Anaplastic Lymphoma Kinase Mutations Research Articles

Abstract LB414: Novel ALK/PIK3CA cooperativity in head and neck squamous cell carcinoma (HNSCC) progression

Abstract Recent genomic characterization studies have demonstrated the genetic heterogeneity of head and neck squamous cell carcinoma (HNSCC). Yet, most of these genetic aberrations remain undruggable. Anaplastic Lymphoma Kinase (ALK) is somatically mutated in ~3.6% of HNSCC (18/504 cases, TCGA-HNSCC Firehose cohort, USA), and the majority of these ALK-mutated HNSCC cases are of advanced stage diseases. Here, we hypothesized that ALK mutations might plausibly contribute to HNSCC progression and could be potentially druggable. Unexpectedly, analysis of HNSCC genomic events revealed a novel co-occurrence relationship between ALK and PIK3CA genomic aberrations in TCGA-HNSCC Firehose cohort (Odds Ratio=2.41; N=522 samples with CNV data), with subsequent validation in an independent metastatic MSK-MetTropism HNSCC cohort (Odds Ratio=5.12; N=412). Consistent with this newly identified co-occurrence relationship, we showed that all 7 HNSCC-relevant ALK point mutations were potent drivers for HNSCC invasiveness and clonogenic growth only in corroboration with PIK3CA aberrations in HNSCC cell models, highlighting the functional importance of this novel ALK/PIK3CA cooperativity in HNSCC, likely reminiscent of ALK/MYCN cooperativity in neuroblastoma. Further, by clonogenic assay, we demonstrated pharmacologic vulnerabilities of all 7 ALK point mutations (including extracellular domain mutations) to ALK targeting by ceritinib in FaDu cell background with endogenous PIK3CA amplification. Strikingly, we were able to capture, for the first time, spontaneous emergence of a druggable ALK somatic mutation directly from a PIK3CA-altered HNSCC patient-derived culture (PDC) upon forced acquisition of anoikis-resistance ex vivo (PDC-25-AR), demonstrating co-evolution of ALK/PIK3CA aberrations in HNSCC likely associated with cancer progression. Patient-derived culture-xenograft (PDCx) of PDC-25-AR displayed remarkable sensitivity to ALK targeting, PI3K targeting, as well as their combination, with induction of apoptosis in these tumors compared to vehicle control. In conclusion, ALK aberrations corroborate with PIK3CA aberrations to promote HNSCC progression, yet, these events are potentially druggable in advanced HNSCC. Citation Format: Lan Wang, Helen Hoi-Yin Chan, Angel Yee Lok Fung, Yuen-Keng Ng, Patrick Kwok-Shing Ng, Yuxiong Su, Jason Ying Kuen Chan, Peony Hiu Yan Poon, Thomas Chun Kit Yeung, Chi Man Tsang, Hoi-Lam Ngan, Wenying Piao, Yuchen Liu, Jack Nimitz Smith, Tin Yu Samuel Law, Ee Ling Kong, Melissa Sue Ann Chan, Chun Ho Law, Sze-Man Chan, Zoey Wing Yee Liu, Joshua Chung-Hymn Ng, Gordon B. Mills, Vivian W.Y. Lui. Novel ALK/PIK3CA cooperativity in head and neck squamous cell carcinoma (HNSCC) progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB414.

Abstract 987: Novel ALK mutations in EML4::ALK+ NSCLC resistant to TKIs identified by liquid biopsy

