On the Intersection of Metabolism and Infectious Diseases

by Kevin C. Ma (Harvard College, 2017)

During most of the New England Symposium, our attention was focused on how metabolism and its pathways converge with and impact tuberculosis, on the scale of molecular genetics to epidemiology. These talks highlighted important new areas of potential interdisciplinary research efforts and suggested metabolism-related therapeutics for continuing the fight against infectious diseases. But it was in the last presentation with Denise Faustman, Associate Professor at MGH, that we saw that the flow of ideas can go both ways: the results of efforts in understanding and treating tuberculosis can provide new therapeutic avenues for the treatment of autoimmune and metabolic disorders, like Type 1 Diabetes.

In 2001, Professor Faustman and colleagues reported in the Journal of Clinical Investigation that the treatment of non-obese diabetic mice with Complete Freund’s Adjuvant (CFA) coupled with islet transplantation or splenocyte injection appeared to cure Type 1 Diabetes (1). CFA is an adjuvant which contains oils and killed Mycobacterium tuberculosis, and has been shown to induce immune response and production of TNFα, an inflammatory signaling protein (2). This work extended earlier research done which implicated TNFα as a potent tool to counter the autoimmune attack (3).

The result of these and numerous other studies implicated TNFα as a protein that is able to induce cellular death specifically in autoimmune T cells (7). Because TNFα cannot be safely administered into humans at high doses, Faustman employed an alternative approach: administration of the BCG vaccine. The BCG vaccine, originally used to combat tuberculosis, had been known to induce TNF for several decades (5). In 2012, Faustman published the results of an initial, proof of concept clinical trial using the BCG vaccine to treat late stage diabetic patients. Her results showed markedly improved insulin secretion in BCG patients; however, these results were transient and not permanent reversals (6).

Further work will be needed to better understand whether or not BCG can be an effective and long-lasting treatment. Professor Faustman is currently raising funds for a Phase II clinical trial, and offers extensive background information as well as a page for frequently asked questions on her lab page (8). We note that her findings are not without controversy: follow up research done by the Benoist-Mathis lab at Harvard questioned some initial findings, including whether or not the infusion of spleen cells was necessary for reversing Type 1 Diabetes, and criticized Faustman’s work for not including a control (9). This follow up study, Faustman’s reply (10) and other technical comments will give the interested reader a more balanced view of this approach. Finally, it is important to note that TNFα is not, by any means, a panacea for all autoimmune related diseases. In fact, as one audience member noted following Faustman’s presentation, elevated levels of TNFα are actually implicated in the pathogenesis of systemic lupus erythematosus (11).

While further research will be needed to determine if BCG is truly an effective therapeutic agent, it is clear that researchers working to tackle metabolic, autoimmune, and infectious diseases have much to benefit in collaborations with one another.

References:

1. Ryu et al., Reversal of established autoimmune diabetes by restoration of endogenous β cell function. J Clin Invest. 2001;108(1):63–72.

2. Rabinovitch, A, Suarez-Pinzon, WL, Lapchak, PH, Meager, A, Power, RF. Tumor necrosis factor mediates the protective effect of Freund’s adjuvant against autoimmune diabetes in BB rats. J Autoimmun 1995. 8:357-366.

3. Ulaeto, D, Lacy, PE, Kipnis, DM, Kanagawa, O, Unanue, ER. A T-cell dormant state in the autoimmune process of nonobese diabetic mice treated with complete Freund’s adjuvant. Proc Natl Acad Sci USA 1992. 89:3927-3931.

4. S. Kodama, W. Kühtreiber, S. Fujimura, E. A. Dale, D. Faustman, Science 302, 1223 (2003).

5. Rahman MM, McFadden G (2006) Modulation of tumor necrosis factor by

microbial pathogens. PLoS Pathog 2: e4.

6. Faustman, D. L. et al. Proof-of-concept, randomized, controlled clinical trial of Bacillus–Calmette–Guerin for treatment of long-term type 1 diabetes. PLoS ONE 7, e41756 (2012).

7. Ban L, Zhang J, Wang L, Kuhtreiber W, Burger D, et al. (2008) Selective death of autoreactive T cells in human diabetes by TNF or TNF receptor 2 agonism. Proc Natl Acad Sci U S A 105: 13644–13649.

