DEVELOPMENT AND EVALUATION OF LAPATINIB DITOSYLATE SELF-NANO-EMULSIFYING DRUG DELIVERY SYSTEMS

DEVELOPMENT AND EVALUATION OF LAPATINIB DITOSYLATE SELF-NANO-EMULSIFYING DRUG DELIVERY SYSTEMS

Abdullah Khan ^{* 1}, S. Roshan ², K. Anandarajagopal ¹ and B. Tazneem ²

KPJ Healthcare University College ¹, Nilai, Negeri Sembilan, Malaysia.

Deccan School of Pharmacy ², Aghapura, Hyderabad - 500001, Telangana, India.

ABSTRACT: The main objective of the study is to develop gelucire 44/14 based lapatinib ditosylate (LPT) SNEDDS for the improvement of solubility and drug release. Pseudo tertiary phase diagram constructed for varying oil, surfactant, and co-surfactant combinations. Fifteen LPT SNEDDS formulations were prepared with anticipated component ratios (gelucire 44/14, peceol, and capmul MCM) based on monophasic region are subjected to thermodynamic physical stability test. Formulations that cleared stability tests were evaluated for % transmission, drug content, and in-vitro drug release analysis. The optimized formulation analyzed for particle size and zeta potential, followed by FTIR and SEM analysis. Formulation F13 with a maximum drug release of 98.96% in 60 min higher than 38.90±3.25% of pure drug is considered an optimized formulation. The average particle size and zeta potential of the F13 were 72.5 nm and -13.4 mV, respectively. The FTIR and SEM studies do not indicate any drug excipient interaction and confirm uniform drug distribution. The formulation subjected to accelerated stability study is proved to be stable over a period of six months. The results suggested that LPT SNEDDS formulations are an encouraging alternative for improved solubility and drug release of LPT.

Lapatinib ditosylate, SNEDDS, Metastatic breast cancer (MBC), Solubility, Gelucire 44/14

INTRODUCTION: Breast cancer is the most spotted cancer amongst women in India. Metastatic breast cancer (MBC) is the most radical stage of breast cancer that occurs when the disease starts spreading to other parts of the body ¹. In the recent past, various new agents contributed to the improvements in the treatment outcome, including taxanes, anthracyclines, antimetabolites, aromatase inhibitors, and antibodies. MBC ranks as the number one cancer among Indian females with a rate as high as 25.8 per 100,000 women and mortality of 12.7 per 100,000 women, according to health ministry ^{2, 3, 4}.

Cancer is an atypical molecular circuitry linking signaling pathways that regulate cellular growth. ERBB receptors, also known as epidermal growth factor receptors (EGFR) play a vital role in breast and another cancer signaling. These receptors are vital in organ development and regulating cellular differentiation and morphogenesis in tissues. LPT is a tyrosine kinase inhibitor that inhibits the receptors ErbB1 and ErbB2 receptor growth. The validation for LPT as an anticancer drug is that the cordon of the tyrosine kinase activity of ErbB1 or ErbB2 is estimated to hinder the transforming activity that grades from overexpression of these receptors ^{5, 6, 7}.

LPT, an active oral anti-cancer agent belongs to class II of BCS classification with deprived water solubility and encompassing the first-pass metabolism, is considered for SNEDDS formulation to improve its solubility and drug release where a drug is incorporated into lipid carrier and produce as self-nanoemulsion, which was stabilized by surfactants and co-surfactants.

Lipid-based self-nano emulsifying drug delivery systems (SNEDDS) have gained back the prominence of nanoemulsions by modest alterations for attaining enhanced drug delivery. Thus the current study was aimed to develop gelucire 44/14 based LPT SNEDDS for the improvement of solubility and dissolution rate.

MATERIALS AND METHODS:

Materials: LPT is gifted by Hetero Labs Limited, Hyderabad. Capric glyceride, cremophor EL, gelucire 33/01, gelucire 44/14, glyceryl monosterate, and oleic acid are procured from Gattefosse, Mumbai. Labrasol, tween 20, brij 35, capmul CMC, and acrysol K-160 were obtained from BASF, Mumbai. Kolliphor PS-80, kolliphor HS-15, and peceol were purchased from Yarrow Chemical Products, Mumbai.