Abstract The identification of Anaplastic Lymphoma Kinase (ALK) chromosomal translocations led to the development of effective targeted therapies in a subset of Non-Small Cell Lung Cancer (NSCLC) patients. Several drugs are currently in clinical practice for advanced ALK+ NSCLC. However, resistance remains a challenge in these patients. The possibility to analyze circulating tumor DNA (ctDNA) with non-invasive methods can greatly improve prognostic capabilities, by allowing early detection of relapse and of drug-resistant mutations from peripheral blood. We report a series of 15 consecutive relapsed NSCLC patients (12 ALK+, 3 ROS1+) treated at the Fondazione IRCCS-San Gerardo dei Tintori (Monza, Italy) that were investigated by liquid biopsy at tyrosine kinase inhibitor (TKI) failure, through amplicon deep sequencing of the ALK/ROS1 kinase domains. Median DNA yield was 4.8 ng per ml of plasma and mean fragment size was 176 bp. Mutational hotspot coding regions of ALK and ROS1 genes were amplified by high-fidelity PCR and sequenced. Mean coverage of called variants was 24,687x. The threshold for mutation calling was set at 0.6%, after correcting for mapping quality, minimum coverage, and strand bias. Analysis of background signal on healthy control DNA identified 0.0024 mismatches per mapped base. Overall, plasma ctDNA analysis detected an ALK mutation in 7/18 relapse samples from 7/15 patients. No ROS1 mutations were found in ROS1 fusion-positive patients. Among 12 patients carrying an EML4::ALK fusion, 10 variants potentially driving resistance were found. Along with mutations that were previously identified (L1196M/G1202R, E1154K, F1174L), novel ALK variants were detected such as L1198R, T1211I, C1235R, C1237Y, L1196P. Mutations were characterized for sensitivity to ALK inhibitors in a BaF3-EML4::ALK cell model, using cell proliferation assays and western blotting to evaluate the effects on ALK phosphorylation. C1237Y and L1196P mutants appeared to resist all TKIs; hence, we further tested their sensitivity to novel investigational drugs, zotizalkib and repotrectinib. Both mutants demonstrated resistance, suggesting that these two mutations confer broad resistance to TKIs. Molecular modelling of C1237Y and L1196P mutants was run to elucidate the mechanisms of resistance. The ADP-C1237Y complex showed a much smaller root-mean-square deviation than ADP-WT, indicating a higher affinity/activity of the mutant. L1196 interacts hydrophobically with a methyl group of lorlatinib, P1196 is predicted to lose such interaction, while ATP binding is not affected. The presence of C1237Y and L1196P variants, is a relevant finding due to their resistance, even after the exposure to the fourth-generation inhibitor zotizalkib. The clonal selection of these mutations, made by second/third generation ALK TKIs may lead to a complete refractory status, which would then render TKIs useless in a sequential therapeutic strategy. Citation Format: Federica Malighetti, Matteo Villa, Elisa Sala, Geeta Sharma, Giulia Arosio, Maria Gemelli, Chiara Manfroni, Diletta Fontana, Nicoletta Cordani, Raffaella Meneveri, Alfonso Zambon, Rocco Piazza, Fabio Pagni, Diego Cortinovis, Luca Mologni. Novel ALK mutations in EML4::ALK+ NSCLC resistant to TKIs identified by liquid biopsy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 987.

Abstract 3357: Identifying mechanisms of acquired resistance to lorlatinib utilizing a genome-wide CRISPR-Cas9 screen

Abstract Survival rates for neuroblastoma (NB), a predominantly pediatric solid tumor arising from the sympathoadrenal neural crest, have improved incrementally from 25% to 50% over three decades with dramatic escalation in therapy intensity. Gain-of-function mutations in anaplastic lymphoma kinase (ALK), of which 85% occur at residues F1174, F1245, and R1275, confer inferior survival compared to patients with wild-type ALK. Lorlatinib, a third generation ALK inhibitor, has completed Phase 1 testing in the NANT Consortium and has moved to frontline therapy in the Children’s Oncology Group. The robust and sustained activity observed with minimal toxicity is transient in relapse patients harboring MYCN amplification and ALK activation. We aim to elucidate and target mechanisms of acquired resistance that emerge from lorlatinib treatment. We utilized the Brunello library, a genome-wide CRISPR-CAS9 loss of function screen targeting 19,114 genes (4 sgRNA per gene), which allows for a high confidence of detection of hits. We screened four clinically representative cell lines: Kelly (ALKF1174L, MYCN-amplified, p53P117T), SMS-SAN (ALKF1174L, MYCN-amplified), LAN5 (ALKR1275Q, MYCN-amplified), and SY5Y (ALKF1174L, MYCN-nonamplified). sgRNAs were ranked according to the degree of depletion or enrichment. We validated top depleted targets through cell-based assays and immunoblotting. We identified several actionable targets across multiple NB cell lines that, when depleted, increase the efficacy of lorlatinib, including FGFR2, BCL2L1, SOX11, SMARCA2, and SMARCD2. Genetic knockout of FGFR2 led to a 40% reduction (p<0.05) in KO cells upon lorlatinib treatment compared to control. PROTAC degradation of the anti-apoptotic BCL-xL (BCL2L1) revealed the combination of lorlatinib and the PROTAC led to a decrease in IC50 in vitro. Additionally, we identified conserved core components of the SWI/SNF complex, a subfamily of ATP-dependent chromatin remodelers, as well as SOX11, a regulator of the SWI/SNF complex and transcription factor of the core regulatory circuitry in adrenergic high-risk NB. In conclusion, we identified actionable targets across four NB cell lines that, when depleted, increase the efficacy of lorlatinib. Depletion of FGFR2 led to increased sensitivity to lorlatinib, however enzymatic inhibition showed minimal effect. Future investigation into dual targeting of ALK and FGFR2 through novel alternative methods is warranted. Furthermore, we observed a synergistic relationship between lorlatinib and a BCL-xL PROTAC in vitro. Ongoing in vivo studies will test the combinatorial efficacy in lorlatinib-naïve and resistant models. Finally, we are investigating epigenetic remodeling by the SWI/SNF complex and its regulators in NB cells during lorlatinib treatment through targeted genetic manipulation of SOX11 to understand its role as a master transcriptional regulator of MYCN and FGFR2. Citation Format: Joshua Renn Kalna, Smita Matkar, Emily O'Drisoll, Skye Balyasny, Matteo Calafatti, Mark Gerelus, Dave Groff, Tina Acholla, Colleen E. Casey, Paul Kamitsuka, Grant Li, Steven J. Pastor, Gabriela Witek, Kateryna Krytska, Jarrett Lindsay, Ophir Shalem, Yael P. Mossé. Identifying mechanisms of acquired resistance to lorlatinib utilizing a genome-wide CRISPR-Cas9 screen [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3357.