8. http://www.faustmanlab.org/

9.  J. Nishio et al., Science 311, 1775 (2006).

10. Faustman DL. Permanent reversal of diabetes in NOD mice. Science 2007; 317:196.

11. D. Kollias et al. Current Directions in Autoimmunity, vol. 5, pp. 30–50, 2002.

The Dance Between Bacterial Metabolism and Host Metabolism

by Skye Fishbein (Harvard, G1, GSAS/HSPH)

“There is an immunological carnival in adipose tissue,” remarked Gokhan Hotamisligil, in his Centennial speech at the symposium. Not a tuberculosis researcher by trade, he expanded on his research endeavors in the field of immunometabolism, and peaked my interested in how the host metabolism may influence the immune response. Lipid chaperones control immunometabolic disease such as asthma and atherosclerosis. He also reported that tuberculosis infection induces expression of an adipocyte lipid chaperone. It has been proposed elsewhere that adipose tissue could be a reservoir for M. tuberculosis[1, 2]. Additionally, lipid chaperones affect the lipid content within a macrophage. In the context of the mycobacterium, macrophage metabolism is a key determinant of disease outcome. As a microbiologist, I wondered about the bacterial metabolism during infection.

If the macrophage cannot clear M. tuberculosis, granuloma formation occurs, and often times, the mycobacterium will shift into a quiescent growth phase of its life cycle in the intracellular niche of the macrophage. While this conference focused on the conjunction of host metabolic syndrome and tuberculosis, it also highlighted many questions and research directions concerning bacterial metabolism, and the host influence on this. Some presented posters suggested that M. tuberculosis may take up exogenous nutrients for biosynthetic purposes, from hexose to cholesterol, that aid in M. tuberculosis survival. M. tuberculosis also uses certain lipoproteins to sequester triacylglycerol, as mentioned by one poster presenter. Multiple posters highlighted the lipid metabolism in M. tuberculosis, with implications extending to both virulence and in vivo survival. One poster even hinted at the possibility of a class of enzymes that M. tuberculosis employs, called serine hydrolases, that may degrade lipids for usable nutrient sources[3]. These aspects of bacterial metabolism reveal vulnerable points in the intracellular lifestyle that we as a research community should use to our advantage in engineering strategies to target both the active and quiescent pathogen.

The symposium as a whole reminded me of the equal host and pathogen contribution to tuberculosis infection. What is the major source of nutrition for M. tuberculosis over the course of the infection, and does it change? What role does the host play in this source of nutrition? These are all questions, whose answers could lead the field towards multiple disease intervention strategies. Either by altering the human immune response to disease, or by cutting off the bacilli’s main source of nutrition, there are likely a wealth of drug targets and metabolic modulators that could be developed. We are only at the surface of understanding the synergy between the host metabolism and the bacterial metabolism in M. tuberculosis pathogenesis.

 

1.         Erol, A., Visceral adipose tissue specific persistence of Mycobacterium tuberculosis may be reason for the metabolic syndrome. Med Hypotheses, 2008. 71(2): p. 222-8.

2.         Neyrolles, O., et al., Is adipose tissue a place for Mycobacterium tuberculosis persistence? PLoS One, 2006. 1: p. e43.

3.         New England Tuberculosis Symposium Abstract Book, 2013

Note: All conclusions from poster presentations made here were discovered either through poster viewing or questioning poster presenters.

 

Tackling Infectious Diseases with Synthetic Biology

by Kevin C. Ma (Harvard College 2017)

Synthetic biology focuses on the application of engineering principles to living systems in order to encode new behaviors and functionalities by integrating new biological parts, or by manipulating endogenous ones. For the past decade, extensive work was done to characterize defined genetic networks, such as switches and oscillators1,2. Despite the challenges of the field, synthetic biologists were able to develop new tools to assist with the construction of their artificial circuits, on the level of both gene and genome3-7. The practical benefits of synthetic biology are immense, ranging from biofuels to antibiotic production to diagnostics. The role of synthetic biology in therapeutics is still unresolved, but the great potential of this engineering approach has already been demonstrated in innovative projects such as bacteria engineered to specifically invade cancer cells8.

