Loading

International Journal of Plant Biology & Research

Anti-Growth Activity of Tanshinone IIA towards Gefitinib. -Sensitive and -Resistant NonSmall Cell Lung Cancer Cells

Short Communication | Open Access

  • 1. Department of Faculty of Pharmaceutical Sciences, Set sunan University, Japan
  • 2. Department of Institute of Health Sciences, University of Sunstar Inc, Japan
  • 3. Department of Pharmaceutical Analysis and Standardization, National Institute of Medicinal Materials, Vietnam
+ Show More - Show Less
Corresponding Authors
Fumiaki Ito, Institute of Health Sciences, Sunstar Inc., Takatsuki, Osaka 569-1195, Japan. Tel: 81-72-682-5555; Fax: 81-72-681-8202
Abstract

Lung cancer is the most common fatal malignancy among all cancers worldwide, and non-small cell lung cancer (NSCLC) accounts for 85% of all lung cancers. Existing therapies for NSCLC include tyrosine kinase inhibitors (TKIs) such as gefitinib and erlotinib, which target the epidermal growth factor receptor (EGFR) tyrosine kinase. In this report, we studied the anti-proliferative activity of tanshinone IIA (Tan IIA) from Radix Salvia miltiorrhiza against gefitinib-sensitive and -resistant NSCLC cells. Tan IIA effectively inhibited the growth of both gefitinib-sensitive NSCLC cells harboring the EGFR mutation of exon 19 (PC-9) and gefitinib-resistant NSCLC cells (H1650, H358, AY-01, and A549). We also identified another type of NSCLC cells (PC-14), which were resistant to both TKI and Tan IIA. H1650 and AY-01 harbor an EGFR mutation of exon 19, but are resistant to gefitinib. H358 and A549 harbor a KRAS mutation but no EGFR mutation. Therefore, Tan IIA is a candidate compound for the development of anti-cancer drugs targeting a variety of gefitinib-sensitive and -resistant NSCLC.

Citation

Takeuchi K, Ito F, Thuong PT (2014) Anti-Growth Activity of Tanshinone IIA towards Gefitinib. -Sensitive and -Resistant NonSmall Cell Lung Cancer Cells . Int J Plant Biol Res 2(3): 1020.

Keywords

•    Epidermal growth factor receptor
•    Gefitinib
•    Erlotinib
•    Tyrosine kinase inhibitor
•    Non-small cell lung cancer
•    Tanshinone IIA

ABBREVIATIONS

EGFR: Epidermal Growth Factor Receptor; TKI: Tyrosine Kinase Inhibitor; NSCLC: Non-Small Cell Lung Cancer; NAC: N-Acetyl-L- Cysteine; ROS: Reactive Oxygen Species; Tanshinone IIA: Tan IIA

INTRODUCTION

Lung cancer is the most common fatal malignancy among all cancers worldwide, and its incidence has gradually increased. Non-small cell lung cancer (NSCLC) accounts for 85% of all lung cancers and affects millions of patients each year worldwide. Existing therapies for NSCLC include tyrosine kinase inhibitors (TKIs), including gefitinib and erlotinib, which target EGFR tyrosine kinase [1, 2].

Nearly 90% of the EGFR mutations detected in NSCLC are small in-frame deletions at exon 19 (Del746-750) or the missense mutation L858R in exon 21, replacing leucine with arginine [3]. The use of TKIs in NSCLC patients harboring these EGFR gene mutations has been associated with dramatic response rates and improved progression-free survival.

Activating alterations in a variety of potential oncogenic driver genes other than EGFR have also been identified in NSCLC [4]. Moreover, acquired resistance to gefitinib and erlotinib occurs inevitably in almost all NSCLC patients [5]. Therefore, the intrinsic and acquired forms of resistance to TKIs remain a persistent complication for targeted therapies of lung cancer.

Compounds extracted from natural sources have been introduced into the chemotherapy of various human cancer cells. Among them, the terpenes represent a treasure house of potential agents for cancer treatment. Tanshinone IIA (Tan IIA), an active diterpenoid in the dried root of Salvia miltiorrhiza, shows anti-proliferative activity toward various human cancer cells including NSCLC cells [6, 7]. However, the effect of Tan IIA on gefitinib-resistant NSCLC cells is not clearly known. The results of this present study show that Tan IIA is a promising compound for the development of anti-cancer drugs targeting gefitinib-resistant NSCLC.