Discovery of novel anaplastic lymphoma kinase (ALK) and histone deacetylase (HDAC) dual inhibitors exhibiting antiproliferative activity against non-small cell lung cancer

A series of novel benzimidazole derivatives were designed and synthesised based on the structures of reported oral available ALK inhibitor and HDAC inhibitor, pracinostat. In enzymatic assays, compound 3b, containing a 2-acyliminobenzimidazole moiety and hydroxamic acid side chain, could inhibit both ALK and HDAC6 (IC50 = 16 nM and 1.03 µM, respectively). Compound 3b also inhibited various ALK mutants known to be involved in crizotinib resistance, including mutant L1196M (IC50, 4.9 nM). Moreover, 3b inhibited the proliferation of several cancer cell lines, including ALK-addicted H2228 cells. To evaluate its potential for treating cancers in vivo, 3b was used in a human A549 xenograft model with BALB/c nude mice. At 20 mg/kg, 3b inhibited tumour growth by 85% yet had a negligible effect on mean body weight. These results suggest a attracting route for the further research and optimisation of dual ALK/HDAC inhibitors.

Updated overall survival and circulating tumor DNA analysis of ensartinib for crizotinib-refractory ALK-positive NSCLC from a phase II study.

The initial phase II stuty (NCT03215693) demonstrated that ensartinib has shown clinical activity in patients with advanced crizotinib-refractory, anaplastic lymphoma kinase (ALK)-positive non-small cell lung cancer (NSCLC). Herein, we reported the updated data on overall survival (OS) and molecular profiling from the initial phase II study. In this study, 180 patients received 225 mg of ensartinib orally once daily until disease progression, death or withdrawal. OS was estimated by Kaplan-Meier methods with two-sided 95% confidence intervals (CIs). Next-generation sequencing was employed to explore prognostic biomarkers based on plasma samples collected at baseline and after initiating ensartinib. Circulating tumor DNA (ctDNA) was detected to dynamically monitor the genomic alternations during treatment and indicate the existence of molecular residual disease, facilitating improvement of clinical management. At the data cut-off date (August 31, 2022), with a median follow-up time of 53.2 months, 97 of 180 (53.9%) patients had died. The median OS was 42.8 months (95% CI: 29.3-53.2 months). A total of 333 plasma samples from 168 patients were included for ctDNA analysis. An inferior OS correlated significantly with baseline ALK or tumor protein 53 (TP53) mutation. In addition, patients with concurrent TP53 mutations had shorter OS than those without concurrent TP53 mutations. High ctDNA levels evaluated by variant allele frequency (VAF) and haploid genome equivalents per milliliter of plasma (hGE/mL) at baseline were associated with poor OS. Additionally, patients with ctDNA clearance at 6 weeks and slow ascent growth had dramatically longer OS than those with ctDNA residual and fast ascent growth, respectively. Furthermore, patients who had a lower tumor burden, as evaluated by the diameter of target lesions, had a longer OS. Multivariate Cox regression analysis further uncovered the independent prognostic values of bone metastases, higher hGE, and elevated ALK mutation abundance at 6 weeks. Ensartinib led to a favorable OS in patients with advanced, crizotinib-resistant, and ALK-positive NSCLC. Quantification of ctDNA levels also provided valuable prognostic information for risk stratification.

A Review on Anaplastic Lymphoma Kinase (ALK) Rearrangements and Mutations: Implications for Gastric Carcinogenesis and Target Therapy.

Gastric adenocarcinoma is a complex disease with diverse genetic modifications, including Anaplastic Lymphoma Kinase (ALK) gene changes. The ALK gene is located on chromosome 2p23 and encodes a receptor tyrosine kinase that plays a crucial role in embryonic development and cellular differentiation. ALK alterations can result from gene fusion, mutation, amplification, or overexpression in gastric adenocarcinoma. Fusion occurs when the ALK gene fuses with another gene, resulting in a chimeric protein with constitutive kinase activity and promoting oncogenesis. ALK mutations are less common but can also result in the activation of ALK signaling pathways. Targeted therapies for ALK variations in gastric adenocarcinoma have been developed, including ALK inhibitors that have shown promising results in pre-clinical studies. Future studies are needed to elucidate the ALK role in gastric cancer and to identify predictive biomarkers to improve patient selection for targeted therapy. Overall, ALK alterations are a relevant biomarker for gastric adenocarcinoma treatment and targeted therapies for ALK may improve patients' overall survival.

