Drug Design and Discovery: Translational Biomedical Science Varies Among Countries

Drug design and discovery is an innovation process that translates the outcomes of fundamental biomedical research into therapeutics that are ultimately made available to people with medical disorders in many countries throughout the world. To identify which nations succeed, exceed, or fail at the drug design/discovery endeavor—more specifically, which countries, within the context of their national size and wealth, are “pulling their weight” when it comes to developing medications targeting the myriad of diseases that afflict humankind—we compiled and analyzed a comprehensive survey of all new drugs (small molecular entities and biologics) approved annually throughout the world over the 20‐year period from 1991 to 2010. Based upon this analysis, we have devised prediction algorithms to ascertain which countries are successful (or not) in contributing to the worldwide need for effective new therapeutics.     

Five cardiac hormones decrease the number of human small-cell lung cancer cells

BACKGROUND: Four peptide hormones of a family of six hormones, i.e. atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), C-natriuretic peptide (CNP), long acting natriuretic peptide (LANP), vessel dilator and kaliuretic peptide, significantly decrease the number of adenocarcinoma cells in culture. The present investigation was designed to determine whether these peptide hormones' effects are specific to adenocarcinomas or whether they might decrease the number of cancer cells of a different type of cancer, i.e. small-cell lung cancer. METHODS AND MATERIALS: These six hormones were evaluated for their ability to decrease the number and/or proliferation of human small-cell lung cancer cells in culture for 24, 48, 72, and 96 h. RESULTS: Within 24 h, vessel dilator, LANP, kaliuretic peptide, ANP and their intracellular mediator cyclic GMP, each at 1 microM, decreased the number of small-cell lung cancer cells by 63% (P < 0.001), 21% (P < 0.05), 30% (P < 0.05), 39% (P < 0.05), and 31% (P < 0.05), respectively. There was no proliferation in the 3 days following this decrease in cell number. These same hormones decreased DNA synthesis 68% to 82% (P < 0.001). Brain natriuretic peptide and CNP did not decrease the number of small-cell lung cancer cells or inhibit their DNA synthesis at 1 microM or 10 microM concentrations. Dose-response curves revealed that at 100 microM, the vessel dilator decreased 92% of the cancer cells in 24 h while BNP had no effect, but CNP caused a 39% decrease. Western blots revealed that the natriuretic peptide receptors A- and C- were present in these cancer cells. CONCLUSIONS: Five peptide hormones significantly decrease the number of human small-cell lung cancer cells within 24 h and inhibit their proliferation for at least 96 h. Their mechanism of doing so involves inhibition of DNA synthesis mediated in part by cyclic GMP.     

Novel therapeutic approach for cancer using four cardiovascular hormones

BACKGROUND: The atrial natriuretic peptide (ANP) gene synthesizes four cardiovascular hormones, i.e. vessel dilator, long-acting natriuretic peptide, kaliuretic peptide and ANP, which decrease the number of human pancreatic adenocarcinoma cells in culture by 65%, 47%, 37%, and 34%, respectively. METHODS AND MATERIALS: None of the cardiovascular hormones has been investigated to determine whether they inhibit the growth of cancers in vivo. These four hormones were evaluated for their ability to inhibit the growth of human pancreatic adenocarcinomas in athymic mice. RESULTS: Vessel dilator (139 ng min(-1) kg(-1) of body weight) infused for 14 days completely stopped the growth of human pancreatic adenocarcinomas in athymic mice (n = 14) with a decrease in their tumour volume, while the tumour volume increased 69-fold (P < 0.001) in the placebo (n = 30)-treated mice. When these peptide hormones (each at 1.4 microg min(-1) kg(-1) body weight) were infused for 4 weeks, vessel dilator, long-acting natriuretic peptide and kaliuretic peptide decreased tumour volume after 1 week by 49%, 28%, and 11%, respectively, with a one- and 20-fold increase in the tumour volume in ANP- and placebo-treated mice. Cyclic GMP (2.4 microg min(-1) kg(-1) body weight) inhibited after 1 week the growth of this cancer 95%. CONCLUSIONS: These results suggest that these peptide hormones have useful anticancer properties, as they each inhibited the growth of the human pancreatic adenocarcinomas in vivo and three of the four peptide hormones decreased the volume of the tumours (up to 49%, i.e. vessel dilator). Part of their mechanism of action appears to be mediated by cyclic GMP.     

