Toxic and carcinogenic constituents in Camel Snus and other U.S. smokeless tobacco products

Irina Stepanov, PhD Associate Professor School of Public Health University of Minnesota

FDA Tobacco Products Scientific Advisory Committee Meeting September 13-14, 2018, Silver Spring, MD Disclosure

Some of the research included in this presentation was supported by grants form NCI and through the NIH Tobacco Regulatory Science Program (R01-CA180880). The content is solely the responsibility of the author and does not represent the official views of the NIH or FDA. Outline

 New Product Watch Project  Recent data on moist snuff  Products marketed as snus New Product Watch

Web-based national monitoring network Monitors - State tobacco program staff and their community partners Six regions; three locations per region New Product Watch

2006 – 2007 NPW Round I NPW Round II NPW Round 2010 2011 III 2012 – 2013 Carcinogens in smokeless tobacco products

Group Constituents

Tobacco-specific N-nitrosamines NNN, NNK Volatile N-nitrosamines NDMA,NPYR, NPIP, NMOR, NDELA Nitrosamino acids NSAR Polycyclic aromatic hydrocarbons BaP, DBahA, BaA, BbF, BjF, BkF, DBaiP, IcdP, 5MC, NAP Metals and metalloids As, Be, Cd, Co, Cr VI, Pb, Ni, Po-210 Aldehydes Formaldehyde, acetaldehyde Inorganic salts Nitrate, nitrite Fermentation-related compounds Ethyl carbamate Mycotoxins Aflatoxin, ochratoxin Other plant material Areca nut

Stanfill and Stepanov, Ch. 3 in Smokeless Tobacco and Public Health: A Global Perspective (NIH Publication No. 14‐7983; 2014) Tobacco-specific N-nitrosamines (TSNA)

 Specific to tobacco exposure (formed from tobacco alkaloids)  Systemic, organ-specific carcinogens  Cancers of target organs are most strongly associated with tobacco use  Classified by IARC as carcinogenic to humans (Group 1)

Hecht, Chem. Res. Toxicol., 1998;11(6):559-603; IARC vol. 89, 2007 Yuan et al., Cancer Research, 2009;69:2990-2995; Yuan et al., Carcinogenesis, 2011;32:1366-1371 Product pH and unprotonated (free) nicotine Constituents and characteristics monitored in the New Product Watch

 Pouch size  Moisture content,  pH  Nicotine  Unprotonated nicotine  NNN  NNK Selected data: focus on time trends

Pouch weight Moisture content pH

Stepanov Laboratory (manuscript in preparation) Selected data: focus on time trends

Total nicotine, Free nicotine, Free nicotine, mg/dry wt mg/dry wt mg/portion

Stepanov Laboratory (manuscript in preparation) Selected data: focus on time trends

NNN, µg/dry wt NNK, µg/dry wt NNN+NNK, µg/portion

Stepanov Laboratory (manuscript in preparation) Comparisons with the U.S. moist snuff data Sum of NNN and NNK

2010

2011

Stepanov and Hatsukami, Tobacco Regulatory Science. 2016;2(1):9-30 Comparison of NNN data from recent analyses (per gram tobacco)

14

12

10

8

6 µg/gproduct

4 Marlboro Snus 2

0 Camel Snus Moist snuff Camel Snus ~1.6 ug/g dry weight NNN + ~0.5 ug/g dry weight NNK (NPW III)

Stepanov Laboratory (manuscript in preparation) Other key constituents in recent analyses: Total and free nicotine

Total nicotine Free nicotine

18 10

16 9 8 14 7 12 6 10 5 mg/g 8 mg/g 4 6 3

4 2

2 1

0 0 1 9 1 9 17 25 33 41 49 57 65 73 81 89 97 17 25 33 41 49 57 65 73 81 89 97 105 113 121 129 137 145 153 161 169 177 185 105 113 121 129 137 145 153 161 169 177 185 Camel Snus Camel Snus

Stepanov Laboratory (manuscript in preparation) − Other key constituents: NO2 N N In tobacco N O Nornicotine and nitrite H N N and endogenously NNN nornicotine in users

Nornicotine Nitrite

0.6 0.9

0.8 0.5 0.7

0.4 0.6

0.5 0.3 µg/g mg/g 0.4

0.2 0.3

0.2 0.1 0.1

0.0 0.0 1 1 10 19 28 37 46 55 64 73 82 91 10 19 28 37 46 55 64 73 82 91 100 109 118 127 136 145 154 163 172 181 100 109 118 127 136 145 154 163 172 181 Camel Snus Camel Snus