Solubility Studies: The solubility of LPT in different oils (glyceryl monosterate, gelucire 33/01, gelucire 44/14, oleic acid and acrysol K-160), surfactants (capric glyceride, kolliphor PS-80, peceol, kolliphor HS-15,cemophor El and span 80), co-surfactants (capmul MCM, brij-35, PEG-600, tween-20 and labrasol) were examined by adding excess amount of LPT (approximately 10mg) with 2 ml each. The samples were further centrifuged at 8,000 rpm for 20-30 min at 4 °C.

The supernatant removed, and the drug concentration was determined by UV at 309 nm. An oil, surfactant, and co-surfactant containing more drug content are selected for formulating SNEDDS for LPT ⁸.

% Transmittance: From the preliminary solubility studies, the selected vehicles were mixed at varying ratios. Firstly S_mix (surfactant: co-surfactant) of different ratios (1:1, 2:1, and 3:1) are prepared, and later, oil: S_mix ratios ranging from 1:9 to 9:1 are prepared in 2 ml Eppendorf tubes and thoroughly shaken and kept aside. A fixed volume of water (100 ml) was added to the calculated amount of vehicles (0.1 ml) and checked for phase separation and turbidity visually. Solutions with no phase separation and clear appearance were separated and used for checking transmittance using a UV spectrophotometer at 309 nm. Maximum transmittance values indicate the formation of small nanosize particles. Ratios showing a percentage transmittance value more than 80 were considered for plotting pseudo ternary phase diagram for representing self-emulsification region ⁹.

Construction of Pseudo Ternary Phase Diagrams: Transmittance values of more than 80 indicate formation of nanoemulsion; hence these ratios were noted and selected for plotting pseudo ternary phase diagram. Phase diagrams are plotted using CHEMIX software in which two ratios were fed and calculated; it shows am emulsification region which is shaded and from which 15 ratios for formulating LPT SNEDDS were chosen ¹⁰.

Development of LPT SNEDDS Formulations: The LPT SNEDDS formulations prepared using gelucire 44/14 as oil phase and peceol and capmul MCM used as surfactant and co-surfactant respectively (Table 1). LPT (250 mg) was added to oil into the glass vial and heated to 40ºC. The oily mixture was mixed thoroughly with surfactant and co-surfactant and sonicated for 15 min.

TABLE 1: FORMULATION TRIALS OF LPT SNEDDS

Thermodynamic Stability Test: The stable samples were centrifuged for 5 min at 3000 rpm and examined for any phase separation. The stability of the formulation is checked by centrifugation study and freeze-thawing test ¹¹.

Drug Content: The extent of drug present in 0.2 ml of formulation is estimated by UV spectrometer. The solution filtered, diluted with 0.1N HCl, and analyzed at λ_max 309 nm against blank ^11.

% Transmittance of SNEDDS: Nanoemulsions are optical isotropic solutions of oil, surfactant, and co-surfactants and thermodynamically stable. Hence investigation of optical clarity is conducted for the determination of isotopic parameter. The LPT SNEDDS reconstituted with distilled water. The resulting emulsion was visually examined for the presence of any turbidity. The percentage transmittance of the samples measured using UV spectrophotometer at 309 nm ^{12, 13}.

In-vitro Dissolution Studies of LPT SNEDDS Formulations: The SNEDDS whose weight equivalent to 250 mg of LPT was filled into hard gelatin capsules with 0.1N HCl with 2% polysorbate 80 as dissolution media, stirred at 37°C at 50 rpm rate. At pre-set time intervals, 5 ml of sample withdrawn, filtered through 0.45 μm membrane filter, and analyzed spectophoto-metrically at 309 nm ^{14, 15}.

Characterization of LPT SNEDDS:

Drug-Excipient Compatibility Analysis: Drug excipient compatibility was checked by FTIR spectroscopy (Shimadzu FTIR 8400S, Japan). Eight scans were recorded at 4 cm⁻¹ resolution within a frequency range between 400–4000 cm⁻¹.

Determination of Particle Droplet Size: The droplet size imitates the uniformity of particles, which in turn depicts the size distribution of nanoemulsion. The droplet size of LPT SNEDDS formulations is evaluated by Photon correlation spectroscopy (Malvern Instrument UK) that measures the size range of 10 to 5000 nm ¹⁶. All measurements were done in triplicate from three independent samples.

Determination of Zeta Potential: The stability of nanoemulsion is directly correlated to surface charge magnitude. The zeta potential of the dilute LPT SNEDDS formulation homogenized was evaluated with a zeta meter system measured with Zetasizer. (Malvern, Nano Series ZS90, Malvern Instruments, Ltd., UK). All studies were conducted in triplicate at 25°C ¹⁷.

