasive and dysplastic preinvasive areas) is not always due to artefact and is invariably related to MSI-high status in the areas of loss. An interesting aspect to this study is the presence of MSH6 somatic mutations irrespective of whether MSH6 IHC staining was intact or lost.
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http://dx.doi.org/10.1002/cjp2.120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6463865PMC
April 2019

Proficiency Testing of Standardized Samples Shows Very High Interlaboratory Agreement for Clinical Next-Generation Sequencing-Based Oncology Assays.

Arch Pathol Lab Med 2019 04 30;143(4):463-471. Epub 2018 Oct 30.

From the Departments of Pathology and Laboratory Medicine & Genetics, Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill (Dr Merker); the Departments of Pathology (Drs Devereaux and Montgomery, and Mr Smail) and Genetics (Dr Montgomery), Stanford University School of Medicine, and the Biomedical Informatics Program (Mr Smail), Stanford University, Stanford, California; the Department of Pathology, Massachusetts General Hospital (Dr Iafrate), and the Department of Pathology, Brigham and Women's Hospital (Drs Kim and Lindeman), Harvard University, Boston; the Departments of Pathology and Clinical Laboratory Genetics, The University Health Network and the University of Toronto, Toronto, Ontario, Canada (Dr Kamel-Reid); the Department of Pathology, Walter Reed National Military Medical Center, Bethesda, Maryland (Dr Moncur); PierianDx, St Louis, Missouri (Dr Nagarajan); the Department of Global Medical Affairs, Roche, Tucson, Arizona (Dr Portier); the Departments of Hematopathology (Dr Routbort) and Pathology, Genomic Medicine, and Translational Molecular Pathology (Dr Lazar), he University of Texas MD Anderson Cancer Center, Houston (Dr Routbort); the Department of Pathology, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia (Dr Surrey); and Proficiency Testing, College of American Pathologists, Northfield, Illinois (Ms Vasalos).

Context.—: Next-generation sequencing-based assays are being increasingly used in the clinical setting for the detection of somatic variants in solid tumors, but limited data are available regarding the interlaboratory performance of these assays.

Objective.—: To examine proficiency testing data from the initial College of American Pathologists (CAP) Next-Generation Sequencing Solid Tumor survey to report on laboratory performance.

Design.—: CAP proficiency testing results from 111 laboratories were analyzed for accuracy and associated assay performance characteristics.

Results.—: The overall accuracy observed for all variants was 98.3%. Rare false-negative results could not be attributed to sequencing platform, selection method, or other assay characteristics. The median and average of the variant allele fractions reported by the laboratories were within 10% of those orthogonally determined by digital polymerase chain reaction for each variant. The median coverage reported at the variant sites ranged from 1922 to 3297.

Conclusions.—: Laboratories demonstrated an overall accuracy of greater than 98% with high specificity when examining 10 clinically relevant somatic single-nucleotide variants with a variant allele fraction of 15% or greater. These initial data suggest excellent performance, but further ongoing studies are needed to evaluate the performance of lower variant allele fractions and additional variant types.
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http://dx.doi.org/10.5858/arpa.2018-0336-CPDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6910717PMC
April 2019

The mutational landscape of accelerated- and blast-phase myeloproliferative neoplasms impacts patient outcomes.

Blood Adv 2018 10;2(20):2658-2671

Department of Medical Oncology and Hematology and.