Anaplastic Lymphoma Kinase (ALK) Inhibitors Show Activity in Colorectal Cancer With ALK Rearrangements: Case Series and Literature Review.

Anaplastic lymphoma kinase (ALK) rearrangement is a well-known driver oncogene detected in approximately 5% of non-small cell lung cancer. However, ALK rearrangement is much less frequent in other solid tumors outside the lungs, such as colorectal cancer (CRC); thus, the optimal management of CRC with ALK rearrangements has yet to be established. In this report, we describe 2 cases of ALK-positive CRC, both of which benefited from ALK tyrosine kinase inhibitor (ALK-TKI) therapy. Case 1 was a postoperative patient with poorly differentiated colon adenocarcinoma, who was diagnosed with metastatic relapse shortly after surgery. Both fluorouracil, leucovorin, and oxaliplatin (FOLFOX) and bevacizumab combined with 5-fluorouracil, l-leucovorin, and irinotecan (FOLFIRI) proved ineffective against the disease. The patient was then treated with ensartinib, as the CAD-ALK fusion gene was detected by genomic analysis. The patient was initially treated with ensartinib monotherapy for 9 months, then with ensartinib combined with local radiotherapy and fruquintinib for another 4 months for isolated hilar hepatic lymph node metastasis. The patient experienced disease progression with an acquired ALK G1202R resistance mutation that responded well to lorlatinib. Case 2 involved a 72-year-old man with advanced colon cancer (pT4bN2aM1b, stage IV) harboring an EML4-ALK fusion. The patient underwent resection of the right colon tumor due to intestinal obstruction, but the disease continued to progress after 12 courses of FOLFIRI and bevacizumab chemotherapy. However, the patient responded remarkably well to alectinib. Our report emphasizes the importance of gene detection in the treatment of malignant tumors, and the significance of ALK mutations in CRC.

Mutant anaplastic lymphoma kinase inhibitor identification by integrated in silico approaches

ABSTRACT Non-small-cell lung cancer (NSCLC) is the primary form of lung cancer globally and remains a leading cause of mortality. Anaplastic lymphoma kinase (ALK) mutations, such as I1171N + L1198H, have been discovered to confer resistance to current ALK inhibitors, reducing their therapeutic effectiveness. Addressing drug resistance necessitates exploring selective inhibitors for innovative therapeutic approaches. In this study, a structure-based pharmacophore model, using ALK-approved inhibitors, was developed to screen an In-house database for potential mutant ALK inhibitors. Compounds with requisite pharmacophoric features were evaluated for binding potential against the I1171N + L1198H ALK mutant phenotype. Selected hits underwent assessment for chemical reactivity, and dynamics stability. The study identified five chemical scaffolds (NS1-5) with favorable binding modes and pharmacokinetic properties. The conformational ensembles featured the average RMSD values, ranging from 0.4 to 0.6 nm. RMSF analysis revealed consistent side chain fluctuations with reduced flexibility, while Rog analysis indicated convergence of most complexes. NS1 and NS5, in particular emerged as promising candidates, exhibiting remarkable performance than others, with binding free energies of −210.12 ± 9.94 and −163.68 ± 11.14 kcal/mol, respectively. These findings thus suggest further exploration and optimisation of NS1 and NS5 for mutant ALK inhibitors for the treatment of NSCLC.

Cumulative incidence and risk factors of brain metastases in metastatic non-small cell lung cancer without baseline brain metastasis: Pooled analysis of individualized patient data from IMpower130, IMpower131, and IMpower150.

The objective of this study was to explore the abilities of atezolizumab plus chemotherapy in preventing brain metastases (BMs) among metastatic non-small cell lung cancer (NSCLC) without initial BMs, as well as the risk factors of BMs. Individual patient data from three trials involving first-line atezolizumab for metastatic NSCLC (IMpower130, IMpower131, and IMpower150) were pooled. Among patients without baseline BMs and without epidermal growth factor receptor(EGFR) and/or anaplastic lymphoma kinase (ALK) mutations, those receiving atezolizumab+chemotherapy±bevacizumab were classified as the atezolizumabplus chemotherapy group and those receiving placebo+chemotherapy±bevacizumab were classified as the chemotherapy group. The cumulative incidences of BM (CI-BMs) between the two groups were compared. Other factors associated with the CI-BM were analyzed by Cox regression analyses. With a median follow-up of 17.6 months (range, 0.03-33.64 months), 74 (3.1%) of the 2380 enrolled patients developed BMs, including 50 (3.1%) and 24 (3.0%) in the atezolizumab plus chemotherapy group (n=1589) and the chemotherapy group (n=791), respectively. The CI-BMs at 6, 12, and 24months were 1.7%, 2.8%, and 3.3%, respectively. After taking competing risk events into account, there was no significant difference in the CI-BMs between the two groups (p=.888). Nevertheless, the use of bevacizumab and the histology of nonsquamous NSCLC were found to be independently associated with the risk of BMs. In patients with metastatic EGFR/ALK wild-type NSCLC without baseline BMs, adding atezolizumab in the first-line treatment might not reduce the CI-BM. However, the administration of bevacizumab may reduce the risk of BMs.

