SHR‐A1403, a novel c‐mesenchymal‐epithelial transition factor (c‐Met) antibody‐drug conjugate,overcomes AZD9291 resistance in non‐small cell lung cancer cells overexpressing c‐Met

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI) have been used as the first‐line treatment of non‐small cell lung cancers (NSCLC) harboring EGFR‐activating mutations, but acquired resistance is ubiquitous and needs to be solved urgently. Here, we introduce an effective approach for overcoming resistance to the EGFR‐TKI, AZD9291, in NSCLC cells using SHR‐A1403, a novel c‐mesenchymal‐epithelial transition factor (c‐Met)‐targeting antibody‐drug conjugate (ADC) consisting of an anti‐c‐Met monoclonal antibody (c‐Met mAb) conjugated to a microtubule inhibitor. Resistant cells were established by exposing HCC827 to increasing concentrations of EGFR‐TKI. c‐Met was found to be overexpressed in most resistant cells. AZD9291 resistance was partially restored by combination of AZD9291 and crizotinib only in resistant cells overexpressing phospho‐c‐Met, which synergistically inhibited c‐Met‐mediated phosphorylation of the downstream targets ERK1/2 and AKT. In resistant cells overexpressing c‐Met, neither crizotinib nor c‐Met mAb was able to overcome AZD9291 resistance. In contrast, SHR‐A1403 strongly inhibited proliferation of AZD9291‐resistant HCC827 overexpressing c‐Met, regardless of the levels of c‐Met phosphorylation. SHR‐A1403 bound to resistant cells overexpressing c‐Met was internalized into cells and released associated microtubule inhibitor, resulting in cell‐killing activity that was dependent on c‐Met expression levels only, irrespective of the involvement of c‐Met or EGFR signaling in AZD9291 resistance. Consistent with its activity in vitro, SHR‐A1403 significantly inhibited the growth of AZD9291‐resistant HCC827 tumors and caused tumor regression in vivo. Thus, our findings show that SHR‐A1403 efficiently overcomes AZD9291 resistance in cells overexpressing c‐Met, and further indicate that c‐Met expression level is a biomarker predictive of SHR‐A1403 efficacy.     

Identification of pro- and mature brain-derived neurotrophic factor in human saliva

Objective

Growth factors, including brain-derived neurotrophic factor (BDNF), are polypeptides that are involved in the maintenance, survival, and death of central and peripheral cells. Numerous growth factors have been identified in saliva and are thought to promote wound healing and maintenance of the oral epithelium. The aim of this study was to determine if BDNF is also found in human saliva.

Methods

Whole, unstimulated saliva samples (n = 30) were analyzed by SDS-PAGE and Western blot using an anti-human BDNF antibody. Proteolytic cleavage products were similarly assessed following the incubation of pooled saliva with N-glycanase F and plasmin. Subjects were also genotyped for the BDNF Val66Met single nucleotide polymorphism (SNP).

Results

These experiments revealed the presence of immunoreactive bands at 14, 32 and 34 kDa, corresponding to mature (mBDNF) and proBDNF, as well as a truncated pro-form at 24 kDa. Not every sample contained all forms of BDNF. Treatment with N-glycanase and plasmin reduced the size of the higher molecular weight bands, confirming the glycosylated pro-form of BDNF. mBDNF was detected significantly less often in subjects with the Val66Met SNP, compared to those without the polymorphism (χ² = 4.05; P < 0.05).

Conclusions

While the function of salivary BDNF still requires elucidation, these findings suggest that it may be possible to use saliva in lieu of blood in future studies of BDNF and the Val66Met polymorphism.     

The brain-derived neurotrophic factor (BDNF) polymorphism Val66Met is associated with neither serum BDNF level nor response to selective serotonin reuptake inhibitors in depressed Japanese patients

Background

We investigated the relationship between a brain-derived neurotrophic factor (BDNF) polymorphism (Val66Met) and the clinical response of patients with major depressive disorder to selective serotonin reuptake inhibitors (SSRIs; here, paroxetine and sertraline). In addition, serum BDNF levels in these patients were considered together with the clinical response.

Methods

A total of 132 patients who met the DSM-IV criteria for major depressive disorder were enrolled in the study. 54 of these patients were male and 78 were female (age range, 20-74 years; mean ± S.D., 51 ± 15). The patients' clinical improvement was evaluated using the 17-item Hamilton Rating Scale for Depression (HAMD-17) before (T0) and at 8 weeks after the administration of SSRI treatment (T8). Patients with at least a 50% decrease in the HAMD-17 score were classified as responders.

Results

No correlation was observed between the BDNF Val66Met polymorphism and response to SSRIs or between the BDNF Val66Met polymorphism and serum BDNF levels at T0. An inverse correlation was found between serum BDNF levels and HAMD-17 scores at T0.