There is a paucity of data regarding the impact of mutations on outcomes in accelerated-phase (AP) and blast-phase (BP) myeloproliferative neoplasms (MPNs). Moreover, it is unknown whether mutational status affects survival, as seen in chronic-phase MPNs. Therefore, we performed a retrospective analysis of all patients treated at our institution with AP/BP MPNs (N = 122; AP = 14; BP = 108) to comprehensively describe the mutational profile and correlate with clinical outcomes. Targeted sequencing with a 54-gene panel was performed. Forty-four patients were treated with intensive therapy, 27 with nonintensive therapy, and 51 with best supportive care (BSC). The most common mutation was , occurring in 55% of subjects; was found in 13% of patients and in 6%. Thirty-two (26%) patients were triple negative. Other frequently mutated genes were (30%), (25%), (22%), (20%), and (17%). Mutations in 1, 2, 3, and ≥4 genes were seen in 15%, 13%, 25%, and 46% of patients, respectively. There was no difference in survival between patients treated with intensive vs nonintensive therapy, and the benefit of intensive therapy was limited to patients who were able to undergo transplantation. was the only individual mutation to correlate with shorter overall survival (hazard ratio, 1.89; = .03). In the multivariate analysis, mutated , ≥4 mutations, low albumin, increased peripheral blood blasts, ≥3 cytogenetic abnormalities, and BSC were associated with shorter survival. In conclusion, mutational data enhance the understanding of patients with AP/BP MPN who are likely to benefit from current therapeutic options.
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http://dx.doi.org/10.1182/bloodadvances.2018021469DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6199664PMC
October 2018

Genomic Classifier for Guiding Treatment of Intermediate-Risk Prostate Cancers to Dose-Escalated Image Guided Radiation Therapy Without Hormone Therapy.

Int J Radiat Oncol Biol Phys 2019 01 29;103(1):84-91. Epub 2018 Aug 29.

Princess Margaret Cancer Centre, University Health Network, Ontario, Canada; Division of Radiation Oncology, National Cancer Centre Singapore, Singapore; Division of Medical Sciences, National Cancer Centre Singapore, Singapore; Oncology Academic Program, Duke-NUS Medical School, Singapore.

Purpose: The National Comprehensive Cancer Network (NCCN) has recently endorsed the stratification of intermediate-risk prostate cancer (IR-PCa) into favorable and unfavorable subgroups and recommend the addition of androgen deprivation therapy (ADT) to radiation therapy (RT) for unfavorable IR-PCa. Recently, more accurate prognostication was demonstrated by integrating a 22-feature genomic classifier (GC) to the NCCN stratification system. Here, we test the utility of the GC to better identify patients with IR-PCa who are sufficiently treated by RT alone.

Methods And Materials: We identified a novel cohort comprising 121 patients with IR-PCa treated with dose-escalated image guided RT (78 Gy in 39 fractions) without ADT. GC scores were derived from tumors sampled in diagnostic biopsies. Multivariable analyses, including both NCCN subclassification and GC scores, were performed for biochemical failure (prostate-specific antigen nadir + 2 ng/mL) and metastasis occurrence.

Results: By NCCN subclassification, 33 (27.3%) and 87 (71.9%) of men were classified as having favorable and unfavorable IR-PCa, respectively (1 case unclassifiable). GC scores were high in 3 favorable IR-PCa and low in 60 unfavorable IR-PCa. Higher GC scores, but not NCCN risk subgroups, were associated with biochemical relapse (hazard ratio, 1.36; 95% confidence interval [CI], 1.09-1.71] per 10% increase; P = .007) and metastasis (hazard ratio, 2.05; 95% CI, 1.24-4.24; P = .004). GC predicted biochemical failure at 5 years (area under the curve, 0.78; 95% CI, 0.59-0.91), and the combinatorial NCCN + GC model significantly outperformed the NCCN alone model for predicting early-onset metastasis (area under the curve for 5-year metastasis of 0.89 vs 0.86 [GC alone] vs 0.54 [NCCN alone]).

Conclusions: We demonstrated the accuracy of the GC for predicting disease recurrence in IR-PCa treated with dose-escalated image guided RT alone. Our findings highlight the need to evaluate this GC in a prospective clinical trial investigating the role of ADT-RT in clinicogenomic-defined IR-PCa subgroups.
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http://dx.doi.org/10.1016/j.ijrobp.2018.08.030DOI Listing
January 2019

Additional germline findings from a tumor profiling program.

BMC Med Genomics 2018 Aug 9;11(1):65. Epub 2018 Aug 9.

Division of Medical Oncology and Hematology, Princess Margaret Cancer Centre, 610 University Ave, Toronto, ON, M5G 2M9, Canada.

