Phyto-Research

Phyto-Research

Translational Research On COVID

During the COVID-19 infec7on, the SARS-CoV-2 binds to the Angiotensinconver7ng enzyme 2 (ACE2) receptors present on the host cell surface which undergo endocytosis, and the virus enters inside the cell. Besides serving as a cell entry site for SARSCoV-2, ACE2 plays a very cri7cal role in regula7ng the balance between the ACE2 driven an7- inflammatory ac7ons and ACE/angiotensin AT1 receptors driven pro-inflammatory effects. There are two main challenges in developing drugs against the infec7on. First, the role of ACE2 is not completely understood and hence no drugsthat specifically target it. Second, as a consequence of SARS-CoV2 interac7on with ACE2, renin-angiotensin system (RAS) involved in the func7oning of vital organs like the heart, kidney, brain, and lungs is damaged. The primary targets in developing an7viral drugs for COVID-19 are ACE2, RNA-dependent RNA polymerase (RdRp), ORF8 and the specific enzymes involved in the viral and cellular gene expression. In silico docking studies revealed phytocons7tuents that interact with ACE2, AT1, and AT2 and thus may suppress the imbalance between ACE's counter-regulatory func7ons by interac7ng with RAS. The phytocons7tuents may also have inhibitory ac7ons on SARSCoV-2 replica7on and improving the oxygen-carrying capacity of haemoglobin. Thus, phytocons7tuents may have mul7ple sites of ac7on, protec7ng and countering the inflammatory damage and may be useful both as prophylac7c and treatment for COVID-19. Severely ill COVID-19 pa7ents may benefit from the diverse modulatory ac7on of these phytocons7tuents on the host RAS, viral RdRp and viral ORF8 targets.

Covid as a pandemic has created significant morbidity and mortality asking for a solution. Atrimed Pharmaceuticals, being specialized in drug development based on plant based molecules, has a systematic approach for drug development. The steps include target identification, Lead generation, lead optimization, ligand target binding in silico studies, MD Simulation, in vitro studies, in vivo studies and human trials.

spike glycoprotein, RdRp, 3CLPro, PLPro,E-Protein and M-Protein are the drug targets which have been identified and worked upon.  Understanding of plant chemistry and computational chemistry takes significant role in lead generation. Since we have a library of plant molecules, initial screening is done by looking for simulation studies for target ligand binding. Lead optimization is carried out by making changes at chemical moieties to simplify the synthesis, reduce the size or improve the binding. Subsequent  retrospective similarity search can identify the potential plants. A poly pharmacology approach to identify a molecule for multiple targets or, multiple molecules in single plant towards same or multiple targets or multiple molecules in multiple plants for different or same targets are some of the strategies to be adopted. All these strategies need to be reconfirmed in in vitro as well as in vivo experiments before trying them in human beings.

Through good understanding of medicinal chemistry, analytical chemistry and Synthesis, team of chemists can contribute for getting physical samples for experiments.

Thus a translational work between traditional medicine practitioners, computation biologists, medicinal chemists, synthetic chemists, Microbiologists/Molecular biologists/Biotechnologists, veterinarians/Animal pharmacologists and finally clinical pharmacologists leads to drug development at Atrimed Pharmaceuticals. The drug development for Covid is currently at Phase-1 and will resume to Phase 2 & 3 soon.

COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARSCoV-2), currently has a detrimental impact on human health, community and economy. SARS-CoV-2-infected lung +ssue ini+ally fails to induce a range of immune cell-recrui+ng molecules, including several interferons, sugges+ng that leukocytes are ineffec+vely recruited to infected lung shortly afer infec+on. The host cell entry receptor ACE2 has been iden+fied as an interferon target gene. Thus, even when interferons are upregulated, upregula+on of ACE2 expression may aggravate infec+on. These observa+ons suggest that baseline levels of leukocytes, already present/ growing in the lung prior to infec+on, may be important for coordina+ng an effec+ve early immune response. The baseline expression levels of the SARS-CoV-2 host cell entry receptor ACE2 and the levels of seven types of leukocytes (involved in triggering an acute an+-viral cellular immune response) were inves+gated in 1,927 human lung +ssue samples by “in silico flow cytometry". Baseline levels of CD8+ T cells, res+ng natural killer (NK) cells and ac+vated NK cells are significantly lower in lung +ssues with high expression of the SARSCoV-2 host cell entry receptor ACE2. Reduced CD8+ T cell counts also predict poor COVID-19 pa+ent survival. Elevated ACE2 expression increases sensi+vity to corona virus infec+on, accompanied with a lower ability to mount a rapid innate immune response at early stages. Finally, it may contribute to the substan+al varia+on in COVID-19 clinical presenta+on, ranging from asymptoma+c to severe respiratory and other symptoms.

Antivirals act in many ways to eliminate a virus, one of them being, blocking the interaction of the virus with host cells. One such interaction is initiated by Haemagglutinin-esterases (HEs). They are a group of viral envelope proteins that can cause the reversible attachment of the virus to host cells. HEs occur in influenza C, toro-, and coronaviruses, including SARS-CoV-2. A study of the crystal structure formed by the fusion of HE with host proteins was performed to show exactly how the HE binds to the host cell. It was found that in SARS-CoV-2, the HE-host protein binding causes the molecules to undergo radical steric changes and adopt opposite orientations leading to strong binding. This means, the virus can bind very strongly to the host cell and it cannot be easily detached again. But molecules can be developed/synthesized, that can block this interaction and thus prevent the virus from attaching to host cells so strongly. This will prevent viral RNA from entering the host cell and causing infection. This opens up potential avenues for the development of broad-spectrum antiviral drugs that can not only help with SARS-CoV-2 but also with other viruses that have similar HE-host cell interactions.

