Preliminary Study Of Diagnosis And Treatment In Acute Lung Injury, And Pharmacogenetic Assessment Of Leukopenia After Platinum Chemotherapy In Non-small Cell Lung Cancer Patients

Acute lung injury (ALI) and its severe form, acute respiratory distress syndrome (ARDS), are associated with high mortality and morbidity in critically ill patients. However, the possible mechanisms of ALI and ARDS are unclear. Moreover, there is no specific the biomarker of ALI/ARDS. Metabonomic is the scientific study of chemical processes involving metabolites, and it is a well-developed platform for studying systematic biology and clinical diagnosis. Mechanical ventilation (MV) is the only known effective and cornerstone therapy used to support patients with ALI or ARDS. However, MV itself can cause a type of acute lung damage termed ventilator-induced lung injury (VILI). Currently, no approved drug has been clinically applied.Pharmacogenetic assessment is a new method to find indicators objectively evaluating pharmacologic responses. Lung cancer is one of the leading causes of cancer death in many countries, of which non-small cell lung cancer (NSCLC) accounts for approximately 80% of all related deaths. Standard treatment for NSCLC involves chemotherapy with platinum (cisplatin or carboplatin), in combination with navelbine, gemcitabine or paclitaxel. These platinum-based regimens bring modest benefits, but also adverse effects which may cause distressing symptoms and prevent further therapies. Matrix metalloproteinase-2 (MMP-2) plays an important role in hematopoietic recovery after chemotherapy-induced myelosuppression. MMP-2 polymorphisms may affect the incidence of leukopenia in advanced non-small cell lung cancer (NSCLC) patients receiving first-line platinum-based chemotherapy.These works include three parts as described below.Part I:1H NMR spectroscopy and pattern recognition analysis of ALI mice lung tissue for characteristic metabolitesObjective:In this study, we investigated a metabonomic approach to evaluate the prognosis of LPS-induced ALI by 1H NMR spectroscopy.Methods:The mice model of ALI was established by intratracheal instillation of LPS (5mg/kg) for 4 hours with the saline in Sham group (Sham group) mice. The mice of dexamethasone treatment group (LPS+DXM group) were treated with DXM intraperitoneally while LPS intratracheally. After 4h, morphology, the wet/dry ratio, protein concentration in bronchial alveolar lavage fluid (BALF), the expression of tumor necrosis factor (TNF)-a in lung tissue and BALF, myeloperoxidase (MPO) activity and content of ATP were measured.1H NMR spectroscopy in combination with pattern recognition methods was applied to study these mice lung tissues samples.Results:LPS induced lung injury was characterized by inflammation and increased microvascular permeability. After stimulation by LPS, the concentration of TNF-a in lung tissue and BALF, and MPO activity were significantly increased, while the concentration of ATP were decreased, all of which were significantly prevented by DXM. With the application of first principal component (PC 1), sham group and LPS group could be differentiated from each other, but sham group and LPS+DXM group could not. To further examin the composition of each group, valine (Val,δ1.06), lactate (Lac,δ1.33), acetate (Ace,δ1.91), creatine (Cre,δ3.93) composite the most parts in all the groups, the difference in each group showed statistically significance. However, the concentration of phosphocholine and glycerophosphocholine (GPCPC,δ3.22), creatinine (Crn,δ3.07), succinatc (Suc,δ2.40) though changed, but showed no statistically significance.Conclusions:For the first time, our study, found four novel metabolites including valine, lactate, acetate, creatine which are changed markedly in ALI mice, moreover, could be prevented by DXM. These metabolites may have a potentially role in indicating the pathogenic processes of ALI/ARDS and response to a therapeutic intervention.PartⅡ:The effects of curcumin on ventilator induced lung injury in rats and associated mechanismsObjective:Curcumin (CUR) can inhibit inflammation and oxidative stress in previous studies. The present study aims at investigating potential effects and mechanism of CUR on ventilator-induced lung injury in rats.Methods:50 specified-pathogens free male Sprague-Dawley rats were randomized into five groups:Group Sham:Sham-operated rats were used as time-matched controls. Rats