Background: Matched tumor-normal sequencing, applied in precision cancer medicine, can identify unidentified germline Medically Actionable Variants (gMAVS) in cancer predisposition genes. We report patient preferences for the return of additional germline results, and describe various gMAV scenarios delivered through a clinical genetics service.

Methods: Tumor profiling was offered to 1960 advanced cancer patients, of which 1556 underwent tumor-normal sequencing with multigene hotspot panels containing 20 cancer predisposition genes. All patients were provided with an IRB-approved consent for return of additional gMAVs.

Results: Of the whole cohort 94% of patients consented to be informed of additional germline results and 5% declined, with no statistically significant differences based on age, sex, race or prior genetic testing. Eight patients were found to have gMAVs in a cancer predisposition gene. Five had previously unidentified gMAVs: three in TP53 (only one fulfilled Chompret's Revised criteria for Li-Fraumeni Syndrome), one in SMARCB1 in the absence of schwannomatosis features and one a TP53 variant at low allele frequency suggesting an acquired event in blood.

Conclusion: Interest in germline findings is high among patients who undergo tumor profiling. Disclosure of previously unidentified gMAVs present multiple challenges, thus supporting the involvement of a clinical genetics service in all tumor profiling programs.
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http://dx.doi.org/10.1186/s12920-018-0383-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6085686PMC
August 2018

The prognostic effect of single and multiple cancer-related somatic mutations in resected non-small-cell lung cancer.

Lung Cancer 2018 09 19;123:22-29. Epub 2018 Jun 19.

Department of Medical Oncology and Hematology, Princess Margaret Cancer Centre, University Health Network and the University of Toronto, Toronto, Ontario, Canada.

Introduction: Somatic mutations are becoming increasingly important biomarkers for treatment selection and outcome in patients with non-small-cell lung cancer (NSCLC). The role of multiple somatic mutations in early-stage NSCLC is unclear.

Methods: Tissue from 214 patients with resected NSCLC at the Princess Margaret Cancer Centre was analyzed by next-generation sequencing by Mi-SEQ or Sequenom multiplex platforms. Associations between mutation status, baseline patient characteristics and outcomes (disease-free survival (DFS) after surgical resection and overall survival (OS)) were investigated.

Results: Somatic mutations were identified in 184 patients with resected stage I-III NSCLC: None (n = 30), single (n = 101) and multiple (≥2, n = 83). Multiple mutations were significantly associated with younger age (p = 0.0006), female sex (p = 0.012), smoking status (p = 0.002) and adenocarcinoma histology (p = 0.0001).TP53, KRAS and EGFR were the most common mutations. TP53 mutation was the most frequent co-mutation occurring in 72% of patients with multiple mutations. In resected stage I-III patients, multiple mutations were significantly associated with worse DFS (HR = 2.56, p = 0.003) but not OS on univariate analysis. Patients with KRAS and EGFR mutations were also associated with shorter DFS (HR = 2.52, p = 0.016 and HR = 4.37, p = 0.001 respectively) but no OS difference. TP53 mutation was associated with both shorter DFS (HR = 2.21, p = 0.02) and OS (HR = 3.08, p = 0.02). In subgroup univariate analysis, poorer DFS was associated with multiple mutations (p = 0.0015), EGFR (HR = 3.14, p = 0.006), and TP53 (HR = 2.46, p = 0.018) in patients with stage I disease.

Conclusion: The presence of known somatic mutations is associated with worse DFS in resected NSCLC. The differences are both statistically significant and clinically relevant. The presence of EGFR, KRAS and TP53 mutations was also associated with adverse outcomes. Larger datasets are required to validate whether mutational status is an independent prognostic factor in early stage NSCLC.
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http://dx.doi.org/10.1016/j.lungcan.2018.06.023DOI Listing
September 2018

Lung cancer in never smokers from the Princess Margaret Cancer Centre.

Oncotarget 2018 Apr 27;9(32):22559-22570. Epub 2018 Apr 27.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Introduction: Lung cancer in never smokers represents a distinct epidemiological, clinical, and molecular entity.