Prognostic factors for invasive mucinous adenocarcinoma of the lung: systematic review and meta-analysis

BackgroundInvasive mucinous adenocarcinoma of the lung (IMA) is a unique and rare subtype of lung adenocarcinoma with poorly defined prognostic factors and highly controversial studies. Hence, this study aimed to comprehensively identify and summarize the prognostic factors associated with IMA.MethodsA comprehensive search of relevant literature was conducted in the PubMed, Embase, Cochrane, and Web of Science databases from their inception until June 2023. The pooled hazard ratio (HR) and corresponding 95% confidence intervals (CI) of overall survival (OS) and/or disease-free survival (DFS) were obtained to evaluate potential prognostic factors.ResultsA total of 1062 patients from 11 studies were included. In univariate analysis, we found that gender, age, TNM stage, smoking history, lymph node metastasis, pleural metastasis, spread through air spaces (STAS), tumor size, pathological grade, computed tomography (CT) findings of consolidative-type morphology, pneumonia type, and well-defined heterogeneous ground-glass opacity (GGO) were risk factors for IMA, and spiculated margin sign was a protective factor. In multivariate analysis, smoking history, lymph node metastasis, pathological grade, STAS, tumor size, and pneumonia type sign were found to be risk factors. There was not enough evidence that epidermal growth factor receptor (EGFR) mutations, anaplastic lymphoma kinase (ALK) mutations, CT signs of lobulated margin, and air bronchogram were related to the prognosis for IMA.ConclusionIn this study, we comprehensively analyzed prognostic factors for invasive mucinous adenocarcinoma of the lung in univariate and multivariate analyses of OS and/or DFS. Finally, 12 risk factors and 1 protective factor were identified. These findings may help guide the clinical management of patients with invasive mucinous adenocarcinoma of the lung.

Overcoming Central β-Sheet #6 (Cβ6) ALK Mutation (L1256F), TP53 Mutations and Short Forms of EML4-ALK v3/b and v5a/b Splice Variants are the Unmet Need That a Re-Imagined 5th-Generation (5G) ALK TKI Must Deliver.

Despite the development and approval of seven anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) spanning over three "generations" since the discovery of ALK fusion positive (ALK+) non-small cell lung cancer (NSCLC), there remains intrinsic and acquired resistances to these approved TKIs. Currently, a fourth-generation (4G) ALK TKI, NVL-655, is being developed to attack some of the unmet needs such as compound resistance mutations in cis. However, EML4-ALK variant 3 and TP53 mutations are intrinsic genomic alterations that negatively modulate efficacy of ALK TKIs. Potentially, in the shifting landscape where lorlatinib should be the first-line ALK TKI of choice based on the CROWN trial, the central β-sheet #6 (Cβ6) mutation ALK L1256F will be the potential acquired resistance mutation to lorlatinib which may be resistant to current ALK TKIs. Here we opine on what additional capacities a putative fifth-generation (5G) ALK TKI will need to possess if it can be achieved in one single molecule. We propose randomized trial schemas targeting some of the intrinsic resistance mechanisms that will lead to approval of a prototypic fifth-generation (5G) ALK TKI and actually be beneficial to ALK+ NSCLC patients rather than just design a positive pivotal superiority trial for the sole purpose of drug approval.

Multisequence MRI-based radiomics signature as potential biomarkers for differentiating KRAS mutations in non-small cell lung cancer with brain metastases