Currently, treatment options for COVID-19 remain limited, with the only evidence-based therapies being remdesivir and dexamethasone. Despite the use of these therapies, clinical outcomes remain dismal. Few recent trials emphasised that the inhaled route could provide better antiviral outcomes, as it enables maximal delivery of the active drug to the biological focus of SARS-CoV-2 infection, the respiratory epithelium, aka, the lungs. In a clinical trial, the safety and efficacy of an inhaled anti-inflammatory molecule, was investigated, in COVID-19 patients. Previous studies established that the molecule, called SNG001 was well tolerated in patients with asthma and COPD. This study showed that SNG001 seems to be well tolerated in COVID-19 patients and is associated with improvement and rapid recovery. Thus, treating patients admitted to hospital with COVID-19, with SNG001 should be explored further in a phase 3 trial to establish the timing of this treatment in critically ill patients with COVID-19 who have evidence of active viral infection in the lungs.

The virus that causes the current COVID-19 pandemic, SARS-CoV2, is a single stranded positive sense RNA virus that is closely related to severe acute respiratory syndrome corona virus (SARS-CoV). A drug developed against SARS-CoV, called Ivermectin, is a drug that can inhibit the transport of viral components to the host nucleus, thus preventing virus replication. As the viruses are very similar, it may well be effective against SARS-CoV-2. Human cells were infected with SARS-CoV-2 and treated with Ivermectin to find that it cleared all virus by 48 h. No toxicity of ivermectin was observed. It was also determined that ivermectin has antiviral action against the SARS-CoV-2 clinical isolate (in vitro), with a single dose able to control viral replication within 24–48 h in our system. This raises the possibility that ivermectin could be a useful antiviral to limit SARS-CoV-2, and is worthy of further consideration as a possible SARS-CoV-2 antiviral drug for further trial.

Effective antiviral agents are needed to treat severe acute respiratory syndrome-associated coronavirus-2 (SARS-CoV-2)infection. Interferons, alone or in combination with other interventions, are partly effective against animal coronaviruses, mouse hepatitis virus, and transmissible gastroenteritis virus; a few data on human studies report a limited response of interferon by the intranasal route. A study performed recently, assessed the antiviral potential of recombinant interferons against two clinical isolates, from Frankfurt and Hong Kong patients, replicated in human cells in the lablab. Interferon β was five to ten times more effective in neutralizing the virus. Interferon α effectively inhibited SARS-CoV-2 replication, but with a selectivity index 50–90 times lower than that for interferon β. Interferon γ was slightly better than interferon α in some human cell cultures, but was completely ineffective in other human cell cultures. Results showed that interferons inhibit SARS-CoV-2 replication in vitro. Interferon β was most potent, showing prophylactic or preventive protection as well as antiviral or curative potential after infection. Interferon β could be the drug of choice, alone or in combination with other antiviral drugs, in the treatment of COVID-19.

The outbreak of SARS-CoV-2 warrants the search for antiviral compounds to treat the disease. At present, no specific treatment has been identified for the infection. The antiviral potential of glycyrrhizin against two clinical isolates of SARS-CoV-2 was assessed and found to be highly potent, with a selectivity index of 67. Additionally, glycyrrhizin inhibits adsorption and penetration of the virus by the host cell. The mechanism of glycyrrhizin’s activity against SARS-CV is unclear. Glycyrrhizin affects cellular signaling pathways such as protein kinase C and nuclear factor NFκB. Preliminary results show that glycyrrhizin inhibits virus replication and has previously been used to treat patients with HIV-1 and chronic hepatitis C virus. Infrequent side-effects such as raised blood pressure and hypokalemia were reported in some patients after several months of glycyrrhizin treatment. Treatment of SARS-CoV-2 should only be needed for a short time. Since the side-effects of this compound are known and can be controlled, proper monitoring could lead to effective use of glycyrrhizin as a treatment for SARS-CoV-2. Even when high doses of glycyrrhizin were used in clinical trials, this compound had very few toxic effects and the drug was reported to be clinically effective. Future research is required before it can be used as a potential therapy against SARS-CoV-2.

In silico screening of phytotherapeutics for anti psoriasis activity

Thymoquinone (TMQ), a complex plant molecule is the main acLve ingredient of Nigella seeds. TMQ can scavenge or remove free radicals which are molecules in our body that can cause harm to us by destroying or inhibiLng the acLvity of various anL-oxidants. It has been shown by experiments in human skin cells that TMQ can help treat psoriasis. However, TMQ is easily degraded inside our body before it can act. Thus, to increase its stability inside our body so that it can act for a longer Lme, experiments were done in which they were placed inside nanoparLcle carriers. These carriers are less than 70nm in size and can enter the psoriaLc skin very efficiently and release the TMQ where they can act. Studies have been done in human skin cells as well as in mice with psoriasis, to show that this strategy works very well. Results indicate a clear reducLon in the level of nitric oxide and cytokines such as IL-2, IL-6, IL-1β, TNF-α, whereas in vivo results indicated improvement in the phenotypic, histopathological features and reduced level of IL-17 and TNF-α in psoriaLc skin. This study concludes that TMQ lipospheres could be used in the management of psoriasis.