G6PI检测在类风湿关节炎诊断中的价值

目的：探讨葡萄糖-6-磷酸异构酶（G6PI）在类风湿性关节炎（RA）诊断中的价值。方法统计分析2011年8月至2013年8月该院共收治的130例 RA 患者的临床资料。同时,选取85例非 RA 的其他风湿病患者作为非 RA 组,60例健康者作为对照组。结果RA 组患者的 G6PI、抗环瓜氨酸肽（CCP）、类风湿因子（RF）水平均比非 RA 组和对照组高,差异有统计学意义（P ＜0．05）;非 RA 组患者 RF 水平比对照组高,差异有统计学意义（P ＜0．05）;RA 组患者 G6PI、抗 CCP 抗体、RF 检测的阳性率分别为67．70％（88／130）、52．30％（68／130）、75．38％（98／130）,均高于非 RA 组和对照组相应检测指标的阳性率,差异有统计学意义（P ＜0．05）;非 RA 组患者 RF 水平比对照组高,差异有统计学意义（P ＜0．05）;G6PI 检测用于 RA 诊断的灵敏度、阴性预测值和约登指数均比抗 CCP 抗体和 RF 高,差异有统计学意义（P ＜0．05）。RA 患者的 G6PI 和抗 RF 水平呈正相关（r＝0．732,P ＜0．05）。结论G6PI 检测在类风湿关节炎的诊断中具有较高的价值。     

抗环瓜氨酸肽抗体对类风湿性关节炎发病的预测价值

目的　探讨抗环瓜氨酸肽 (CCP)抗体对类风湿性关节炎 (RA)发病的预测价值。方法　对 14 6例新发病的未分化关节炎 (UA)患者随访 3年 ,观察RA发病率 ,检测血清抗CCP抗体和类风湿因子 (RF)。结果　 39%的UA患者(5 7/ 14 6 ) 3年中发展为RA ,其中包括 2 3 7%的抗CCP抗体阴性患者和 93 8%抗CCP抗体的阳性患者 (比数比OR为4 8 3,95 %可信区间为 16 3～ 137 6 ,P <0 0 1)。抗CCP抗体与RF具有相关性。结论　抗CCP抗体的出现是UA患者发生RA的高度危险因素 ,对预测RA的发生具有重要价值。     

Four peptide hormones' specific decrease (up to 97%) of human prostate carcinoma cells

BACKGROUND: Mortality from prostate cancer remains a significant problem with current treatment(s), with an expected 30 350 deaths from prostate cancer in 2005. Long-acting natriuretic peptide, vessel dilator, kaliuretic peptide and atrial natriuretic peptide have significant anticancer effects in breast and pancreatic adenocarcinomas. Whether these effects are specific and whether they have anticancer effects in prostate adenocarcinoma cells has not been determined. MATERIALS AND METHODS: These peptide hormones were evaluated to determine if they have specific anticancer effects in human prostate adenocarcinomas. RESULTS: Dose-response curves revealed a significant (P < 0.05) decrease in human prostate cancer number with each tenfold increase in the concentration from 1 microM to 1000 microM (i.e. 1 mM) of these four peptide hormones. There was a 97.4%, 87%, 88% and 89% (P < 0.001 for each) decrease in prostate cancer cells secondary to vessel dilator, long-acting natriuretic peptide, kaliuretic peptide and atrial natriuretic peptide, respectively, at their 1-mM concentrations within 24 h, without any proliferation in the 3 days following this decrease. These same hormones decreased DNA synthesis from 68% to 89% (P < 0.001). When utilized with their respective antibodies their ability to decrease prostate adenocarcinoma cells or inhibit their DNA synthesis was completely blocked. Western blots revealed that for the first time natriuretic peptide receptors (NPR) A- and C- were present in prostate cancer cells. CONCLUSIONS: These results indicate that these peptide hormones' anticancer effects are specific. Furthermore, they have very potent effects of eliminating up to 97% of prostate cancer cells within 24 h of treatment.     