MATERIALS AND METHODS

Tan IIA

Salvia miltiorrhiza Bunge was cultivated and collected in high mountainous Sapa, Laocai, Vietnam. The dry roots of S. miltiorrhiza (0.3 kg) were extracted at room temperature for 1 week with methanol (2L × 3 times). The combined methanol extract was exhaustively evaporated under reduced pressure to obtain a residue (35.1 g), which was then suspended in water and successively partitioned with n-hexane and dichloromethane. Following removal of the solvents in vacuo, n-hexane (Hx, 7.3 g) and dichloromethane (CH2 Cl2 , 9.6 g) residues were obtained. The CH2 Cl2 residue (8.0 g) was chromatographed on a silica gel column with n-hexane-ethyl acetate (10:1, 9:1, 8:1, … 1:1) as the eluent to yield 8 fractions (F.1 - F.8). Fraction 2 was then applied to a Sephadex LH20 column and eluted with methanol to yield 5 sub fractions (F.2.1 - F.2.5). The red-colored fraction (F.2.2) was chromatographed on a silica gel column using n-hexane-CH2 Cl2 (1:1) as the eluting solvent to yieldTan IIA (65.1 mg).

Tan IIA: red amorphous powder; mp 202-204o C; UV (MeOH) λmax: 224, 251, 269, 352, 458 nm; IR (KBr) νmax: 1152, 1280, 1460, 1578, 1668, 2935; ESI-MS m/z 295 [M+H] + ; 1 H- and 13C-NMR spectra were in accordance with previous reports [8, 9].

Chemical Reagents

AG1478, [4-(3-chloroanilino)-6,7-dimethoxyquinazoline], was purchased from Calbiochem-Novabiochem Corp. (San Diego, CA). RPMI1640 was from Nissui Pharmaceutical Co., Ltd (Tokyo, Japan). N-acetyl-L-cysteine (NAC) was obtained from Calbiochem, Merck, Darmstadt, Germany. Hoechst 3342 Solution was obtained from Dojindo (Kumamoto, Japan). 2’,7’-Dichlorodihydrofluorescein diacetate (DCF-DA) was purchased from Invitrogen Molecular Probes (USA).

Cell culture

Human non-small cell lung cancer (NSCLC) cells PC-9 and PC14 were obtained from Tokyo Medical University (Tokyo, Japan). Gefitinib-resistant NSCLC cells H358 and H1650 were purchased from ATCC. All these cell lines were cultivated in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS) in 5% CO2 at 37o C in a fully humidified atmosphere.

Cloning of AG1478-resistant cell line from PC-9 cells

PC-9 cells were seeded at a density of 1.0 x 105 cells/60 mm dish and cultured for 2 days. The cells were then exposed to 500 nM AG1478 for 48 h. The surviving cells were suspended in RPMI1640 medium containing 5% FBS and 500 nM AG1478 and plated at 0.5 cell/well into a 96-well microplate. After 48 h of incubation, the culture medium was replaced with AG1478- free growth medium, and incubation was continued to obtain AG1478-resistant cell line, designated as AY-01.

Growth inhibition assay (WST-1 assay)

PC-9, PC-14, H358, H1650 or AY-01 cells were seeded at a density of 2 x 103 cells/well into a 96-well microplate and cultured for 48 h. The cells were then treated with various concentrations of Tan IIA or AG1478. After incubation for 2 days, growth inhibition was quantified by a colorimetric assay using the WST-8 reagent according to the manufacturer’s instructions (Dojindo Laboratories, Kumamoto, Japan).

ROS level assay

Intracellular ROS in Tan IIA-treated cells was measured by using the oxidation sensitive fluorescent prove DCF-DA as described previously [10]. The intensity of the fluorescence was detected by using a Cellomics Array Scan VTI HCS Reader (Thermo Scientific, Finland) at an excitation wavelength of 515 nm and an emission wavelength of 549 nm.