Results: Most 712 never smoking lung cancer patients were female (72%) with a median age at diagnosis of 62.2 years (18-94). Caucasians (46%), East Asians (42%), adenocarcinoma histology (87%) and presentation with metastatic disease at diagnosis (59%) were common. Of 515 patients with available archival tissue, the most common identified single mutations were (52.2%), followed by (7.5%), (2.3%), (1.3%), (1%), (0.4%), (0.4%), (0.4%), (0.2%), (0.2%), and (0.2%); 8% tumors had multiple mutations, while 25.8% had none identified. Median overall survival (mOS) was 42.2 months (mo) for the entire cohort. Patients with mutations in their tumors had significantly better mOS (69.5 mo) when compared to those without (31.0 mo) (HR = 0.59; 95% CI: 0.44-0.79; < 0.001). Earlier stage ( < 0.001), adenocarcinoma histology ( = 0.012), good performance status ( < 0.001) and use of targeted therapy ( < 0.001) were each independently associated with longer survival. Patients with -translocation-positive tumours have significantly longer OS compared to those without any mutations ( = 0.0029) and to those with other and null mutations ( = 0.022).

Conclusions: Lung cancer in never smokers represents a distinct clinical and molecular entity characterized by a high incidence of targetable mutations and long survival.

Methods: We analyzed retrospectively the data from electronic patient records of never smokers diagnosed with lung cancer treated at the Princess Margaret Cancer Centre (Toronto) between 1988-2015 to characterize demographic and clinical features, pathology, molecular profile (using hotspot or targeted sequencing panels), treatment and survival.
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http://dx.doi.org/10.18632/oncotarget.25176DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5978248PMC
April 2018

Clinical Utility of Next-generation Sequencing in the Management of Myeloproliferative Neoplasms: A Single-Center Experience.

Hemasphere 2018 Jun 4;2(3):e44. Epub 2018 May 4.

Medical Oncology and Hematology, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Although next-generation sequencing (NGS) has helped characterize the complex genomic landscape of myeloid malignancies, its clinical utility remains undefined. This has resulted in variable funding for NGS testing, limiting its accessibility. At our center, targeted sequencing (TAR-SEQ) using a 54-gene NGS myeloid panel is offered to all new patients referred for myeloid malignancies, as part of a prospective observational study. Here, we evaluated the diagnostic, prognostic, and potential therapeutic utility of clinical grade TAR-SEQ in the routine workflow of 179 patients with myeloproliferative neoplasms (MPN). Of 13 patients with triple negative (TN) MPN, who lacked driver mutations in , , and , TAR-SEQ confirmed clonal hematopoiesis in 8 patients. In patients with intermediate-risk myelofibrosis (MF), TAR-SEQ helped optimize clinical decisions in hematopoietic cell transplant (HCT)-eligible patients through identifying a high molecular risk (HMR) mutation profile. The presence of an HMR profile favored HCT in 9 patients with intermediate-1 risk MF. Absence of an HMR profile resulted in a delayed HCT strategy in 10 patients with intermediate-2 risk MF, 7 of which were stable at the last follow-up. Finally, TAR-SEQ identified patients with various targetable mutations in (4%), spliceosome genes (28%), and (7%). Some of these patients can be potential candidates for future targeted therapy trials. In conclusion, we have demonstrated that TAR-SEQ improves the characterization of TN MPN, can be integrated in clinical practice as an additional tool to refine decision making in HCT, and has the potential to identify candidates for future targeted therapy trials.
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http://dx.doi.org/10.1097/HS9.0000000000000044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6745993PMC
June 2018

Translation of Knowledge to Practice-Improving Awareness in NSCLC Molecular Testing.

J Thorac Oncol 2018 07 11;13(7):1004-1011. Epub 2018 Mar 11.