A recent study inves7gated the possible stress-associated disturbances in lipid metabolism in lymphocytes of pa7ents with psoriasis Vulgaris (Ps) and psoria7c arthri7s (PsA). The outcomes suggested that the development of psoriasis is firmly associated with oxida7ve stress ensuing in altered lipid metabolism and cytokine produc7on, leading to the development of pro-inflammatory precondi7oning of the immune cells. Mononuclear cells addi7onally ac7vate pro-inflammatory states and enhance pro-oxida7ve, stressful condi7ons. Thus, ROS and enzyme-dependent changes in mononuclear cell phospholipids
can be considered as important processes associated with the pathophysiology of psoriasis. The levels of non-enzyma7c lipid metabolites associated with oxida7ve stress were higher in PsA. In the case of the enzyma7c metabolism of lipids, enhanced levels of endocannabinoids were observed in both forms of psoriasis, while reduced expression of their receptors and
ac7vi7es of phospholipases were detected only in PsA, sugges7ng the failure of the an7- the inflammatory mechanism and this may be an important factor leading to the transforma7on of Ps into a more severe form of PSA. In conclusion, monitoring the level of phospholipases can aid in predic7on of the progression of Ps into PsA if done complementary to the other parameters of lipid metabolism and the levels of respec7ve cytokines.

Psoriasis is a noncontagious autoimmune disease. Psoriasis is an immune-mediated chronic skin disease characterized by epidermal hyperproliferation, and intraepidermal accumulation of neutrophils and dermal inflammatory cell infiltrates that are composed of dendritic cells and T cells. In psoriasis, the diseased areas are typically red, dry, itchy, and scaly. Psoriasis varies in severity from small, localized patches to complete body coverage. Injury to the skin can trigger psoriatic skin changes at that spot, which is known as the Koebner phenomenon. It has an estimated global prevalence of 2–4% [2]. Males are more likely to develop psoriasis compared to females [3]. As desquamative erythema can affect any skin site, psoriasis profoundly impairs the patients’ quality of life, treatment satisfaction and adherence, and socioeconomic stability [4].

Psoriasis is also significantly comorbid with other autoimmune diseases, such as bullous pemphigoid [5]. Psoriasis is frequently associated with cardiovascular diseases, metabolic diseases, and renal disorders [6,7,8]. Cancer risk is slightly higher in patients with psoriasis [9]. The topical application of steroids and vitamin D3 analogs inhibits psoriatic inflammation and normalizes epidermal differentiation [10]. Systemic treatments, such as methotrexate, cyclosporine, phototherapy, and the phosphodiesterase 4 inhibitor apremilast, are useful for patients with extensive lesions [11].

Proinflammatory cytokines are commonly called immunoregulatory cytokines produced predominantly by activated macrophages and are involved in the up-regulation of inflammatory reactions and hence they favor inflammation. The net effect of an inflammatory response is determined by the balance between proinflammatory and anti-inflammatory cytokines. That the tumor necrosis factor-α (TNF-α) and IL-23/IL-17A axes appear to be major drivers in the pathogenesis of psoriasis is underscored by the excellent response of psoriasis to biologics targeting TNF-α, IL-23, and IL-17A, although a difference exists in their efficacy [11]. Anti-TNF-α/IL-23/IL-17A inhibitors successfully improve psoriatic arthritis [12]. Reductions in comorbid cardiovascular events and systemic inflammation have been reported in patients with psoriasis treated with anti-TNF/IL23/IL17 molecules.

Interleukin (IL)-1 and tumor necrosis factor (TNF) are the major proinflammatory cytokines involved, inflammation and tissue destruction, and also the inducers of endothelial adhesion molecules. Proinflammatory cytokine-mediated inflammation is a cascade of gene products usually not produced in healthy persons. IL-1, IL-6, IL-17, IL-12/23, and TNF are the major proinflammatory cytokines in psoriatic conditions, and hence, we are exclusively focused on inhibiting them using plant-derived small molecule inhibitors. The product under development for topical application is likely to contain the phyto-molecule which can significantly inhibit the pro-inflammatory cytokines IL-17, IL-12/23, and TNF and deliver the anti-psoriatic activity.

Our work on psoriasis towards attaining this polypharmacological activity, we employed computer aided drug discovery approach at Atrimed to screen the plant secondary metabolites as proinflammatory cytokine inhibitors and accelerated the classical route of drug discovery.
Drug target identification and validation Psoriasis associated protein-protein interaction (PPI) networks Target proteins of drug molecules are classified into a primary target and off-targets. The former is the desired target, whereas the latter could lead to adverse drug reactions or unexpected beneficial effects in drug repositioning. Therefore, comprehensive analysis throughout primary targets and off-targets on a genome- wide-scale is crucial in drug discovery. The in silico approach is expected to improve the research productivity in this field.
The protein-protein interactions were obtained from the STRING database and PPI networks were reconstructed around differentially expressed genes (DEGs). The visualization and topological analysis of the PPI network was performed via Cytoscape. Hub proteins were identified using the CytoHubba plugin which provides a user-friendly interface to analyze the topology of protein-protein interaction networks, such as human, yeast, fly, etc.
Apart from the identification of essential nodes, the betweenness and centrality score based subnetwork can be reconstructed utilizing CytoHubba in Cytoscape. These essential nodes in the reconstructed network may serve as candidates of drug-targets for developing novel therapy of human diseases.

Molecular Docking

As a part of the study, we have developed the plant secondary metabolites database which holds the 3D co-ordinate information of more than 4Lakh plant molecules. The the plant secondary metabolites in the database were screened against proinflammatory cytokine using molecular docking technique and the potential inhibitors are screened. The phytochemicals screened are possessing significant interaction with L-17, IL-12/23 and TNF with fairly acceptable bonded interactions. The screened molecules are isolated, purified and analysed experimentally to confirm the inhibitory potential of screened molecules.

Drug targets for topical application

References:

1.Boehncke WH, Schön MP. Psoriasis. Lancet. 2015 Sep 5; 386(9997):983-94.