Urodilatin and four cardiac hormones decrease human renal carcinoma cell numbers

BACKGROUND: Mortality from renal-cell cancer remains a significant problem with an estimated 12,600 deaths in the United States in 2005 even with current treatment(s) of surgery, chemotherapy, radiation and immunotherapy. Four cardiac natriuretic peptides, that is, atrial natriuretic peptide, vessel dilator, long-acting natriuretic peptide and kaliuretic peptide have significant anti-cancer effects in breast, pancreatic, prostate and colon adenocarcinomas. MATERIALS AND METHODS: These four peptide hormones plus brain natriuretic peptide (BNP), C-natriuretic peptide (CNP) and urodilatin, a peptide hormone formed in the kidney by a different post-translational processing of the atrial natriuretic peptide prohormone, were evaluated for their anti-cancer effects in renal carcinomas. RESULTS: Dose-response curves revealed a significant (P < 0.0001) decrease in human renal carcinoma cells with each 10-fold increase in concentration from 1 microm to 100 microm of five of these peptide hormones. There was an 81%, 74%, 66%, 70% and 70% elimination within 24 h in renal carcinoma cells secondary to vessel dilator, kaliuretic peptide, urodilatin, atrial natriuretic peptide and long-acting natriuretic peptide, respectively (P < 0.0001 for each), whereas BNP had no effect and CNP decreased renal cancer cell number by 10% (P = 0.04) at their 100 microm concentrations. Three days after treatment with these peptide hormones, the cancer cells began to proliferate again. The four cardiac hormones and urodilatin decreased DNA synthesis from 65-84% (P < 0.00001), whereas BNP and CNP decreased DNA synthesis 3% and 12% (both non-significant). Western blots revealed for the first time natriuretic peptide receptors (NPR)-A, -B and -C were present in the renal cancer cells. CONCLUSIONS: These results indicate that urodilatin and the four cardiac hormones have potent anti-cancer effects by eliminating up to 81% of renal carcinoma cells within 24 h of treatment.     

Binding interactions between peptides and proteins of the class II major histocompatibility complex

The activation of helper T cells by peptides bound to proteins of the class II Major Histocompatibility Complex (MHC II) is pivotal to the initiation of an immune response. The primary functional requirement imposed on MHC II proteins is the ability to efficiently bind thousands of different peptides. Structurally, this is reflected in a unique architecture of binding interactions. The peptide is bound in an extended conformation within a groove on the membrane distal surface of the protein that is lined with several pockets that can accommodate peptide side-chains. Conserved MHC II protein residues also form hydrogen bonds along the length of the peptide main-chain. Here we review recent advances in the study of peptide-MHC II protein reactions that have led to an enhanced understanding of binding energetics. These results demonstrate that peptide-MHC II protein complexes achieve high affinity binding from the array of hydrogen bonds that are energetically segregated from the pocket interactions, which can then add to an intrinsic hydrogen bond-mediated affinity. Thus, MHC II proteins are unlike antibodies, which utilize cooperativity among binding interactions to achieve high affinity and specificity. The significance of these observations is discussed within the context of possible mechanisms for the HLA-DM protein that regulates peptide presentation in vivo and the design of non-peptide molecules that can bind MHC II proteins and act as vaccines or immune modulators.     