Division of Medical Oncology, Princess Margaret Cancer Centre, Toronto, Ontario, Canada. Electronic address:

Background: Molecular testing in advanced lung cancer is standard in guiding treatment selection. However, population-wide implementation of testing remains a challenge. We developed a knowledge translation intervention to improve understanding among diagnostic specialists about molecular testing and appropriate diagnostic sampling in lung cancer.

Methods: Specialty-specific education programs were developed from existing literature and input from Canadian leaders in lung pathology, respirology, interventional radiology, thoracic surgery, radiation oncology, and medical oncology. The programs, including key messages, review of current data, existing guidelines, group discussion, and participant feedback, were administered at provincial and national specialty meetings. Participant knowledge was assessed before and after the intervention by using anonymous questionnaires. Molecular (EGFR) testing rates in Ontario were also evaluated before and after the intervention period.

Results: Ten programs were administered to diagnostic specialists, including respirologists, pathologists, thoracic surgeons, radiologists, radiation oncologists, and medical oncologists, with completion of 255 preintervention and 219 postintervention surveys. At baseline, 30% were unsure of tissue handling methods for molecular testing, 20% chose an incorrect technique, and half were unfamiliar with how to initiate testing. After intervention, specialist knowledge improved regarding tissue handling and appropriate fixation techniques and uncertainty decreased from 30% to 2% (p < 0.001). A 12% increase (relative increase 57%) in molecular (EGFR) testing requests in Ontario was observed over the intervention period (p = 0.0032).

Conclusions: Significant knowledge gaps exist among diagnostic specialists regarding molecular testing and targeted therapy in lung cancer. This initiative significantly improved understanding of the importance and methods of successful molecular testing and correlated with increased testing rates.
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http://dx.doi.org/10.1016/j.jtho.2018.03.005DOI Listing
July 2018

AML refractory to primary induction with Ida-FLAG has a poor clinical outcome.

Leuk Res 2018 05 20;68:22-28. Epub 2018 Feb 20.

Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada. Electronic address:

We evaluated outcomes of 100 patients with high risk AML treated with Ida-FLAG induction as first-line therapy. 72 achieved remission with one cycle; 19 did not. High risk cytogenetics and TP53 mutations were associated with failure to achieve remission. In those reaching remission, allogeneic bone marrow transplantation was associated with better relapse-free and overall survival. Those not achieving remission with induction therapy were extremely unlikely to reach remission with further therapy and had a dismal prognosis. Exploratory molecular analysis confirmed persistence of the dominant genetic mutations identified at diagnosis. Ex vivo chemosensitivity did not demonstrate significant differences between responders and non-responders. Thus, Ida-FLAG induction has a high chance of inducing remission in patients with high risk AML. Those achieving remission require allogeneic transplantation to achieve cure; those not achieving remission rarely respond to salvage chemotherapy and have a dismal outcome. Alternatives to conventional chemotherapy must be considered in this group.
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http://dx.doi.org/10.1016/j.leukres.2018.02.012DOI Listing
May 2018

Impact of genomic alterations on outcomes in myelofibrosis patients undergoing JAK1/2 inhibitor therapy.

Blood Adv 2017 Sep 8;1(20):1729-1738. Epub 2017 Sep 8.

Department of Medical Oncology and Hematology.

In myelofibrosis (MF), driver mutations in or impact survival and progression to blast phase, with the greatest risk conferred by triple-negative status. Subclonal mutations, including mutations in high-molecular risk (HMR) genes, such as and have also been associated with inferior prognosis. However, data evaluating the impact of next-generation sequencing in MF patients treated with JAK1/2 inhibitors are lacking. Using a 54-gene myeloid panel, we performed targeted sequencing on 100 MF patients treated with ruxolitinib (n = 77) or momelotinib (n = 23) and correlated mutational profiles with treatment outcomes. Ninety-nine patients had at least 1 mutation identified, 46 (46%) had 2 mutations, and 34 (34%) patients had ≥3 mutations. Seventy-nine patients carried a mutation in 14 patients had mutations in 6 patients had an mutation, and 2 patients were triple negative. No mutation was significantly associated with spleen or anemia response. A high Dynamic International Prognostic Scoring System score and pretreatment transfusion dependence were associated with a shorter time to treatment failure (TTF), and this association retained significance on multivariable analysis. Patients with (hazard ratio [HR], 1.86; = .03) and mutations (HR, 2.94; = .009) and an HMR profile (HR, 2.06; = .01) had shorter TTF. On multivariate analysis, or mutations were independently associated with shorter TTF and overall survival. These findings help identify patients unlikely to have a durable response with current JAK1/2 inhibitors and provide a framework for future studies.
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http://dx.doi.org/10.1182/bloodadvances.2017009530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5728340PMC
September 2017