In particular, synthetic biology may prove to be a valuable tool in today’s crucial fight against infectious diseases, such as cholera. Cholera is caused by the bacterium V. cholerae and its secretion of the cholera toxin, a protein complex which causes epithelial cells to secrete solutes into the lumen of the intestine. Water flows out as a result, giving rise to the severe diarrhea characteristic of the disease. Cholera results in over a hundred thousand deaths each year, and remains a pressing challenge for developing countries. Synthetic biology may offer a solution – in one study, researchers at Cornell engineered probiotic E. coli to inhibit the development of virulence in cholera by expressing a signaling molecule, CAI-19. CAI-1 is secreted by V. cholerae when it reaches high densities, and it signals the organism to end the expression of virulence factors such as the cholera toxin and a pilus that allows the bacterium to attach to the intestinal wall. A probiotic strain of E. coli was therefore engineered to permanently express an enzyme responsible for synthesizing CAI-1; the goal was to “hijack” natural pathways of virulence reduction in order to prevent V. cholerae from exerting its harmful effects. When these engineered probiotics were administered to infant mice, the survival rate of V. cholerae was reduced by 70%, likely due to the inability of V. cholerae to effectively colonize the intestine.

Synthetic biology can also assist global health efforts indirectly by assisting with drug discovery. Ethionamide is an antibiotic used to combat tuberculosis infection by acting as an NAD derivative; it is converted to an active state by an endogenous M. tuberculosis enzyme, EthA.12 However, resistance has arisen in the form of EthR, a protein which represses the translation of EthA. To combat this, researchers in Switzerland and France created a mammalian genetic circuit in order to screen for compounds that could inhibit EthA in addition to assessing the compound’s cytotoxicity and permeability.11 This circuit is designed to screen for compounds which interfere with EthR binding to its operator site, and is similar to conventional two hybrid screens. Researchers fused EthR to a transactivating domain controlling expression of human placental secreted alkaline phosphatase (SEAP); SEAP expression can then be assayed through various means. When screened against a library, these engineered mammalian cells offer an innovative way to screen for drug candidates. The authors found several compounds which reduced SEAP expression (presumably through EthR binding interference), including 2-phenylethyl-butyrate, a food additive. These results suggest molecules applicable to combating resistant TB.

These are but a few of the numerous examples of innovative ways in which synthetic biology is being applied to tackle challenges in global health10. However, as with other studies, there remain important caveats: in the cholera study, the continuous administration of the probiotic bacterium, for example, may present a logistical challenge. Furthermore, the authors were unable to assay the duration of protection – a short duration necessitating frequent re-administrations of the engineered strain would also pose financial problems. In the tuberculosis study, it is important to consider whether this assay can be applied in a high throughput manner, and whether suggested compounds may detrimentally affect long term gene expression in mammalian cells. Nevertheless, the power and potential of synthetic biology is clear. With new, innovative tools and approaches being developed each day, synthetic biologists are poised to tackle the big challenges facing our world.

References:

1. Gardner, T. S., Cantor, C. R. & Collins, J. J. Construction of a genetic toggle switch in Escherichia coli. Nature 403, 339–342 (2000).

2. Elowitz, M. B. & Leibler, S. A synthetic oscillatory network of transcriptional regulators. Nature 403, 335–338 (2000).

3.Carr, P. A. & Church, G. M. Genome engineering. Nature Biotech. 27, 1151–1162 (2009).

4. Wang, H. H. et al. Programming cells by multiplex genome engineering and accelerated evolution. Nature 460, 894–898 (2009).

5. Ellis, T., Adie, T. & Baldwin, G. S. DNA assembly for synthetic biology: from parts to pathways and beyond. Integr. Biol. 3, 109–118 (2011).

6. Esvelt KM, Carlson JC, Liu DR. A system for the continuous directed evolution of biomolecules. Nature 472, 499–503 (2011).

7. D. G. Gibson, L. Young, R. Y. Chuang, J. C. Venter, C. A. Hutchison and H. O. Smith, Enzymatic assembly of DNA molecules up to several hundred kilobases, Nat. Methods, 6, 343–5 (2009).

8. J. C. Anderson, E. J. Clarke, A. P. Arkin, C. A. Voigt, Environmentally controlled invasion of cancer cells by engineered bacteria. J. Mol. Biol. 355, 619 (2006).

9. F. Duan and J. C. March. Engineered bacterial communication prevents Vibrio cholerae virulence in an infant mouse model. Proc. Natl. Acad. Sci. U.S.A. 107, 11260 (2010).

10. Ruder WC, Lu T and Collins JJ. Synthetic biology moving into the clinic. Science 333: 1248-1252 (2011).

11. Weber W, et al. A synthetic mammalian gene circuit reveals antituberculosis compounds. Proc Natl Acad Sci USA;105:9994–8 (2008).

12. Wang F, et al. Mechanism of thioamide drug action against tuberculosis and

leprosy. J Exp Med 204:73–78 (2007).