Four peptide hormones decrease the number of human breast adenocarcinoma cells

BACKGROUND: A family of six hormones, i.e. atrial natriuretic peptide, brain natriuretic peptide, C-natriuretic peptide, long-acting natriuretic peptide, vessel dilator, and kaliuretic peptide's main known biologic properties are sodium and water excreting and blood pressure lowering. METHODS AND MATERIALS: These six hormones, each at their 1-microm concentrations, were evaluated for their ability to decrease the number and/or proliferation of breast adenocarcinoma cells in culture for 24, 48, 72, and 96 h. RESULTS: Within 24 h, vessel dilator, long-acting natriuretic peptide, kaliuretic peptide, atrial natriuretic peptide and 8-bromo-cyclic GMP, a cell-permeable analogue of their intracellular mediator cyclic GMP (each at 1 microm), decreased the number of breast adenocarcinoma cells 60%, 31%, 27%, 40%, and 31%, respectively. There was no proliferation in the 3 days following this decrease in breast adenocarcinoma cell number. These same hormones decreased DNA synthesis 69% to 85% (P < 0.001). Brain natriuretic peptide and CNP did not decrease the number of breast adenocarcinoma cells or inhibit their DNA synthesis. Vessel dilator, long-acting natriuretic peptide, kaliuretic peptide and 8-bromo-cyclic GMP (each at 1 microM) decreased the number of cells in the S phase of the cell cycle by 62%, 33%, 50%, and 39%, respectively (all P < 0.05). Natriuretic peptide receptors-A and -C were present in the breast adenocarcinoma cells. CONCLUSIONS: Four peptide hormones significantly decrease the number of human breast adenocarcinoma cells within 24 h and inhibit the proliferation of these cells for at least 96 h. Their mechanism of doing so involves inhibition of DNA synthesis and a decrease in cells in the S phase of the cell cycle mediated in part by cyclic GMP.     

Antimicrobial peptides: therapeutic potentials

The increasing appearance of multidrug-resistant pathogens has created an urgent need for suitable alternatives to current antibiotics. Antimicrobial peptides (AMPs), which act as defensive weapons against microbes, have received great attention because of broad-spectrum activities, unique action mechanisms and rare antibiotic-resistant variants. Despite desirable characteristics, they have shown limitations in pharmaceutical development due to toxicity, stability and manufacturing costs. Because of these drawbacks, only a few AMPs have been tested in Phase III clinical trials and no AMPs have been approved by the US FDA yet. However, these obstacles could be overcome by well-known methods such as changing physicochemical characteristics and introducing nonnatural amino acids, acetylation or amidation, as well as modern techniques like molecular targeted AMPs, liposomal formulations and drug delivery systems. Thus, the current challenge in this field is to develop therapeutic AMPs at a reasonable cost as well as to overcome the limitations.     

Alteration of a Single Hydrogen Bond between Class II Molecules and Peptide Results in Rapid Degradation of Class II Molecules after Invariant Chain Removal

To characterize the importance of a highly conserved region of the class II β chain, we introduced an amino acid substitution that is predicted to eliminate a hydrogen bond formed between the class II molecule and peptide. We expressed the mutated β chain with a wild-type α chain in a murine L cell by gene transfection. The mutant class II molecule (81βH⁻) assembles normally in the endoplasmic reticulum and transits the Golgi complex. When invariant chain (Ii) is coexpressed with 81βH⁻, the class II–Ii complex is degraded in the endosomes. Expression of 81βH⁻ in the absence of Ii results in a cell surface expressed molecule that is susceptible to proteolysis, a condition reversed by incubation with a peptide known to associate with 81βH⁻. We propose that 81βH⁻ is protease sensitive because it is unable to productively associate with most peptides, including classII–associated invariant chain peptides. This model is supported by our data demonstrating protease sensitivity of peptide-free wild-type I-A^d molecules. Collectively, our results suggest both that the hydrogen bonds formed between the class II molecule and peptide are important for the integrity and stability of the complex, and that empty class II molecules are protease sensitive and degraded in endosomes. One function of DM may be to insure continuous groove occupancy of the class II molecule.     