Guidance Statement On BRCA1/2 Tumor Testing in Ovarian Cancer Patients.

Semin Oncol 2017 06 15;44(3):187-197. Epub 2017 Sep 15.

Queen's University Belfast, Belfast, UK. Electronic address:

The approval, in 2015, of the first poly (adenosine diphosphate-ribose) polymerase inhibitor (PARPi; olaparib, Lynparza) for platinum-sensitive relapsed high-grade ovarian cancer with either germline or somatic BRCA1/2 deleterious mutations is changing the way that BRCA1/2 testing services are offered to patients with ovarian cancer. Ovarian cancer patients are now being referred for BRCA1/2 genetic testing for treatment decisions, in addition to familial risk estimation, and irrespective of a family history of breast or ovarian cancer. Furthermore, testing of tumor samples to identify the estimated 3%-9% of patients with somatic BRCA1/2 mutations who, in addition to germline carriers, could benefit from PARPi therapy is also now being considered. This new testing paradigm poses some challenges, in particular the technical and analytical difficulties of analyzing chemically challenged DNA derived from formalin-fixed, paraffin-embedded specimens. The current manuscript reviews some of these challenges and technical recommendations to consider when undertaking BRCA1/2 testing in tumor tissue samples to detect both germline and somatic BRCA1/2 mutations. Also provided are considerations for incorporating genetic analysis of ovarian tumor samples into the patient pathway and ethical requirements.
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http://dx.doi.org/10.1053/j.seminoncol.2017.08.004DOI Listing
June 2017

Comparison of Laboratory-Developed Tests and FDA-Approved Assays for BRAF, EGFR, and KRAS Testing.

JAMA Oncol 2018 06;4(6):838-841

Walter Reed National Military Medical Center, Bethesda, Maryland.

Importance: The debate about the role of the Food and Drug Administration (FDA) in the regulation of laboratory-developed tests (LDTs) has focused attention on the analytical performance of all clinical laboratory testing. This study provides data comparing the performance of LDTs and FDA-approved companion diagnostics (FDA-CDs) in proficiency testing (PT) provided by the College of American Pathologists Molecular Oncology Committee.

Objective: To compare the analytical performance of LDTs and FDA-CDs on well-characterized PT samples and to compare the practice characteristics of laboratories using these assays.

Design, Setting, And Participants: This comparison of PT responses examines the performance of laboratories participating in the College of American Pathologists PT for 3 oncology analytes for which both FDA-CDs and LDTs are used: BRAF, EGFR, and KRAS. A total of 6897 PT responses were included: BRAF (n = 2524; 14 PT samples), EGFR (n = 2216; 11 PT samples), and KRAS (n = 2157, 10 PT samples). US Food and Drug Administration companion diagnostics and LDTs are compared for both accuracy and preanalytic practices of the laboratories.

Main Outcomes And Measures: As per the College of American Pathologists PT standards, results were scored and the percentages of acceptable responses for each analyte were compared. These were also broken down by the specific variants tested, by kit manufacturer for laboratories using commercial reagents, and by preanalytic practices.

Results: From analysis of 6897 PT responses, this study demonstrates that both LDTs and FDA-CDs have excellent performance overall, with both test types exceeding 97% accuracy for all 3 genes (BRAF, EGFR, and KRAS) combined. Rare variant-specific differences did not consistently favor LDTs or FDA-CDs. Additionally, more than 60% of participants using an FDA-CD reported adapting their assay from the approved procedure to allow for a greater breadth of sample types, minimum tumor content, and instrumentation, changing the classification of their assay from FDA-CD to LDT.