Tuberculosis: What the Past Can Teach Us

by Natasha Rodgers (Harvard College 2015)

Tuberculosis has one of the worst cumulative mortality in the history of infectious disease.  It is estimated that in the past two centuries a billion people have died from tuberculosis1.  Recently, the understanding and management of tuberculosis changed dramatically due to a number of major achievements.  How these advances shaped the current approach in tackling tuberculosis, demonstrates the tension between a singular global approach and tailored control measures.

The advent of antibiotics provided the foundation for scientifically-driven treatments of tuberculosis.  Until the discovery of streptomycin in 1944, treatment options were limited.  Throughout the 1950s and 60s, a number of other antibiotics, including isoniazid and rifampicin, were identified.  As our understanding of the bacterium expanded and better treatment options became available, management was revolutionized.  In the 1980‘s the DOTS program, Direct Observation Treatment Short-course, was introduced.  While DOTS entails standards for TB diagnosis, monitoring and drug supply, a critical pillar of the DOTS strategy is a globally standardized supervised treatment regimen for TB.  It works. When a particular program reaches WHO targets of 70% detection and 85% cure rate, then the mortality is projected to decrease 12% It is an efficient template program that has lasting effects globally. However worldwide programs ensuring drug availability without consideration of a locality’s diagnostic capabilities or resistance management programs, allowed drug resistance to emerge. MDR-TB and now XDR-TB have arisen repeatedly and in some places become functionally untreatable.

DOTS is an incredibly powerful tool that changed the way we manage tuberculosis.  However the subsequent decades have also revealed the issues of one solution for everyone based disease control. Lessons from drug resistance speak to the importance of local approaches.

While once unthinkable, recently dramatic progress has recently been made in the TB field that makes more local approaches feasible.  First, new antibiotics are becoming available for TB. In 2012, the FDA approved a new antibiotic, bedaquiline, which has a novel mechanism of action, attacking the ATP synthase proton pump.  This is the first truly innovative therapy to be developed in over forty years.  There is promising research indicating other classes of chemicals as future therapies; bedaquiline will hopefully be the start of a significant phase of novel drug roll out.  In addition, the genomic revolution has led to a game changing new diagnostic for TB. Previous diagnostics for MDR-TB using traditional methods took months, allowing patients to return to their communities untreated.  Rapid diagnosis of TB and of drug resistance promises the ability to quickly get patients on therapy tailored to match their TB strain.

Today, there are high burdens of tuberculosis in India, China and South Africa, all of which have disparate risk factors and may require differential management.  For tuberculosis in the 21st century; understanding the changing risks across different countries and altering programs to suit could have a massive impact on the global burden of tuberculosis.

Bibliography

1. Ryan, F. Tuberculosis: The Greatest Story Never Told; Swifth Publishers: Bromsgrove, Worcestershire, U.K., 1992.

2. Chan. E D, Iseman M D.Current medical treatment for tuberculosis. BMJ. 2002 November 30;325(7375). 1282–1286.

3. Dye C, Garnett GP, Sleeman K, Williams BG. Prospects for worldwide tuberculosis control under the WHO DOTS strategy. Directly observed short-course therapy. Lancet. 1998 Dec 12;352(9144):1886-91.

4. Keshavjee, S.Farmer P.  Tuberculosis, Drug Resistance, and the History of Modern Medicine. New England Journal Med 2012; 367:931-93

Alex Sakula. Robert Koch: Centenary of the Discovery of the Tubercle Bacillus, 1882. Can Vet J. 1983 April; 24(4): 127–131.

Reichman LB, Tanne JH. Timebomb : The global epidemic of multi-drug-resistant tuberculosis. New York: McGraw-Hill; 2002

Colditz GA, Brewer TF, Berkey CS, Wilson ME, Burdick E, Fineberg HV, et al. Efficacy of BCG vaccine in the prevention of tuberculosis. Meta-analysis of the published literature. JAMA. 1994 Mar 2;271(9):698-702.

Bates JH, Stead WW. The history of tuberculosis as a global epidemic. Med Clin North Am. 1993 Nov;77(6):1205-17.

http://www.who.int/vaccine_research/diseases/tb/vaccine_development/bcg/en/.  Website Accessed 22.9.2013

http://pubs.acs.org/subscribe/archive/mdd/v05/i02/html/02timeline.html.  Website Accessed 22.9.2013

http://www.news-medical.net/health/History-of-Tuberculosis.aspx.  Website Accessed 22.9.2013

http://www.who.int/tb/features_archive/xpertprojectlaunch/en/index.html Website Accessed 26.9.13

The Delicate Balance Between Metabolic and Communicable Diseases

by Alexandra Rojek (Harvard College 2015)

When we think of infectious, or communicable diseases, images of deadly plagues and widespread contagion may come to mind. On the other hand, when we think of metabolic disease the symptoms that seem most relevant are ones that contribute to increased risk of heart disease, stroke, and diabetes. Superficially, it may not seem as if there is much to connect these two classes of disease, yet as we learn more about their deeper molecular mechanisms, we see a convergence on shared principles emerging.