2. Michalek IM, Loring B, John SM . A systematic review of worldwide epidemiology of psoriasis. J Eur Acad Dermatol Venereol. 2017 Feb; 31(2):205-212.

3. Ito T, Takahashi H, Kawada A, Iizuka H, Nakagawa H. Epidemiological survey from 2009 to 2012 of psoriatic patients in Japanese Society for Psoriasis Research. Japanese Society For Psoriasis Research. J Dermatol. 2018 Mar; 45(3):293-301.

4. Ichiyama S, Ito M, Funasaka Y, Abe M, Nishida E, Muramatsu S, Nishihara H, Kato H, Morita A, Imafuku S, Saeki H. Assessment of medication adherence and treatment satisfaction in Japanese patients with psoriasis of various severities. J Dermatol. 2018 Jun; 45(6):727-731.

5. Furue K, Ito T, Tsuji G, Kadono T, Nakahara T, Furue M. Autoimmunity and autoimmune co-morbidities in psoriasis. Immunology. 2018 May; 154(1):21-27.

6. Bayaraa B, Imafuku S. Relationship between environmental factors, age of onset and familial history in Japanese patients with psoriasis. J Dermatol. 2018 Jun; 45(6):715-718.

7. Chiu HY, Chang WL, Shiu MN, Huang WF, Tsai TF. Psoriasis is associated with a greater risk for cardiovascular procedure and surgery in patients with hypertension: A nationwide cohort study. J Dermatol. 2018 Dec; 45(12):1381-1388.

8. Furue M, Kadono T.  “Inflammatory skin march" in atopic dermatitis and psoriasis. Inflamm Res. 2017 Oct; 66(10):833-842.

9. Lee JH, Kim HJ, Han KD, Kim HN, Park YM, Lee JY, Park YG, Lee YB. Cancer risk in 892 089 patients with psoriasis in Korea: A nationwide population-based cohort study. J Dermatol. 2019 Feb; 46(2):95-102.

10. Imafuku S, Zheng M, Tada Y, Zhang X, Theng C, Thevarajah S, Zhao Y, Song HJ. Asian consensus on assessment and management of mild to moderate plaque psoriasis with topical therapy. J Dermatol. 2018 Jul; 45(7):805-811.

11. Masutaka Furue, Kazuhisa Furue, Gaku Tsuji, and Takeshi Nakahara. Interleukin-17A and Keratinocytes in Psoriasis. Int J Mol Sci. 2020 Feb; 21(4): 1275.

12. Yamamoto T, Ohtsuki M, Sano S, Morita A, Igarashi A, Okuyama R, Kawada A, working group of the epidemiological survey in the Japanese Society for Psoriasis Research. Switching biologics in the treatment of psoriatic arthritis in Japan. J Dermatol. 2019 Mar; 46(3):e113-e114.

Psoriasis is a chronic inflammatory disease mediated by immuno-regulatory dysfunc+on and other factors. CD4+T cells and their related cytokines play an important role in the pathogenesis of psoriasis, a chronic, recurrent, inflammatory skin disease. In this study, the expression of IL-35 in pa+ents with psoriasis was examined and analyzed its rela+onship with the other cytokines to assess its clinical significance in diagnosing or monitoring this disease. Protein and mRNA levels of specified markers were measured by ELISA and qRT-PCR, respec+vely. Plasma IL-35 concentra+ons were lower in pa+ents with psoriasis than in healthy individuals. mRNA +ters were lower in peripheral blood mononuclear cells from pa+ents with psoriasis than in those from healthy individuals. Pearson correla+on analysis showed that, in psoria+c pa+ents, plasma IL-35 expression correlated nega+vely with concentra+ons of INF-γ, tumor necrosis factor-alpha, IL-23, −17, and −22, and the Psoriasis Ac+vity and Severity Index and posi+vely with concentra+ons of transforming growth factor beta and IL-10. In summary, IL-35 may mediate pathogenesis of psoriasis probably by inhibi+ng expression of Th1/Th17-related cytokines and inducing expression of Treg-related cytokines, such as TGF-β and IL-10. Sta+s+cally significant associa+ons between low IL-35 concentra+ons and the cri+cal cytokines involved in the pathogenesis of psoriasis were iden+fied. IL-35 is a poten+al biomarker for monitoring or diagnosing psoriasis; this possibility should be inves+gated further.

Psoriasis is an inflammatory skin disease characterized by a significant elevation in the concentration of pro-inflammatory cytokines, including TNF-α. Chinese herbal medicine (CHM) formulation ‘psoriasis 1’, which comprises 13 Chinese herbs, has successfully treated a large number of patients, by improving skin lesions. Although, the underlying molecular mechanism of action unknown. Prior studies have demonstrated that the NF-κB and STAT signalling pathways are involved in the gene expression of certain cytokines during psoriasis. The effect of ‘psoriasis 1’ on these signaling pathways was studied and it was found that ‘psoriasis 1’ inhibits these pathways and thus lowers the levels of cytokines TNF-α, IFN-γ, IL-22, IL-17C, IL-1β, and IL-4. These studies were done both in human white blood cells as well as human skin cells. These results suggest that similar effects might be observed in animals, and, also in humans. Further experiments are currently underway, and if they are positive, then ‘psoriasis 1’ can be developed into a potent anti-psoriatic topical drug.