Recent advances in the development of a chemically synthesised anti-malarial vaccine

Introduction: Obtaining an effective antimalarial vaccine has represented one of the biggest public health challenges over the last 50 years. Despite efforts by many laboratories around the world using whole-organism, recombinant proteins and genome-based approaches, the results have been disappointing. One of the main problems when designing an antimalarial vaccine is the poor immunogenicity induced by the functionally relevant and conserved protein regions of the parasite.

Areas covered: This review focuses on the logical and rational methodology followed to identify Plasmodium falciparum conserved functional regions with the ability to bind to target cells conserved high activity binding peptides (cHABPs) and the physicochemical and immunological characteristics that should be taken into account for modifying them into highly immunogenic and protection-inducing peptides (mHABPs) into highly immunogenic and protection-inducing in Aotus monkeys.

Expert opinion: The functional approach taken to develop a fully protective, minimal subunit-based, multiantigenic, multistage and synthetic peptide-based antimalarial vaccine has shown promising results. The clear relationship observed between mHABPs structure and their immunological properties highlights the challenges and opportunities arising from this methodology, as well as the universal principles and rules derived therefrom.     

Orphan G protein‐coupled receptors: targets for new therapeutic interventions

With the completion of the human genome, many genes will be uncovered with unknown functions. The ‘orphan’ G protein coupled receptors (GPCRs) are examples of genes without known functions. These are genes that exhibit the seven helical conformation hallmark of the GPCRs but that are called ‘orphans’ because they are activated by none of the primary messengers known to activate GPCRs in vivo. They are the targets of undiscovered transmitters and this lack of knowledge precludes understanding their function. Yet, because they belong to the supergene family that has the widest regulatory role in the organism, the orphan GPCRs have generated much excitement in academia and industry. They hold much hope for revealing new intercellular interactions that will open new areas of basic research which ultimately will lead to new therapeutic applications. However, the first step in understanding the function of orphan GPCRs is to ‘deorphanize’ them, to identify their natural transmitters. Here we review the search for the natural primary messengers of orphan GPCRs and focus on two recently deorphanized GPCR systems, the melanin‐concentrating hormone (MCH) and prolactin‐releasing peptide (PrRP) systems, to illustrate the strategies applied to solve their function and to exemplify the therapeutic potentials that such systems hold.     

Venoms as a platform for human drugs: translating toxins into therapeutics

Introduction: An extraordinarily diverse range of animals have evolved venoms for predation, defence, or competitor deterrence. The major components of most venoms are peptides and proteins that are often protease-resistant due to their disulfide-rich architectures. Some of these toxins have become valuable as pharmacological tools and/or therapeutics due to their extremely high specificity and potency for particular molecular targets. There are currently six FDA-approved drugs derived from venom peptides or proteins.

Areas covered: This article surveys the current pipeline of venom-derived therapeutics and critically examines the potential of peptide and protein drugs derived from venoms. Emerging trends are identified, including an increasing industry focus on disulfide-rich venom peptides and the use of a broader array of molecular targets in order to develop venom-based therapeutics for treating a wider range of clinical conditions.

Expert opinion: Key technical advances in combination with a renewed industry-wide focus on biologics have converged to provide a larger than ever pipeline of venom-derived therapeutics. Disulfide-rich venom peptides obviate some of the traditional disadvantages of therapeutic peptides and some may be suitable for oral administration. Moreover, some venom peptides can breach the blood brain barrier and translocate across cell membranes, which opens up the possibility of exploiting molecular targets not previously accessible to peptide drugs.     