Stepanov Laboratory (manuscript in preparation) Minor alkaloids and β-carbolines: potential contribution to addictiveness

 Minor alkaloids: ◦ binding to nicotine receptors ◦ enhancement of nicotine effects

 β-Carbolines: ◦ MAO inhibitors ◦ Plasma levels increase 10-fold in 5 minutes after smoking a cigarette ◦ Measured in brain Anatabine and anabasine data from recent analyses (per gram tobacco)

Anatabine, mg/g Anabasine, mg/g

0.25 0.20

0.18

0.20 0.16

0.14

0.15 0.12

0.10

0.10 0.08

0.06

0.05 0.04

0.02

0.00 0.00 1 9 1 9 17 25 33 41 49 57 65 73 81 89 97 17 25 33 41 49 57 65 73 81 89 97 105 113 121 129 137 145 153 161 169 177 105 113 121 129 137 145 153 161 169 177 Camel Snus Camel Snus

Stepanov Laboratory (manuscript in preparation) Harman and norharman data from recent analyses (per gram tobacco)

Harman, ng/g Norharman, ng/g

900 30000

800 25000 700

600 20000 Camel Snus 500 15000 400

300 10000

200 Camel Snus 5000 100

0 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 1 2 3 4 5 6 7 8 9 1011121314151617181920

Marlboro Snus Marlboro Snus

Stepanov Laboratory (manuscript in preparation) Particular importance of NNN in smokeless tobacco products

N N O N NNN

Specific to tobacco

Potent oral and esophageal carcinogen in laboratory animals

Evidence for carcinogenicity in humans

J.-M. Yuan et al, Carcinogenesis 32(9): 1366-1371 (2011); I. Stepanov et al. Int J Cancer 134:2278-2283 (2014) Reported relative risks associated with smokeless tobacco use in various countries

6 c US 5 Sweden

4 d India b

3 a OR b b 2 b a a a b b b b 1

0

Oral cancer Larynx Esophageal cancer

Average NNN levels in moist snuff (our laboratory): ~1.7 µg/g product in recent analyses ~2.5 µg/g product 10 years ago

aBoffetta, et al. Lancet Oncology, 9: 667-675, 2008; bLee & Hamling, BMC Medicine, 7: 36, 2009; cIARC Vol 83, 2004; dPednekar et al. Cancer Causes Control, 22: 859-868, 2011 There is snus, Snus, and “snus” U.S. Snus

Constituent Content (2013)* Nitrite (µg/g) 1.1 (1.0 - 1.1) Product NNN NNK NNN + NNK (µg/g) 0.47 (0.46 - 0.48) Marlboro Snus 0.6 0.4 NDMA (ng/g) <0.6 Camel Snus 1.0 0.6 B(a)P (ng/g) <0.6 Skoal Snus 1.6 0.2 Cadmium (µg/g) 0.28 (0.28 - 0.29) Stepanov Laboratory (2015) *http://www.swedishmatch.com/en/Snus-and-health/GOTHIATEK/GOTHIATEK-standard/

Swedish snus

NNN: 25 ug/g NNK: 2.6 ug/g

Indian “snus”

Stepanov et al. Tobacco Control 2014

Stepanov Laboratory (manuscript in preparation) Summary

 Constituent profile of Camel Snus has been evolving since its first introduction to the market  Current levels of NNN in Camel Snus are comparable to the levels found in many popular moist snuff brands  The increase in NNN levels in Camel Snus occurred while NNN levels in some major moist snuff brands were declining  Available data on other constituents show comparable (minor alkaloids, nitrite, nitrate, harman, metals) or lower (norharman, PAH) levels between Camel Snus and moist snuff  Products marketed as snus vary substantially in their constituent profiles and other characteristics Acknowledgements

Collaborators University of Minnesota Other Dorothy Hatsukami Lois Biener, University of Massachusetts Stephen Hecht Amy Nyman, Georgia State University Peter Villalta Yen-Yi Ho, University of South Carolina Jian-Min Yuan, University of Pittsburgh Stepanov laboratory members Mark Parascandola, National Cancer Institute Katrina Yershova, Galina Yakovlev, Prakash Gupta, Healis-Sekhsaria Institute for Vipin Jain, Anshu Jain Public Health, India

Funding NCI contract HHSN261201200392P R01 CA180880 R01 CA129534 R01 CA144034 R01 CA-81301 CA-77598 (Mass-Spectrometry) Global Programs and Strategy (UMN) Minnesota Masonic Charities Thank you