RESULTS AND DISCUSSION

Tan IIA was earlier shown to have anti-proliferative activity toward various human cancer cells including NSCLC cells and colon cancer cells [6,7]. We first tested the cytotoxic activity of Tan IIA towards 4 human colorectal cancer cell lines and 2 human NSCLC cell lines. The compound inhibited the growth of all of the colorectal cancer cell lines (Caco-2, LS180, LoVo, and HT-29), and its concentrations required to inhibit 50% of the cell growth (IC50) were higher than 30 µM (data not shown). On the other hand, the IC50 of Tan IIA against NSCLC cell line PC-9 was less than 1 µM; whereas another NSCLC cell line PC-14 was resistant to Tan IIA up to a concentration of 50 µM (Figure 1).

TKIs gefitinib and erlotinib, which competitively block binding of ATP to the tyrosine kinase domain of EGFR, are effective therapeutic agents for NSCLC treatment. PC-9 cells are a gefitnib-sensitive human cell line with an exon 19 deletion (delE746-A750) in their EGFR, and PC-14 cells are a gefitinibresistant one without EGFR mutations. Therefore, the sensitivity to Tan IIA in PC-9 cells and PC-14 cells may be associated with the responsiveness of both cell lines to TKIs.

To study whether the responsiveness to TKIs and that to Tan IIA are related to each other, we isolated AG1478-resistant cell line AY-01 from PC-9 cells (Figure 2A). AG1478 is a selective inhibitor of EGFR tyrosine kinase, and its quinazoline-containing structure is closely similar to that of both TKIs gefitinib and erlotinib. Next, AY-01 cells were used together with 2 other TKI-resistant NSCLC cell lines, H358 and H1650, to test the anti-growth activity of Tan IIA. H1650 cells express an exon 19 deletion of their EGFR gene and harbor no known EGFR TKIresistance mechanism other than functional PTEN loss. On the other hand, H358 cells harbor a heterozygous activating KRAS G12C mutation, but no EGFR mutation. As shown in (Figure 2B), all of the TKI-resistant NSCLC cell lines were sensitive to the growth inhibitory action of Tan IIA; although H1650 cells were much more sensitive to Tan IIA than were the other 2 cell lines. Therefore, there was no association between TKI-sensitivity and growth inhibition by Tan IIA.

A secondary mutation in EGFR (T790M) and amplification of MET have been identified as major mechanisms of acquired resistance to TKIs [11]. Mutational activation of KRAS is also associated with primary resistance toward gefitinib [12]. KRAS encodes a small GTP-binding protein involved in many cellular processes including proliferation, differentiation, and apoptosis. Its mutations are detected in more than 25% of lung adenocarcinomas, and activating mutations in EGFR and KRAS seem to be mutually exclusive. Since H358 cells harboring a KRAS mutation were sensitive to the growth inhibitory action of Tan IIA (Figure2B), it is of importance to examine the effect of Tan IIA on A549 lung adenocarcinoma cells, which harbor a homozygous activating KRAS G12S mutation and wild-type EGFR. Microscopic observation revealed that 10 µΜ Tan IIA completely inhibited the growth of A549 cells (data not shown). Thus, Tan IIA exerted antigrowth activity against NSCLCs carrying not only EGFR mutations but also KRAS mutations. Although we identified another type of NSCLC cells (PC-14), which were resistant to both TKI and Tan IIA, Tan IIA should be considered as a potential agent for the chemotherapeutic treatment of a variety of gefitinib-sensitive and -resistant NSCLC.

NAD (P)H: quinone oxidoreductase catalyzes the twoelectron reduction of a variety of quinone substrate, using both NADH and NADPH as electron donors. Tan IIA is a phenanthrene quinone derivative. So, to explore if the growth of NSCLCs was inhibited by Tan IIA through the generation of reactive oxygen species (ROS), we treated PC-9 and PC-14 cells with Tan IIA. As shown in (Figure 3A), the compound increased the intensity of the fluorescence in PC-9 cells, but not in PC-14 cells. Therefore, it is likely that DCF-DA was oxidized and converted into fluorescent 2’, 7’-dichlorofluorescein by ROS in Tan IIA-sensitive PC-9 cells. Next, we used N-acetyl cysteine (NAC), ROS scavenger, to counteract the production of ROS and examined its effect on the Tan IIA-induced growth inhibition. Treatment of PC-9 cells with 5 mM NAC abolished the Tan IIA -induced growth inhibition (Figure 3B). Chiu et al. [13] previously demonstrated that Tan IIA-induced apoptosis of A549 cells is associated with ROS production. Furthermore, Liu et al. [6] reported that Tan IIA induces cell death of A549 NSCLC cells via a NAD(P)H:quinone oxidoreductase-initiated and ROS-mediated apoptotic pathway. Taken together, all evidence suggests that Tan IIA is likely to exert its inhibitory activity towards a variety of NSCLC cells via increased ROS production.

