TEST:

Oncomine™ Comprehensive Assay v3M

The OCAv3 assay on the GeneXus System provides a rapid and reliable method for CGP of solid tumors from FFPE samples, enabling the identification of clinically actionable mutations for personalized oncology. Whereas limitations exist in homopolymer regions and coverage variability through all targets, the assay's turnaround time, low input material requirement, and overall performance when using optimal cut-offs support its adoption in clinical practice.

SNAQ-SEQ copy loss detection was based on estimating the exon abundance of ATM, BRCA1, BRCA2, and PALB2 genes, converting abundance into copies per cell and then detecting genes with significantly lower copy number. The CNV detection comparison between OCAv3 with and without SNAQ indicated that analysis with SNAQ was more robust, giving a result for every gene in every sample. Further, there were copy losses detected by OCAv3 that were not supported by SNAQ analysis.

Segmental CNV detection is feasible with the OCAv3 platform. Identified thresholds resulted in 100% sensitivity and specificity, although 20% to 30% of CDKN2A and +7/-10 cases require FISH reflex. If validated, this solution affords an efficient strategy in terms of hands-on-time, cost, and tissue use for combined SNV and CNV detection, particularly in a resource-constrained setting.

In this real-world clinical dataset, WTS performed significantly worse than amplicon-based sequencing for detection of clinically meaningful fusions, predominantly due to a high QNS rate. It did not significantly increase the detection rate of novel fusions. Although amplicon-based methods may have limitations due to the inability to detect novel fusions, this can be overcome with design for additional targets, and is balanced by a superior ability to test a wide range of clinical samples without failure.

Our technical criteria for NGS, using leftovers in formalin-fixed and paraffin-embedded blocks after routine pathology diagnosis, were met by around half of EUS-FNBs. Almost all EUS-FNBs fulfilling the technical criteria yielded a successful NGS analysis.

We were able to successfully conduct NGS studies in paraffin-embedded material archived for 10 years using both middlesized (OPA) and large (OCA v3) NGS panels. The identified S3FB1 mutation helps to explain the onset of metastases, even though the patient had spindle cell predominant UM with BAP-1 preservation, which are normally associated with decreased metastases risk. Our case highlights the value of using large NGS panels to unravel the molecular landscape of UM and further understand this unique eye cancer.

Conclusions : In our cohort, KRAS -mutant patients appear to have a different metastatic pattern depending on their KRAS mutation subtype. In addition, KRAS G12C and KRAS G12V -mutant patients seem to have better prognosis and response to chemoimmunotherapy than KRAS G12D -mutant patients.

Yet, to our knowledge, we present herein the first case of ALK + NSCLC rapidly progressing on 1 st line Brigatinib treatment due to de novo MET- amplification. The recognition of this mechanism of ALK-TKI resistance by FISH, especially in NGS-negative cases, should be considered before initiating 1 st line treatment. This is of clinical importance, as effective combined therapy with ALK-TKI and MET-inhibitor is feasible.

With the present treatment possibilities, OCAv3 panel testing provided higher diagnostic yield due to better coverage. Our study emphasizes that WES, although offering the potential to identify novel findings in genes not covered by OCAv3, might overlook variants in genes relevant for OC.

We reported the frequency of HRD and MMR somatic mutations in a large real-world cohort of consecutive primary and metastatic PCs, providing a realistic estimate of the proportion of patients that could benefit from PARP or immune checkpoint inhibitors. Analysis of HR genes beyond BRCA1 and BRCA2 significantly increased the pool of patients that would be eligible for PARP inhibitor trials. BRCA1/BRCA2 and MMR mutations were more frequent in metastatic lesions suggesting these gene defects to be strong metastatic drivers.

According to the current guidelines, the assignment of endometrial carcinomas into the copy number high molecular group is based on p53 immunohistochemistry. We found discrepancies in p53 status between the two methods in 9.1% of our patients. This could be explained by multiclassifier phenotype in 13 patients.