Metabolic disease is linked to the rising obesity epidemic, and the list of the conditions that it influences is broad: hypertension, coronary heart disease, osteoarthritis, and especially type 2 diabetes. An individual’s body mass index (BMI) and their fat tissue distribution are also key contributors to risk, where a higher BMI and abdominal obesity constitute the highest risk category. It is not intuitively obvious, however, why obesity would lead to such conditions1. In recent years, the search for a connection between obesity and the resulting metabolic diseases has resulted in the concept of ‘metainflammation’, or the constant low-level immune response present in obese individuals2.

The immune system has evolved a complex response to deal with infection that is largely dependent on the inflammatory response. TNFα, one of the many signaling molecules essential to this part of innate immunity, is known to regulate other immune cells and propagate the inflammatory response. TNFα is one of the immune system’s signaling molecules, or cytokines, significantly upregulated in the tissues of obese individuals1. Signaling molecules, including TNFα, are also induced upon infection by foreign bugs such as Mycobacterium tuberculosis, and are crucial to the control of infection of macrophages3.

Cytokines like TNFα play an essential role in controlling infection and without them our immune system would not be able to fully develop and respond to such infections. However, when unregulated, cytokines can lead to tissue damage and necrosis. This delicate balance of the immune system response is crucial to our health, and in individuals with increased body fat, their immune system is consistently stimulated, factors such as TNFα are produced consistently, metabolic processes are disrupted, and what we define as metabolic diseases arise2.

At a time when the developing world is increasingly experiencing a rise in metabolic diseases due to nutritional and lifestyle changes leading to obesity, it is crucial to further probe this connection between metabolic disease and the immune system. When the critical balance maintained between these two systems is disturbed, it creates the potential for metabolic diseases to emerge. Since inflammation represents a critical convergence between communicable and metabolic disease, we can use the existing vast reserve of knowledge about the inflammatory response to explore solutions to metainflammation linked to the current worldwide obesity epidemic. By examining the connections between metabolic and communicable diseases through the shared principle of inflammation, we can better understand what the natural balance between these two systems is, how that balance is disturbed in the diseased state, and what solutions we can envision to restore that natural balance.

  1. R. J. Genco, S. G. Grossi, A. Ho, F. Nishimura, Y. Murayama, A proposed model linking inflammation to obesity, diabetes, and periodontal infections. J Periodontol 76, 2075 (Nov, 2005).
  2. C. N. Lumeng, A. R. Saltiel, Inflammatory links between obesity and metabolic disease. J Clin Invest 121, 2111 (Jun, 2011).
  3. S. J. Sasindran, J. B. Torrelles, Mycobacterium Tuberculosis Infection and Inflammation: what is Beneficial for the Host and for the Bacterium? Front Microbiol 2, 2 (2011).

Welcome New England TB Community

New England boasts the highest density of research focused on tuberculosis biology and disease in the country.  Groups in the area investigate the molecular and genetic epidemiology of tuberculosis, mycobacterial genomics, epigenetics, and metabolism, high-throughput chemical biology, lipid biochemistry, the immunological response to M. tuberculosis, TB diagnostics, vaccinology, implementation research, and more.

The purpose of the New England TB Symposium is to bring together people interested in tuberculosis and mycobacterial research, foster collaboration and showcase areas of active investigation.  We look to incorporate a range of viewpoints and perspectives from experts to students to the lay public on the state of the TB field and the possibilities for its future.  Each year the Symposium highlights a special topic related to tuberculosis research.  This year, motivated by the changing epidemiology of infectious diseases, our theme is TB in the 21st Century: The Convergence of Infectious and Metabolic Diseases.

To further the discussion, we have asked many of the talented undergraduate students in the New England community to write blog pieces based on their perceptions of infectious  and metabolic diseases as we head into a new era of both domestic and global public health.

If you have an interest in writing a blog piece for the New England TB community, please contact Alicair Peltonen (apeltone@hsph.harvard.edu).