Our laboratory is focused on the development and testing of cancer immunotherapy and non-immune-based cancer therapies. Since most cancer deaths occur by metastases (primary tumors can often be removed by surgery, chemotherapy, or radiation), our therapies are focused on the treatment of metastases. We have developed various therapies using different novel approaches, in preclinical mouse tumor models with metastatic breast and pancreatic cancer. For instance, we use an attenuated bacterium Listeria monocytogenes as a platform for the delivery of anticancer agents to the tumor microenvironment and into tumor cells such as radioactivity, tumor-associated antigens, or small molecules like alphagalactosylceramide, or we kill tumor cells through cryoablation by freezing and thawing tumor cells, combined with various adjuvants targeting myeloid-derived suppressor cells (MDSC) such as stimulator of interferon genes (STING)-ligand cyclic di-guanylate (c-di-GMP, Curcumin, and AMD3100. Since MDSC play a major role in immune suppression in the tumor microenvironment, MDSC are an important target in cancer immunotherapies.  We also focus on the age factor since most cancer patients are old and elderly react less efficient to vaccines than young adults. The MDSC are present in blood of patients and mice with cancer. This MDSC population is strongly increased in the tumor microenvironment particularly at older age, and contributes to the age-related T cell unresponsiveness.

Listeria-based cancer vaccines

Attenuated Listeria monocytogenes is a weakened facultative anaerobic bacterium (non-toxic and non-pathogenic) and has been used to deliver antigens into antigen-presenting cells. We developed various Listeria-based constructs expressing tumor-associated antigens including Mage-b, Survivin, p53 etc and tested these constructs in mice with metastatic breast and pancreatic cancer, and demonstrated a significant reduction in metastases and tumor growth. In addition, we have further improved the efficacy of the Listeria-Mage-b vaccine with help of MDSC-targeting adjuvants like c-di-GMP and Curcumin. However, in 2009 our lab discovered that the Listeria infects and kills tumor cells by the generation of reactive oxygen species (ROS) through the activation of the NADPH-oxidase pathway, and left healthy tissues unharmed (Fig 1). Based on this tropism for the tumor microenvironment we started using Listeria as a platform for the selective delivery of anti-cancer agents to the tumor microenvironment. For more detail see Kim et al. Cancer Res 2009; Chandra et al, Cancer Immunology Research, 2014.

Mechanisms that contribute to the selective survival and multiplication of Listeria in the tumor microenvironment

We have analyzed potential mechanisms explaining why Listeria survived and multiplied in the TME and not in healthy tissues. We discovered that Listeria is protected from immune clearance in the TME through strong immune suppression, but is rapidly killed in healthy tissues that lack immune suppression. In addition, we found that MDSC play an important role in the selective delivery and survival of Listeria in the tumor microenvironment. MDSC are selectively attracted by the primary tumor through the production of attractants such as granulocyte macrophage-colony stimulating factor (GM-CSF), interleukin (IL)-6, A100. Listeria infects, survives and multiplies in MDSC of tumor-bearing mice, and is protected from immune clearance because of the immune suppressive character of MDSC. We have shown that Listeria, once at the tumor site, infects (and kills) tumor cells directly or spreads from MDSC into tumor cells through the cell-to-cell spread mechanism specific for Listeria (Fig 2). For more detail see Quispe-Tintaya et al, PNAS 2013; Chandra et al, BJC, 2013.

Radioactive Listeria for the treatment of pancreatic cancer

Ninety six percent of patients diagnosed with pancreatic cancer have only 6 months to live, despite aggressive treatments. This underlines the urgent need for new effective therapies. In collaboration with Dr. Ekaterina Dadachova (Department of Radiation, Einstein), we developed a radioactive Listeria for the treatment of pancreatic cancer, by coupling ¹⁸⁸Rhenium to anti-Listeria antibodies followed by incubation with Listeria bacteria. This resulted in the synergistic destruction of cancer cells through Listeria-induced ROS and through ionizing radiation of the ¹⁸⁸Re. The number of metastases was reduced by 50% in mice treated with Listeria alone, and by 90% in mice treated with Listeria-¹⁸⁸Re. This correlated with the accumulation of radioactivity in the metastases. This was the first time that a live attenuated bacterium was successfully used to deliver radioactivity selectively to the tumor microenvironment (Fig 3).  The potential of the radioactive Listeria for the treatment of pancreatic cancer was discussed in several high profile journals like Science, Nature, as well as lay Journals like The Economist, Forbes Magazine and many others. Currently, we are testing Listeria with other radioisotopes. For more detail see Quispe-Tintaya et al, PNAS 2013.