Based on reflex molecular analysis performed on consecutive advanced/metastatic BCs, we found FGFR alterations to be present in approximately 17% of UCs, with FGFR3 being more frequently altered than FGFR1 and FGFR2 genes. These patients could gain clinical benefits from receiving a FGFR inhibitor such as erdafitinib which is an approved drug to treat FGFR-altered BCs lacking response to chemotherapy. Future studies will include a larger cohort to assess the correlation of FGFR alterations with histologic subtypes of invasive UC.

Our study provides novel insights on the role of rare uKRAS mutations in CRC, which seems to be correlated with a more aggressive clinical behavior compared to cKRAS ones. Further investigations are required to confirm these hypothesis generating results.

Due to disease progression, lenvatinib was initiated...Zoledronic acid was initiated for the treatment of the skeletal metastases, and palliative external beam radiation (2000 cGy in 5 fractions) was administered to the sternal lesion to alleviate the bone pain... The NBN gene encodes for the protein nibrin, a member of the MRE11/RAD50 complex that is involved in DNA repair. PVs in NBN are commonly identified in non-thyroidal malignancies such as breast and lung carcinomas. While NBN PVs are rarely reported in DTCs, as per our knowledge, this is the first described case of OTC with a PV in NBN as the molecular driver, manifesting with an aggressive disease course.

These results demonstrate that the TCR repertoire is associated with characteristics, genetic alterations, and clinical outcomes. Monitoring changes in the TCR repertoire may serve as a prognostic biomarker for breast cancer patients.

We assessed the expression of CEACAM5 by IHC on clinical biopsy/resection samples of consecutively diagnosed NSCLC from September 2022 to June 2023 at our institution, using the anti-CEACAM5 clone 769 antibody assay protocol developed for tusamitamab ravtansine clinical trials... Our data showed that approximately 20% of routinely diagnosed clinical NSCLC samples had high CEACAM5 expression by IHC. The interrater reliability on the determination of high CEACAM5 expression was moderate among the three pathologists. We highlighted the challenging aspects in the evaluation of this biomarker.

LB NGS resulted in a higher detection rate for BRCA1/2 alterations in comparison to conventional TT NGS (20% vs 9%). Ideally, BRCA1/2 testing should be based on both approaches to identify all patients with mPC eligible for PARP inhibitor therapy.

This was an ad hoc analysis of a KSCC1301 randomized phase II trial in which patients with untreated resectable LARC were randomly assigned to receive S-1 and oxaliplatin or folinic acid, 5-fluorouracil, and oxaliplatin as NAC...NAC was associated with increased expression of ICMs and TILs. Rectal cancer could be susceptible to combined immunotherapy with chemotherapy.

In summary, PTEN IHC expression was retained in the majority of PTEN- and POLE-mutated ECs with frequent missense mutations in codon 130. Some codon 130 mutations (e.g., R130Q) are known to be associated with protein stability. Nearly half of PTEN-mutated MMR-d ECs were associated with PTEN expression (retained, reduced or subclonal loss).

In summary, our results indicate that PTEN IHC staining is present in majority of ECs with four common dominant-negative PTEN mutations (retained or less commonly reduced). While recent literature suggests complementary use of PTEN IHC and sequencing to assess PTEN status in EC, these results further highlight that IHC may not be helpful in assessment of PTEN abnormality in cases with dominant-negative PTEN mutations.

The frequency of Tier 1/2 HRD in metastatic PC is twice that of primary PC (p=0.001). No association was identified between CC/IDC morphology and HRD in primary cases, or between HRD and site of metastases. Additional work exploring less frequent HR genes in primary and metastatic PC is ongoing.

"Five genes showed a different mutational status across samples and/or panels: 4 discordant results involved NBF samples. In conclusion, acid-deprived fixatives (GAF and ADF) guarantee the highest DNA preservation/sequencing performance, thus allowing more complex molecular profiling of tissue samples."