BMS-986165 is a small molecule developed at the Pharma company BMS for treating psoriasis. It inhibits cell signalling pathways implicated in the pathophysiology of psoriasis, specially the expression of the pro-inflammatory cytokine IL-23 which is known to be one of the reasons behind many of the symptoms of psoriasis. A phase-2, double-blind trial of BMS-986165, was conducted in adults with moderate-to-severe psoriasis. The selectivity of BMS-986165 allows for targeting the additional cytokine pathways of IL-12 and some interferons, all implicated in psoriasis. Patients were given the drug orally at a dose of 3 mg every other day, 3 mg daily, 3 mg twice daily, 6 mg twice daily, or 12 mg daily or the placebo. The primary end point was a 75% or greater reduction from baseline in the Psoriasis Area and Severity Index (PASI) score at week 12. Results indicated a therapeutic benefit of oral BMS-986165 which was comparable to ustekinumab (a monoclonal antibody that inhibits IL-23 and IL-12). The dose of 3 mg daily and higher resulted in greater clearing of psoriasis over a period of 12 weeks. Larger, longer-period trials of this drug are necessary to determine its safety and durability.

Wnt5a, a protein, is made in excess, in psoriasis lesions. It is believed to control the multiplication and death of human skin cells – keratinocytes. The levels of Wnt5a, were tested in psoriatic samples and also a healthy control skin samples. Cell cycle activity and the proliferation and apoptosis of skin cells were assessed. Results suggested that Wnt5a levels were highly increased in psoriatic lesions, suggesting that this leads to inflammatory responses, which are very important for the onset of psoriasis. Without Wnt5a, there is suppressed proliferation and induced apoptosis in skin cells and no onset of psoriasis. These findings conclude that knockdown of Wnt5a suppresses the proliferation of keratinocytes and induces apoptosis However, further studies are needed to elucidate the precise function of Wnt5a in psoriasis pathogenesis. This protein may however prove to be a strong target for the development of effective drugs against psoriasis.

Psoriasis is thought to be initiated by abnormal interactions, involving inflammatory molecules called cytokines, between keratinocytes (skin cells) and immune cells. However, the role of the cytokine IL-35 in psoriasis remains unclear. To determine this, levels of IL-35 in three well-known psoriasis models were studied. The models are: a human keratinocyte skin cell line (HaCaT), a transgenic (Tg) mouse model, and an imiquimod induced psoriasis mouse model. First, it was observed that IL-35 suppressed the expression of IL-6 and CXCL8 in HaCaT cells. Secondly , human IL-35 showed potent immunosuppressive effects on both mouse models. 

In the Tg model, results indicated that several types of pro-inflammatory cytokines were significantly reduced, whereas IL-10 (an anti-inflammatory cytokine) was remarkably induced by IL-35. Mainly, this study found that IL-35 decreased the total number of macrophages and ratio of different types of macrophages, which has not been reported previously. In addition, compared to standard steroids, IL-35 showed long-term therapeutic efficacy. These results strongly indicate that IL-35 has potential for development as a new therapeutic strategy for psoriasis.

IL-36 is abundantly expressed in psoriatic skin, where it interferes with keratinocyte cornification/maturation, thus preventing proper skin formation. It also induces the release of antimicrobial peptides and chemokines from white blood cells like neutrophils and Th17 lymphocytes leading to persistence of psoriatic symptoms. IL-38 present in psoriatic skin and circulating in blood are reduced in psoriatic patients and in other skin diseases similar to it. In psoriasis, the balance of IL-36/IL-38 serum levels is in favour of IL-36 and is closely associated with increased disease severity. Interestingly, IL-38 levels increase with anti-IL-17A antibody treatment and positively correlates with the therapeutic efficacy of secukinumab, an anti-IL-17A antibody, in psoriatic patients. The down-regulation of IL-38 expression is strictly related to worsening of psoriasis and increase in levels of inflammatory cytokines IL-36, IL-17, and IL-22. Finally, administration of recombinant full-length IL-38 counteracts the biological processes induced by IL-36 in human skin cells in mice with psoriasis. Thus IL-38 seems to have a protective and therapeutic role in psoriasis treatment. 

Wound Healing

Healing of wounds requires high levels of molecules called prostaglandins or PGs in our body, specially one parLcular PG called PGE2. But this molecule is very quickly degraded in our body by an enzyme called 15- PGDH. So, inhibiLon of this enzyme, will lead to beSer and faster wound healing. A large study with 98 plants was conducted to look for molecules that could inhibit 15-PGDH. 5 top inhibitors were chosen from an in silico study for tesLng in human skin cells (HaCaT cells). The human skin cells were wounded by scratching them and then they were treated with the plant molecules and levels of PGE2 were checked, as well as, the Lme taken for the scratch to heal. It was found that a molecule called EEAH, from the plant Artocarpus heterophillus, the common jackfruit, has very good wound healing ability. It causes very high levels of PGE2 to be maintained both inside and outside human skin cells to heal scratches very quickly. The molecule is safe, non-toxic and can be used in a topical formulaLon.

The objec+ve of a recent study (Yeh et al. 2017) was to es+mate the wound healing poten+al and therapeu+c mechanism of Artocarpin (ARTO), a prenylated flavonoid purified from the plant Artocarpus communis. Immunohistochemical staining of neutrophils and macrophages and mouse cytokine array analysis demonstrated that ARTO accelerates inflammatory progression and subsequently decreases persistent inflamma+on. ARTO increases collagen produc+on and increases human fibroblast prolifera+on and migra+on by ac+va+ng the P38 and JNK pathways. Moreover, ARTO increases the prolifera+on and migra+on of human kera+nocytes through the ERK and P38 pathways and augments human endothelial cell prolifera+on and tube forma+on through the Akt and P38 pathways. The data in the study suggests that ARTO enhances skin wound healing, possibly by accelera+ng the inflammatory phase and by increasing myofibroblast differen+a+on, prolifera+on, and migra+on of fibroblasts and kera+nocytes, collagen synthesis and matura+on, re-epithelializa+on, and angiogenesis. These findings indicate that ARTO has poten+al as a therapeu+c agent for the treatment of skin wounds.