Synergy among antibacterial peptides and between peptides and small-molecule antibiotics

Antimicrobial peptides are key components of native immunity. Unlike most common small-molecule antibiotics, antimicrobial peptides have been refined by evolution to work synergistically within the host environment. The mechanisms of synergy are complex and specific, and it is only now that they are beginning to be understood. Thus, antimicrobial peptides are top candidates to broaden our limited therapeutic arsenal, and are very well suited to be used in synergic combinations with currently available antibiotics. In order for this promising strategy to become a reality, however, some key steps in basic and translational research need to be improved. These include the standardization and critical evaluation of testing and quantification methods, the characterization of the molecular mechanism of action, the study of indirect antibacterial activity such as immune-response modulation, and several other aspects that will be presented and discussed, with a focus primarily on antibacterial therapy. There will be a special focus on advances and innovations that might significantly improve the future perspectives of antimicrobial peptides from a therapeutic point-of-view.     

A role for ring‐closing metathesis in medicinal chemistry: Mimicking secondary architectures in bioactive peptides

Synthetically versatile and easy to carry out, Ring‐Closing Metathesis (RCM) constitutes an attractive chemical tool, easily amenable for multiple substrates in mild conditions. In medicinal chemistry, the use of RCM has been especially prolific during the last few years. An important application that has benefited from this reaction is the stabilization of spatial conformations in bioactive peptides, since their 3D arrangements play relevant roles in biomolecular recognition processes. RCM reaction is being widely used to introduce conformational constraints into small peptides, through the generation of cyclic structures from appropriate linear precursors. As an alternative to strategies like disulfide or lactam‐bridged cyclizations, RCM shows the additional advantage of generating hydrocarbon bridges, less prone to metabolic degradation, and metabolically more stable, which could benefit their pharmacokinetic properties. Particularly remarkable is the application of RCM to the preparation of small peptide modulators able to mimic epitopes identified as hotspots within the surface contact areas in protein‐protein interactions (PPIs). This review deals with the replacement of S‐S and thioether linkages of cyclic peptides by C‐C‐bridges and with the stabilization of peptide secondary architectures (α‐helix, β‐hairpins, β‐turns) through RCM, as a useful strategy for the modulation of therapeutically relevant signaling pathways. © 2010 Wiley Periodicals, Inc. Med Res Rev 31: 677‐715, 2011     

Haemodynamic and renal effects of urodilatin bolus injections in patients with congestive heart failure

Urodilatin (ANF(95-126)) is an analogue of the atrial natriuretic factor (ANF(99-126)), which has been isolated from human urine. Recently we have shown in healthy volunteers, that intravenous bolus injections of synthetic urodilatin produce more pronounced reductions of pulmonary arterial pressure than ANF(99-126). To compare haemodynamic and renal effects of synthetic urodilatin with those of ANF(99-126) in congestive heart failure (CHF), 12 patients (66.3 +/- 1.4 years) received either two high dose intravenous bolus injections of 4 micrograms kg-1 bw Urodilatin (URO) at a 30 min interval (n = 6) or the same doses of ANF(99-126) (n = 6). Prior to i.v. URO, no URO immunoreactivity was found in human plasma (specific RIA, no crossreactivity to ANF). Similar to ANF, the increase in diuresis (1.4 +/- 0.7 to 3.7 +/- 1.6 ml min-1) and natriuresis (169 +/- 114 to 430 +/- 197 mumol min-1) was moderate after URO in CHF. During the 90 min study period, mean plasma cyclic GMP levels increased much more after URO (by 53.4 +/- 15.1 nM) than after ANF (by 13.1 +/- 3.0 nM; P = 0.04). In contrast to ANF, i.v. bolus injections of URO produced sustained haemodynamic effects in CHF lasting up to 90 min: The average (0-90 min) reduction of systemic vascular resistance was more pronounced after URO (-578 +/- 148) than after ANF (-204 +/- 65 dyn*s*cm-5, P = 0.04).(ABSTRACT TRUNCATED AT 250 WORDS)