Listeria incorporated with alphagalactosylceramide

In collaboration with Dr. Steven Porcelli (Department Microbiology and Immunology, Einstein), we incorporated alphagalactosylceramide (αGC) into the Listeria bacteria simply during culture. This method was originally developed for mycobacteria. αGC is a marine sponge that activates natural killer T cells (NKT) cells, which in turn stimulates other immune cells like natural killer (NK) cells and T cells. We demonstrated that Listeria expressing tumor-associated antigen Mage-b incorporated with αGC created an immune-stimulating environment that attracted the NKT cells to the metastases, resulting in improved activation of CD8 T cells to Mage-b and a dramatic reduction in the number of metastases in a mouse model of metastatic breast cancer (4T1). For a mchanistic overview see Fig 4. For more detail see Singh et al, BJC 2014.

Cryoablation combined with adjuvants

Cryoablation involves killing of tumor cells through freezing and thawing, resulting in recruitment of tumor-specific T cells.  Since MDSC strongly inhibits these T cells we have the cryoablation combined with MDSC-targeting adjuvants like STING ligand c-di-GMP, Curcumin, and AMD3100. c-di-GMP reduces the number of MDSC and converts a subpopulation of MDSC into an immune-stimulating phenotype producing IL-12 (stimulates T cells). Curcumin reduces IL-6 produced by MDSC and breast tumors. IL-6 strongly inhibits T cells in the tumor microenvironment. AMD3100 is a small molecule that prevents the interaction of CXCR4 on MDSC and stromal cell-derived factor (sdf-1) on tumor cells. Currently, these combination therapies are under investigation in mice with metastatic breast cancer in collaboration with the Anticancer Fund (Brussels, Belgium). Preliminary results are extrmely promising. Cryoablation and Meriva (Curcumin derivate) significantly delayed the onset of metastases and eliminated completely the primary tumor, prolonged the the survival rate compared to the control groups in correlation with improved CD8 T cell responses to multiple tumor-associated antigens. For more detail see Chandra et al, OnoImmunology 2015.

Feasibility of cancer vaccination at older age

Cancer is a disease of the elderly. When cancer becomes metastatic, it often needs aggressive second-line treatment, for which the options are limited. This is particularly challenging for frail, elderly cancer patients in which comorbidity plays an antagonistic role. Immunotherapy is the most promising and benign option for preventing or curing metastatic cancer in such patients. Unfortunately, cancer immunotherapy is less effective at old than at young age, due to T cell unresponsiveness, especially in the tumor microenvironment (TME). Various causes have been described for T cell unresponsiveness at old age, such as lack of naïve T cells at older age, deficiency in the upregulation of co-stimulatory molecules on aged dendritic cells (DCs), and most recently, the increase in the population of MDSC in the TME of old compared to young mice, among other age-related immune impairments. As mentioned above, Listeria has an intimate relationship with MDSC. We have shown that Listeria-based vaccination was equally effective in young and old mice with metastatic breast cancer by targeting MDSC. The Listeria killed the tumor cells directly through ROS, and Listeria-activated T cells killed the infected tumor cells presenting the Listeria antigens. For more detail see Chandra et al, BJC, 2013.

