TEST:

VENTANA ALK (D5F3) CDx Assay

Our inhouse panel was associated with rapid TAT and faster initiation of systemic treatment. Approximately 30% of patients would not need tissue sent for NGS based on inhouse detection of a potentially actionable mutation.

The NGS testing revealed EML4::ALK fusion in three, STRN::ALK fusion in two, and ITSN2::ALK fusion in one. In conclusion, although ALK-rearranged PTCs have been associated with neutral gene expression profile from a BRAF-RAS scoring perspective, the "RAS-like" nuclear features were more commonly identified in this series, resulting in frequent indeterminate diagnosis of preoperative FNA.

ALK was highly expressed in NB. ALK expression was not related to several clinical and histopathological features. More studies are needed to elucidate the association between ALK expression and ALK gene status and to investigate disease progression, especially the oncogenesis of ALK-positive NB.

P1/2; N=12; Recruiting; Sponsor:University of Colorado, Denver

Immunostaining is a robust method for ALK-rearrangement testing, with fluorescence in situ hybridization adding value in the rare equivocal stained case. ROS1-rearrangement testing is more cost-effective if immunohistochemistry is followed by fluorescence in situ hybridization after excluding EGFR-mutant and ALK-rearranged adenocarcinomas.

Following corticosteroid treatment and discontinuation of alectinib, clinical presentations and CT scan gradually improved, but the primary lung lesions enlarged in the regular follow-up. The administration of a first-generation ALK inhibitor crizotinib was then initiated and the disease was stable for 25 months without recurrence of ILD.

Immunohistochemistry is a useful tool for confirming the Spitz histogenesis of a melanocytic tumour when it demonstrates the presence of expression of a kinase fusion. Our data suggest that the ALK-1 antibody is significantly less sensitive than ALK-D5F3 and ALK-5A4 antibodies for the detection of ALK-rearranged Spitz tumors and we do not recommend its use for this purpose.

Immunohistochemistry-based assays provide a valid and reasonably priced alternative, especially in settings where molecular confirmatory tests are neither offered nor accessible. Given high interassay and molecular concordance, we propose that the novel Dako OTI1A4 assay can be reliably used to identify cases with ALK rearrangement.

Conclusion Our study confirms that compared to IHC, NGS is a more accurate first line testing platform for the detection of consensus ALK fusion status. However access to IHC may be required to mitigate the higher tissue requirements of NGS and access to FISH may be beneficial to resolve atypical results.

P1/2; Not yet recruiting --> Recruiting | Initiation date: Dec 2023 --> Jul 2023

Gene co-occurrence survival analysis showed that mPFS of pts harboring FAT3/FAT4 mutations were significantly shorter than those without FAT3/FAT4 mutations (8.9 mo vs 16.5 mo, p=0.012). Conclusions Clinical utility of ctDNA was demonstrated, not only at baseline by identifying fusion types and subgroups of ALK+ NSCLC pts that may benefit more from SAF-189s, but also at progression by tracking ALK-resistant mutations.

Ventana ALK (D5F3) immunohistochemistry improved the accuracy of pathologic assessment after neoadjuvant ALK-TKIs for ALK-positive NSCLC. It is recommended to use Ventana ALK D5F3 immunohistochemistry to help confirm MPR and PCR in future pathological assessment work.

In this regard, Dako-CD246 proves to yield concordant results with Ventana-D5F3, which is already promptly used in routine pathology practice. The present study being the first in the field, given its high inter-assay and molecular concordance, we propose that novel Dako-CD246 assay can be reliably used in identifying cases for ALK-targeted therapy.

P3; Trial primary completion date: Apr 2023 --> Apr 2024

P1/2; N=12; Not yet recruiting; Sponsor:University of Colorado, Denver

P2; Trial completion date: Aug 2023 --> Dec 2024

In this case series, all six ALK -rearranged PTCs demonstrated a predominant follicular growth pattern and five of them were diagnosed as infiltrative follicular variant. The nuclear features were subtle and might be present in only focal areas, resulting in a significant rate of false negative diagnosis (33%, 2/6) in pre-operative FNA.

We identified a subset of microcystic/reticular schwannomas with NONO::TFE3 fusions. TFE3 and ALK-D5F3 immunohistochemistry appear to be reliable surrogates for fusion status.

P3; Recruiting --> Active, not recruiting

P2; Trial completion date: Aug 2022 --> Aug 2023

In Taiwan, the patients with thoracic IMT were young, and had early stages of disease status. ALK rearrangement and NTRK3 overexpression were the major genetic alterations. Different partners of ALK arrangement could be detected.