β-sitosterol, a common steroid in many plants, can promote wound healing by promoting cell growth and activating multiple signalling pathways, including wound healing pathways. To find potent wound healing small molecules, β-sitosterol derivatives were designed and synthesised. Their potential to inhibit Na+/K+-ATPase, an enzyme that in turn inhibits many cell signalling pathways, was investigated. Among them, 3 compounds showed improved inhibitory activity on Na+/K+- ATPase. It was found that by inhibiting Na+/K+-ATPase, these molecules induce cell proliferation, migration and soluble collagen production in damaged human tissue. These compounds were also shown to accelerate wound healing in wounded rats. Further studies indicated that these compounds can activate a molecule called extracellular signal-regulated kinase (ERK) protein, which is a master-switch that can turn on many cell signalling pathways, including the ones involved in wound healing. These molecules were also found to be very highly potent and selective, meaning, their side-effects will be less. These derivatives of βsitosterol can thus be potential candidates for wound healing agents

Periploca forrestii (PF) is a herb used in traditional folk medicine for treating traumatic injuries and wound healing. To study how it brings about these effects, the total extract of PF and its fractions were evaluated in in vitro wound healing assays using mouse cells. Cell proliferation and migration, two important events in healing, were evaluated. Collagen production was also determined. Wound healing in rats treated with PF extracts was also investigated with the wounded skin being examined on day 6 and day 12 post treatment. A few key cell signalling events involved in wound healing were also studied. Results suggest PF and its extracts can promote cell proliferation, migration and increase collagen production. A PF hydrogel also promotes wound healing in rats starting day 3 post treatment already. The extracts hasten healing, leading to skin that looks almost like normal sooner. The extracts also promote cell signalling involved in cell proliferation and migration. All of these results demonstrate that PF extracts promote wound healing by promoting beneficial cell signalling and should be studied further.

The wound healing properties of the gel and whole-leaf extract of Aloe vera, Aloe ferox and Aloe marlothii, are well known. Nuclear magnetic resonance (NMR) spectroscopy was used to identify all the molecules in them. A cell viability study was performed to determine their effect on human skin cells and found to be completely safe. The gel and extract were then tested in an in vitro wound healing assay on skin cells to see if they help heal wounds faster. After 48 hours, all the aloe gel and extracts caused almost complete wound closure, displaying 98.07% (A. marlothii whole-leaf), 98% (A. vera gel), 97.2% (A. marlothii gel), 96% (A. vera whole-leaf), 94% (A. ferox gel) and 81.3% (A. ferox whole-leaf) wound closure, respectively. The gel materials of all the three aloe species exhibited significantly faster wound healing actions when compared to their respective whole-leaf extracts at 32 hours. So, gels and topicals with these extracts can be safely used to expedite wound healing.

β-Sitosterol, is a common steroid that can be identified in a variety of plants and its ability to promote wound healing is well known. Both sodium and potassium ions (found in common salt) are involved in transmission of signals from cell to cell with instructions for all body functions, including wound healing. There are enzymes in our body that can block the functioning of these ions which can lead to improper wound healing. Thus, in order to promote proper and quick wound healing, such enzymes must be blocked. β-sitosterol and its derivatives are potent inhibitors of such enzymes. Among the derivatives, 3 compounds showed best inhibition. One of the 3 compounds induced cell proliferation, migration and soluble collagen production in human cells as well as injured rats, thus promoting wound healing. The compound directly binds and blocks the enzymes thereby, leading to good wound healing. These results prove β-sitosterol and its derivatives can serve as a potential candidate for accelerating wound healing.

The extract of Boswellia serrata or Sallaki has wound healing potential for diabetic foot ulcer and is used in traditional medicine and Ayurveda for this purpose. α-Boswellic acid isolated from the plant was tested in diabetic rats with foot ulcers. The treatment was administered to the rats for 16 days (400 mg/kg) leading to significantly increased rate of wound contraction and healing at the wound area. RT-PCR analysis revealed that α-Boswellic acid can inhibit TNF-α, IL-1β, IL-6 and enhance levels of TGF-β1, VEGF, and collagen-1 which aid in healing. In conclusion, standardised extracts of B. serrata exert its wound healing potential via orchestrating mechanisms, which include the inhibition of inflammatory cytokines and enhancing growth factors, collagen synthesis and angiogenesis, to accelerate wound healing in diabetic foot ulcer in rats. Further testing may prove to be useful in treating the same condition in humans.

Acne

Commensal microorganisms in the skin microbiota like Staphylococcus epidermidis (S. epidermidis) are known to fight pathogens like Propionibacterium acnes (P. acnes), associated with progression of acne vulgaris. It is known that many skin microorganisms can mediate fermenta7on of glycerol, which is produced in the skin. A recent study of microbial interac7ons in skin has revealed new details on how S. epidermidis contributes to controlling the growth of P. acnes. S. epidermidis ferments glycerol to produce succinic acid, one of four short chain faOy acids produced during fermenta7on. Succinic acid can effec7vely inhibit the growth and inflamma7on caused by P. Acnes, both in vitro as well as in vivo, in mice treated either with intra-lesional injec7ons or topical applica7on over the lesions. With recent evidence of fermenta7on based control of P. acnes by S. epidermidis, the role of poten7al new probio7cs against acne and other skin diseases.