were pretreated with a single intraperitoneal dose of vehicle 2 h before anesthesia. Group VILI:rats were subjected to high Vt ventilation; Group VILI+CUR-50:rats were subjected to high Vt ventilation and were pretreated with CUR (50mg/kg) 2 h before anesthesia, Group VILI+CUR-200:rats were subjected to high Vt ventilation and were pretreated with CUR (200mg/kg) 2 h before anesthesia, Group VILI+DXM:rats were subjected to high Vt ventilation and were pretreated with DXM (5mg/kg) 2 h before anesthesia. After 4h of ventilation, morphology, wet/dry ratio, protein concentration in bronchial alveolar lavage fluid (BALF), nuclear factor (NF)-κB activity, the expression of tumor necrosis factor (TNF)-α, (macrophage inflammatory protein)MIP-2, myeloperoxidase (MPO) activity, malondialdehyde (MDA) and total anti-oxidative capability (TAOC) were tested.Results:Mechanical Ventilation induced lung injury was characterized by inflammation in morphology and increased microvascular permeability. Pretreatment with CUR (200 mg/kg) or DXM markedly attenuated Ventilation induced inflammation, pulmonary edema, and decreased protein concentration in BALF. Mechanical ventilation could induce the NF-κB activity, pretreatment with CUR (200 mg/kg) or DXM markedly attenuated the NF-κB activity. The concentration of TNF-α, MIP-2 in BALF were significantly increased after mechanical ventilation, accompanied with increased levels of lipid peroxidation and xanthine oxidase (XO) activity, but the total anti-oxidative capability(TAOC) were decreased in VILI group. The increases in NF-κB, the expressions of TNF-α, MIP-2, MPO and malondialdehyde (MDA) were significantly suppressed by administration of CUR (200 mg/kg) or DXM. Pretreatment with CUR (200 mg/kg) significantly attenuated XO activity and increased TAOC.Conclusions:Rats showed obvious lung injury after mechanical ventilation for 4h with 25ml/kg tidal volume. Pretreatment with CUR 2h before mechanical ventilation would attenuate VILI induced by high tidal volume ventilation, which was similar to the effects of steroid. CUR mediated VILI attenuating is probably through inhibiting NF-κB activity, suppressing the expression of inflammatory cytokines and improving oxidative stress. Thus, CUR may have potential therapeutic role for improving the outcome of ventilator-induced lung injury.PartⅢ:Pharmacogenetic assessment of leukopenia after platinum chemotherapy in non-small cell lung cancer patientsObjective:In this study, we investigated the association between MMP-2 polymorphisms and the incidence of leukopenia in advanced non-small cell lung cancer (NSCLC) patients receiving first-line platinum-based chemotherapy.Methods:We used Illumina iSelect HD BeadChip to genotype the 16 polymorphisms in 1004 stage III and IV NSCLC patients treated with platinum combination chemotherapy (including 221 treated with cisplain-navelbine,195 with cisplatin-gemcitabine,190 with carboplatin-paclitaxel, and 108 with cisplatin-paclitaxel). Information about grade 3 or 4 leukopenia was available.Results:In the subgroupanalysis for patients receiving gemcitabine in combination with cisplatin treatment, the variant homozygotes of MMP-2 rs1477017(P=0.027), rs1293424 (P=0.047)lpolymorphism were associated with a significantly increased risk of grade 3 or 4 leukopenia toxicity.In the subgroupanalysis for patients receiving carboplatin-paclitaxel treatment, the variant homozygotes of MMP-2 rs243844 (P=0.04) 1polymorphism were associated with a significantly increased risk of grade 3 or 4 leukopenia toxicity.In the subgroupanalysis for patients receiving cisplain-navelbine treatment, the variant homozygotes of MMP-2 rs1477017 (P=0.023),rs17301608(P=0.041),rs243836(P=0.005)andrs7201(P=0.0004) lpolymorphism were associated with a significantly increased risk of grade 3 or 4 leukopenia toxicity.Consistent with these results of single-locus analysis, the haplotype and diplotype analyses revealed a protective effect of the haplotype "AA" on the risk of grade 3 or 4 leukopenia toxicity(P=0.002). The haplotype"GC" were associated with a significantly increased risk of grade 3 or 4 leukopenia toxicity, but we did not observed significant associations between MMP-2 polymorphisms and platinum sensitivity.Conclusions:Our study, for the first time, provides evidence for the predictive role of MMP-2 polymorphisms on severe chemotherapy-related leukopenia among platinum-treated advanced NSCLC Chinese patients.