Determination of Shape and Surface Morphology: Shape and surface morphology LPT SNEDDS scanned with scanning electron microscopy (JEOL JEM 2100 F, USA) ¹⁸.

Stability Studies: The LPT SNEDDS formulations were filled in gelatin capsules, and stability studies conducted stability chambers. The final dosage form was subjected to stability studies for 6 months at 40±2°C/75±5% RH as mentioned according to ICH guidelines (Thermo lab, Mumbai, India) ¹⁹.

RESULTS AND DISCUSSION:

Solubility Studies: From the solubility studies, it was evident that medium-chain triglycerides were highly influencing the solubility of the drug; these are obtained from a vegetable source and are resistant to oxidation. Gelucire 44/14, being a medium-chain triglyceride with high solvent capacity, was showing a high degree of solubility of about 232.11mg/ml for LPT when compared to other oils. Surfactants and co-surfactants with high HLB values were useful in solubilizing LPT; these are most commonly used as they are more stable and non-toxic in the formulation. From the solubility study, peceol-81.22 mg/ml (surfactant) and capmul MCM-80.99 (co-surfactant) were showing high solubilizing efficiency for LPT Fig. 1-3.

Pseudo Ternary Phase Diagram: Pseudo-ternary phase diagram constructed to ascertain the micro emulsion regions for optimizing the concentrations of oil, surfactant, co-surfactant, and their mixing ratios. This diagram comprises oil, water, and S_mix, at each corner in the phase triangle. The oil (gelucire 44/14), surfactants (peceol) with co-surfactant (peceol) at different ratios (1:1, 2:1, 3:1) were dispersed at varying weight ratios oil and S_mix ranging from 1:9 to 9:1 in different vials followed by vertexing and water titration (Fig. 4).The translucent and low viscosity nanoemulsion area is presented as a shaded area in the phase diagrams. This shaded area in phase diagram represents self-emulsification region and thus ratios were extracted from this region for formulating LPT SNEDDS.

FIG. 4: TERNARY PHASE DIAGRAM OF GELUCIRE 44/14, PECEOL AND CAPMUL MCM

Thermodynamic Stability Studies of LPT SNEDDS: Prolonged storage of SNEDDS might result in precipitation of drug; hence the stability of a formulation is checked by centrifugation and freeze-thawing test. The stability studies conducted indicated no significant phase separation after centrifugation and no effect of temperature variation on the physical appearance of the LPT SNEDDS formulations after freeze-thawing Table 2.

TABLE 2: THERMODYNAMIC STABILITY STUDIES OF LPT SNEDDS FORMULATIONS

TABLE 3: % TRANSMISSION AND DRUG CONTENT OF LPT SNEDDS

Data  from  each  response  for  the  observed  values  is  presented  in  mean ± SD, n=3

The % Transmittance Measurement and Drug Content of LPT SNEDDS: The % transmittance of the 15 formulations post-dilution at diverse time intervals studied. The values ranged from 72.17% to 99.18%. The results indicated that an increase in surfactant to cosurfactant ratio increased % transmittance, thus increasing the optical clarity of the formulation. The highest transmission observed for F13 with S_mix ratio of 3:1 is also indicative of nanometer range particles.

The drug content of the formulated LPT SNEDDS was found to be in the range of 90.12 – 99.32 %, with maximum drug content observed for F13 (99.32%) Table 3.

In-vitro Dissolution of LPT SNEDDS: The dissolution rate from all the formulations varied between 90.33±5.02 - 98.96±5.28% for 60 min, which was a higher almost three fold increase when compared to pure drug release (38.90±3.25% in 60 min) indicates formation of small nanosize particles with increased surface area forming nano-emulsion. The formulation, when it comes in contact with the dissolution media, immediately form nanoemulsion whose interfacial tension is reduced by surfactants and co-surfactants; hence the ratio with more surfactant and co-surfactant showed a high degree of drug release, and also in general as the particle size is small with more surface area, more is the absorption and hence more is the drug release, and out of all formulation F13 was showing maximum drug release of 98.96±5.28% at the end of 60min with more S_mix ratio Fig. 5, 6 and 7.