Celestrol, a triterpene derived from the Chinese herb Trypterygium wilfordii, has been reported to induce apoptosis in 3 gefitinib resistant NSCLC cells lines, H1650, H1975, and H2228 [14]. As described above, H1650 has an activating deletion on exon 19 of the EGFR gene, but is resistant to gefitinib. H1975 harbors a secondary mutation (T790M) in EGFR in addition to the L858R activating mutation. On the other hand, H2228 contains wild-type EGFR and EML4-ALK fusion mutation. Therefore, natural compounds with a terpene structure, such as Tan IIA and celestrol, could serve as lead candidates for the development of therapeutic agents against a variety of gefitinib-resistant NSCLC.

In a previous study, we characterized the molecular mechanism by which ent-kaurane diterpenoids (eKDs) from Croton tonkinensis induce apoptosis of colorectal cancer cell lines, a study in which we examined 10 eKDs with slightly different structures [10].

In addition to Tan IIA, we have also isolated tanshinone I and cryptotanshinone. Since these 3 tanshinones possess slightly different structures one from another (Figure 4), a comparative study of their anti-growth activity could be useful to understand the molecular basis of their action towards NSCLC cells. Especially, it is interesting to compare the effect of these 3 tanshinones on the growth of PC-14 cells, whose driver mutation genes leading to gefitinib resistance are not known.

CONCLUSION

We studied the anti-growth activity of Tan IIA against 6 NSCLC cell lines, which included gefitinib-sensitive PC-9 cells and gefitinib-resistant PC-14, AY-01, H358, H1650, and A549 cells. Growth of all the cells, except PC-14 cells, was inhibited by Tan IIA. Therefore, Tan IIA will be of help to develop new anti-cancer drugs for the treatment of a variety of gefitinib-sensitive and -resistant NSCLC.

ACKNOWLEDGEMENTS

This work was supported by JSPS KAKENHI Grant Numbers 18590088, 23590098 and by the Ministry of Science and Technology of Vietnam (No 37/2010/H?-N?T). We thank Miss A. Nagai, Miss M. Okayama, and Mr. D. Hashimura for their technical assistance.

REFERENCES

1. Fukuoka M, Wu YL, Thongprasert S, Sunpaweravong P, Leong SS, Sriuranpong V, et al. Biomarker analyses and final overall survival results from a phase III, randomized, open-label, first-line study of gefitinib versus carboplatin/paclitaxel in clinically selected patients with advanced non-small-cell lung cancer in Asia (IPASS). J Clin Oncol. 2011; 29: 2866-2874.

2. Pérez-Soler R , Chachoua A, Hammond LA, Rowinsky EK, Huberman M, Karp D, Rigas J. Determinants of tumor response and survival with erlotinib in patients with non--small-cell lung cancer. J Clin Oncol. 2004; 22: 3238-3247.

3. Lynch TJ , Bell DW, Sordella R, Gurubhagavatula S, Okimoto RA, Brannigan BW, Harris PL. Activating mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib. N Engl J Med. 2004; 350: 2129-2139.

4. Roh MS. Molecular pathology of lung cancer: current status and future directions. Tuberc Respir Dis (Seoul). 2014; 77: 49-54.

5. Engelman JA , Jänne PA. Mechanisms of acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors in nonsmall cell lung cancer. Clin Cancer Res. 2008; 14: 2895-2899.

6. Liu F , Yu G, Wang G, Liu H, Wu X, Wang Q, Liu M. An NQO1-initiated and p53-independent apoptotic pathway determines the anti-tumor effect of tanshinone IIA against non-small cell lung cancer. PLoS One. 2012; 7: e42138.

7. Su CC , Chen GW, Kang JC, Chan MH. Growth inhibition and apoptosis induction by tanshinone IIA in human colon adenocarcinoma cells. Planta Med. 2008; 74: 1357-1362.