Conclusions

These results suggest that the BDNF Val66Met polymorphism is independent of both the response to SSRI treatment and serum BDNF levels. The findings in the present study reconfirm that the serum BDNF level is a state biomarker for depression.     

Association of brain-derived neurotrophic factor (BDNF) Val66Met polymorphism with Parkinson's disease in a Greek population

Brain-derived neurotrophic factor (BDNF) enhances survival of dopaminergic neurons in the substantia nigra, whereas in patients with Parkinson's disease (PD), the expression of BDNF mRNA is decreased, thus making BDNF a candidate gene for PD susceptibility. The association between BDNF Val66Met polymorphism and PD has been evaluated in several studies with controversial results. Thus, we determined the distribution of BDNF Val66Met polymorphism in 184 Greek patients with sporadic PD and 113 control participants using polymerase chain reaction-restriction fragment length polymorphism, and explored the association of the polymorphism with certain clinical parameters of the disease. Our results do not support a major role for the BDNF Val66Met polymorphism in PD in the Greek population.     

The BDNF Val66Met polymorphism is an independent risk factor for high lethality in suicide attempts of depressed patients

Some authors have reported an association of BDNF Val66Met polymorphism with suicidal behavior and/or clinical aspects of suicidal attempts. We evaluated, here, the impact of BDNF Val66Met polymorphism on the clinical characteristics of suicide attempts. The study was conducted on a cohort of 120 consecutive patients who were admitted to the Emergency Hospital of Porto Alegre, Brazil, due to a suicide attempt. Variables of univariate analyses were included in a logistic regression model to test whether the risk factors had independent effect. In univariate analyses, sex, BDNF genotype, intent and method of suicide attempt were all risk factors for high lethality in suicide attempts. After logistic regression analysis, male sex (O.R. = 3.03; 95% C.I = 1.34–6.84; 0.008) and the presence of BDNF 66Met allele (O.R. = 2.62; 95% C.I = 1.04–6.57; 0.04) were significantly and independently associated with the high lethality in suicide attempts. The present study showed that BDNF 66Met allele is an independent predictor of high lethality in suicide attempts of depressed patients. This finding is important because it might allow earlier identification of patients at high risk for suicide, perhaps providing better tools for clinical care of these patients in the future.     

Anomalous inhibition of c-Met by the kinesin inhibitor aurintricarboxylic acid

c-Met [the hepatocyte growth factor (HGF) receptor] is a receptor tyrosine kinase playing a role in various biological events. Overexpression of the receptor has been observed in a number of cancers, correlating with increased metastatic tendency and poor prognosis. Additionally, activating mutations in c-Met kinase domain have been reported in a subset of familial cancers causing resistance to treatment. Receptor trafficking, relying on the integrity of the microtubule network, plays an important role in activation of downstream targets and initiation of signalling events. Aurintricarboxylic acid (ATA) is a triphenylmethane derivative that has been reported to inhibit microtubule motor proteins kinesins. Additional reported properties of this inhibitor include inhibition of protein tyrosine phosphatases, nucleases and members of the Jak family. Here we demonstrate that ATA prevents HGF-induced c-Met phosphorylation, internalisation, subsequent receptor trafficking and degradation. In addition, ATA prevented HGF-induced downstream signalling which also affected cellular function, as assayed by collective cell migration of A549 cells. Surprisingly, the inhibitory effect of ATA on HGF-induced phosphorylation and signalling in vivo was associated with an increase in basal c-Met kinase activity in vitro. It is concluded that the inhibitory effects of ATA on c-Met in vivo is an allosteric effect mediated through the kinase domain of the receptor. As the currently tested adenosine triphosphate competitive tyrosine kinase inhibitors (TKIs) may lead to tumor resistance (McDermott U, et al., Cancer Res 2010;70:1625-34), our findings suggest that novel anti-c-Met therapies could be developed in the future for cancer treatment.     