Lab Members Einstein 2009 and 2015

Dinesh Chandra, PhD (Faculty Associate)

Arthee Jahangir, PhD (Graduate student, received PhD in 2014)

Wilber Quispe-Tintaya, PhD (Post doc)

Manisha Singh (Graduate student, received PhD in 2013)

Ilyssa Ramos (Technician)

Denise Asufu-Adjei (Technician)

PHD or PhD/MD students

Jackie Coley (PhD student)

Marika Osterbur (MD/PhD student)

Karin Skalina (MD/PhD student)

Summer students

Alice Kwon

Swathi Krishnan

Lukman Solola

James Gibson

Marianna Vitiello

Rodrigo Alves Da Silva

Most recent peer-reviewed publications relevant to the field of cancer vaccination and cancer therapies selected out of 56

Kim SH, Castro F, Paterson Y, Gravekamp C. High efficacy of a Listeria-based vaccine against metastatic breast cancer reveals a dual mode of action. Cancer Res. 2009; 69(14): 5860-5866. PMID 19584282

Castro F, Leal B, Denny A, Bahar R, Lampkin S, Reddick R, Lu S, and Gravekamp C. Vaccination with Mage-b DNA induces CD8 T cell responses at young but not at old age in mice with metastatic breast cancer. British Journal of Cancer, 2009; 101:1329-1337. PMID 19826426

Quispe-Tintaya W*, Chandra D*, Jahangir A, Harris M, Casadevall A, Dadachova E, and Gravekamp C. A non-toxic radioactive Listeria^at is a highly effective therapy against metastatic pancreatic cancer. PNAS, 2013; 110(21):8668-73. PMID: 23610422.*Both authors contributed equally to the manuscript.

Singh M, RamosI, Asafu-AdjeiD, Quispe-TintayaW, ChandraD, JahangirA, ZangX, AggarwalBB, and Gravekamp. C. Curcumin Improves the Therapeutic Efficacy of Listeria^at-Mage-b Vaccine in mice with Triple Negative Breast Cancer (TNBC) in correlation with improved T cell responses in blood. Cancer Medicine, 2013; 2(4): 571-582 PMID: 24156030

Chandra D*, Quispe-Tintaya W, Jahangir A*, Singh M, and Gravekamp C. Myeloid-derived suppressor cells have a central role in attenuated Listeria monocytogenes-based immunotherapy against metastatic breast cancer in young and old mice. Brit J Cancer 2013, 108(11): 2281-90. PMID: 23640395. *Both authors contributed equally to the manuscript.

Chandra D, Quispe-TintayaW, Jahangir A, Asafu-AdjeiD, Ramos I, Sintim, HO, Zhou J, Hayakawa, Y, Karaolis D, and Gravekamp C. STING ligand c-di-GMP improves cancer vaccination against metastatic breast cancer. Cancer Immunology Research, 2014; 2(9): 901-910. (DOI: 10.1158/2326-6066.CIR-13-0123). PMID: 24913717

Singh M, Quispe-Tintaya, W, Chandra D, Jahangir A, Venkataswamy MM, Ng TW, Sharma S, Carreno LJ, Porcelli SA, and Gravekamp C. Direct incorporation of the NKT cell activator a-galactosylceramide into a recombinant Listeria monocytogenes improves breast cancer vaccine efficacy. British Journal of Cancer 2014; (doi: 10.1038/bjc.2014.486). PMID: 25314062

Valdor R, Mocholi E, Botbol Y, Guerrero-Ros I, Chandra D, Koga H, Gravekamp C, Cuervo AM, Macian F. Chaperone-mediated autophagy regulates T cell responses through targeted degradation of negative regulators of T cell activation. Nat Immunol. 2014 Nov;15(11):1046-54. doi: 10.1038/ni.3003. PMID: 25263126

Chandra D, Jahangir A, Cornelis F, Rombauts K, Meheus L, Jorcyk CL, and Gravekamp C. Cryoablation and Meriva has strong therapeutic effect on triple negative breast cancer. OncoImmunology. 2015. In Press. Doi: 10.1080/2162402X.2015.1049802.