8. Onitsuka M, Fujiu M, Shinma N, Maruyama HB. New platelet aggregation inhibitors from Tan-Shen; radix of Salvia miltiorrhiza Bunge. Chem Pharm Bull (Tokyo). 1983; 31: 1670-1675.

9. Lee SY , Choi DY, Woo ER. Inhibition of osteoclast differentiation by tanshinones from the root of Salvia miltiorrhiza bunge. Arch Pharm Res. 2005; 28: 909-913.

10. Thuong PT, Khoi NM, Ohta S, Shiota S, Kanta H, Takeuchi K, et al. ent-Kaurane diterpenoids from Croton tonkinensis induce apoptosis in colorectal cancer cells through the phosphorylation of JNK mediated by reactive oxygen species and dual-specificity JNK kinase MKK4. Anticancer Agents Med Chem. 2014; 14: 1051-1061.

11. Bean J, Brennan C, Shih JY, Riely G, Viale A, Wang L, et al. MET amplication occurs with or without T790M mutations in EGFR mutant lung tumors with acquired resistance to gefitinib or erlotinib. Proc Natl Acad Sci U S A. 2007; 104: 20932-20937.

12. Pao W, Wang TY, Riely GJ, Miller VA, Pan Q, Ladanyi M, Zakowski MF. KRAS mutations and primary resistance of lung adenocarcinomas to gefitinib or erlotinib. PLoS Med. 2005; 2: e17.

13. Chiu TL , Su CC. Tanshinone IIA induces apoptosis in human lung cancer A549 cells through the induction of reactive oxygen species and decreasing the mitochondrial membrane potential. Int J Mol Med. 2010; 25: 231-236.

14. Fan XX, Li N, Wu JL, Zhou YL, He JX, Liu L, et al. Celastrol induces apoptosis in gefitinib-resistance non-small cell lung cancer cells via caspase-dependent pathways and Hsp90 client protein degradation. Molecules. 2014. 19; 3508-3522.