Conclusion Tumor-based NGS frequently provides clinically significant information. It holds the potential to reveal specific therapeutic targets and contribute to clinical decision-making

Our study depicted the mutational patterns of BRCA1, BRCA2, and PALB2 in Taiwanese breast cancer patients through tumor-only sequencing. This highlights the growing importance of BRCA1/2 and PALB2 alterations in breast cancer susceptibility risk and the treatment of index patients. We also emphasized the need to meticulously annotate variants in cancer-driver genes as well as actionable mutations across multiple databases.

Key eligibility criteria include allowing hydroxyurea cytoreduction to a WBC <10,000/mm3 for proliferative disease and excluding pts with need for emergent disease-directed therapy as well as prior therapy for myeloid malignancy other than hydroxyurea or thrombopoietic/erythropoietin stimulating agents. Planned exploratory studies include the use of: 1) RNA-seq to evaluate if differences in baseline transcriptional programs appear to be predictive of response to ASTX727 and ASTX727+venetoclax, 2) next generation (Oncomine Comprehensive Assay (OCAv3)/whole exome sequencing to uncover biomarkers predictive of response/outcome, validate risk modeling to stratify these pts and detect/monitor measurable residual disease (MRD) on serial marrow samples that can predict depth of response with translation in improvement in event-based endpoints, and 3) circulating tumor DNA as a non-invasive means (i.e., without need for a marrow evaluation) of prospective MRD detection for prognostic use. This ongoing, randomized study may inform frontline therapeutic options, provide prospective data for venetoclax combination after HMA failure, and generate striking correlative science for pts with "high risk" MDS/MPN.

Fusions are overall uncommon events in endometrial carcinoma. We present a rare case of an FGFR3::TACC3 fusion in a case of endometrial carcinoma. In our institutional cohort, we demonstrated that FGFR3::TACC3 fusions are most frequently identified in high-grade gliomas, but can be seen in other tumor types as well.

HER2 status was determined by IHC/FISH and NGS in 45 cases: 19 serous, 18 high-grade (unspecified histologic type, HGU), 5 endometrioid 1 carcinosarcoma, 1 undifferentiated and 1 gastric- type. IHC-FISH HER2 was positive in 11/45 (24.4%) of cases, all serous or HGU. NGS identified 3 cases as amplified (concordant) and 1 non-pathogenic SNV.

The most prevalent mutant genes were TP53 (68.3%) and KRAS (53.5%), while the PIK3CA (39.4%), AKT1 (45.9%), and ERBB2 (17.1%) mutations constituted biomarkers for FDA-approved therapeutics. Our study showed that tumor tissue should be the source of actionable mutation detection for early breast cancers, considering that the concordance rate between tumor and liquid biopsy was only one-quarter.

Case 1: 73-year-old woman, January 2014 retroperitoneal leiomyosarcoma of the ureter - R0 en bloc resection with partial resection of the ureter and left hemicolectomy - Transcriptome-guided adjuvant treatment: Sorafenib and Riluzole due to GRM1 gene overexpression - Scapular recurrence with bone infiltration in Sept...Treatment with Tazemetostat in December 2022...- Neoadjuvant chemotherapy and radiotherapy treatment, R0 wide surgery with adequate preservation of functionality and IORT - Early chest wall dermal/subcutaneous recurrence (resected) after initiation of immunotherapy – Nivolumab – due to a favorable profile for immunotherapy in tumor transcriptome - Genome of recurrence: BRAF V600E mutation. Treatment with Dabrafenib + Trametinib... STS are tumors with complex surgical and oncological management and a poor response to conventional chemotherapy protocols. Given its heterogeneity, precision oncology through molecular profiling with next-generation massive sequencing (panels for tumor genome and transcriptome) allows the development of personalized therapeutic strategies in patients with STS, and opens up the possibilities for targeted treatments for molecular alterations, immunotherapy and the development of new agents.

Accordingly, the patient continued Osimertinib combined with Carboplatin/Pemetrexed and with additional palliative irradiation due to a new symptomatic spinal cord compression at L3. Acquired EML4-ALK fusion represents an extremely rare (1%) acquired mechanism of Osimertinib-resistance, which enables further effective and feasible therapy by combining EGFR- and ALK-TKI. Rebiopsies, when possible, are of noteworthy value even in heavily pretreated and fragile patients.