Characterization of LPT SNEDDS:

Drug Excipient Interactions by FTIR Spectroscopy: The IR spectra peaks of the LPT pure drug and the optimized SNEDDS formulation F13 spectra were observed at the same wavenumbers as shown in Fig. 8 and 9. The peaks at 1120–1250 cm^-1 represent C-SO₂ stretching frequency, 3050–3150 cm^-1 represent N-H group, and 1350–1470 cm^-1 belongs C=N. The strong band at 682 cm^-1 corresponds to C-Cl stretching, while the band at 1166 cm^-1 corresponds to C-F stretching frequency. The presence of these predominant peaks in all the spectra indicates no significant interaction takes place between drug and formulation excipients.

FIG. 8: FTIR SPECTRUM OF LPT PURE DRUG

FIG. 9: FTIR SPECTRUM OF FORMULATION F13 OF LPT SNEDDS

Particle Size Analysis of LPT SNEDDS: Smaller particle size facilitates larger interfacial surface area for drug absorption and hence improving bioavailability and also governs the drug release effectively. The average particle size of LPT SNEDDS formulation (F13) is 72.5 nm that is within the nanometric range of < 100 nm that attributed to the stability of the oil droplets and localization of the surfactant molecules at the oil-water interface. This small nanorange particle size of F13 SNEDDS formulation provided more surface area for dissolution and thus enhanced the solubility and facilitated high drug release Fig. 10.

FIG. 10: PARTICLE SIZE ANALYSIS OF LPT SNEDDS FORMULATION F13

Zeta Potential of LPT SNEDDS: The emulsification efficiency of the formulation depends on the polarity of the emulsion droplet. Zeta potential value correlates to the stability of colloidal dispersions and the drug release efficiency of the formulation. For nanoparticles, higher zeta potential values infer stability and higher cellular uptake. The zeta potential value > 5 mV provides excellent stability; thus, in the present study, the zeta potential values of F13 is -13.7 mV that is the constraint for particle stability Fig. 11.

FIG. 11: ZETA POTENTIAL OF LPT SNEDDS OPTIMIZED FORMULATION F13

Scanning Electron Microscopy (SEM) for LPT SNEDDS: Scanning electron microscopy evaluated the morphology of SNEDDS formulation. The data showed a spherical shape for the optimized SNEDDS (F13) formulation with uniform and relatively narrow particle distribution. The images clearly indicate the spherical nature of nanoemulsions, with no signs of coalescence. The nanoemulsion droplets appeared as bright spheres with a smoother surface. The droplets appeared distinct, with no signs of drug precipitation conjecturing the stability of the formulation Fig. 12.

FIG. 12: SEM IMAGES OF LPT OPTIMIZED FORMULATIONS (F13)

Stability Studies: The stability of optimized LPT SNEDDS formulation studied for six months at 40 ±2 °C / 75±5 % RH according to ICH guidelines. The formulation was packed in hard gelatin capsules for stability studies. The studies indicated no significant variation in drug release and the drug content, physical and emulsifying properties Table 4.

TABLE 4: STABILITY STUDY OF OPTIMIZED FORMULATION (F13) STORED AT 40 ±2°C /75 ±5 %RH

Data  from  each  response  for  the  observed  values  is  presented  in  mean ± SD (n=3)

CONCLUSION: LPT SNEDDS was formulated with gelucire 44/14, peceol, and capmul MCM. The developed formulations were subjected to different evaluation parameters. Formulation F13 was found to be the best final optimized one based on the evaluation parameters. The in-vitro dissolution of the F13 formulation was highest among all other formulations of 98.96%, with reduced particle size and enhanced solubilizing capacity than other formulations and pure drugs. The results showed that the globule size of the F13 formulation was in the nanometer range (72.5 nm) with zeta potential -13.7 mV, which indicated reduced particle size and proved good stability. The SEM studied of F13 indicated the narrow uniform distribution of a drug in the formulation and found it to be stable for 6 months. Hence these investigations advocated the suitability of SNEDDS loaded LPT in the enhancement of solubility and drug release.