Received : 16 Oct 2014
Accepted : 21 Nov 2014
Published : 24 Nov 2014
Journals
Annals of Otolaryngology and Rhinology
ISSN : 2379-948X
Launched : 2014
JSM Schizophrenia
Launched : 2016
Journal of Nausea
Launched : 2020
JSM Internal Medicine
Launched : 2016
JSM Hepatitis
Launched : 2016
JSM Oro Facial Surgeries
ISSN : 2578-3211
Launched : 2016
Journal of Human Nutrition and Food Science
ISSN : 2333-6706
Launched : 2013
JSM Regenerative Medicine and Bioengineering
ISSN : 2379-0490
Launched : 2013
JSM Spine
ISSN : 2578-3181
Launched : 2016
Archives of Palliative Care
ISSN : 2573-1165
Launched : 2016
JSM Nutritional Disorders
ISSN : 2578-3203
Launched : 2017
Annals of Neurodegenerative Disorders
ISSN : 2476-2032
Launched : 2016
Journal of Fever
ISSN : 2641-7782
Launched : 2017
JSM Bone Marrow Research
ISSN : 2578-3351
Launched : 2016
JSM Mathematics and Statistics
ISSN : 2578-3173
Launched : 2014
Journal of Autoimmunity and Research
ISSN : 2573-1173
Launched : 2014
JSM Arthritis
ISSN : 2475-9155
Launched : 2016
JSM Head and Neck Cancer-Cases and Reviews
ISSN : 2573-1610
Launched : 2016
JSM General Surgery Cases and Images
ISSN : 2573-1564
Launched : 2016
JSM Anatomy and Physiology
ISSN : 2573-1262
Launched : 2016
JSM Dental Surgery
ISSN : 2573-1548
Launched : 2016
Annals of Emergency Surgery
ISSN : 2573-1017
Launched : 2016
Annals of Mens Health and Wellness
ISSN : 2641-7707
Launched : 2017
Journal of Preventive Medicine and Health Care
ISSN : 2576-0084
Launched : 2018
Journal of Chronic Diseases and Management
ISSN : 2573-1300
Launched : 2016
Annals of Vaccines and Immunization
ISSN : 2378-9379
Launched : 2014
JSM Heart Surgery Cases and Images
ISSN : 2578-3157
Launched : 2016
Annals of Reproductive Medicine and Treatment
ISSN : 2573-1092
Launched : 2016
JSM Brain Science
ISSN : 2573-1289
Launched : 2016
JSM Biomarkers
ISSN : 2578-3815
Launched : 2014
JSM Biology
ISSN : 2475-9392
Launched : 2016
Archives of Stem Cell and Research
ISSN : 2578-3580
Launched : 2014
Annals of Clinical and Medical Microbiology
ISSN : 2578-3629
Launched : 2014
JSM Pediatric Surgery
ISSN : 2578-3149
Launched : 2017
Journal of Memory Disorder and Rehabilitation
ISSN : 2578-319X
Launched : 2016
JSM Tropical Medicine and Research
ISSN : 2578-3165
Launched : 2016
JSM Head and Face Medicine
ISSN : 2578-3793
Launched : 2016
JSM Cardiothoracic Surgery
ISSN : 2573-1297
Launched : 2016
JSM Bone and Joint Diseases
ISSN : 2578-3351
Launched : 2017
JSM Bioavailability and Bioequivalence
ISSN : 2641-7812
Launched : 2017
JSM Atherosclerosis
ISSN : 2573-1270
Launched : 2016
Journal of Genitourinary Disorders
ISSN : 2641-7790
Launched : 2017
Journal of Fractures and Sprains
ISSN : 2578-3831
Launched : 2016
Journal of Autism and Epilepsy
ISSN : 2641-7774
Launched : 2016
Annals of Marine Biology and Research
ISSN : 2573-105X
Launched : 2014
JSM Health Education & Primary Health Care
ISSN : 2578-3777
Launched : 2016
JSM Communication Disorders
ISSN : 2578-3807
Launched : 2016
Annals of Musculoskeletal Disorders
ISSN : 2578-3599
Launched : 2016
Annals of Virology and Research
ISSN : 2573-1122
Launched : 2014
JSM Renal Medicine
ISSN : 2573-1637
Launched : 2016
Journal of Muscle Health
ISSN : 2578-3823
Launched : 2016
JSM Genetics and Genomics
ISSN : 2334-1823
Launched : 2013
JSM Anxiety and Depression
ISSN : 2475-9139
Launched : 2016
Clinical Journal of Heart Diseases
ISSN : 2641-7766
Launched : 2016
Annals of Medicinal Chemistry and Research
ISSN : 2378-9336
Launched : 2014
JSM Pain and Management
ISSN : 2578-3378
Launched : 2016
JSM Women's Health
ISSN : 2578-3696
Launched : 2016
Clinical Research in HIV or AIDS
ISSN : 2374-0094
Launched : 2013
Journal of Endocrinology, Diabetes and Obesity
ISSN : 2333-6692
Launched : 2013
Journal of Substance Abuse and Alcoholism
ISSN : 2373-9363
Launched : 2013
JSM Neurosurgery and Spine
ISSN : 2373-9479
Launched : 2013
Journal of Liver and Clinical Research
ISSN : 2379-0830
Launched : 2014
Journal of Drug Design and Research
ISSN : 2379-089X
Launched : 2014
JSM Clinical Oncology and Research
ISSN : 2373-938X
Launched : 2013
JSM Bioinformatics, Genomics and Proteomics
ISSN : 2576-1102
Launched : 2014