In our cohort, the most frequent KRAS mutation found was G12C and the most frequent co-mutation was TP53. We found a high response rate and a long PFS of KRAS mutant patients treated with chemoimmunotherapy or immunotherapy. Further prospective studies are needed to derive definitive conclusions.

The patient initially received 1st line Erlotinib with remission maintained more than 4 years, despite an early dose reduction due to skin toxicity. Despite loss of the original compound EGFR mutation, the rebiopsy revealed a new pathogenic and druggable driver (BRAF p.V600E), which gave the patient a new possibility of further treatment. Yet, replacing Osimertinib with a BRAF/MEK-Is combination resulted only in mixed response, suggesting that some lesions might have remained, at least in part, dependent on the original EGFR mutations. Indeed, re-challenge with Osimertinib in reduced dose together with continuation of Dabrafenib and Trametinib resulted in SD and reflected spatial and temporal heterogeneity of NSCLC.

In conclusion, we present a mutational landscape of actionable genes in CRC for the SA population in patients without a familial history of CRC. Additionally, we address the clinical relevance of low variant allele frequency variants. A comprehensive analysis of population-specific molecular markers for CRC can provide insights into the disease progression, with an aim of optimizing individualized therapeutic options and treatment regimens in the management of CRC in the SA population.

With this approach, the detection rate of NTRK fusions was 20 times higher (5.57%) by only screening 323 patients than the largest cohort in the literature (0.30%) comprising several hundred thousand patients. Based on our findings, we propose a multiparametric strategy (ie, "supervised tumor-agnostic approach") when pathologists start searching for NTRK fusions.

NERoNE evaluates data from NGS profiling of three OU mapping key actionable molecular alterations in a large population of aNSCLC also accounting for PDL1 status. This preliminary analysis will be the starting point for further evaluations in order to improve the personalized approach to aNSCLC. >*For 15 pts PDL1 expression was not known.

As more comprehensive biomarker testing became available, more AGAs are identified with an increase in TAT from biopsy to sign-out and treatment start. Despite our institutional policy of reflex testing, future endeavours will be focus on ways to reduce TAT. >

Palbociclib was used in 80%, ribociclib in 17% and abemaciclib in 3%, mostly as early lines (1L 65%, 2L 22%). Pathogenic gBRCA1/2-PALB2 and higher aneuploidy were associated with shorter survival. Profiling of aneuploidy, HRD, TMB and APOBEC in all pts is ongoing including 55 baseline ctDNA samples and 361 samples from the GENIE BPC cohort. Results will account for FDR, left truncation and optimal cutoffs.

Our results revealed the mutational frequencies of BRCA1, BRCA2, and PALB2 with a commercial targeted sequencing panel. The prevalence of BRCA1, BRCA2, and PALB2 genomic alterations were 3%, 5%, and 8%, respectively. Co-occurrence among these 3 genes was observed.

P=N/A; Trial completion date: Dec 2022 --> Dec 2024 | Trial primary completion date: Dec 2022 --> Dec 2024

Overall, targetable gene fusions have a prevalence of 7%. Moreover, 38% of overall cases harbor mutations in genes coding for tyrosine kinases potentially targetable and/or have defects in the MMR that claim a potential role for immunotherapy.

Targeted sequencing with Oncomine Comprehensive Assay V3 did not reveal particular mutational patterns between the two Luminal A groups. Nevertheless, radiomic analysis of mammographic images of the cancer, as well as single cell phenotyping of the tumor microenvironment with protein multiplexing will be incorporated to further characterise elements that are phenotypically different, and potentially identify mechanistic drivers contributing to late recurrence in Luminal A cancers.

Our study demonstrated that amplicon-based NGS assays can be routinely applied to clinical samples to detect amplification. However, they are not sufficiently sensitive in detecting amplification when tumor celluarity is less than 30%, which is a major limitation that should be clearly stated in NGS reporting.