Chandra D, Selvanesan CB, Yuan Z, Libutti SK, Koba W, Beck A, Casadevall A, Dadachova E, Gravekamp C. 32-Phosphorus selectively delivered by Listeria to pancreatic cancer demonstrates strong therapeutic effect. Oncotarget. 2017 Feb 6. doi: 10.18632/oncotarget.15117. PMID: 28186976

Revskaya E, Jiang Z, Morgenstein A, Bruchertseifer F, Sesay M, Walker S, Fuller S, Lebowitz MS, Gravekamp C, Ghanbari HA, and Dadachova E. A radiolabeled fully human aspartyl (asparaginyl) beta-hydroxylase is a promising agent for imaging and therapy of metastatic breast cancer. Cancer Biother Radiophar. 32 (2): 57-65, 2017. PMID: 27907230

Jahangir A, Chandra D, Quispe-Tintaya W, Singh M, and Gravekamp C. Immunotherapy with Listeria reduces metastatic breast cancer in young and old mice through different mechanisms. OncoImmunology, 2017. http://www.tandfonline.com/eprint/dnkC4emkjuSeYUeJtZGz/full. PMID: 28932647

Most recent invited publications relevant to the field of cancer vaccination at older age selected out of 17

Gravekamp C. 2009. Cancer Immunotherapy and aging: Lessons from the mouse. In: Handbook on Immunosenescence: basic understanding and clinical applications, Eds Fulop T, Pawelec G, Franceschi C, and Hirokawa K. Springer. Pp1217-1243.

Gravekamp C, Kim SH, and Castro F. Cancer vaccination: manipulation of immune responses at old age. Mechanism of Ageing and Development 2009:130(1-2): 67-75. PMID 18561984

Gravekamp C. The importance of the age factor in cancer vaccination at older age. Cancer Immunology and Immunotherapy 2009; 58(12): 1969-1977. PMID 19259666.

Pawelec G, Lustgarten J, Ruby C, and Gravekamp C. Impact of aging on cancer immunity and immunotherapy. Cancer Immunol Immunother 2009: 58: 1907-1908. PMID 19787858

Gravekamp C, and Paterson Y. Harnessing Listeria monocytogenes to target tumors. Cancer Biol and Ther 9: 1-9, 2010. PMID 20139702.

Gravekamp C. The impact of aging on cancer vaccination. COII 2011; 23: 555-560. PMID:21763118.

Gravekamp C. Cancer vaccination in older age. Interdiscip Top Gerontol. 2013;38:28-37. PMID: 23503513.

Chandra D and Gravekamp C. Myeloid-derived suppressor cells: cellular missiles to target tumors. Oncoimmunology 2013; 1;2(11):e26967. PMID: 24427545

Gravekamp C and Jahangir A. Is cancer vaccination feasible at older age? Exp Gerontol 2014 Jun;54C:138-144. PMID: 24509231.

Gravekamp C and Chandra C. Targeting STING pathways for the treatment of cancer. OncoImmunology, 2015. In press. doi: 10.4161/2162402X.2014.988463.

Kohanski RA, Deeks SG, Gravekamp C, Halter JB, High K, Hurria A, Fuldner R, Green P, Huebner R, Macchiarini F, Sierra F. Reverse geroscience: how does exposure to early diseases accelerate age-related decline in health? Ann N Y Acad Sci. 2016 Dec;1386(1):30-44. doi: 10.1111/nyas.13297. Epub 2016 Dec 1.

PMID: 27907230

RESEARCH IMPACT

Commentary on paper about Radioactive Listeria by Quipe-Tintaya et al. in PNAS 2013. Stritzker J, and Szalay AA. Single-agent combinatorial cancer therapy. www.pnas.org/cgi/doi/10.1073/pnas.1305832110.

Websites of Journals and magazines that discussed the impact of the radioactive Listeria for therapeutic treatment of pancreatic cancer:

http://www.nature.com/nrgastro/journal/vaop/ncurrent/full/nrgastro.2013.81.html

http://news.sciencemag.org/sciencenow/2013/04/radioactive-microbes-nuke-tumor-.html

http://txchnologist.com/post/48768770496/radioactive-metal-hitches-ride-on-bacteria-to-fight

http://www.economist.com/news/science-and-technology/21576628-surprising-new-way-kill-tumours-three-wrongs-make-right

https://www.sciencenews.org/article/microbes-can-redeem-themselves-fight-disease