JSM Chemistry
ISSN : 2334-1831
Launched : 2013
Journal of Trauma and Care
ISSN : 2573-1246
Launched : 2014
JSM Surgical Oncology and Research
ISSN : 2578-3688
Launched : 2016
Annals of Food Processing and Preservation
ISSN : 2573-1033
Launched : 2016
Journal of Radiology and Radiation Therapy
ISSN : 2333-7095
Launched : 2013
JSM Physical Medicine and Rehabilitation
ISSN : 2578-3572
Launched : 2016
Annals of Clinical Pathology
ISSN : 2373-9282
Launched : 2013
Annals of Cardiovascular Diseases
ISSN : 2641-7731
Launched : 2016
Journal of Behavior
ISSN : 2576-0076
Launched : 2016
Annals of Clinical and Experimental Metabolism
ISSN : 2572-2492
Launched : 2016
Clinical Research in Infectious Diseases
ISSN : 2379-0636
Launched : 2013
JSM Microbiology
ISSN : 2333-6455
Launched : 2013
Journal of Urology and Research
ISSN : 2379-951X
Launched : 2014
Journal of Family Medicine and Community Health
ISSN : 2379-0547
Launched : 2013
Annals of Pregnancy and Care
ISSN : 2578-336X
Launched : 2017
JSM Cell and Developmental Biology
ISSN : 2379-061X
Launched : 2013
Annals of Aquaculture and Research
ISSN : 2379-0881
Launched : 2014
Clinical Research in Pulmonology
ISSN : 2333-6625
Launched : 2013
Journal of Immunology and Clinical Research
ISSN : 2333-6714
Launched : 2013
Annals of Forensic Research and Analysis
ISSN : 2378-9476
Launched : 2014
JSM Biochemistry and Molecular Biology
ISSN : 2333-7109
Launched : 2013
Annals of Breast Cancer Research
ISSN : 2641-7685
Launched : 2016
Annals of Gerontology and Geriatric Research
ISSN : 2378-9409
Launched : 2014
Journal of Sleep Medicine and Disorders
ISSN : 2379-0822
Launched : 2014
JSM Burns and Trauma
ISSN : 2475-9406
Launched : 2016
Chemical Engineering and Process Techniques
ISSN : 2333-6633
Launched : 2013
Annals of Clinical Cytology and Pathology
ISSN : 2475-9430
Launched : 2014
JSM Allergy and Asthma
ISSN : 2573-1254
Launched : 2016
Journal of Neurological Disorders and Stroke
ISSN : 2334-2307
Launched : 2013
Annals of Sports Medicine and Research
ISSN : 2379-0571
Launched : 2014
JSM Sexual Medicine
ISSN : 2578-3718
Launched : 2016
Annals of Vascular Medicine and Research
ISSN : 2378-9344
Launched : 2014
JSM Biotechnology and Biomedical Engineering
ISSN : 2333-7117
Launched : 2013
Journal of Hematology and Transfusion
ISSN : 2333-6684
Launched : 2013
JSM Environmental Science and Ecology
ISSN : 2333-7141
Launched : 2013
Journal of Cardiology and Clinical Research
ISSN : 2333-6676
Launched : 2013
JSM Nanotechnology and Nanomedicine
ISSN : 2334-1815
Launched : 2013
Journal of Ear, Nose and Throat Disorders
ISSN : 2475-9473
Launched : 2016
JSM Ophthalmology
ISSN : 2333-6447
Launched : 2013
Journal of Pharmacology and Clinical Toxicology
ISSN : 2333-7079
Launched : 2013
Annals of Psychiatry and Mental Health
ISSN : 2374-0124
Launched : 2013
Medical Journal of Obstetrics and Gynecology
ISSN : 2333-6439
Launched : 2013
Annals of Pediatrics and Child Health
ISSN : 2373-9312
Launched : 2013
JSM Clinical Pharmaceutics
ISSN : 2379-9498
Launched : 2014
JSM Foot and Ankle
ISSN : 2475-9112
Launched : 2016
JSM Alzheimer's Disease and Related Dementia
ISSN : 2378-9565
Launched : 2014
Journal of Addiction Medicine and Therapy
ISSN : 2333-665X
Launched : 2013
Journal of Veterinary Medicine and Research
ISSN : 2378-931X
Launched : 2013
Annals of Public Health and Research
ISSN : 2378-9328
Launched : 2014
Annals of Orthopedics and Rheumatology
ISSN : 2373-9290
Launched : 2013
Journal of Clinical Nephrology and Research
ISSN : 2379-0652
Launched : 2014
Annals of Community Medicine and Practice
ISSN : 2475-9465
Launched : 2014
Annals of Biometrics and Biostatistics
ISSN : 2374-0116
Launched : 2013
JSM Clinical Case Reports
ISSN : 2373-9819
Launched : 2013
Journal of Cancer Biology and Research
ISSN : 2373-9436
Launched : 2013
Journal of Surgery and Transplantation Science
ISSN : 2379-0911
Launched : 2013
Journal of Dermatology and Clinical Research
ISSN : 2373-9371
Launched : 2013
JSM Gastroenterology and Hepatology
ISSN : 2373-9487
Launched : 2013
Annals of Nursing and Practice
ISSN : 2379-9501
Launched : 2014
JSM Dentistry
ISSN : 2333-7133
Launched : 2013
Author Information X