BackgroundKirsten rat sarcoma virus (KRAS) has evolved from a genotype with predictive value to a therapeutic target recently. The study aimed to establish non-invasive radiomics models based on MRI to discriminate KRAS from epidermal growth factor receptor (EGFR) or anaplastic lymphoma kinase (ALK) mutations in lung cancer patients with brain metastases (BM), then further explore the optimal sequence for prediction. MethodsThis retrospective study involved 317 patients (218 patients in training cohort and 99 patients in testing cohort) who had confirmed of KRAS, EGFR or ALK mutations. Radiomics features were separately extracted from T2WI, T2 fluid-attenuated inversion recovery (T2-FLAIR), diffusion weighted imaging (DWI) and contrast-enhanced T1-weighted imaging (T1-CE) sequences. The maximal information coefficient and recursive feature elimination method were used to select informative features. Then we built four radiomics models for differentiating KRAS from EGFR or ALK using random forest classifier. ROC curves were used to validate the capability of the models. ResultsThe four radiomics models for discriminating KRAS from EGFR all worked well, especially DWI and T2WI models (AUCs: 0.942, 0.942 in training cohort, 0.949, 0.954 in testing cohort). When KRAS compared to ALK, DWI and T2-FLAIR models showed excellent performance in two cohorts (AUCs: 0.947, 0.917 in training cohort, 0.850, 0.824 in testing cohort). ConclusionsRadiomics classifiers integrating MRI have potential to discriminate KRAS from EGFR or ALK, which are helpful to guide treatment and facilitate the discovery of new approaches capable of achieving this long-sought goal of cure in lung cancer patients with KRAS.

Immunohistochemical expression of anaplastic lymphoma kinase in neuroblastoma and its relations with some clinical and histopathological features.

Anaplastic lymphoma kinase (ALK) mutations have been identified as a prominent cause of some familial and sporadic neuroblastoma (NB). ALK expression in NB and its relationship with clinical and histopathological features remains controversial. This study investigated ALK expression and its potential relations with these features in NB. Ninety cases of NB at the Department of Pathology, University of Medicine and Pharmacy at Ho Chi Minh City, Viet Nam from 01/01/2018 to 12/31/2021, were immunohistochemically stained with ALK (D5F3) antibody. The ALK expression and its relations with some clinical and histopathological features were investigated. The rate of ALK expression in NB was 91.1%. High ALK expression (over 50% of tumor cells were positive with moderate-strong intensity) accounted for 65.6%, and low ALK expression accounted for 34.4%. All the MYCN-amplified NB patients had ALK immunohistochemistry positivity, most cases had high ALK protein expression. The undifferentiated subtype of NB had a lower ALK-positive rate than the poorly differentiated and differentiated subtype. The percentages of ALK positivity were significantly higher in more differentiated histological types of NB (p = .024). There was no relation between ALK expression and: age group, sex, primary tumor location, tumor stage, MYCN status, clinical risk, Mitotic-Karyorrhectic Index, prognostic group, necrosis, and calcification. 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.

Impacts of probiotics on the efficacies of immune checkpoint inhibitors with or without chemotherapy for patients with advanced non-small-cell lung cancer.

The relationships between the therapeutic effects of immune checkpoint inhibitors (ICIs) and the intestinal flora have attracted increasing attention. However, the effects of oral probiotics on the efficacies of ICIs used to treat non-small-cell lung cancer (NSCLC) remain unclear. We investigated the effects of probiotics on the efficacies of ICIs in patients treated with and without chemotherapy. We investigated patients with advanced NSCLC on ICI monotherapy or combination ICI and chemotherapy using the Okayama Lung Cancer Study Group Immunotherapy Database (OLCSG-ID) and the Okayama Lung Cancer Study Group Immunochemotherapy Database (OLCSG-ICD). In total, 927 patients (482 on ICI monotherapy, 445 on an ICI + chemotherapy) were enrolled. Most were male, of good performance status, smokers, and without epidermal growth factor receptor (EGFR)/anaplastic lymphoma kinase (ALK) mutations. Probiotics were administered to 19% of patients on ICI monotherapies and 17% of those on ICIs + chemotherapy. Of the former patients, progression-free survival (PFS) and overall survival (OS) were significantly better in the probiotics group (PFS 7.9 vs. 2.9 months, hazard ratio [HR] 0.54, p < .001; OS not attained vs. 13.1 months, HR 0.45, p < .001). Among patients receiving ICI and chemotherapy, there were no significant differences in PFS between those on probiotics and not but OS was significantly better in the probiotics group (PFS 8.8 vs. 8.6 months, HR 0.89, p = .43; OS not attained vs. 22.6 months, HR 0.61, p = .03). Patients on probiotics experienced better outcomes following ICI treatment.