The bacteria that cause acne live mainly on the skin, but, under the right condiLons, they can enter the skin and cause serious infecLons inside our body as well. Acne bacteria have the property to sLck to surfaces, like orthopedic implants at shoulder, knee and hip joints, cerebrovascular shunts, pacemakers and breast implants, causing serious infecLons because of their ability to form biofilms around the metal surfaces of the implants. Their increasing resistance to the anLbioLc clindamycin normally used for treatment, makes the infecLon hard to treat quickly, thus requiring surgery as well as 3-6 months of anLbioLc treatment to completely eliminate the infecLon. Recently, many studies have been done with bacterial cultures isolated from such infected implants to idenLfy the exact strain of acne bacteria and accordingly treat it with anLbioLcs like rifampicin to which it is highly suscepLble. This has decreased the treatment Lme and side-effects. However, more studies are required to prove this.

Propionibacterium acnes, recently reclassified as Cu4bacterium acnes, is the primary resident of the pilosebaceous follicles, but contrary to previous beliefs, an increase in its prolifera+on is not the cause of acne vulgaris. Acne is caused more by a subset of C. acnes strains like IA1 which are acne-associated phylotypes, aided by a hyperseborrheic environment. Also, these strains have varied poten+al for biofilm forma+on, ini+a+ng inflamma+on, and for virulence. These strains are also more resistant to regula+on by S. epidermidis which can inhibit both their growth as well as their inflammatory poten+al. Small molecules, such as levulinic acid, are able to inhibit porphyrin biosynthesis in these strains without disrup+ng the growth of health-associated strains and are thus aWrac+ve drug candidates for the treatment of acne. Using an+-biofilm compounds like myrtucommulones, can help to deconstruct acne biofilms and restore an+bio+c sensi+vity even in resistant strains. Individualized acne therapies that supplement the skin microbiota with probio+cs seem to be the future of acne therapy.

Acne vulgaris, or simply, acne, is the most common skin disease and affects more than 85% of teenagers. While moderate acne is usually treated with topical antibacterials, severe acne often requires oral antibiotic treatment. Some of the most commonly used antibiotics are doxycycline, minocycline, azithromycin, clindamycin, etc. either alone or in combination and sometimes, in combination with oral contraceptives, especially in females. In recent times, however, there is increased concern with antibiotic resistance to long-used antibiotics like clindamycin and erythromycin prompting the use of first and second-generation tetracyclines like doxycycline which can have considerable side-effects like photosensitivity, bone and tooth impairment, esophagitis (ulcers in the throat) etc. Minocycline and Sarecycline were developed to avoid the side-effects of doxycycline but they have their own problems with vertigo and hyperpigmentation. Azithromycin, a macrolide antibiotic, is much safer, even for pregnant women, but needs to be taken for much longer durations for effectiveness and as such is only a second-line option for acne. There is thus an imminent need for safe and effective antibiotics, with minimal side-effects for acne treatment.

Passion fruit is used commercially for consumption and in beverages. This plant exhibits various pharmacological properties and possesses a complex phytochemistry. In recent years, this plant has been shown to have potential antimicrobial activity. The seeds contain a high amount of piceatannol, which exhibits an inhibitory effect on Propionibacterium acnes. Seed extract was prepared by maceration and the agar disc diffusion method was used to evaluate the inhibitory effect, in comparison with clindamycin and erythromycin. After 24 hours, all concentrations of the extract showed an inhibitory effect against P. acnes, which was comparable to Clindamycin, or even better, at some concentrations. The extract is also safe as determined by an MIC test. We can conclude that passion fruit seed extract may be used in formulations to control acne

The bacteria that cause acne, also have an effect on other conditions like sarcoidosis and ulcerative colitis. Acne bacteria occur not only in skin but also the bowel, eye and mouth, and in these other places, it is suspected, acne bacteria can trigger both sarcoidosis (eye, skin granulomas) and ulcerative colitis (bloody bowel). This is supported also by the fact that acne bacteria occur abundantly also in people who dont suffer from acne. After many studies, it has been found that, all 3 diseases have very similar immune dysregulation, meaning, the same components of the immune system go wrong, in all these disease conditions. Lung and cardiac tissue from sarcoidosis patients have been found to have acne bacteria, specifically the C1 and ST26 strains. A PCR-based diagnostic test for sarcoidosis is being developed based on testing for presence of acne bacterial RNA. Similarly, it has been found that an increase in acne bacteria in the gut microbiome, can lead to ulcerative colitis. A fair conclusion can be made that there is some definitive link between acne bacteria and these disease conditions.

Acne causing bacteria are a key pathogen involved in the development and progression of acne and associated inflammation. The leaves of wild bitter melon have many medicinal applications in traditional medication, including acne control. A methanolic extract of the leaves was prepared and processed further to obtain various fractions with different bioactive molecules. These were tested on live acne-induced inflammation in human white blood cells and also in mice. The hexane fraction exerted the most potent anti-inflammatory effect, suppressing acne-induced interleukin-8 (IL-8) production by 36% in both white blood cells and mice. The ethanol fraction also exhibited IL-8 inhibition by 20% in mice. 24 bioactive compounds in the ethanol fraction were identified, specially β-ionone, which reduced the production of IL-1β and IL-8 up to 40% in white blood cells as well as mouse ear. Thus, β-ionone is a potential anti-inflammatory agent for modulating in vitro and in vivo acne-induced inflammatory responses.