Inhibition of Met/HGF receptor and angiogenesis by NK4 leads to suppression of tumor growth and migration in malignant pleural mesothelioma

NK4 exhibits two distinct biological actions: antagonistic inhibition of hepatocyte growth factor (HGF) through binding to the Met/HGF receptor, and antiangiogenic action through binding to perlecan. Here, the anti‐tumor effect of NK4 on malignant pleural mesothelioma was investigated. Of the 7 human malignant mesothelioma cell lines (ACC‐Meso‐1, ACC‐Meso‐4, EHMES‐1, EHMES‐10, H28, H2052 and JMN‐1B), only EHMES‐10 cells formed subcutaneous tumors when implanted into mice. For EHMES‐10 cells, HGF facilitated invasion of the cells in collagen gel, whereas NK4 and neutralizing anti‐HGF antibody suppressed the HGF‐induced invasion. In addition, NK4 but not anti‐HGF antibody suppressed proliferation of EHMES‐10 cells in collagen, suggesting that the suppression by NK4 was independent of the HGF‐Met pathway. In the subcutaneous tumor model, recombinant adenovirus‐mediated intratumoral expression of NK4 inhibited tumor growth, while the invasive characteristic of tumor cells was not observed. Analysis of Met receptor tyrosine phosphorylation, proliferation, apoptosis and blood vessels in the tumor tissues indicated that the inhibitory effect of NK4 expression might be primarily caused by the inhibition of tumor angiogenesis. In all the 7 mesothelioma lines, HGF stimulated Met tyrosine phosphorylation, and this was associated with enhanced cell migration. HGF‐dependent Met activation and migration were inhibited by NK4. Since malignant pleural mesothelioma represents an aggressive neoplasm characterized by extensive invasive growth, suppression of invasive growth has therapeutic value. Thus, the simultaneous inhibition of the HGF‐Met pathway and angiogenesis by NK4 for treatment of malignant pleural mesothelioma is significant, particularly to attenuate migration and invasive growth.     

MiR-130a Overcomes Gefitinib Resistance by Targeting Met in Non-Small Cell Lung Cancer Cell Lines

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and the most common cause oflung cancer death. Currently, the epidermal growth factor receptor inhibitor gefitinib is used for its treatment;however, drug resistance is a major obstacle. Expression of Met has been associated with both primary andacquired resistance to gefitinib, but the mechanisms regulating its expression are not fully understood. Recently,miRNAs such as miR-130a have been shown to play a role in gefitinib resistance, but importance in NSCLC andrelationships with Met have not been fully explored. Here we show that miR-130a is over-expressed in gefitinibsensitiveNSCLC cell lines, but is low in gefitinib-resistant NSCLC cell lines. Moreover, miR-130a expressionwas negatively correlated with that of Met. Further analysis revealed that over-expression of miR-130a increasedcell apoptosis and inhibited proliferation of NSCLC cells treated with gefitinib, whereas lowering the expressionof miR-130a decreased cell apoptosis and promoted cell proliferation after treatment with gefitinib in bothgefitinib-sensitive and -resistant NSCLC cell lines, suggesting that miR-130a overcomes gefitinib resistance.We also demonstrated that miR-130a binds to the 3’-UTR of Met and significantly suppresses its expression.Finally, our results showed that over-expressing Met could “rescue” the functions of miR-130a regarding cellapoptosis and proliferation after cells are treated with gefitinib. These findings indicate that the miR-130a/Metaxis plays an important role in gefitinib resistance in NSCLC. Thus, the miR-130a/Met axis may be an effectivetherapeutic target in gefitinib-resistant lung cancer patients.     

CACNA1A-p.Thr501Met mutation associated with familial hemiplegic migraine: a family report

Background and aimsHemiplegic migraine (HM) is a rare form of migraine characterized by the presence of a motor and other types of aura. HM can be sporadic or familial. Familial hemiplegic migraine (FHM) is an autosomal dominant disorder, classified into 3 subtypes, based on the gene involved (CACNA1A in FHM1, ATP1A2 in FHM2 and SCN1A in FHM3). The clinical presentation is highly heterogeneous and some attacks may be severe.We report the clinical characteristics and genetic analysis of 12 patients belonging to a family with CACNA1A-p.Thr501Met gene mutation.MethodsWe screened for mutations in CACNA1A gene 15 patients belonging to the same family. The exonic sequences of CACNA1A were analyzed using a Tru-seq® Custom Amplicon (TSCA) (Illumina Inc., San Diego, CA) targeted capture and paired end library kit. Sanger sequencing was used to confirm CACNA1A variants and segregation analysis.ResultsCACNA1A-p.Thr501Met mutation was found in 12 of the 15 patients screened, which was compatible with the diagnosis of FHM1.Attacks of hemiplegic migraine were reported by 10 of the 12 subjects (83.33%). Only one subject developed persistent mild cerebellar symptoms and none of the subjects developed cerebellar atrophy.DiscussionThe variant p.Thr501Met was described previously in association with episodic ataxia and rarely with FHM related to cerebellar symptoms. FHM1 has a broad clinical spectrum and about half of the families have cerebellar involvement. In our study, only one patient developed persistent cerebellar deficits.These data suggest that CACNA1A-p.Thr501Met mutation can occur prevalently as hemiplegic migraine.Supplementary InformationThe online version contains supplementary material available at 10.1186/s10194-021-01297-5.