Therapeutic benefit of the dual ALK/FAK inhibitor ESK440 in ALK-driven neuroblastoma

Neuroblastoma (NB) is a predominantly pediatric cancer with greater than 90% of cases arising in children under the age of five. More than half of patients have metastases detected at diagnosis, and high-risk disease is associated with five-year survival rates of only 50–60 %. Standard therapy involves highly toxic chemotherapy, surgery, radiation, and immunotherapy, and less toxic, more specific targeted therapies are urgently needed. Genomic studies have identified common driver aberrations in high-risk NB, such as MYCN amplification. In addition, a proportion of high-risk patients harbor amplification or activating mutations in anaplastic lymphoma kinase (ALK), and co-occurrence of ALK mutations and MYCN amplification have been associated with aggressive disease. In this study, we analyzed the efficacy of a Phase Ia-cleared, orally bioavailable dual ALK and focal adhesion kinase (FAK) inhibitor, ESK440, in multiple preclinical NB models. ESK440 potently inhibited proliferation of NB cell lines, with increased sensitivity in cell lines harboring ALK aberrations. ALK, FAK, and downstream target activation were rapidly decreased upon ESK440 treatment, and this was associated with impaired cellular migration and invasion. Importantly, ESK440 treatment also decreased MYCN levels. NB cell line and patient-derived xenograft studies showed significant reduction in tumor growth in ESK440-treated mice with no signs of toxicity. In certain NB models, ESK440 showed comparable or enhanced efficacy to lorlatinib, another clinical ALK inhibitor, and a lorlatinib-resistant cell line (COG-N-561 LR) retained sensitivity to ESK440. These preclinical results indicate that ESK440 is a promising targeted agent for ALK-driven NB and support future clinical studies to evaluate its efficacy in NB patients.

Camrelizumab combined with apatinib and nanoparticle albumin-bound paclitaxel in lung adenocarcinoma (CAPAP-lung): a single-arm phase II study

Platinum-doublet chemotherapy plus immunotherapy has been the standard of care for the first-line treatment of advanced non-small cell lung cancer lacking actional driver mutations. However, optimization of drug combinations is still needed to find a better balance between therapeutic efficacy and safety in the immunotherapy era. We aimed to investigate the efficacy and safety of platinum-free albumin bound paclitaxel (nab-paclitaxel) combined with camrelizumab and apatinib as first-line treatment for patients with advanced lung adenocarcinoma. In this multicenter open-label, single-arm phase II trial, patients with systemic treatment-naïve advanced lung adenocarcinoma without epidermal growth factor receptor (EGFR) or anaplastic lymphoma kinase (ALK) mutations received a rational-based combination of camrelizumab (200mg intravenously, day one), apatinib (250mg, q.d., five continuous days per week), and nab-paclitaxel (135mg/m2 intravenously, days one and eight) every three weeks for four to six cycles in China. Patients with controlled disease were maintained with camrelizumab and apatinib. The primary end point was progression-free survival (PFS). This trial is registered with ClinicalTrials.gov (No. NCT04459078). Between August 26, 2020 and May 20, 2022, 64 patients were enrolled. The median PFS was 14.3 (95% CI: 9.9, not reached) months. The confirmed objective response rate was 64.1% (95% CI: 51.1, 75.7). The grade 3-4 hematologic treatment-related adverse events (TRAEs) were decreased neutrophil count (14.1%), decreased white blood cell count (7.8%), and anemia (3.1%). The most common non-hematologic TRAEs of grade 3-4 were increased alanine transaminase (18.8%) and aspartate transaminase (15.6%). No treatment-related death occurred. The quality of life was on average not clinically meaningful worse through treatment cycle14. Nab-paclitaxel plus camrelizumab and apatinib showed clinically meaningful anti-tumor activity and manageable safety, with few hematologic toxicities, and might be a potential treatment option in patients with advanced lung adenocarcinoma lacking EGFR/ALK mutations. Heath Research Foundation of Chinese Society of Clinical Oncology, Hunan Provincial Natural Science Foundation of China, Hunan Cancer Hospital Climb Plan, Sister Institution Network Fund of The University of Texas MD Anderson Cancer Center, The Science and Technology Innovation Program of Hunan Province, and Suzhou Sheng Diya Biomedical Co., Ltd, a subsidiary of Jiangsu Hengrui Pharmaceuticals Co., Ltd. (Shanghai, China).

Pembrolizumab-induced Stevens-Johnson syndrome/Toxic Epidermal Necrolysis in a Vietnamese patient with nonsmall-cell lung cancer

Chemoimmunotherapy is an effective therapy for an individual with nonsmall-cell lung cancer (NSCLC) without anaplastic lymphoma kinase or epidermal growth factor receptor mutations. However, it can also be related to adverse cutaneous reactions such as Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN) with high morbidities and mortality rates. We present a case of a 65-year-old male with NSCLC who underwent first-line chemotherapy with paclitaxel, carboplatin, and pembrolizumab, which was later followed by a second cycle of the same therapies. The patient developed a fever and rash 12 days after the second cycle. Pembrolizumab was strongly suspected as the culprit medication because cutaneous reactions to this drug have been frequently reported and threw other medications used as less likely candidates. This is the first case reported in Vietnam of SJS/TEN related to pembrolizumab and contributes to our knowledge of severe skin reactions associated with immune checkpoint inhibitors.