Conclusions: This study demonstrates the high degree of accuracy and comparable performance of both LDTs and FDA-CDs for 3 oncology analytes. More significantly, the majority of laboratories using FDA-CDs have modified the scope of their assay to allow for more clinical practice variety, rendering them LDTs. These findings support both the excellent and equivalent performance of both LDTs and FDA-CDs in clinical diagnostic testing.
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http://dx.doi.org/10.1001/jamaoncol.2017.4021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6145687PMC
June 2018

Impact of multi-gene mutational profiling on clinical trial outcomes in metastatic breast cancer.

Breast Cancer Res Treat 2018 Feb 24;168(1):159-168. Epub 2017 Nov 24.

Division of Medical Oncology and Hematology, Princess Margaret Cancer Centre, 7-723 700 University Avenue, Toronto, Canada.

Purpose: Next-generation sequencing (NGS) has identified recurrent genomic alterations in metastatic breast cancer (MBC); however, the clinical utility of incorporating routine sequencing to guide treatment decisions in this setting is unclear. We examine the frequency of genomic alterations in MBC patients from academic and community hospitals and correlate with clinical outcomes.

Methods: MBC patients with good performance status were prospectively recruited at the Princess Margaret Cancer Centre (PM) in Canada. Molecular profiling on DNA extracted from FFPE archival tissues was performed on the Sequenom MassArray platform or the TruSeq Amplicon Cancer Panel (TSACP) on the MiSeq platform. Clinical trial outcomes by RECIST 1.1 and time on treatment were reviewed retrospectively.

Results: From January 2012 to November 2015, 483 MBC patients were enrolled and 440 were genotyped. At least one somatic mutation was identified in 46% of patients, most commonly in PIK3CA (28%) or TP53 (13%). Of 203 patients with ≥ 1 mutation(s), 15% were treated on genotype-matched and 9% on non-matched trials. There was no significant difference for median time on treatment for patients treated on matched vs. non-matched therapies (3.6 vs. 3.8 months; p = 0.89).

Conclusions: This study provides real-world outcomes on hotspot genotyping and small targeted panel sequencing of MBC patients from academic and community settings. Few patients were matched to clinical trials with targeted therapies. More comprehensive profiling and improved access to clinical trials may increase therapeutic options for patients with actionable mutations. Further studies are needed to evaluate if this approach leads to improved clinical outcomes.
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http://dx.doi.org/10.1007/s10549-017-4580-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5847065PMC
February 2018

Association of Ipilimumab With Safety and Antitumor Activity in Women With Metastatic or Recurrent Human Papillomavirus-Related Cervical Carcinoma.

JAMA Oncol 2018 07 12;4(7):e173776. Epub 2018 Jul 12.

Drug Development Program, Division of Medical Oncology, Princess Margaret Cancer Centre, Toronto, Ontario, Canada.

Importance: Based on evidence of human papillomavirus (HPV)-induced immune evasion, immunotherapy may be an attractive strategy in cervical cancer. Ipilimumab is a fully humanized monoclonal antibody that blocks cytotoxic T-lymphocyte antigen-4 (CTLA-4), which acts to downregulate the T-cell immune response.

Objective: To assess the safety and antitumor activity of ipilimumab in recurrent cervical cancer.

Design, Setting, And Participants: A multicenter trial was designed for patients with metastatic cervical cancer (squamous cell carcinoma or adenocarcinoma) with measurable disease and progression after at least 1 line of platinum chemotherapy. A run-in safety cohort using ipilimumab, 3 mg/kg, every 21 days for 4 cycles in 6 patients was followed by a phase II cohort of ipilimumab, 10 mg/kg, every 21 days for 4 cycles and then 4 cycles of maintenance therapy every 12 weeks for patients demonstrating radiologic response or stabilization. Immune correlative studies were performed on peripheral blood before and after therapy on archival tissue and fresh tumor obtained prior to registration and 7 days after cycle 2. The study was conducted from December 3, 2012, to September 15, 2014. The data were analyzed from April 2016 to June 2016 and in July 2017.