ALK-positive patients experience clinical benefit from pemetrexed-based chemotherapy possibly due to lower thymidylate synthase (TS) levels...D5F3 and 5A4 clones have the highest percentage of agreement. TS levels are significantly lower in FISH-positive patients.

An Archer FusionPlex panel assay demonstrated a fusion involving both exons 4 from the JAZF1 and BCORL1 genes. The JAZF1::BCORL1 has not, to the best of our knowledge, been previously reported in a benign/low-grade mesenchymal uterine lesion.

P2; Active, not recruiting --> Completed | Trial completion date: May 2022 --> Jul 2021

Patients with advanced ALK-positive (by immunohistochemistry) NSCLC were randomized 1:1 to alectinib 600 mg or crizotinib 250 mg, twice daily. Reasonable concordance between FACT and immunohistochemistry was observed; both methods are valuable in identifying ALK-positive patients, separately or concurrently. Alectinib was found to have superior PFS in the plasma ALK-positive population, as in the ITT population.

P2; Recruiting --> Active, not recruiting

"Moreover, the patient achieved an admirable response to alectinib, with a clinical evaluation of complete response (CR). In summary, our findings expand the spectrum of ALK fusion patterns and provide robust evidence for the precise administration of alectinib in the future."

P2; N=12; Not yet recruiting; Sponsor:Yonsei University

Overall, D5F3 showed greater stain intensity and proportion staining than ALK1 in ALK-positive ALCL cases, which is especially helpful in limited samples. Caution and consideration of orthogonal ALK testing types is recommended, especially for cases with weak or focal staining.

Four ALK-positive cases were male and ALK-positive patients ranged from 42-55 years with median 48. Despite the incidence of ALk-AdC is uncommon, it might be suspected in lesions of lower age group especially with mucinous foci.

P3; Trial primary completion date: Jan 2022 --> Apr 2023

We unexpectedly found that this GIST patient showed ALK (D5F3) overexpression and harbored a novel fusion CDC42BPB exon 24-ALK in exon 20.

Non-CB cytology smears are highly suitable for multiplex FISH analysis with 100% concordance with FFPE FISH and/or ALK D5F3 companion diagnostics assay results.

Outcomes of patients with ALK IHC-positive/FISH-positive and ALK IHC-positive/FISH-uninformative NSCLC were similar to the overall ALEX population. These results suggest that Ventana ALK IHC is a standard testing method for selecting patients for treatment with alectinib.

The use of combined ALK-ROS FISH allows for excellent demonstration of ALK and ROS1 rearrangements in the same section using the same criteria validated in single gene break apart FISH probes. ROS1 protein is commonly overexpressed in ALK rearranged NSCLC and may represent cross reactivity with ALK or a related protein.

Moreover, CNS protection also needed to be taken into consideration in the adjuvant setting. IA IB II III 2year DFS 96.6% (95%CI: 91.1%-98.7%）94.0% (95%CI: 77.9%-98.5%) 79.2% (95%CI: 60.8%-89.6%) 46.8% (95%CI: 32.7%-59.7%) 3year DFS 96.6%（95%CI：91.1%-98.7%）87.3% (95%CI: 62.4%-96.1%) 63.2% (95%CI: 40.3%-79.3%) 30.7% (95%CI: 15.7%-47.1%)

Twenty-seven treatment outcomes from 20 cases were assessed and cases expressing more than half of the specific predictive proteins were sensitive to matching therapeutic agents and showed longer progression-free survival than the other cases (p < 0.001). SRM showed a potential role in therapeutic decision making in NSCLC patients with ambiguous ALK test results.

We report here 4 sarcomas with ALK rearrangement, including a novel fusion TMEM217-ALK . Two of these cases had a 5'ALK loss in addition to the 3'ALK rearrangement, and both had much longer PFS on Crizotinib therapy . The underlying mechanism deserved further investigation .

This single-center study exhibited a large subset of small lung biopsies from a Chinese institution and demonstrated that applying the 2015 WHO classification for small lung biopsies can help predict the mutational status of primary lung carcinomas.

Twenty-two patients were alive with no evidence of tumor, four patients had tumor recurrences (three patients were treated with crizotinib and were alive with tumor), and three patients died of the disease (including two patients with EIMS). IMTs show a wide morphologic spectrum, and should be differentiated form a variety of benign or malignant tumors. Immunohistochemistry (ALKp80, ALKD5F3) and FISH (ALK break-apart probe) could assist the diagnosis of IMT, with NGS recommended for the atypical cases.

P3, N=246, Recruiting, Takeda | Trial completion date: Dec 2022 --> May 2024

An ALK IHC "equivocal" interpretation category should be incorporated into practice. Atypical patterns of ROS1 and genomic heterogeneity need to be evaluated further for any clinical relevance.