Anaplastic lymphoma kinase inhibitors-a review of anticancer properties, clinical efficacy, and resistance mechanisms.

Fusions and mutations of anaplastic lymphoma kinase (ALK), a tyrosine kinase receptor, have been identified in several neoplastic diseases. Rearranged ALK is a driver of tumorigenesis, which activates various signaling pathway associated with proliferation and survival. To date, several agents that target and inhibit ALK have been developed. The most studied ALK-positive disease is non-small cell lung cancer, and three generations of ALK tyrosine kinase inhibitors (TKIs) have been approved for the treatment of metastatic disease. Nevertheless, the use of ALK-TKIs is associated with acquired resistance (resistance mutations, bypass signaling), which leads to disease progression and may require a substitution or introduction of other treatment agents. Understanding of the complex nature and network of resistance mutations may allow to introduce sequential and targeted therapies. In this review, we aim to summarize the efficacy and safety profile of ALK inhibitors, describe off-target anticancer effects, and discuss resistance mechanisms in the context of personalized oncology.

Development of Two Novel One-Step and Green Microwell Spectrophotometric Methods for High-Throughput Determination of Ceritinib, a Potent Drug for Treatment of Anaplastic Lymphoma Kinase-Positive Non-Small-Cell Lung Cancer.

Background and Objectives: Ceritinib (CER) is a potent drug of the third-generation tyrosine kinase inhibitor class. CER has been approved for the treatment of patients with non-small-cell lung cancer (NSCLC) harboring the anaplastic lymphoma kinase (ALK) mutation gene. In the literature, there is no green and high-throughput analytical method for the quantitation of CER in its dosage form (Zykadia® capsules). This study describes, for the first time, the development and validation of two novel one-step and green microwell spectrophotometric methods (MW-SPMs) for the high-throughput quantitation of CER in Zykadia® capsules. Materials and Methods: These two methods were based on an in microwell formation of colored derivatives upon the reaction of CER with two different benzoquinone reagents via two different mechanisms. These reagents were ortho-benzoquinone (OBQ) and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), and their reactions proceeded via condensation and charge transfer reactions, respectively. The reactions were carried out in 96-well transparent plates, and the absorbances of the colored reaction products were measured with an absorbance microplate reader at 540 and 460 nm for reactions with OBQ and DDQ, respectively. The optimum conditions of reactions were established, their molar ratios were determined, and reaction mechanisms were postulated. Under the refined optimum reaction conditions, procedures of MW-SPMs were established and validated according to the guidelines of the International Council on Harmonization. Results: The limits of quantitation were 6.5 and 10.2 µg/well for methods involving reactions with OBQ and DDQ, respectively. Both methods were applied with great reliability to the determination of CER content in Zykadia® capsules and their drug uniformity. Greenness of the MW-SPMs was evaluated using three different metric tools, and the results proved that the two methods fulfil the requirements of green analytical approaches. In addition, the simultaneous handling of a large number of samples with microvolumes in the proposed methods gave them the advantage of a high-throughput analysis. Conclusions: The two methods are valuable tools for rapid routine application in pharmaceutical quality control units for the quantitation of CER.

Three Innovative Green and High-Throughput Microwell Spectrophotometric Methods for the Quantitation of Ceritinib, a Potent Drug for the Treatment of ALK-Positive Non-Small Cell Lung Cancer: An Application to the Analysis of Capsules and Drug Uniformity Testing.

Ceritinib (CER) is a potent drug that has been recently approved by the Food and Drug Administration for the treatment of patients with non-small cell lung cancer harboring the anaplastic lymphoma kinase mutation gene. The existing methods for the quality control of CER are very limited and suffer from limited analytical throughput and do not meet the requirements of the green analytical principles. This study presented the first-ever development and validation of three innovative green and high-throughput microwell spectrophotometric methods (MW-SPMs) for the quality control of CER in its dosage form (Zykadia® capsules). These MW-SPMs were based on the formation of colored N-vinylamino-substituted haloquinone derivatives of CER upon its reactions with each of chloranil, bromanil, and 2,3-dichloro-1,4-naphthoquinone in the presence of acetaldehyde. The optimized procedures of the MW-SPMs were established, and their analytical performances were validated according to the ICH. The linear range of the MW-SPMs was 5-150 µg/mL, with limits of quantitation of 5.3-7.6 µg/mL. The accuracy and precision of the MW-SPMs were proved, as the average recovery values were 99.9-101.0%, and the relative standard deviations did not exceed 1.8%. The three methods were applied to the determination of CER content in Zykadia® capsules and drug content uniformity testing. The greenness of the MW-SPMs was proved using three different metric tools. In addition, these methods encompassed the advantage of high-throughput analysis. In conclusion, the three methods are valuable tools for convenient and reliable application in the pharmaceutical quality control units for CER-containing capsules.