ACKNOWLEDGEMENT: Nil

CONFLICTS OF INTEREST: Nil

REFERENCES:

1. Atalay G, Cardoso F and Awada A: Novel therapeutic strategies targeting the epidermal growth factor receptor (EGFR) family and its downstream effectors in breast cancer. Ann Oncol 2003; 14: 1346-63.
2. Yarden Y: Biology of HER2 and its importance in breast cancer. Oncology 2001; 61: 1-13.
3. Stern DF: Tyrosine kinase signalling in breast cancer: ErbB family receptor tyrosine kinases. Breast Cancer Res 2000; 2: 176-83.
4. Nahta R, Hortobágyi GN and Esteva FJ: Growth factor receptors in breast cancer: potential for therapeutic intervention. The Oncologist 2003; 8: 5-17.
5. Mendelsohn J and Baselga J: Status of epidermal growth factor receptor antagonists in the biology and treatment of cancer. J Clin Oncol 2003; 21: 2787-99.
6. Zhou H, Kim YS and Peletier A: Effects of the EGFR/HER2 kinase inhibitor GW572016 on EGFR- and HER2-overexpressing breast cancer cell line proliferation, radio sensitization, and resistance. Int J Radiat Oncol Biol Phys 2004; 58: 344-52.
7. Konecny GE, Pegram MD and Venkatesan N: Activity of the dual kinase inhibitor LPT (GW572016) against HER-2-overexpressing and trastuzumab-treated breast cancer cells. Cancer Res 2006; 66: 1630-39.
8. Miryala V and Kurakula M: Self-nano emulsifying drug delivery system (SEDDS) for oral delivery of atorvastatin-Formulation and bioavailability studies. JDDT 2013; 3: 131-42.
9. Mahapatra AK, Murthy PN, Swadeep B and Prasad R: Self-emulsifying drug delivery systems (SEDDS): An update from formulation development to therapeutic strategies. Int J Pharm Tech Res 2014; 6: 545-68.
10. Shanmugam S, Baskaran R and Balakrishnan P: Solid selfnanoemulsifying drug delivery system (S-SNEDDS) containing phosphatidylcholine for enhanced bioavailability of highly lipophilic bioactive carotenoid lutein. Eur J Pharm Biopharm 2011; 79: 250-57.
11. Rajinikanth PS, Keat NW and Garg S: Self-nanoemulsifying drug delivery systems of valsartan: Preparation and in-vitro characterization. International Journal of Drug Delivery 2012; 4: 153-63.
12. Setthacheewakul S, Mahattanadul S, Phadoongsombut N, Pichayakorn W and Wiwattanaptapee R: Development and Characterization of Solid Self-emulsifying Drug Delivery System of Cilnidipine. Eur J Pharm Biopharm 2010; 76: 475-85.
13. Fahmy UA, Ahmed OA and Hosny KM: Development and evaluation of avanafil selfnanoemulsifying drug delivery system with rapid onset of action and enhanced bioavailability. AAPS Pharm SciTech 2015; 16: 53-58.
14. Zhao Y, Wang C and Chow AHL: Self-nanoemulsifying drug delivery system (SNEDDS) for oral delivery of Zeodary essential oil: formulation and bioavailability studies. Int J Pharm 2010; 383: 170-77.
15. Khoo SK, Humberstone AJ and Porter JH: Formulation design and bioavailability assessment of lipidic self-emulsifying formulations of halofantrine. Int J Pharm 1998; 167: 155-64.
16. Balakrishnan P, Lee BJ and Oh DJ: Enhanced oral bioavailability of dexibuprofen by a novel solid self-emulsifying drug delivery system (SEDDS). Eur J Pharm Biopharm 2009; 72: 539-45.
17. Gupta S, Chavhan S and Sawant KK: Self-nanoemulsifying drug delivery system for adefovir dipivoxil: design, characterization, in vitro and ex vivo evaluation. Coll Surf A: Physiochem Eng Aspects 2011; 392: 145-55.
18. Surendra S, Roshan I and Nagori B: Formulation and Evaluation of Self-emulsifying drug delivery System of Etoricoxib. Asian J Pharm Clin Res 2017; 10: 367-72.
19. Singh SK, Verma PRP and Razdan B: Glibenclamide-loaded selfnanoemulsifying drug delivery system: development and characterization. Drug Dev Ind Pharm 2010; 36: 933-45.
    

    ---

    How to cite this article:

    Khan A, Roshan S, Anandarajagopal K and Tazneem B: Development and evaluation of lapatinib ditosylate self-nanoemulsifying drug delivery systems. Int J Pharm Sci & Res 2021; 12(4): 2492-99. doi: 10.13040/IJPSR.0975-8232.12(4).2492-99.

    ---

    All © 2013 are reserved by the International Journal of Pharmaceutical Sciences and Research. This Journal licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License.

    ---