Acne bacteria play an important role in the pathogenesis and progression of the skin disorder acne. The methanolic extract of Helichrysum or curry plant, was investigated for its ability to target and inhibit bacterial growth and its virulence factors associated with acne progression. The extract exhibited potent antimicrobial activity against C. acnes (ATCC 6919) at very low doses. The extract showed high specificity against C. acnes cell aggregation, thus preventing biofilm formation, a crucial virulence factor in persistence of acne. Mature C. acnes biofilms were also disrupted by the extract, which also reduced IL-1α, IL-8 cytokine levels in C. acnes-induced human keratinocytes (HaCaT). The extract finally inhibited hyaluronidase enzyme activity, which is responsible for hyaluronan degradation and scar formation at the site of acne. This study provides scientific validation for the traditional use of curry plant as an ointment for pimples, not only due to its ability to control C. acnes proliferation but also due to its inhibitory activity on various targets associated with bacterial virulence leading to acne progression.

Pain Management

Burn pain can be severe and even cause death. The Magnolia tree, a Chinese flowering plant, produces a molecule called Honokiol, which has pain relieving properLes. To test these, mice were inflicted with burn wounds and then treated with a honokiol extract preparaLon for 3 days along with standard opiod pain medicaLon. It was found that the combinaLon of honokiol and opiod is very effecLve in reducing the pain sensaLon, heat in the wound area and oedema/swelling at the wound area. Honokiol also succeeded in reducing weight loss and spontaneous pain behaviour in the mice. It also significantly balanced blood electrolytes and general profile. Honokiol has these effects by significantly decreasing the expression of molecules called cytokines which cause inflammaLon, a process that can cause pain to persist. Honokiol treatment also maintains anL-oxidant levels in blood which aid in reducing inflammaLon and promoLng pain management. It can be concluded on the base of the results that honokiol has a significant analgesic/pain relieving acLvity through its acLon on cytokines. It also has a protecLve role against burn damage by promoLng levels of anLoxidants.

The plant Swertia chirata (Chirayata) is commonly used for the treatment of liver infections, inflammation, abdominal pain, and bacterial infection in Ayurveda. However, the bioactive components present in this plant with anti-inflammatory activities are fairly unknown. A recent study yielded two main xanthones from whole-plant extracts of Swertia, called, Bellidifolin and Swerchirin. Their anti-inflammatory and pain-relief properties were determined in mouse cells in vitro. The assays demonstrated that both compounds inhibit the production of the pro-inflammatory cytokines IL-6 and TNF-α in mouse cells, while Bellidifolin also inhibits the production of prostaglandin E2 (PGE2) by suppressing the protein expression of COX-2 in mice cells, thereby leading to pain relief. The potent suppressive effects of this molecule suggest its potential use as a preventive therapeutic candidate for the management of pain relief in humans, upon further testing and trials.

The Centers for Disease Control and Prevention have called for a drastic cut in opioid prescription and use for chronic pain conditions, and thus, it is imperative to discover alternative non-opioid based strategies. Towards this goal, a library of natural products was screened in search of pharmacological inhibitors of voltage-gated ion channels. These channels, found in nerve cells, play a big role in pain sensation. The screening led to the discovery of molecules isolated from plants, Croton californicus and Eremocarpus setigerus, that inhibit such ion channels in neurons, specifically sodium channels. The molecules, called HDA and HTA, could reverse chronic pain behaviour in rats with chronic pain. The results illustrate the therapeutic potential for HDA and HTA for chronic pain management and could represent an option that may yield novel specific sodium channel antagonists for pain relief

Tendinopathy is the degeneration of tendons, leading to painful joints. Tendon cells produce proinflammatory molecules that induce pain and tissue deterioration. Presently used nonsteroidal anti-inflammatory drugs or ‘pain-killers’ are soothing but have side-effects, thus the need for better pain management. A combination of avocado-soybean extracts, rich in various plant metabolites, might help manage such pain. Tenocytes (tendon cells) were first induced into stress and then treated with avocado-soybean extract. The cells were then studied under the microscope and further analysed to reveal that pain inducing protein COX-1 expression was significantly reduced by the treatment. This suggests that the combination may help attenuate deleterious inflammation and reduce pain in tendons.

Common cancer medicines like oxaliplatin often cause chronic neuropathic pain during treatment. Such pain is difficult to treat and responds poorly to common painkillers, which represents a clinical challenge. (+)-Borneol, a molecule present in the essential oil of plants, is used for analgesia and anesthesia in traditional Chinese medicine. Although borneol can reduce acute pain models, little is known about its effect on chronic pain and how it relieves pain. A recent study found that (+)-borneol cured pain induced by cancer medicine oxaliplatin in a mouse model. In addition, (+)-borneol blocked the action of pain transmitters called ankyrin 1 agonist in further tests on those mice showing exactly how it works to relieve pain. Repeated treatment with (+)-borneol did not lead to tolerance or resistance to the medication and did not affect body weight and movement of the mice so it is safe. (+)-Borneol may thus be a useful analgesic in the management of neuropathic pain.

Several species of the bush Buddleja (lilac) have been used in the management of various health conditions including pain and inflammation.. Male rats and mice, with artificially induced pain, were randomly divided into different groups and treated with various doses of plant extract of Buddleja. The analgesic/pain-relief and anti-inflammatory activities were assessed in the animals post treatment. The methanolic and hexane extracts of Buddleja exhibited significant inhibition of pain in mice and attenuated reaction time of the animals. Pure compounds BdI-2, BdI-H3 and BH-3 isolated from Buddleja produced significant analgesic activity in in rats, upto 70%. The analgesic and anti-inflammatory effect of the plant extract and isolated pure compounds were comparable to diclofenac sodium. The findings from this study suggest that Buddleja possesses analgesic/pain-relief and anti-inflammatory properties and thus it could serve as a potential novel source of compounds effective for treatment of pain and inflammatory conditions.