Main Outcomes And Measures: The primary end points were safety and objective response rate. Immune analyses were performed on blood and tumor tissue.

Results: A total of 42 women (median age, 49 years; range, 23-78 years) were enrolled (29 [69%] squamous cell cervical cancer and 13 [31%] adenocarcinoma; 37 [93%] of 40 patients with tissue available for analysis had HPV-positive confirmation; there was no archival tissue for 2 women). Grade 3 toxic effects included diarrhea in 4 patients, 3 of whom had colitis. Of 34 patients evaluated for best response (Response Evaluation Criteria in Solid Tumors, version 1.1), 1 patient had partial response and 10 had stable disease. The median progression-free survival and overall survival were 2.5 months (95% CI, 2.1-3.2 months) and 8.5 months (95% CI, 3.6-not reached; 1 patient was still alive), respectively. Intratumoral pretreatment CD3, CD4, CD8, FoxP3, indoleamine 2,3-dioxygenase, and programmed cell death ligand 1 (PD-L1) expression was not predictive of benefit and did not significantly change with treatment. Multicolor flow cytometry on peripheral lymphocytes revealed a treatment-dependent increase of inducible T-cell costimulator, human leukocyte antigen-antigen D related, and PD-1 during initial treatment, which returned to baseline during maintenance.

Conclusions And Relevance: Ipilimumab was tolerable in this population but did not show significant single-agent activity. Immune changes were induced by anti-CTLA-4 therapy but did not correlate with clinical activity. Changes in these markers may guide further treatment strategies.
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http://dx.doi.org/10.1001/jamaoncol.2017.3776DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6145732PMC
July 2018

Molecular Profiling of Patients With Advanced Colorectal Cancer: Princess Margaret Cancer Centre Experience.

Clin Colorectal Cancer 2018 03 23;17(1):73-79. Epub 2017 Oct 23.

Division of Medical Oncology and Hematology, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada; Department of Medicine, University of Toronto, Toronto, ON, Canada. Electronic address:

Background: Molecular aberrations in KRAS, NRAS, BRAF, and PIK3CA have been well-described in advanced colorectal cancer. The incidences of other mutations are less known. We report results of molecular profiling of advanced colorectal cancer in an academic cancer center.

Patients And Methods: Patients with advanced colorectal were enrolled in an institution-wide molecular profiling program. Profiling was performed on formalin-fixed paraffin embedded archival tissues using a customized MassArray panel (23 genes, 279 mutations) or the Illumina MiSeq TruSeq Cancer Panel (48 genes, 212 amplicons, ≥ 500× coverage) in a Clinical Laboratory Improvement Amendments-certified laboratory. PTEN was determined by immunohistochemistry.

Results: From March 2012 to April 2014, 245 patients were enrolled. At least one mutation was found in 54% (97/178) and 91% (61/67) of patients using MassArray or MiSeq platforms, respectively (P < .01). Of all patients, KRAS G12/13 mutation was identified in 39%, and non-G12/13 KRAS, BRAF, or NRAS mutations were present in 9%, 6%, and 4%, respectively. Other common mutations included TP53 (68.7%), APC (41.8%), and PIK3CA (13.5%). Co-mutation with KRAS, NRAS, or BRAF was found in 75% of patients with PIK3CA mutation. Of 106 patients with known PTEN immunohistochemistry status, 16% were negative. A higher average number of mutations were observed in right versus left colorectal cancer (P < .01), with 13 of 14 BRAF mutations located in right colon cancer.

Conclusion: Mutations are common in advanced colorectal cancer. Right colon cancers harbor more genetic aberrations than left colon or rectal cancers. These aberrations may contribute to differential outcomes to anti-epidermal growth factor receptor therapy among patients with right colon, left colon, or rectal cancers.
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http://dx.doi.org/10.1016/j.clcc.2017.10.010DOI Listing
March 2018